The number of metastatic lymph nodes (LNs) and the ratio of metastatic LN (LR) have been reported as predictors of recurrence in papillary thyroid carcinoma (PTC), while the role of LR or the number of metastatic LNs on the clinical response remains unclear.We aimed to compare the prognostic value of LR and the number of metastatic LNs on clinical response in PTC.A total of 384 PTC patients with LN metastases were enrolled in this study, all of whom underwent total or near total thyroidectomy and subsequent radioiodine ablation.After a mean follow-up of 25.7 months, response to initial therapy was classified as excellent response (ER), indeterminate, biochemical incomplete or structural incomplete response. The scatter diagram and receiver operating characteristic (ROC) curve were respectively employed to identify and compare the clinical value of the number of metastatic LNs and LR for predicting ER in different number of dissected LNs (DLNs). Multivariate analyses were further performed to explore the indicator for ER.P = .02), whereas it turns opposite in those with >10 DLNs. In the multivariate analysis, LR  rather than the number metastatic LNs  was an independent indicator for ER in addition to preablative-stimulated thyroglobulin  among patients with ≤10 DLNs. Although in patients with >10 DLNs, the number of metastatic LNs  turned to be independent factor for ER, apart from ps-Tg  and sex .ER tended to be more concentrate in patients with lower LR and lower number of metastatic LNs in scatter diagram. Although in patients with ≤10 DLNs, LR presented higher area under the ROC curve than the number of metastatic LNs in predicting ER (LR: 0.687, LNs, 0.556, LR appears to be a better negative predictor for ER than the number of metastatic LNs in PTC patients with ≤10 DLNs, whereas the number of metastatic LNs is superior to LR in those with >10 DLNs. Our recent research indicated that 10 or more dissected LNs (DLNs) might reflect adequate intraoperative evaluation and carry incremental value in achieving better response. Although for LR, which refers to the involved ratio of metastatic LNs, it may afford more information regarding the intraoperative evaluation range. One Korean study showed that LR is a significant risk factor for predicting poor prognosis. In 2015 American Thyroid Association (ATA), a new therapeutic response system is proposed to specify the recurrence and mortality in patients after both total/near total thyroidectomy and radioiodine (RAI) therapy, which appears to be a more objective ongoing assessment in predicting the clinical prognosis. It remains uncertain whether LR holds more value for both therapeutic response and clinical prognosis in PTC patients when comparing with the number of metastatic LNs. This study aimed to elucidate the predictive value of LN metastasis for clinical outcome in terms of LR and the number of metastatic LNs.Lymph node (LN) metastasis occurs in up to 90% of patients with papillary thyroid carcinoma (PTC).22.1We retrospectively studied a total of 985 patients diagnosed with PTC, patients underwent postoperative RAI ablation between October 2012 and December 2014 in Chinese Academy of Medical Science, all of whom underwent total or near total thyroidectomy with LN dissection. Among them 601 patients were excluded because of any of the following reasons: with initial distant metastases; with no evidence of LN metastasis on histology; (3) inadequate follow-up information for at least 6 months/incomplete follow-up records after 1st RAI therapy. Furthermore, written consent was obtained from the patients in the present study.2.2Total or near total thyroidectomy was usually performed in patient who meets either of the follows: primary tumor >1 cm; presence of multiple or bilateral lesions; extrathyroid extension; suspicious metastatic LNs during the preoperative evaluation or surgery. RAI ablation was employed following levothyroxine (LT4) withdrawal or with no LT4 therapy after surgery, accompanied with a low-iodine diet for at least 2 weeks as per our normal clinical protocol. Dose of RAI ranged from 1.11 GBq (30 mCi) to 5.55 GBq (150 mCi) was delivered according to tumor stage and likelihood of recurrence as per risk stratification. Routine medical examinations before RAI ablation included serum thyrotropin (TSH), preablative-stimulated thyroglobulin (ps-Tg), anti-Tg antibody (TgAb), chest computed tomography (CT), neck ultrasound (US), and diagnostic RAI whole-body scan (DxWBS), if necessary. Posttherapy RAI whole-body scan was performed 5 to 8 days after therapeutic RAI administration.2.3After initial therapy, patients took LT4 for TSH suppression after 3 days. Patients also received regular follow-up with examinations, such as neck US and DxWBS as well as serum-stimulated thyroglobulin level with TgAb performed 6 to 12 months after RAI therapy. Additional diagnostic imaging studies, such as chest CT and whole body fluoro-deoxyglucose-PET were performed if needed. Considering the status of LN evaluation, patients were divided into 2 cohorts according to the number of DLNs.2.4 Patients who achieved ER should accord with the criteria below: either suppressed Tg <0.2 ng/mL or stimulated Tg <1 ng/mL, US without evidence of disease, and negative cross-sectional and/or nuclear medicine imaging (if performed). The criteria for IDR is defined that if patients had any of the following: nonspecific findings on imaging studies with faint uptake in thyroid bed on RAI scanning, suppressed Tg ≥0.2 ng/mL but <1 ng/mL, stimulated Tg ≥1 but <10 ng/mL, or TgAb stable or declining in the absence of structural or functional disease. BIR was defined with either of the following criteria: negative imaging and suppressed Tg >1 ng/mL, or stimulated Tg >10 ng/mL, or rising TgAb values. SIR represented the patients who had structural or functional evidence of disease with any Tg level and/or TgAb.Patients response after initial RAI ablation were classified into 4 types: excellent (ER), indeterminate (IDR), biochemical incomplete (BIR), and structural incomplete (SIR) according to the