Lactobacillus johnsonii (L. johnsonii) in a rat model of PNALD. Total parenteral nutrition (TPN)-fed rats were used to assess the role of L. johnsonii in liver steatosis, bile acid metabolism, gut microbiota, and hepatocyte apoptosis. We observed a depletion of L. johnsonii that was negatively correlated with the accumulation of glycochenodeoxycholic acid (GCDCA), a known apoptosis inducer, in rats subjected to TPN. L. johnsonii attenuated TPN-induced liver steatosis by inhibiting fatty acid synthesis and promoting fatty acid oxidation. TPN resulted in a decrease in bile acid synthesis and biliary bile secretion, which were partially restored by L. johnsonii treatment. The gut microbial profile revealed depletion of pathogenic bacteria in L. johnsonii-treated rats. L. johnsonii treatment reduced both hepatic GCDCA levels and hepatocyte apoptosis compared with the TPN group. In vitro, L. johnsonii treatment inhibited GCDCA-induced hepatocyte apoptosis via its bile salt hydrolase (BSH) activity. Our findings suggest that L. johnsonii protects against liver steatosis, bile acid dysregulation, and hepatocyte apoptosis in TPN-fed rats.Parenteral nutrition (PN), a vital therapy for patients with intestinal failure, can lead to the development of parenteral nutrition-associated liver disease (PNALD). In this study, we aimed to investigate the role of  Parenteral nutrition-associated liver disease (PNALD) encompasses a spectrum of liver diseases, including cholestasis, steatosis, fibrosis, and eventually cirrhosis. These conditions affect individuals with intestinal failure who depend on extended periods of parenteral nutrition (PN) . The occLactobacillus and Bifidobacterium are commonly used in the treatment of gastrointestinal diseases such as inflammatory bowel disease, necrotizing enterocolitis, and short bowel syndrome (SBS) with controversial results [Lactobacillus johnsonii (L. johnsonii) is a potentially beneficial bacterium from the genus Lactobacillus that may regulate metabolically related diseases [L. johnsonii supplementation in mice with nonalcoholic fatty liver disease (NAFLD) resulted in improved insulin resistance, reduced fatty acid synthesis, and attenuation of mitochondrial abnormalities. Importantly, a negative correlation was reported between the duration of PN and the proportion of Lactobacillus in patients with SBS [Lactobacillus depletion. In this study, we aimed to determine whether L. johnsonii can attenuate PNALD its the underlying mechanisms.Previous studies have suggested that total parenteral nutrition (TPN) leads to a significant loss of microbial diversity and alterations in the gut microbial profile in animals and infants ,5. Probi results . Lactobadiseases ,8. Xin ediseases  showed twith SBS , suggestL. johnsonii was reduced in TPN-fed rats, a well-recognized animal model of PNALD [L. johnsonii attenuated TPN-induced liver steatosis, bile acid dysregulation, and hepatocyte apoptosis. Mechanistically, L. johnsonii inhibited TPN-induced hepatocyte apoptosis by deconjugating glycochenodeoxycholic acid (GCDCA) [L. johnsonii in regulating bile acid metabolism can be attributed to the increased ileal expression of Nr1h4  and its target gene, fibroblast growth factor 19 (Fgf19). Our findings provide insights into L. johnsonii as a novel therapeutic strategy in PNALD.In this study, we revealed that the abundance of of PNALD . Supplem (GCDCA) , a knownL. johnsonii was donated by Shanghai Jiaoda Onlly Co., Ltd. . Rats in the TPN group were orally gavaged with either L. johnsonii containing 5 × 107 CFU/day (TPN+ L. johnsonii group) or PBS (TPN group). All animal studies were approved by the Experimental Animal Care and Use Committee of Shanghai Jiao Tong University (No. XHEC-F-2019-003).Male Sprague–Dawley rats at the aged 4 weeks were purchased from Shanghai Laboratory Animal Center  and infused with 0.9% saline with a standard chow diet (control group) or TPN for 7 days as previously described ,13. Befo® III 1st strand cDNA synthesis supermix , and real-time qPCR was conducted with SYBR Premix  as previously described [L. johnsonii was normalized to 18S and 16S, respectively.Bacterial genomic DNA was extracted from rat stool samples and colon mucosa as previously described . TRIzol escribed . Primer Protein levels of cleaved caspase-3 in rat livers were determined by Western blot analysis. Proteins extracted from livers were subjected to SDS-PAGE gels and electrotransferred to polyvinylidene fluoride (PVDF) membranes. Nonspecific binding sites were blocked in 5% bovine serum albumin at room temperature for 1 h. Membranes were subsequently probed with primary antibodies at 4 °C overnight, then incubated with the appropriate peroxidase-conjugated secondary antibody. Cleaved caspase-3 monoclonal antibody  and Cyp7a1 antibody  were used as primary antibodies, and horseradish-peroxidase-conjugated anti-rabbit IgG  was used as a secondary antibody. Densitometry of immunoblot analysis was performed using ImageJ software as previously described .Liver and serum bile acids were measured by a liquid chromatography tandem mass spectrometry (LC-MS) system . Briefly, blood samples were obtained and centrifuged to collect serum for analysis. For liver tissues, approximately 50 mg of lyophilized, homogenized tissue was weighed and grounded with methanol to precipitate protein. After vortexing and centrifuging at 4 °C, the supernatant was mixed with water and filtered before being analyzed . A totalhttp://www.arb-silva.de, accessed on 16 December 2019) [L. johnsonii were deposited in the NCBI Sequence Read Archive (SRA) under Bioproject PRJNA1004452.DNA from rat feces was extracted using a TIANamp stool DNA kit  following the manufacturer’s instructions. The V3-V4 regions of the 16S ribosomal DNA gene were amplified and purified for sequencing using an Illumina MiSeq instrument . Raw sequencing data was decomposed and filtered for quality control. Operational taxonomic units (OTUs) were clustered at 97% similarity, and taxonomic classification was performed using UCLUST software  with theer 2019) . The funL. johnsonii, detected by agarose gel electrophoresis, and quantified by a Qubit 2.0 fluorometer. Libraries for nanopore sequencing were constructed with an insert size of 10 kb. DNA was fragmented to a size of 350 bp to construct libraries for the Illumina NovaSeq platform. The whole genome of L. johnsonii was sequenced on the Nanopore PromethION platform and Illumina NovaSeq PE150 at Beijing Novogene Bioinformatics Technology Co., Ltd . PE150 data and nanopore data were assembled and subjected to genome component prediction. For bacteria, the GeneMarkS program was used to retrieve the related coding gene. Raw sequence reads for L. johnsonii were deposited in the NCBI (SRA) associated with Bioproject PRJNA990471.Bacterial genomic DNA was extracted from Formalin-fixed liver tissue was embedded in paraffin and cut to a thickness of 5 μm for H&E staining. To assess lipid accumulation, liver sections were embedded into OCT compound, sliced to a thickness of 8 μm, and stained with Oil red O  as previously described ,15. CellAspartate aminotransferase (AST), alanine aminotransferase (ALT), total bile acids, and direct bilirubin in serum were measured using commercially available detection kits . Hepatic triglyceride levels were measured using a triglyceride assay kit  as previously described . HepaticL. johnsonii proteins were prepared using the sonication method. Incubation was carried out in 0.1 M sodium phosphate containing 0.1 mg/mL L. johnsonii protein, 10 mM GCDCA, and 100 μM caffeic acid phenethyl ester  as previously described [w/v) trichloroacetic acid (1:1 ratio). The supernatants were collected after centrifuging for GCDCA quantification by LC-MS and in vitro experiments.escribed . The mixHepG2 cells were cultured with Dulbecco’s Modified Eagle Medium, which was supplemented with 10% fetal bovine serum and 1% penicillin/streptomycin. Then, 1% bacterial supernatant was added to cell culture medium and incubated for 6 h. The final concentration for GCDCA in the cell culture medium was 100 μM. Caspase 3/7 activity was measured using an Apo-ONE homogeneous caspase-3/7 assay kit .t test or Mann–Whitney–Wilcoxon rank-sum test for highly skewed distributions. The significance of multiple groups was evaluated using one-way ANOVA with Tukey’s post hoc test. Spearman’s correlation was performed for correlation analysis. Statistical analysis was performed with GraphPad Prism (V.8.4.2). A value of p < 0.05 was considered statistically significant.All data are expressed as mean ± SEM unless otherwise stated. Comparisons between two groups were performed using Student’s L. johnsonii in TPN-fed rats , a gene associated with fatty acid synthesis, showed significant upregulation in response to TPN, which was effectively downregulated upon L. johnsonii treatment  were downregulated by TPN, whereas their expression was upregulated after L. johnsonii treatment  were expanded in the TPN group, while the proportions were reduced by L. johnsonii treatment  , Abcc2 , and Abcc3 , was significantly inhibited by TPN, while their expression was upregulated by L. johnsonii treatment  D,E accomCyp7a1) G, both oreatment D. The ilhe ileum F. CollecCyp7a1 expression is known to be suppressed by farnesoid X receptor -induced expression of fibroblast growth factor (Fgf) 19 in the terminal ileum [Nr1h4 and Fgf19 in the ileum was unaffected by TPN, while L. johnsonii significantly upregulated their expression levels , suggesting a possible BSH activity that may contribute to GCDCA deconjugation . After 6 h, the remaining GCDCA concentration was measured by LC-MS/MS. As we expected, we observed a significant reduction in GCDCA levels in the group treated with GCDCA + L. johnsonii, suggesting that L. johnsonii could directly deconjugate GCDCA to CDCA  improved liver steatosis induced by HFD in mice. By using radioactive tracers and LC-MS methods, they showed that LGG and hosts competed for fatty acid absorption in the intestine, resulting in hepatic lipid accumulation and decreased body weight. Due to their ability to ferment sugars, Lactobacillus bacteria are known to produce lactate. Another study conducted by Ritze et al. [L. johnsonii is less studied in metabolic liver diseases. Future studies could aim to unravel the specific roles of L. johnsonii-derived metabolites in PNALD, exploring their mechanisms in both preclinical and clinical settings.Consistent with previous findings from Koelfat et al.  and Wangof PNALD . Here, wg et al.  reportede et al.  showed tL. johnsonii. In line with previous findings, Zhan et al. [L. johnsonii treatment. These findings collectively suggest a potential beneficial role of L. johnsonii in mitigating TPN-induced bile stasis.Previous studies have reported that PN disrupts the enterohepatic circulation, resulting in bile “stasis” and subsequent cholestatic liver injury ,31. The n et al.  analyzedn et al. , who repL. johnsonii treatment promoted both ileal Fxr/Fgf19 signaling and Cyp7a1 expression, suggesting that the impaired regulation of Fxr caused by TPN was not restored by L. johnsonii. Furthermore, the L. johnsonii treatment group exhibited an elevated proportion of taurocholic acid (TCA) and CDCA in the serum  activity and bile resistance, which contribute to its ability to persist and colonize the gut [L. johnsonii. Furthermore, our in vitro experiments demonstrated that L. johnsonii possesses the ability to deconjugate GCDCA. Our study provides valuable insights into the mechanisms underlying PN-induced gut dysbiosis and the promotion of hepatocyte apoptosis. These findings open up new possibilities for the development of therapeutic approaches to manage PNALD.As the most extensively studied form of regulated cell death in PNALD, increased hepatocyte apoptosis has been consistently reported in preclinical studies, as well as in patients with PNALD ,45,46. H the gut . Using wL. johnsonii treatment, serum GCDCA remained unaffected. Further investigations to evaluate the effects of L. johnsonii on the bile acid pool are warranted.This work is subject to several important limitations. First, although most patients who received TPN have SBS , the rats retained the entirety of their bowel. Therefore, careful investigation is critical in translation to human PNALD patients. Second, although hepatic GCDCA was reduced after L. johnsonii in a rat model of PNALD. L. johnsonii supplementation can protect against TPN-induced liver steatosis, bile acid dysregulation, and hepatocyte apoptosis. Mechanistically, L. johnsonii attenuates TPN-induced hepatocyte apoptosis by directly deconjugating GCDCA via its BSH activity. These findings may facilitate the development of therapeutic strategies utilizing probiotics to prevent PNALD pathogenesis.In summary, to the best of our knowledge, this is the first study to demonstrate the protective effects of