reclassification system newly proposed by ATA.2.5 Multivariate logistic regression analyses were further analyzed among patients with ≤10 or >10 DLNs, to find independent prognostic factors for ER, including sex, age, T stage, N status, LR, the number of metastatic LNs, ps-Tg, and dose of RAI. All tests were 2 sided, and a P value <.05 was considered statistically significant. Statistical analyses were performed using the SPSS version 22.0 and MedCalc version 14.8.1.Data were presented as median rang or mean ± standard deviation. Scatter diagram was used to identifying the patients who showed ER under different LR and number of metastatic LNs in different context of DLNs (≤10 or >10). Moreover, receiver operating characteristic (ROC) curve was applied to determine the area under the curve (AUC) to evaluate the value of LR and the number of metastatic LNs in predicting ER in different context of DLNs. Comparison of ROC curve was performed using the method of Hanley and McNeil for the same set of patients.33.1Characteristics of the patient are summarized in Table 3.2P = .001) was an independent negative indicator for ER in addition to Ps-Tg , ps-Tg , and sex  were demonstrated to be the independent predictive factors for ER (Table For this part of patients, who presented ER continued to rise over the course of LR and the number of LNs increasing Fig. B. Contra4,13 Surgical dissection and subsequent selective RAI ablation are therefore recommended as the 1st line protocol, with an aim to reduce regional recurrence in patients with intermediate or high risk of recurrence.,15 Still, even after initial treatment, cervical recurrence could occur in up to 30% of patients. Invasive extent of LN metastases including the number of metastatic LNs and LR has been shown to be 1 of poor factor for recurrence. Study from Leboulleux et al revealed that the 10-year locoregional recurrence rate would rise from 3%, 7 to 21% among patients with the number of metastatic LNs <5, 6 to 10 and >10, respectively. Sugitani I et al reported that the recurrence rate of patients with any metastatic LN >3 cm would be 27%. Based upon these studies, the 2015 ATA guideline 1st takes the number of metastatic LNs and size of LN metastases into account when stratifying the recurrence risk of patients, while without mentioning the context of DLNs. LR has recently emerged as an alternative prognostic indicator, which compromises both the number of metastatic LNs and DLNs. A recent report indicated that the 5-year locoregional recurrence-free survival were 97.1% and 78.8% in patients with LR below or above 0.22, respectively. The range of the neck dissection has been the concern of the surgeons, our recent research indicated 10 or more DLNs seems to carry incremental value in achieving better response. About 10, DLNs, therefore, could be regarded as a reference for proper number for intraoperational LN evaluation. To our knowledge, no reports have valued LR and the number of metastatic LNs in predicting clinical outcome, especially in terms of the new clinical response system raised by 2015 ATA guidelines. In this study, for the 1st time, we tried to compare the prognostic impact between LR and the number of metastatic LNs on clinical response under different LN dissection yield.Though carrying a good 10-year survival rate at 93%, an increased risk of recurrence and compromised survival rate can be observed in PTC patients with nodal involvement.Our study finds that ER tends to occur in lower LR or less number metastatic LNs in regardless the number of DLNs. Moreover, it is worthy to mention that LR, rather than the number of metastatic LNs, is a prognostic indicator for ER in patients with less than 10 DLNs. Conversely, in patients with the number of DLNs more than 10, the number of involved LNs manifested leading role in predicting ER. Multivariate analyses also suggested that the number of involved LNs, rather than LR, became an independent factor for ER. What may contribute to the difference between the 2 circumstances? One possible explanation seems that more DLNs may provide a kind of ‘more adequate LN evaluation,’ so that the number of metastatic LNs could play an informative and objective role in presenting the actual extent of LNs metastasis. In contrast, in patients with fewer DLNs, the number of metastatic LNs might be less objective because of relatively more uncertainty remained in those unresected LNs, in this case, LR appears to be a more comprehensive indicator taking both the number and the extent of LN metastasis into consideration. Thus, this system in combination of LR and the number of metastatic LNs might be helpful for evaluating both LN management and optimal postoperative prognostic system for PTC. ps-Tg is also an independent factor for ER, regardless of the LN dissection yield. Furthermore, age is also an independent factor when more LNs are evaluated, which implies that the older patients are easy to exhibiting non-ER. This is inconsistent with Chéreau et al, who have demonstrated that the patients with advanced age have a negative effect on prognosis. In addition, we observed that the dose of RAI ablation was not significantly related to clinical response in our population, which might provide more evidence to reinforce that 30 mCi is effective enough for remnant ablation in intermediate risk patients.Like the results indicated in our previous work,Although we enrolled and analyzed 384 patients, this study still had several limitations related to sample size as well as its retrospective nature. In addition, number of metastatic LNs detected is dependent on the preoperative assessment, surgical skills, and surgery management strategies.5In conclusion, LR may serve as a better marker for an alternative staging system than the number of LNs in patients with ≤10 DLNs, whereas the number of metastatic LNs models demonstrated superior prognostic value to LR in those with >10 DLNs. Our findings suggest a comprehensive LN evaluation combining both LR and number of metastatic LNs, which might be a new optimal prognostic system for PTC.