The TRP channel superfamily was widely found in multiple species. They were involved in many extrasensory perceptions and were important for adapting to the environment. The migratory locust was one of the worldwide agricultural pests due to huge damage. In this study, we identified 13 TRP superfamily genes in the locust genome. The number of LmTRP superfamily genes was consistent with most insects. The phylogenetic tree showed that LmTRP superfamily genes could be divided into seven subfamilies. The conserved motifs and domains analysis documented that LmTRP superfamily genes contained unique characteristics of the TRP superfamily. The expression profiles in different organs identified LmTRP superfamily genes in the head and antennae, which were involved in sensory function. The expression pattern of different life phases also demonstrated that LmTRP superfamily genes were mainly expressed in third-instar nymphs and male adults. Our findings could contribute to a better understanding of the TRP channel superfamily gene and provide potential targets for insect control. Drosophila melanogaster [Tribolium castaneum [Spodoptera littoralis [Lygus hesperus [Nilaparvata lugens [Dmtrp, including TRPA, TRPV, TRPC, TRPM, and TRPN. The other group of TRP channels consists of the TRPP and TRPML subfamilies, which lack the multiple ankyrin repeats and TRP domains characteristic of one-group channels [Drosophila, named TRPA1, Painless (Pain), Pyrexia (Pyx), and Water witch (Wtrw) [Drosophila brivido proteins (Brv) [T. castaneum, Pediculus humannus, Acyrthosiphon pisum and Caenorhabditis elegans, respectively [Apis mellifera, Solenopsis invicta, Bombyx mori, and Nasonia vitripennis [Transient receptor potential (TRP) channels are members of cation channels, which are evolutionarily conserved proteins that have been widely identified in many model and non-model species, such as nogaster , Triboliastaneum , mice [3astaneum , Spodoptttoralis , Lygus hhesperus , and Nila lugens . Howevera lugens ,8. Membea lugens . Based oa lugens , TRPC (cchannels . The varh (Wtrw) . The TRPh (Wtrw) . Meantimns (Brv) . In contectively ,17,18,19dmtrp mutants that responded to light [N. lugens and D. melanogaster [SlTRPγ has been identified in both pupal and adult male antennae and is essential for insect olfactory in the transduction of S. littoralis [RproIav is related to thermal sensitivity [DmNan-Iav heteromers are silenced under insecticide and prevent insect feeding [DmNan and DmWtrw can also form heteromers and be activated by insecticides. Meanwhile, Wtrw increases the binding affinity of Nan for binding afidopyropen [Drosophila, NompC mutant adults are severely uncoordinated, and mutant larvae display severe defects in behavioral responses to gentle touches [Drosophila antennae [NlNompC [Previous studies have shown that TRP channel superfamily genes have a crucial role in insect growth and development. TRP channels were first discovered in to light . Despitenogaster . SlTRPγ ttoralis . RproIavsitivity . Additio feeding . Howeveropyropen . The Nomopyropen . NompC iopyropen ,26. In Dantennae . NlNompC[NlNompC . Subsequ[NlNompC .T. castaneum [Apolygus lucorum [Thermotaxis and temperature acclimation are connected to three TRPA members, including TRPA1, Painless, and Pyx in astaneum . Since iastaneum . However lucorum . A recen lucorum .osm-9 mutant encoding a protein with structural similarity to the TRP channel superfamily gene cannot be affected by odors in C. elegans [AgTRPA1 is expressed in the antennae and can be activated by citronella in Anopheles gambiae [Chemical sensations can be divided into olfactory and gustatory and are important in feeding and reproduction. These chemicals are sensed by olfactory and gustatory receptors on specific organs of insects, including the head, leg, wing, and antennae. Although TRP channel genes do not directly sense chemicals, they can participate in the transmission of signals from these chemical substances. The  elegans . AgTRPA1 gambiae . In mamm gambiae .L. migratoria is one of the most harmful pests to worldwide agricultural production. Locusts can be either solitarious or gregarious depending on population density [The migratory locust  density . Phenoty density ,39. Rese density . 4-Vinya density . Juvenil density . AlthougL. migratoria. The gene structure of subfamily members is displayed. The expression profile of members was detected. Our studies could provide more helpful information for understanding TRP channel superfamily genes and might offer potential targets for insect control.In this study, we investigated LmTRP superfamily genes in the The migratory locusts were collected from the laboratory of the College of Life Sciences, Hebei University, and reared on fresh wheat seedlings every day. All eggs, nymphs, and adult locusts grew at a greenhouse under a 14 h light and 10 h dark photoperiod at 30 ± 2 °C, 60 ± 10% relative humidity.D. melanogaster, B. mori, and N. lugens were downloaded from the NCBI database (nih.gov)National Center for Biotechnology Information ( (accessed on 17 April 2023)). All accession numbers of the TRP superfamily in this study are listed in L. migratoria were obtained using BLASTP with an e-value < 10−5 and a similarity of than 30% in L. migratoria genomic website (LocustBase-Locust Genome data (accessed on 17 April 2023)) [nih.gov)Conserved Domains Database (CDD) and Resources ( (accessed on 17 April 2023)). The conserved motifs were predicted in the MEME Suite (meme-suite.org)MEME—MEME Suite ( (accessed on 19 April 2023)). TRP superfamily protein sequences from different species were aligned by MUSCLE Wrapper and trimmed by trimAl Wrapper in TBtools. The phylogenetic tree was constructed with the neighbor-joining (NJ) method in MEGA11 [TRP superfamily protein sequences of l 2023)) . The conn MEGA11 . A bootsn MEGA11 .During the feeding process, to examine the expression levels of different developmental stages, samples are randomly collected first-instar nymph, the second-instar nymph, the third-instar nymph, the four-instar nymph, the fifth-instar nymph, and male and female adults, which ranged from after 1 day to 9 days post adult eclosion. The head, wings, and antenna of the four-instar nymph locusts were collected to RNA-Seq. All samples had three biological replicates, and each replicate had more than three locusts.TM Isolation Reagent  following the manufacturer’s protocol. All experimental steps were performed at low temperatures or on ice. The concentration of the extracted RNA was determined using a NanoDrop 2000 spectrophotometer . The RNA integrity was evaluated following 1.5% agarose gel electrophoresis. The 1 μg RNA was used to synthesize first-stand cDNA using HiScript® III All-in-one RT SuperMix Perfect for qPCR  following the manufacturer’s recommendations. The cDNA product was diluted 10 times. All samples were stored at −80 °C in order to further utilize for qRT-PCR.Total sample RNAs were isolated from different developmental stages and adult tissues using RNA-easyTM Isolation Reagent . Total RNA of each sample was quantified and qualified by Agilent 2100 Bioanalyzer , NanoDrop , and 1% agarose gel. A total of 1 μg RNA with a RIN value above 7 was used for the following library preparation. Next-generation sequencing library preparations were constructed according to the manufacturer’s protocol (NEBNext® Ultra™ RNA Library Prep Kit for Illumina®). The poly(A) mRNA isolation was performed using NEBNext Poly(A) mRNA Magnetic Isolation Module  or Ribo-Zero™ rRNA Removal Kit . The mRNA fragmentation and priming were performed using NEBNext First-Strand Synthesis Reaction Buffer and NEBNext Random Primers. First-strand cDNA was synthesized using ProtoScript II Reverse Transcriptase, and the second-strand cDNA was synthesized using Second AxyPrep vStrand Synthesis Enzyme Mix. The purified double-stranded cDNA by AxyPrep Mag PCR Clean-up  was then treated with End Prep Enzyme Mix to repair both ends and add a dA-tailing in one reaction, followed by a T-A ligation to add adaptors to both ends. Size selection of Adaptor-ligated DNA was then performed using AxyPrep Mag PCR Clean-up , and fragments of ~360 bp (with the approximate insert size of 300 bp) were recovered. Each sample was then amplified by PCR for 11 cycles using P5 and P7 primers, with both primers carrying sequences that can anneal to flow cells to perform bridge PCR and P7 primer, carrying a six-base index allowing for multiplexing. The PCR products were cleaned up using AxyPrep Mag. Then libraries with different indices were multiplexed and loaded on an Illumina HiSeq instrument according to the manufacturer’s instructions . Sequencing was carried out using a 2× 150 bp paired-end [p-adjust < 0.05.Total RNA of each sample was extracted using TRIzol Reagent /RNeasy Mini Kit /RNA-easyired-end  configurired-end . The rawhttps://primer3.ut.ee/ (accessed on 30 April 2023)), and all primers were listed in −ΔΔCt method. LmGAPDH was used as internal control. All samples were subjected to three biological replicates.RT-qPCR assays were conducted with SYBR qPCR Master Mix  by using Bio-RAD CFX Connect Real-Time System. The qPCR primers were designed by using Primer 3 (p < 0.05). Data were mean ± SD.All qRT-PCR results were analyzed using GraphPad Prism 6.0 in this study. Significant differences among samples were assessed by one-way ANOVA .To understand the expression pattern of LmTRP superfamily genes, RNA-Seq data from different tissues were used. The head, antennae, and wings are important external organs for sensing environmental changes. the wing A. To clafat body B. 4VA  . Multiply system . TRP mutaptation . DmPain aptation . OR35 cabehavior . AgTRPA1ne cells . This suB. dorsalis are mainly expressed in adults [LmNan, LmIav, LmPain1, and LmTRPγ have higher expression levels in adults. LmPain2, LmPyx, LmTRPM, LmTRPL, LmTRPML, and LmTRP have higher expression levels in the third-instar nymphs than in adults. The third instar is an important developmental stage in locusts, at which there is a greater mortality rate due to changes in the environment leading to some kind of intestinal disease, although we do not have direct evidence. BmTRPA1 is thermosensitive and affects the determination of diapause in the egg stage [LmTRPA1 has a higher expression in eggs as well. These indicate that TRPA1 is important for egg development.Insects are exposed to stimuli brought by varied environments throughout their lives. TRP channel genes in different developmental stages of n adults . Similarn adults . Howevergg stage . LmTRPA1L. migratoria. Phylogenetic tree analysis compared the relationship between locusts and other insects. Furthermore, conserved functional domains, motifs, and gene location analysis were useful for understanding the functional differences among TRP genes. In addition, we also determined the expression pattern of TRP genes in different growth development and tissues. Additionally, these offer a theoretical scientific foundation for potential biological pest control.In conclusion, we identified seven subfamilies that contained thirteen TRP genes in


Vibrio cholerae bacteria, threatens public health worldwide. The organism mentioned above has a significant historical record of being identified as a prominent aquatic environmental pollutant capable of adapting its phenotypic and genotypic traits to react to host patients effectively. This study aims to elucidate the heterogeneity of the sporadic clinical strain of V. cholerae VC01 among patients residing in Silvasa. The study involved conducting whole-genome sequencing of the isolate obtained from patients exhibiting symptoms, including those not commonly observed in clinical practice. The strain was initially identified through a combination of biochemical analysis, microscopy, and 16s rRNA-based identification, followed by type strain-based identification. The investigation demonstrated the existence of various genetic alterations and resistance profiles against multiple drugs, particularly chloramphenicol (catB9), florfenicol (floR), oxytetracycline (tet(34)), sulfonamide (sul2), and Trimethoprim (dfrA1). The pan-genomic analysis indicated that 1099 distinct clusters were detected within the genome sequences of recent isolates worldwide. The present study helps to establish a correlation between the mutation and the coexistence of antimicrobial resistance toward current treatment.Cholera, a disease caused by the  Vibrio cholerae is widespread and has long coevolved with humans. It is the agent that causes cholera, a severe intestinal illness that can result in up to 140,000 fatalities and affects between 1 and 5 million people annually [V. cholerae carries the cholera toxin and has resulted in several pandemics throughout human history. The seventh V. cholerae pandemic continues to this day, and it is even more concerning that it has evolved to present new challenges for treating cholera disease outbreaks [V. cholerae. Over 200 distinct serogroups of V. cholerae can be classified based on the antigenic properties of its “O” antigenic lipopolysaccharide [The pathogen utbreaks . Choleraccharide . In receV. cholerae and their potential for pandemic evolution holds tremendous significance in discerning molecular differentiations in their pathogenicity. V. cholerae exhibits various stages of mechanisms in its repertoire for both colonization and infection. The development of resistance in V. cholerae species is mainly attributed to genetic recombination, genomic rearrangements, and horizontal gene transfer (HGT), which enable modifications to their genomes [V. cholerae called O139 was discovered to be the root of the cholera outbreak in India and Bangladesh [The investigation of pathogenicity in strains of  genomes . In addingladesh . Accordingladesh .V. cholerae strains [V. cholerae protease (PrtV), and a serine protease [V. cholerae access to the CTX toxin [The filamentous CTX bacteriophage (CTX), which encodes the cholera toxin, the TCP pathogenicity island that encodes the TCP pili, a colonization factor and receptor for CTX, and toxR, an essential virulence regulatory gene, have all been emphasized in numerous studies as being necessary for pathogenic  strains . CTX genprotease ,10. ConcTX toxin .V. cholerae. The present investigation pertains to multidrug-resistant (MDR) V. cholerae. Pan-genomic analysis facilitates comprehension of the molecular assessment of particular genes within the genome [V. cholerae pathogenicity and its pandemic potential.Azithromycin, fluoroquinolones, and tetracycline are just a few antimicrobials successfully used to treat cholera victims over time . Howevere genome ,14. The V. cholerae from previously collected rectal swabs, followed by biochemical and serological typing. The disc diffusion method was used to evaluate susceptibility to antibiotics like ampicillin, doxycycline, erythromycin, nalidixic acid, chloramphenicol, ciprofloxacin, sulfamethoxazole-trimethoprim, and tetracycline based on the method suggested by Kirby-Bauer [A female patient, aged 26, was admitted to Silvassa’s tertiary hospital due to severe symptoms of vomiting and diarrhea. A rectal swab was obtained and enriched with alkaline peptone water under the medical officer’s observation. The selective medium, thiosulfate-citrate-bile-sucrose (TCBS) agar, was used to grow by-Bauer .High molecular weight genomic DNA was extracted from the isolate using a QIAamp DNA Mini Kit . The genomic DNA was quantified using a Qubit fluorometer . The quality of the genomic DNA was assessed using 2% agarose gel . The Minhttps://usegalaxy.org.au/, accessed on 30 June 2022). The TrueBac ID system and Type (Strain) Genome Server (TYGS) were used to carry out a whole genome-based taxonomic analysis [https://cge.food.dtu.dk/services/MLST, accessed on 30 June 2022) was utilized to validate further species identification based on type strain-based identification [The genome assembly was constructed using the Canu assembler on the Galaxy Australia webserver , including prophage sequences, insertion sequences, CRISPR-Cas systems, and plasmids [V. cholerae VC01 [plasmids . The ICEplasmids . The onlrae VC01 ,32, whilrae VC01 .https://github.com/tseemann/abricate, accessed on 30 June 2022), One-Codex , Blastn against Virulence factor database(VFDB) [Four approaches determined the presence of acquired resistance genes: Abricate (se(VFDB) , and Resse(VFDB) . All metUsing OrthoFinder 1.1.8, ortho-groups were compiled, and trees based on the presence–absence of ortho-group members were created to understand the highly diverse type of Genomes . The RoaV. cholerae O1 biotype. Irrespective of their biotype, the isolated V. cholerae VC01 strain was resistant to amikacin, gentamicin, co-trimoxazole, and nalidixic acid, while retaining susceptibility towards tetracycline, chloramphenicol, and trimethoprim as observed in the organism. Only some genome sequencing studies of V. cholerae are available from the western region of India. To understand the changes in the genome sequence of the present isolate, we carried out whole genome sequencing of one isolated organism to find differences in the genomic level to compare genomes during pre- and post-COVID-19 situations.A rectal swab sample was collected from patients with acute diarrhoeal disease in a tertiary hospital at Silvassa (India) showing sporadic cholera cases from hospitalized patients. The antimicrobial drug susceptibility exhibits a strain showing agglutination with polyvalent O1 antisera. The preliminary test results based on agglutination show similarity to the V. cholerae ATCC 14035 strain, followed by Vibrio albensis ATCC 14547, Vibrio paracholerae EDC-792T, and Vibrio tarriae 2521-89  Genome Server (TYGS) results, 14 strains were found with a closely related genome sequence for the isolated organism. The nearest match observed was to the  2521-89 .V. cholerae ATCC 14035 strain. These results also show consecutive confirmation using biochemical analysis performed for the isolated strain. Based on the results, the isolate underwent MLST profiling which confirmed its close clustering with V. cholerae  as a reference database, Abricate’s findings revealed 63 antibiotic resistance genes, followed by 28 from the Megares database using 75% identity/coverage. The analysis was also carried out for CARD, the NCBI antimicrobial resistance database, and the Argannot database and showed the presence of 9, 8, and 7 genes, respectively  were identified as conferring resistance to the multidrug efflux drug class. Aminoglycoside, diaminopyrimidine, carbapenem, macrolide, fluoroquinolone, penam, phenicol, elfamycin, and multidrug efflux-based resistance are the main categories of antibiotic resistance that have been reported in the genome sequence of an isolated organism .A resistance gene was detected that belongs to V. cholerae strain VC1786ICE (NCBI Accno: CP028827.1) with a maximum score of 167.5, followed by 165 for V. cholerae VC833 ICE element genomic sequence (Genbank Accno: KC886258.1) and 163.5 for V. cholerae strain VC504 ICE element genomic sequence (Genebank Accno: KC886257.1).ICEfinder predicted an ICE region on the analyzed genome sequence at 278,238–424,664 nucleotides covering 146,427 base pairs  as operoVibrio phage CTXphi (NCBI Acc No: NC_015209.1). However, three incomplete prophage regions belong to E. coli and P. aeruginosa. The co-morbid presence of V. cholerae with other filamentous organisms from clinical samples also reported such transfer [V. cholerae-like organisms.The current study aims to identify the presence of phage sequences within the genetic material of an isolated strain by utilizing the PHAge Search Tool Enhanced Release (PHASTER) . The PHAtransfer . DespiteV. cholerae VC01 strain is related to the V. cholerae PS4 strain as a distinct group, according to phylogenetics based on the pan-genomic study  is a critical member of the bacterial mobile genetic elements, which is integrative to the bacterial chromosome, encodes fully functioning conjugation machinery, and is thus self-transmissible between bacterial cells. The genes present in ICEs, which encode for both antibiotic resistance determinants and virulence factors, can provide the host with selective advantages. ICEfinder offers an efficient way to detect the presence of ICE sequences by comparison with its database [V. cholerae O1 and non-O1/non-O139 from the Indian subcontinent and South America, as well as in Shewanella sp. from North America [V. cholerae O44 that exhibits resistance to clotrimoxazole has been linked to MGIVchHai6 T4SS. The comparative study regarding the ICE and its importance previously revealed that the genome of V. cholerae O1 isolates responsible for the cholera outbreak in Haiti in 2010 contained ICEVchHai1, which carried auxiliary support conferring resistance to trimethoprim (dfrA1), sulfamethoxazole (sul2), streptomycin (strAB), and chloramphenicol (floR) [This present study’s whole genome sequencing and comparative genomics methodology provide insight into the diversity of the V. cholerae strain VC01 isolated from clinical samples. A similar study was conducted to characterize the genome for pathogenic and non-pathogenic in China . Our findatabase . Earlier America . Howeverl (floR) . Similarl (floR) .V. cholerae O1 [V. cholerae genomes were thoroughly investigated. The V. cholerae O1 serogroup classical biotype strains possess the cholera-toxin encoding phage CTX(cla) specific to their biotype, while the El Tor biotype strains carry CTX-1 [The detected prophage sequence from an isolated organism was found in a similar sequence of Phage CTX observed in lerae O1 . This inry CTX-1 . HoweverV. cholerae can acquire resistance to antimicrobial compounds, antibiotic target replacement, and antibiotic target protection, and prompt inactivation of the antibiotic by hydrolysis, preventing access to the target site by changing membrane permeability and actively exporting antibiotics from bacterial cells [V. cholerae. Additionally, the isolate was resistant to pulvomycin, which was horizontally transmitted to V. cholerae. A similar antibiotic resistance mechanism was previously reported in the Uganda study during 2014–2016 [Generally, Norfloxacin, trimethoprim-sulfamethoxazole (TMP-SMX), ciprofloxacin, and azithromycin have been used to treat cholera. The World Health Organization (WHO) recommends the management of cholera with oral rehydration salts in combination with antibiotics such as streptomycin, aminoglycosides, trimethoprim, fosfomycin, fluoroquinolones, sulphonamides, chloramphenicol/florfenicol, and tetracyclines . Usuallyal cells . In the 014–2016 .V. cholerae. Antimicrobial resistance in V. cholerae is also known to be due to the acquisition of resistance genes from closely or distantly related microbial species through horizontal gene transfer [V. cholerae [Although it is presently difficult to predict, the effect of treatment for infectious disease will affect antimicrobial resistance. However, most studies show extensive empirical use in contrast to comparatively infrequent bacterial co-infection and secondary infections . Exposurtransfer . Inadequcholerae ,63.Actinobacillus pleuropneumoniae, Aggregatibacter actinomycetemcomitans, Aliivibrio wodanis, Bacillus subtilis, Escherichia coli, Kingella kingae, Pseudoalteromonas sp., Salmonella enterica, Staphylococcus aureus, Vibrio spp. Similarly, Verma et al. (2019) also reported the cross-species transfer of resistance genes to V. cholerae genomes from 2008 to 2015 [Based on pan-genomic analysis, out of 1099 unique clusters, 22 were identified as having a role in CRISPR (04), drug resistance (07), phage sequences (02), and toxin-related functions (09). The unique clusters  were rep to 2015 . Pan-genV. cholerae resistance to major classes of antibiotics is concerning. An example is the incidence of quinolone resistance genes in V. cholerae O139 strains from China that were conjugative and contributed due to a CIP-resistant plasmid similar to that observed in V. vulnificus [V. cholerae [V. cholerae strain having resistance towards more than three groups of antibiotics using genome analysis [Over time, lnificus . Additiocholerae . All theanalysis . Such tyV. cholerae vaccine can show the same scenario in the future. However, the antibiotic susceptibility data of publicly available genomes were not included in the analysis as the data were not available. Hence, there is a need for genome-based surveillance from all other currently isolated strains that might provide more insight into such antibiotic resistance. The present study provides insight into whole genome sequencing of such isolates from the patient and also can be replicated for future isolates for tracking cholera like infectious diseases.In August 2020, the first round of the OCV campaign was done in the most affected districts of the Littoral, South, and Southwest regions . The numV. cholerae. Pan-genomic profiling offers valuable insights into the presence of additional sequences in V. cholerae through gene transfer, which can confer resistance to multiple antibiotics. The findings demonstrated the need to explore alternative therapeutic approaches for managing multidrug-resistant cholera. In addition, it is beneficial to perform re-efficacy testing on presently utilized vaccines against V. cholerae. This may offer explicit knowledge of the possible existence of epitopes derived from other organisms as a potential site for the development of vaccines. The presence of antibiotic resistance genes in the whole genome and in vitro study of susceptibility towards similar antibiotics shows a correlation of gene expression in the isolated strain. This research has generated interest in further genomic profiling and understanding the dissemination and multidrug resistance using whole genome sequencing of V. cholerae. India, like other Asian countries, carries more asymptomatic V. cholerae incidences every year, leading to increased mutation of multidrug resistance. The concurrent whole genome sequencing provides a detailed understanding of the dissemination and multidrug resistance in such infectious agents.Recently, due to COVID-19 implications, the emphasis on infectious diseases such as cholera shifted, resulting in high fatality rates worldwide due to co-morbid infection with


Silver nanoparticles have been a recent focus of many researchers in dentistry, and their potential uses and benefits have drawn attention in dentistry and medicine. The fabrication and utilization of nanoscale substances and structures are at the core of the rapidly developing areas of nanotechnology. They are often used in the dental industry because they prevent bacteria from making nanoparticles, oxides, and biofilms. They also stop the metabolism of bacteria. Silver nanoparticles (AgNPs) are a type of zero-dimensional material with different shapes. Dentistry has to keep up with changing patient needs and new technology. Silver nanoparticles (AgNPs) can be used in dentistry for disinfection and preventing infections in the oral cavity. One of the most interesting metallic nanoparticles used in biomedical applications is silver nanoparticles (AgNPs). The dental field has found promising uses for silver nanoparticles (AgNPs) in the elimination of plaque and tartar, as well as the elimination of bacterial and fungal infections in the mouth. The incorporation of AgNPs into dental materials has been shown to significantly enhance patients’ oral health, leading to their widespread use. This review focuses on AgNP synthesis, chemical properties, biocompatibility, uses in various dental fields, and biomaterials used in dentistry. With an emphasis on aspects related to the inclusion of silver nanoparticles, this descriptive review paper also intends to address the recent developments of AgNPs in dentistry. Silver (Ag) ions or salts have significant antibacterial capabilities and have been employed in medical applications for years. Sterile gauze, catheterization, and prosthesis are examples of this. Ag’s low toxicity, great biocompatibility with human cells, and constant mobile ions provide long-lasting bactericidal action . Silver S. mutans bacterium” [The term “Nano” derives from the Greek word “Nano”, which signifies dwarf or extremely little . The phrcterium” . There acterium” .Based on their functions, silver nanoparticles have been used in a variety of healthcare disciplines. Silver nanoparticles have functions  that inc6H12O6), hydrazine (N2H4), hydrazine hydrate, ascorbate (C6H7NaO6), ethylene glycol (C2H6O2), N-dimethylformamide (DMF), hydrogen, dextrose, ascorbate, citrate (Turkevich method), and sodium borohydride, can be used to chemically reduce these metal salts (BSS method) [Metallic nanoparticles (NPs) are typically produced using chemical synthesis processes that reduce the metal salts of NPs and disperse them in an organic or aqueous solution. A number of metallic salts are used to make gold, silver, iron, zinc oxide, copper, palladium, platinum, and other analogous metal nanospheres. The nucleation and formation of NPs are controlled by several reaction parameters, such as reaction temperature, pH, concentration, type of precursor, reducing and stabilizing agents, and the molar ratio of surfactant/stabilizer and precursor. Different chemical reductants, such as glucose (C method) . Brust a method) . Previou method) . Because method) ,14,15,16After considerable research and development, dental and medical nanoproducts undergo preclinical in vitro testing to assess their mechanical, toxicological, and immunological properties. Many authorities, such as the National Institute of Occupational Safety and Health and the US Environmental Protection Agency, have nanoparticle risk guidelines . The ethSociety is the client, financier, and policymaker. Thus, nanotechnology’s success depends on public opinion. New sparking technologies have made ethics, morality, and values more sensitive  because Federal and state agencies in the US have developed a four-stage framework for examining and evaluating health issues . These sNew biocompatible dental materials have been developed to protect the mouth’s keratinocytes and oral mucosal tissues . BiocompS. mutans was significantly inhibited without affecting resin adhesiveness [AgNPs improved self-etching adhesives’ antibacterial durability. siveness ,44. AgNPsiveness . AgNP posiveness . In firssiveness . Silver siveness .S. aureus and E. faecalis, as well as 5.25% NaOCl. This irrigation solution can irrigate root canals because AgNPs have a deeper antibacterial effect. [AgNPs boost the adhesive’s flexibility to nano-imperfections and have a short setting time compared to conventional sealers, particularly stiffness, poor solubility in extracellular space, and biochemical affinity to dental tissue. To improve intra-canal drugs, calcium hydroxide with AgNPs was examined regarding Enterococcus faecalis development . It kill effect. S. aureus and E. coli. It also promoted osteoblast-like MG63 cell line development. This study demonstrated AgNPs’ biological action in a novel way. This matters because AgNPs’ physicochemical qualities directly affect surface cytotoxicity [Dental implants are more reliable than removable prostheses or permanent bridges for tooth replacement. Implant success depends on mechanical and biological factors. Biofilm buildup on titanium implants may cause failure despite their biocompatibility. Peri-implant infection might disturb patients. To prevent peri-implant infections, titanium surfaces need antibacterial coatings . A singltoxicity . Biofilmtoxicity . Carboxytoxicity .w/w were tested. AgNPs/HA of 5% w/w had the strongest antibacterial efficacy against different microorganisms [Orthodontic patients frequently acquire white spots and cavities. Plaque builds around orthodontic brackets due to inadequate self-cleaning and acidogenic mouth flora. Oral biofilms may cause bracket caries and demineralization. AgNPs could greatly reduce bacteria adhesion to teeth and demineralization. AgNP-containing resin inhibited bacterial growth due to extended Ag+ ion release. Mechanical properties were unaffected by this strong biocompatibility . Thus, crganisms . Orthodorganisms . The userganisms . Researcrganisms ,61. Thes2). Compression resistance was high for PC/ZrO2 + AgNPs. AgNPs’ porosity-reducing integration may be to blame [Pediatric dentistry uses AgNPs widely and AgNPs are blended with diverse materials to take advantage of their antimicrobial and smaller particle size, which decreases biofilm generation and improves oral health. Research examining AgNPs in composite resins and adhesive systems is lacking . AgNPs ato blame . MTA andto blame . Thus, AOral squamous-cell carcinomas are a major cause of employment disability, as well as catastrophic cosmetic problems in the mouth and face. Both radiotherapy and chemotherapy have the potential to alter homeostasis in a variety of ways. The same is true for the disadvantages of anticancer drugs, which require research on other therapeutic options. Although nanoparticles of various types have been used in medicine and dentistry, their efficiency in treating malignant tumors has yet to be demonstrated. AgNPs are cytotoxic because they can damage DNA and cause apoptosis. However, studies on the efficacy of AgNPs in cancer treatment are scarce. AgNPs reduce the development of a cancer cell line at low doses . BerberiSilver nanoparticles have been used in various dental materials and various uses of AgNPs in dental biomaterials are illustrated in Acrylic polymers derived from poly (methyl methacrylate), also known as PMMA, are utilized in the production of partial and complete dentures used to replace missing teeth. These rough-surfaced resins are more likely to attract potentially dangerous microbes. C. Albicans, one of the most important opportunistic infections, can colonize these resins . Many de2). Compression resistance was high for PC/ZrO2 and AgNPs. AgNPs’ porosity-reducing integration may be to blame [S. mutans. Composite resins and adhesives with AgNPs minimize biofilm and tooth loss. Ag+ ions barely affect polymerization. AgNPs affected light-cured polymerized specimens’ production of important chemicals [S. mutans on their surface and at a longer distance than the QADM bonding agent [AgNPs are blended with diverse materials to take use of their antimicrobial properties, which minimize biofilm and promote oral health. Due to their tiny size, AgNPs can easily permeate cell membranes, causing damage and inactivity. Research examining AgNPs in composite resins and adhesive systems is lacking . This prto blame . MTA andhemicals . AgNPs ang agent . Both chng agent .E. faecalis (NaOCl). As a novel intra-canal irrigant, a solution below 5 mg/L is bactericidal, similar to 5.25% NaOCl, a microorganism-fighting AgNP-containing NaOCl-based irrigant [S. aureus, E. faecalis, and 5.25% NaOCl. Because AgNPs are more antimicrobial, this irrigation solution can irrigate root canals [E. faecalis the least. They are cytocompatible at low doses [E. faecalis in root canal therapy than either were alone. Ca(OH)2 alone outperformed AgNPs. This study found that AgNPs without Ca(OH)2 killed less bacteria. Application technique, particle concentration and size, and AgNP-Ca(OH)2 antagonism may explain these results. Bactericidal irrigation solutions using AgNPs as the last irrigant were tested. As shown, AgNPs killed E. faecalis, as well as NaOCl. Unlike the previous study’s 5.25% concentration, NaOCl at 2.25% was as effective as AgNP’s irrigants. AgNP-containing irrigants eliminate the smear layer, improving penetration. The clinical use of uniradicular teeth was innovative. A better bacterial performance was observed. Bacteria in the root canal after treatment may lead to treatment failure, according to studies. Root canal treatment eliminates germs. AgNPs in root canal filling and intra-canal irrigation are worth studying because most endodontic materials lack antimicrobial characteristics [Root canal materials should destroy these germs to improve tooth prognosis. Root canal fillings should be gutta-percha. Nanosilver-coated gutta-percha tests obturated teeth for microleakages. Nanosilver leaks less than gutta-percha but functions similarly. Eliminating bacteria and pulp tissue prevents canal space contamination. Disinfecting instruments may overlook root canal anatomy. These constraints allow for mechanical devices to clean and sanitize the canal with different irrigants, such as sodium hypochlorite (NaOCl), ethylene diamine tetraacetic acid (EDTA), and chlorhexidine (CHX). Several studies have advocated nanoparticle irrigation systems for root canal lavage . We testirrigant . This Agt canals . CHX andow doses  and demoow doses . Candidaow doses . The comeristics . Silver eristics .Oral biofilms—polymicrobial communities of bacteria and yeasts—cause gum and tooth infections. Biofilm bacteria cause cavities, periodontal disease, and abscesses. Biofilm removal prevents periodontal disease and tooth loss. Mechanical treatment with adjunctive antibacterials is possible. Structurally sound biofilms resist antimicrobials. Thus, antibacterial medication research without resistance is essential. Carboxymethyl cellulose and sodium alginate protect AgNPs from bacteria and biofilms. Carboxymethyl cellulose-capped AgNPs suppress Gram-negative bacteria more than sodium alginate AgNPs. Because Gram-negative bacteria cause most periodontal infections, AgNPs may help to treat them. AgNP size and antibacterial activity of 5 nm AgNPs were most effective against oral anaerobic microorganisms. AgNPs from banana peel extract were synthesized to kill human pathogens .w/w mixes were tested. The 5% w/w AgNPs/HA is the most antimicrobial. The 5% composite (AgNPs/HA) orthodontic adhesives prevent cariogenic bacteria and biofilm growth. Shear bond strength (SBS) of orthodontic adhesive increased by 1%, was maintained by 5% AgNP/HA, and decreased by 10%, suggesting that 5% AgNPs/HA provides appropriate antibacterial and mechanical properties for orthodontic adhesives. Although AgNPs in orthodontics are still in their infancy, there is still the possibility for growth. AgNPs can be employed as nano-vectors for gene transfer to accelerate mandibular growth due to their antibacterial qualities. Future studies should show how these technologies can be utilized. Clinical applications can increase treatment quality and minimize orthodontic care costs [Orthodontic patients acquire more white spots and cavities. Plaque builds around orthodontic brackets due to inadequate self-cleaning and acidogenic mouth flora. Oral biofilm may cause bracket caries and demineralization. Plaque biofilm can be mechanically removed and treated with antibacterial agents and fluoride. Orthodontists utilize AgNPs to prevent bacteria from clinging to teeth and demineralizing enamel. Antibacterial orthodontic band cement was created using AgNPs. White lesion progression was monitored. Resins generate AgNPs in situ. AgNP-containing resin inhibit bacterial growth due to extended Ag+ ion release. Mechanical properties are unaffected by excellent biocompatibility. Thus, orthodontic cements with AgNPs may prevent white spot lesions. Silver nanoparticles (AgNPs) and hydroxyapatite (HA) in orthodontic adhesive have antibacterial properties. AgNPs’ antibacterial properties were used in this investigation because HA’s early enamel lesions demineralize, and 1%, 5%, and 10% re costs .Squamous-cell carcinomas of the head and neck account for 6% of all cancer diagnoses. Nearly 40% of them are derived from the tongue or the floor of the mouth. Those who develop metastatic disease or advanced malignancies frequently do not fare well. Early detection and combinations of radiation, surgical treatments, and chemotherapy have not considerably lowered cancer incidence or mortality. Oral squamous-cell carcinomas are a major cause of employment disability, as well as catastrophic cosmetic problems in the mouth and face. Both radiotherapy and chemotherapy have the potential to alter homeostasis in a variety of ways. The same is true for the disadvantages of these anticancer drugs, showing the need for research on other therapeutic options. Although nanoparticles of various types have been used in medicine and dentistry, their efficiency in treating malignant tumors has yet to be demonstrated. AgNPs are cytotoxic because they can damage DNA and cause apoptosis. They may also have an effect on healthy cells. However, there is a scarcity of studies on the efficacy of AgNPs in cancer treatment. AgNPs reduce the development of cancer cell lines at low doses. Berberine, an all-natural chemical, reduces cell proliferation. These findings imply that AgNPs plus berberine could form an effective anticancer treatment combination. The creation of AgNPs has also been explored from several perspectives. Surface changes enable malignant cells to be specifically targeted. These molecules can also be utilized to detect cancer early, providing clinicians with a valuable tool. Because they specifically kill cancer cells while preserving healthy ones, nanocarriers reduce the common side effects of chemotherapy .S. mutans bacterium” [S. mutans” by either curative or preventative means. The vast majority of persons who were a part of our group did not receive regular updates, and the instruction regarding dental health was insufficient. Because it inhibits the enzymes that are involved in the respiratory chain and interferes with DNA synthesis [S. mutans [The antibacterial (AB) characteristics of metallic silver (Ag) have been recognized for a very long time. Compounds containing silver were applied in a range of treatment approaches for ailments such as ulcers, burns, and wound healing prior to the discovery that bacteria were the source of infection. This was the case both before and after the discovery of antibiotics. Ag possesses a wide variety of advantageous qualities, one of which is potent “antimicrobial” activity against a wide variety of infectious diseases . In rececterium” ,88,89. Tynthesis , silver . mutans ,89. Howe. mutans . AgNPs a. mutans  Research. mutans . These d. mutans ,95. This. mutans ,97. AgNP. mutans ,40,51. D. mutans ,77. Oste. mutans . For dec. mutans ,46,51,52Despite the positive findings of previous research regarding the use of silver nanoparticles (AgNPs) in dental materials, more in-depth, long-term clinical studies are necessary to evaluate their clinical efficacy and overall benefits for the dental health of patients. Because AgNPs have a greater chance of passing through cell membranes, their ability to kill bacteria is enhanced . This isIn the current research, it has been demonstrated that silver nanoparticles possess specific characteristics and can be utilized in a range of dental applications. Although silver nanoparticles have greater potential in the field of dentistry, it has seen limited use in specific disciplines. In addition, it has been shown that silver nanoparticles are biocompatible with mammalian cells, further reassuring the medical community that using silver nanoparticles in dental materials is safe for humans. However, greater research is required to determine how to make the most effective use of silver complexes to assure antibacterial activity without increasing cytotoxicity throughout the various subspecialties of dentistry. This is the case despite the fact that silver complexes have been shown to be effective.


Aegilops accessions distributed among 23 different species using more than 45K single nucleotide polymorphisms identified by genotyping-by-sequencing. The Wheat Genetics Resource Center (WGRC) Aegilops germplasm collection was curated through the identification of misclassified and redundant accessions. There were 49 misclassified and 28 sets of redundant accessions within the four diploid species. The curated germplasm sets now have improved utility for genetic studies and wheat improvement. We constructed a phylogenetic tree and principal component analysis cluster for all Aegilops species together, giving one of the most comprehensive views of Aegilops. The Sitopsis section and the U genome Aegilops clade were further scrutinized with in-depth population analysis. The genetic relatedness among the pair of Aegilops species provided strong evidence for the species evolution, speciation, and diversification. We inferred genome symbols for two species Ae. neglecta and Ae. columnaris based on the sequence read mapping and the presence of segregating loci on the pertinent genomes as well as genetic clustering. The high genetic diversity observed among Aegilops species indicated that the genus could play an even greater role in providing the critical need for untapped genetic diversity for future wheat breeding and improvement. To fully characterize these Aegilops species, there is an urgent need to generate reference assemblies for these wild wheats, especially for the polyploid Aegilops.Genetic diversity found in crop wild relatives is critical to preserve and utilize for crop improvement to achieve sustainable food production amid climate change and increased demand. We genetically characterized a large collection of 1,041  We also examined Aegilops and wheat genomes relationships through Aegilops sequence mapping to the wheat genome and genetic clustering.The Wheat Genetics Resource Center (WGRC) gene bank at Kansas State University has been maintaining myriads of wild wheat accessions under the id wheat  and Ae. tauschii . Thus, t22.1Aegilops species preserved and maintained in the WGRC gene bank    and Ae. periment  were useAe. speltoides and Ae. mutica are partially out-crossing and were maintained through sib-mating multiple plants. Specifically, Ae. mutica accessions consisted of 54 samples from five out-crossing plants bulked together.Most of these species are self-pollinated and were primarily maintained by single seed descent, with exceptions described below. 2.2Ae. mutica, where we sequenced 54 individuals obtained from randomly crossing five plants, because the species is cross-pollinating and it has a low germination rate.The DNA extraction, GBS library preparation, and sequencing were performed as we described in our earlier studies  using twde-novo single nucleotide polymorphism (SNP) calling, reads were demultiplexed using sabre (https://github.com/najoshi/sabre) and adapters were trimmed using fastp  was used as a threshold to confirm genetically identical accessions. Allele matching used homozygous and non-missing sites between two given accessions, and the raw markers were filtered using MAF >0.01, missing <50%, and heterozygous <20% parameters before allele matching. We conducted further examinations of the sets of genetic duplicates to assess their phenotypic similarities, collection sites, and sources of collection.In the second step, the genetically identical accessions were determined using allele matching . However2.4Aegilops populations, which were then used for exploring the population structure and ancestry. For phylogenetic clustering, the genetic distance was computed using the “dist” function in R  and Ae. neglecta  for the presence/absence of the M genome. There are recent cytology-based findings that have questioned the traditional genome symbols of these species  whose genomic compositions are unequivocally recognized as MU or UM. The de-novo variants were first identified for the diploid M genome (Ae. comosa) and U genome (Ae. umbellulata) populations separately, and then the same variants were called on these four tetraploid species. We also constructed the phylogenetic clustering among Ae. columnaris, Ae. neglecta, Ae. geniculata, Ae. biuncialis, and a tetraploid that shares only the U genome  to see their relative positions in the tree.We investigated the traditional genome symbols of  species . To test2.6Aegilops GBS reads to the wheat genome (CS.Ref.v1)   and comp33.1Aegilops species characterized in this study were primarily collected around the Fertile Crescent, Anatolia, central Asia, northern Africa, and southern Europe (Aegilops section  exhibited a much wider distribution from central Asia to northern Africa (Cylindropyrum  were primarily collected from Uzbekistan, Tajikistan, Kazakhstan, Azerbaijan, and Turkey. The species of the Comopyrum  mainly come from Greece, Turkey, and Russia. The Sitopsis (S genome) species  were predominantly collected in Turkey, Israel, Syria, Iraq, and Jordan. The Vertebrata section species  were obtained from central Asia to southern Europe , we retained 46,879 SNPs , Ae. markgrafii (2), and Ae. searsii (1) (The phylogenetic clustering and PCA enabled us to identify and correct the classification of 49 accessions (rsii (1) Figure 23.3.2Ae. tauschii and four members of the Sitopsis section  and Ae. geniculata and Ae. biuncialis (UM), clustered into respective primary clades. Overall, there were three primary clades: (i) the first clade consisted of Ae. speltoides and Ae. mutica; (ii) the second clade has four diploids of Sitopsis (except Ae. speltoides), Ae. tauschii, and D genome polyploids (except Ae. cylindrica); (iii) the third primary clade has all other species, including M, N, C, and U genome diploids and polyploids.The unrooted NJ phylogenetic tree with all tested t groups Figure 3Ae. neglecta and Ae. crassa, were grouped separately by ploidy. The ploidy levels of these genetically clustered sub-groups (6X and 4X) were also verified using chromosome counting (Ae. crassa (The hexaploid (6X) and tetraploid (4X) species within a clade, such as . crassa .Aegilops species commensurate with the phylogenetic analysis. The first and second principal components (PC1 and PC2) explained about 17% and 14% of the variations among the Aegilops, respectively. PC1 separated Ae. speltoides from other polyploids and diploids  and the Emarginata. We also observed that the Ae. mutica (T genome) clustered closer to Ae. speltoides both in PCA and phylogenetic analysis (Sitopsis group and Ae. mutica were further verified by computing pairwise Nei’s FST (Ae. mutica has the closest genetic relationship [lowest FST (0.65)] with Ae. speltoides, closer than any other members of the Sitopsis   and the other had all remaining tetraploids  de-novo reference, the tetraploids distinctly grouped for their genomic compositions. The tetraploid species Ae. pregerina and Ae. kotschyi (US genome), Ae. neglecta and Ae. columnaris , and the UM genome tetraploids Ae. biuncialis and Ae. geniculata formed a separate clade and sub-clades  and Ae. biuncialis (MU) in both phylogenetic clustering and PCA  and U genome (Ae. umbellulata mock reference)  on the U genome, whereas the percentage read alignment of Ae. columnaris and Ae. neglecta on M genome was low (~21%) as compared to the alignment of Ae. geniculata and Ae. biuncialis reads (~38%). We also noticed that a few Ae. comosa segregating loci were mapped for Ae. columnaris (10%) and Ae. neglecta (24%) on the M genome. In contrast, Ae. biuncialis had 50% and Ae. geniculata had 46% M-genome loci. Hence, the proportion of mapped reads and loci also suggested that the Ae. neglecta and Ae. columnaris must have the U genome, but a different second sub-genome than M. Thus, we proposed that Ae. columnaris and Ae. neglecta genome formulas are most likely UX  or UXN in hexaploid form as proposed based on the cytology  followed by Ae. searsii (0.013) and Ae. umbellulata (0.015). Among the diploids, the Ae. speltoides had the highest Nei’s diversity (0.072), which was followed by Ae. mutica (0.053). Among the tetraploids, the Ae. triuncialis had the lowest diversity index (0.032) while the Ae. neglecta had the highest diversity index (0.062). The hexaploid species Ae. vavilovii has the highest Nei’s diversity index value among all 23 species analyzed in the experiment , the highest proportion of sequence reads was mapped onto wheat chromosomes of the D subgenome, followed by those of the A and B subgenomes  (Ae. comosa) (Ae. markgraffii)  and Emarginata (four other) Sitopsis were greater; therefore, the redefinition of the section Sitopsis could be desirable. One of the ideas could be the separation of Ae. speltoides from the rest of the four Sitopsis members and regrouping the Ae. speltoides with Ae. mutica  and are most likely the sub-species of the same species. Also, both of these species are equally distant from Ae. speltoides. The finding is also supported by the latest study, where We also showed that the Ae. speltodies; var. speltoides and ligustica with no population differentiation , variation in repetitive nucleotide sequences, and differences in the karyotype structure C-banding pattern, have been used to confirm the absence of the M genome in Ae. neglecta and Ae. columnaris (Ae. columnaris (UX) and Ae. neglecta (UX or UXN). Furthermore, cytological and genomic evaluation of the X genome is certainly warranted.In this study, we found further evidence that the ignation  have preignation . Severallumnaris . This st4.5Aegilops accessions diversity (Ae. sharonensis (nine accessions) exhibited a higher diversity index (0.019) compared to Ae. umbellulata (58 accessions), and the tetraploid Ae. ventricosa (17 accessions) had a higher diversity index than another tetraploid, Ae. triuncialis (199 accessions)  are genetically closer to the wheat D subgenome (Ae. speltoides (Ae. tauschii (the D-genome lineage), resulting in stronger evolutionary relationships of the U, M, N, and C diploid Aegilops within the D-genome lineage  using over 45K SNPs and constructed a robust phylogenetic tree and the PCA clusters. The population grouping and structuring of this valuable wild wheat species largely align with the traditional nomenclatures at the species level. Moreover, using these high-throughput genome-wide markers, we have confirmed the genome symbols of two tetraploid species that were previously under debate in the literature.In conclusion, this study has unveiled the genomic and genetic relationships among all Aegilops subgenome and wheat subgenomes exhibit unique relationships at the genomic level, warranting further investigation. Notably, Ae. mutica showed unique characteristics, appearing as a sister group of Ae. speltoides, yet displaying a higher number of sequences and variants mapped onto the wheat subgenome D. The genetic and evolutionary relationships among Aegilops and with wheat will become clearer when we have more genomic resources, such as genome assemblies and resequencing data for each Aegilops species. This study offers a comprehensive view of the relative genetic diversities of all 23 species together for the first time. The substantial genetic diversity observed, along with its relative extent in each Aegilops species, presents an opportunity to select species and germplasms as sources of novel alleles for wheat breeding and improvement.Our findings also reveal that each https://datadryad.org/stash/dataset/doi:10.5061/dryad.mgqnk994n.The Raw GBS data, the fastq files, are available in the Sequence Read Archive (SRA) of the National Center for Biotechnology Information (NCBI) under the BioProject accession PRJNA985892. The key file and necessary SNP matrices and the R script files (.rmd) are provided in the dryad public repository which are available with the unique DOI: 10.5061/dryad.mgqnk994n. All data are available in the article or the supplementary files and at the Dryad digital repositories LA: Data curation, Formal Analysis, Investigation, Methodology, Validation, Visualization, Writing – original draft, Writing – review & editing. JR: Data curation, Formal Analysis, Methodology, Resources, Validation, Writing – review & editing. SW: Investigation, Methodology, Writing – review & editing. D-HK: Conceptualization, Formal Analysis, Investigation, Methodology, Resources, Validation, Visualization, Writing – review & editing. BF: Conceptualization, Methodology, Resources, Supervision, Validation, Writing – review & editing. JP: Conceptualization, Funding acquisition, Investigation, Methodology, Project administration, Resources, Supervision, Validation, Writing – original draft, Writing – review & editing.


A-D). The patient denied scalp itch, pain, and was otherwise asymptomatic.A 57-year-old woman with a history of systemic lupus erythematosus presented to a dermatology clinic with 20 years of ongoing hair loss affecting the frontoparietal scalp and crown , A-D. ThA and B).A punch biopsy was performed of the vertex scalp which showed decrease and miniaturization in the number of hair follicles, associated with sebaceous glands, fibrous stellae, and a peri-infundibular lymphocytic infiltrate with few dyskeratotic keratinocytes , A and BThe patient was started on hydroxychloroquine, methotrexate, topical clobetasol, oral minoxidil, and intralesional Kenalog injections at her subsequent visits, with improvement in hair regrowth.Question 1: What is the most likely diagnosis?A.Frontal fibrosing alopecia (FFA)B.Alopecia AreataC.Lupus hairD.Fibrosing alopecia with a pattern distribution (FAPD)E.Discoid lupus erythematosus (DLE)Answers:A.FFA – Incorrect. The clinical presentation does not support this diagnosis. Frontal fibrosing alopecia (FFA) is a lymphocytic cicatricial alopecia characterized by frontotemporal and eyebrow hairloss.B.Alopecia Areata – Incorrect. Diffuse alopecia areata may be a mimicker clinically, however the classic histopathology would not show a scarring alopecia. There would be absence of perifollicular fibrosis and there would be a lymphocytic cell infiltrate around the anagen bulb.C.Lupus hair – Incorrect. Lupus hair is characterized by dry and fragile short hairs on the frontal hair-line in patients with SLE, giving the appearance of vellus hairs on the anterior scalp.D.FAPD – Correct. FAPD is a lymphocytic cicatricial alopecia that has been recently described and has overlapping clinical and histopathologic features of FFA and androgenetic alopecia.E.DLE – Incorrect. Discoid lupus can cause a scarring alopecia characterized by violaceous atrophic plaques with follicular plugging and dyspigmentation. Histopathology shows interface dermatitis, basement membrane thickening, lymphohistiocytic infiltration around vessels and appendages, mucin deposition, and follicular keratotic plugs.Question 2: What are the classic trichoscopic findings in FAPD?A.Loss of follicular openings, hair diameter variability, and perifollicular hyperkeratosisB.Yellow dots and tapered hairsC.Loss of hair follicles, yellow dots, and dystrophic hairsD.Black dots, comma hairs, and scaleE.Arborising telangiectasia, dark brown pigmentation, and follicular plugsAnswers:A.D). Other findings may include perifollicular erythema, hair tufting, and predominance of single hair follicles.Loss of follicular openings, hair diameter variability, and perifollicular hyperkeratosis – Correct. The classic trichoscopic findings of FAPD include perifollicular hyperkeratosis, loss of follicular ostia, and hair diameter variability , D. OtheB.Yellow dots and tapered hairs – Incorrect. Yellow dots and tapered hairs are indicative of alopecia areata, which can also present with dystrophic and fractured telogen roots.C.Loss of hair follicles, yellow dots, and dystrophic hairs – Incorrect. Loss of hair follicles, yellow dots and dystrophic hairs are indicative of dissecting cellulitis, which can also present with follicular pustules at the borders.D.Black dots, comma hairs, and scale – Incorrect. Black dots, comma hairs, and scale are indicative of tinea capitis, which can also present with blotchy pigmentation.E.Arborising telangiectasia, dark brown pigmentation, and follicular plugs – Incorrect. Arborising telangiectasia, dark brown pigmentation, and follicular plugs are indicative of DLE.Question 3: Which of the following is not a recommended therapy for a patient with advanced FAPD at this time?A.Topical corticosteroidsB.Topical minoxidilC.HydroxychloroquineD.Hair transplantationE.Antiandrogen therapyAnswers:A.Topical corticosteroids – Incorrect. Topical corticosteroids are appropriate to use, however when used alone do not appear to affect hair loss but may improve symptoms of FAPD.B.Topical minoxidil – Incorrect. Topical minoxidil is a common treatment used in androgenetic alopecia and also to help thicken miniaturized hairs.C.Hydroxychloroquine – Incorrect. Hydroxychloroquine is commonly used in lichen planopilaris (LPP) and FFA. It has been reported to be successful in a cohort of 4 patients with FAPD in combination with topical clobetasol proprionate and topical 5% minoxidil.D.Hair transplantation – Correct. There is no good evidence to support successful treatment of FAPD with hair transplantation. LPP and FFA have been seen following this procedure in a cohort of 10 patients.E.Antiandrogen therapy – Incorrect. Antiandrogen therapy (cyproterone and finasteride) have been evaluated in case series used in combination with topical corticosteroids and topical 5% minoxidil to improvement hair regrowth in FAPD.None disclosed.


Escherichia coli bacteria [Biosensors are devices composed of a biorecognition part and of a transduction part. Natural biomolecules such as enzymes , antibodbacteria . A multibacteria . After abacteria . A reviebacteria .Non-contact and non-invasive techniques are used for monitoring some behaviors or some health problems. They are also classified in biosensing techniques. A non-invasive measurement of the arterial blood based on a diffused light model has been presented . Infra-r


Lactobacillus brevis NZ4 and Kluyveromyces marxianus SY11 (MFM), the dominant microorganisms isolated from traditional dairy products in western nan. The results indicated that MFM displayed better overall quality than the milk fermented with L. brevis NZ4 (LFM) and K. marxianus SY11 (KFM), respectively. MFM exhibited good sensory quality, more organic acid types, more free amino acids and esters, and moderate acidity and ethanol concentrations. Non-targeted metabolomics showed a total of 885 metabolites annotated in the samples, representing 204 differential metabolites between MFM and LFM and 163 between MFM and KFM. MFM displayed higher levels of N-acetyl-L-glutamic acid, cysteinyl serine, glaucarubin, and other substances. The differential metabolites were mainly enriched in pathways such as glycerophospholipid metabolism, arginine biosynthesis, and beta-alanine metabolism. This study speculated that L. brevis affected K. marxianus growth via its metabolites, while the mixed fermentation of these strains significantly changed the metabolism pathway of flavor-related substances, especially glycerophospholipid metabolism. Furthermore, mixed fermentation modified the flavor and quality of fermented milk by affecting cell growth and metabolic pathways.Traditional fermented milk from the western Sichuan plateau of China has a unique flavor and rich microbial diversity. This study explored the quality formation mechanism in fermented milk inoculated with  Western Sichuan presents unique microbial resources due to its specific geographical and climatic conditions. Naturally fermented yak yogurt contains abundant lactic acid bacteria (LAB) and yeasts, contributing a unique flavor and texture to fermented dairy products on the western Sichuan plateau of China. Screening and utilizing the dominant strains can help develop fermented dairy products with regional characteristics and provide nutritional enrichment.Kluyveromyces marxianus and Saccharomyces cerevisiae [K. unisporus (49.43%) and K. marxianus (26.44%) dominating in Xinjiang, and P. fermentans (41.89%), S. cerevisiae (27.02%), and K. marxianus (24.32%) being the most abundant in Qinghai. K. marxianus is widely present in traditional fermented dairy products. Screening and applying these yeasts as starters or co-starters can help develop new dairy products with local characteristics.Fermented dairy products are popular with consumers due to their rich nutritional value and unique flavor. Commercial fermented milk is mainly produced using LAB as a starter, while traditional products like milk dregs and kefir primarily use yeasts in addition to LAB, such as revisiae ,2,3. Ni revisiae  studied In milk, lactose represents the main carbohydrate. When LAB and yeast are simultaneously inoculated into fermented milk, they can utilize lactose metabolites in complementary ways. Therefore, mixed fermentation is widely used in dairy products ,6. The iL. casei M2-L2 and S. cerevisiae M4-Y2, revealing that mixed fermentation produced more long-chain fatty acids. Higher methionine, phenylalanine, tyrosine, and arginine levels were found in the mixed fermentation system, confirming that amino acids might promote LAB proliferation.Sun  used metTherefore, this study uses quality analysis and metabolomics technology to examine small- and medium-sized molecular metabolite production during LAB and yeast fermentation while analyzing the metabolic pathways involved in the fermentation process using the bioinformatics database  and asseL. brevis NZ4 (OQ660312) and K. marxianus SY11 (MZ144218) were screened from naturally fermented dairy products in western Sichuan, China. Milk was purchased from local pasture, while the UHPLC-Q Exactive HF-X was obtained from Waltham, MA, USA. The pure methanol, acetonitrile, formic acid, and propanol were supplied by Fisher from Waltham, MA, USA.L. brevis NZ4 and K. marxianus SY11 strains were inoculated into the milk and fermented at 36 °C and 28 °C, respectively, until set; after which, they were refrigerated at 4 °C as starters.The activated L. brevis NZ4, K. marxianus SY11, and L. brevis NZ4 + K. marxianus SY11 (1:1) were labeled LFM, KFM, and MFM, respectively. The samples were refrigerated at 4 °C for subsequent examination.The sterilized milk was inoculated with the starters at a 5% ratio and fermented at 35 °C until set. The fermented milks inoculated with Ten panelists were selected and trained to evaluate the tissue status, odor, and taste of the fermented milk. The acidity and ethanol content were determined according to the GB 5009.239-2016 standard of China and a method described by Wang , using pv:v)) containing 0.02 mg/mL of internal standard  was added for metabolite extraction. The extract was frozen and ground for 6 min , followed by low-temperature ultrasound extraction for 30 min . The sample was allowed to stand at −20 °C for 30 min and centrifuged for 15 min ; after which, the supernatant was collected for analysis. The sample metabolites were mixed at equal volumes to prepare a quality control sample (QC).Here, 200 mg of the fermented milk and grinding beads was placed in a centrifuge tube; after which, 400 μL of an extraction solution  was used to separate a 2 μL sample, which was subjected to mass spectrometric detection. Mobile phase A consisted of 95% water +5% acetonitrile (containing 0.1% formic acid), while mobile phase B comprised 47.5% acetonitrile +47.5% isopropanol +5% water (containing 0.1% formic acid). These procedures were performed using methods described by Meng .The mass spectrometric data were collected using a Thermo UHPLC-Q Exactive HF-X Mass Spectrometer equipped with an electrospray ionization (ESI) source operating in either positive or negative ion mode. The optimal conditions included a heating temperature of 425 °C, a capillary temperature of 325 °C, a sheath gas flow rate of 50 arb, an aux gas flow rate of 13 arb, an ion-spray voltage floating (ISVF) of −3500 V in negative mode and 3500 V in positive mode, and a normalized collision energy of 20–40–60 V rolling for MS/MS. The full MS resolution was 60,000, while that of MS/MS was 7500. The data were acquired in Data-Dependent Acquisition (DDA) mode, while detection occurred in a 70–1050 m/z mass range. These procedures were performed using methods described by Fang .The raw LC–MS data were imported into Progenesis QI metabolomics processing software  for baseline filtering, peak recognition, integration, retention time correction, and peak alignment. The mass spectrometric information was matched with the HMDB and Metlin databases to obtain the metabolite data. After searching the database, the data matrix was filled with missing values after removing them using an 80% rule. The response intensity of the sample mass spectrum peak was subjected to sum normalization to obtain the data matrix. The variables with a relative standard deviation (RSD) exceeding 30% in the QC samples were deleted, and the data matrix was obtained via log10 logarithmization for subsequent analysis.p-value of the Student’s t-test. VIP > 1 and p < 0.05 values indicated differential metabolites, which were annotated using the KEGG database to determine the related pathways. Python (Version 3.10) software package scipy.stats was used for pathway enrichment analysis, and Fisher’s exact test was employed to determine the most relevant biological pathways for experimental processing.The preprocessed matrix file was subjected to differential assessment using R software package ropls (Version 1.6.2) for principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA). Differential metabolite selection was based on the variable importance in projection (VIP) value obtained via OPLS-DA and the K. marxianus SY11 displayed a strong acid-producing ability. The low alcohol level in the fermented milk, ranging between 1 and 4 g/L, indicated that the yeast provided a mellow flavor to the fermentation system without causing health or other issues due to high alcohol content. The MFM exhibited a LAB level of 10.35 lg CFU/mL and a yeast content of 9.86 lg CFU/mL, both at a higher order of magnitude, showing that the two microorganisms exhibited excellent proliferation in the fermentation system. The sensory description results indicated that KFM exhibited an uneven, thin texture with bubbles, whey precipitation, an obvious alcoholic flavor, and a bland, sour taste. Therefore, K. marxianus SY11 alone was unsuitable for milk fermentation. LFM displayed a smooth, thick texture with a distinct sour aroma, showing that L. brevis NZ4 was appropriate as a common yogurt starter. MFM showed a smooth, thick texture with micro-bubbles and the moderate alcohol, ester, and sour aroma characteristic of fermented milk in western Sichuan, China. Therefore, combining K. marxianus SY11 and L. brevis NZ4 as a starter culture is suitable for producing local, uniquely flavored dairy products.The sensory description, acidity, ethanol content, and viable LAB and yeast count of the fermented milk are shown in L. brevis and K. marxianus for milk fermentation, while the acids reacted with the alcohol to increase the ester content.The volatile flavor compounds in the three kinds of fermented milk samples detected via GC–MS are shown in L. brevis and K. marxianus fermentation, while acetic acid is produced in large quantities [As shown in antities , which iThe amino acid determination results are shown in 2X values of the PCA model in positive and negative ion modes were 0.537 and 0.555, respectively, indicating high fitting accuracy. The three experimental samples showed significant clustering in the PCA plots in both positive and negative ion modes, indicating excellent repeatability and minimal differences in the sample group.The PCA is shown in Metabolomics detected a total of 9384 peaks in both positive and negative ion modes. Of these, 901 metabolites were annotated via the KEGG and HMDB databases. After removing data with missing values > 20% and QC validation RSD values < 30%, 885 metabolites were ultimately annotated, including 627 for positive ions and 258 for negative ions. KEGG annotated a total of 295 metabolic products, mainly including phospholipids (20 types), fatty acids (14 types), carboxylic acids (12 types), and glycerol phosphate choline (12 types). HMDB annotated a total of 667 metabolites, primarily including lipids (282 species), organic acids (136 species), oxygen-containing organic compounds, such as monosaccharides (70 species), and organic heterocyclic compounds, such as nucleic acids, vitamins, antibiotics (69 species), and benzoids (36 species).2X value of 0.7177, a cumulative R2Y value of 0.991, and a cumulative Q2 value of 0.4027. Negative ion mode produced a cumulative R2X value of 0.754, a cumulative R2Y value of 0.988, and a cumulative Q2 value of 0.3492, indicating that the PLS-DA models displayed excellent fitting and high predictive ability.PLS-DA was used to analyze the differences between the samples, as shown in p-value. VIP values > 1 and p < 0.05 indicated differential metabolites, of which 204 were evident between MFM and LFM, 163 between MFM and KFM, and 151 between LFM and KFM. The metabolites related to the fermented milk flavor  were screened. The 50 most abundant metabolites were selected to prepare the cluster diagram, as shown in OPLS-DA was used to obtain the VIP and Student’s test The relevant metabolites were searched in the KEGG database and enriched in the KEGG pathway. As shown in As shown in L. brevis and K. marxianus fermentation to prepare alcohol-flavored fermented milk.Fermented dairy products are highly popular with consumers worldwide. Yeast can change the quality during the fermentation and ripening of these products. Its high protein decomposition ability can produce flavor-affecting precursors as well as alcohol and carbon dioxide that reduce the pH and inhibit harmful bacterial growth ,23. Ther10 CFU/mL. The yeast content was significantly higher in MFM than in KFM, while the ethanol level was lower. This may be because the ethanol in MFM is utilized by LAB or esterified with other substances. Some studies have shown that ethanol and lactic acid esterification can reduce the damage to their own cells to a certain extent [K. marxianus significantly promoted L. casei proliferation, while L. lactis substantially encouraged K. marxianus growth.MFM and LFM displayed high LAB levels at about 10n extent . The typn extent ,26. LAB n extent  revealedK. marxianus. Since the amino acids produced by K. marxianus were used by L. brevis to maintain growth and metabolism, the free amino acid content was slightly lower in MFM than in KFM. The analysis of the basic physical and chemical indexes showed that L. brevis and K. marxianus eventually improved the quality of fermented milk by promoting cell growth and causing changes in other indexes.The free amino acid content was significantly higher in MFM and KFM than in LFM. The nutritional factors required for LAB growth were relatively abundant, with many amino acids consumed during fermentation. Moreover, the free amino acid content was low in LFM since its protein hydrolase enzyme system was not as rich as MFM, which exhibited a higher amino acid level due to protein hydrolyzation by L. brevis metabolism provided the carbon source for K. marxianus growth. Kadyan [L. plantarum and K. lactis metabolites during single and mixed fermentation. The results indicated a lower sugar content in the mixed fermentation system, possibly improving the sugar utilization rate. Liu [L. paracasei and K. marxianus fermentation significantly changed sucrose metabolism, glycolysis, and other pathways by stimulating sugar utilization by microorganisms. The flavor metabolites in MFM included N-acetyl-L-glutamic acid, cysteine-serine, glaucarubin, maleic acid, and serotonin, with fatty acid metabolism-related substances being the most abundant at a total of 97 types. Tian [K. maximus lipase activity was the highest, and speculated that yeast participated in protein and fat metabolism, accelerating decomposition and producing various fatty and amino acids. This represents the main source of many flavor substances in fermented foods [The flavor substance sources in fermented milk are mostly represented by fat, lactose, and protein as precursors, producing amino acids, sulfur-containing amino acids, peptides, lipids, sugars, and other substances under the action of microorganisms. Since these substances further constitute the flavor components in fermented milk , amino a. Kadyan  used nonate. Liu  revealedes. Tian  tested ted foods . Other sed foods .The selected flavor-related differential metabolites in the KEGG metabolic pathway showed that glycerophospholipid metabolism displayed the highest enrichment. Eight test group substances were enriched into the pathway. Glycerophospholipid metabolism included two sub-pathways, namely phosphatidylcholine and phosphatidylethanolamine biosynthesis, while the substances in the test group were mainly enriched in phosphatidylethanolamine biosynthesis, including four compounds, as shown in (L. sanfranciscensis)Lactobacillus sanfranciscensis  and (S. cerevisiae)Saccharomyces cerevisiae  to ferment sourdough, followed by a decline. It is speculated that the two microorganisms consume a large number of amino acids during the initial fermentation stage, while the lower pH in the dough during the later fermentation stage increases the protease activity in the flour, promoting amino acid degradation. Furthermore, S. cerevisiae and L. sanfranciscensis were exposed to metabolic activity, such as protein hydrolysis, where the protein was decomposed into amino acids, increasing the amino acid content in the dough [Of the other metabolic pathways noted in this experiment, arginine biosynthesis, beta-alanine metabolism, arginine and proline metabolism, and benzoxazinoid biosynthesis exhibited a higher abundance, most of which were related to amino acid synthesis and metabolism. Yang  revealedhe dough ,45.L. brevis and K. marxianus fermentation promotes the metabolism of milk proteins, carbohydrates, and fats. The significant enrichment of differential metabolites in the glycerophospholipid metabolism pathway can explain the mechanism behind the changes in fermented milk quality caused by mixed fermentation.In conclusion, mixed Lactobacillus brevis NZ4 and Kluyveromyces marxianus SY11, the dominant microorganisms isolated from traditional dairy products. MFM displays a better overall quality with good sensory characteristics, more organic acid types, higher levels of free amino acids and volatile substance esters, and lower acidity and ethanol concentrations than LFM and KFM. The differential metabolites in MFM are mainly enriched in pathways such as glycerophospholipid metabolism, arginine biosynthesis, beta-alanine metabolism, arginine, and proline metabolism. The mixed fermentation of these two strains significantly changes the flavor-related substance metabolism pathway. The results will pave the way for industrial production and flavor regulation of local special fermented milk.This study reveals the quality formation in fermented milk inoculated with


Plasmodium knowlesi infection was confirmed by PCR, and treatment with atovaquone/proguanil brought successful recovery. We review the evolving epidemiology of P. knowlesi malaria in the Philippines, specifically within Palawan Island.A 55-year-old man sought treatment for an uncomplicated febrile illness after returning to Canada from the Philippines. A suspected diagnosis of  Results of abdominal computed tomography were unremarkable and of a single-target Plasmodium falciparum histidine-rich protein 2 rapid diagnostic test were negative. Peripheral blood thin smear demonstrated variable intraerythrocytic parasite morphology, including band-like forms suggestive of P. malariae (<0.1% parasitemia) .In February 2023, a 55-year-old man sought care at the emergency department of Vancouver General Hospital, Vancouver, British Columbia, Canada, for daily fevers, headache, and abdominal pain 5 days after returning from a 3-week trip to the Philippines. He stayed mostly in Manila but spent 4 days on Palawan Island in the western Philippines 4 days before his return to Canada; he had not taken malaria chemoprophylaxis. Bloodwork was notable for platelet nadir of 48 × 10P. knowlesi, given the patient’s travel history and blood smear morphology, and subsequently confirmed the infection via species-specific laboratory-developed PCR. Amplicon sequencing on a 118-bp sequence also confirmed P. knowlesi identification. The patient was afebrile at follow-up 4 days after drug therapy, with resolution of thrombocytopenia and symptoms.We suspected a diagnosis of P. knowlesi malaria cases within the Philippines have been concentrated in Cebu Province and, as in this case, Palawan Island , the only monkey species naturally found in the Philippines and natural host reservoirs for P. knowlesi . However, the true burden is likely underestimated, given the lack of routine molecular testing for species confirmation in the Philippines. Because early ring-form trophozoites of P. knowlesi can resemble P. falciparum and developing band-like trophozoites can resemble P. malariae . AP. knowlesi erythrocytic cycle is shortest among Plasmodium spp., which may contribute to rapid development of high parasitemia  to endemic Sabah raises concern of P. knowlesi becoming a predominant species in future years. Ongoing investigations into mosquito behavior implicated with cross-species transmission will help inform appropriate control strategies for P. knowlesi and other simian species . Because of overlapping microscopy features with P. falciparum and P. malariae, molecular confirmation is required to enable early diagnosis and appropriate treatment. Despite gains in control of P. falciparum and P. vivax infection in Southeast Asia, the zoonotic nature of P. knowlesi and rise in cases highlight the need for tailored prevention and control strategies.In summary, Plasmodium knowlesi infection in traveler returning to Canada from Philippines, 2023.More information for