Nuclear industrial accidents and the detonation of nuclear devices cause a variety of damaging factors which, when their impacts are combined, produce complicated injuries challenging for medical treatment. Thus, trauma following acute ionizing irradiation (IR) can deteriorate the IR-induced secondary reactive metabolic and inflammatory impacts to dose-limiting tissues, such as bone marrow/lymphatic, gastrointestinal tissues, and vascular endothelial tissues, exacerbating the severity of the primary injury and decreasing survival from the exposure. Previously we first reported that ghrelin therapy effectively improved survival by mitigating leukocytopenia, thrombocytopenia, and bone-marrow injury resulting from radiation combined with burn trauma. This study was aimed at investigating whether radiation combined with burn trauma induced the cerebro-vascular impairment and intracranial hemorrhage that could be reversed by ghrelin therapy. When B6D2F1 female mice were exposed to 9.5 Gy Cobalt-60 γ-radiation followed by 15% total skin surface burn, cerebro-vascular impairment and intracranial hemorrhage as well as platelet depletion were observed. Ghrelin treatment after irradiation combined with burn trauma significantly decreased platelet depletion and brain hemorrhage. The results suggest that ghrelin treatment is an effective therapy for ionizing radiation combined with burn trauma. The adverse effects of ionizing irradiation (IR) to organs and systems have been known since the early 20th century. However, current public health concerns related to radiation exposure are based upon the extensive development and proliferation of nuclear technologies, radiation and nuclear medicine, and nuclear weapon systems, which overall increase the risk of radiation accidents as well as the imminence of nuclear military conflicts. Therefore, it elevates the frighteningly real possibility of mass casualties from terrorist acts with improvised nuclear and radiological devices.It is widely accepted that irradiation per se at any given dose can affect each biological system. However, there are “limiting” doses, which trigger acute radiation sickness . The acute sickness occurs and propagates due to direct cell lesions and cascades of indirect reactive responses that can ultimately lead to the impairment of sensitive tissues sustaining crucial immunochemical and metabolic homeostasis, breach of biological barriers, post-irradiation sepsis and multiple organ failure (MOF) ,5,6,7,8.in vivo ghrelin production is shown to be dramatically affected by irradiation and CI [The emerging threat from acute radiation exposure evolves a need for the development of new countermeasures and modalities effective in response to and management of radiation injury . In recen and CI . Thus, sn and CI ,26,27,28The data presented in this report demonstrate a decrease in incidents of intracranial hemorrhage in CI animals treated with ghrelin compared to CI animals treated with vehicle that sustains the main hypothesis.Gross pathology assessments of skulls and brains obtained from B6D2F1/J mice revealed that all moribund animals subjected to CI experienced intracranial hemorrhages that varied in the extent, grade, hemorrhage type, and depth of lesions. The scope of intra-axial and extra-axial lesions included (i) intra-parenchymal hemorrhage; (ii) subdural hemorrhage; (iii) subarachnoid hemorrhage; and (iv) hemorrhage to the cranial and the mandibular bones of the skulls . ProjectLevel 1—Characterized by the appearance of isolated intra-parenchymal and subarachnoid hemorrhage in a form of hemorrhagic petechiae observed predominantly in cerebellum, medulla oblongata, pons, midbrain, cerebral cortex, and corpus callosum. The surface of the hemorrhagic lesions constituted less than 1% of the total surface of the brain.Level 2—Characterized by the presence of numerous hemorrhagic petechiae observed predominantly in cerebellum, medulla oblongata, pons, the midbrain, cerebral cortex, corpus callosum, striatum, thalamus, hypothalamus, and the olfactory bulb. The surface of the hemorrhagic lesions constituted 1% to 3% of the total surface of the brain.Level 3—Characterized by the presence of subarachnoid purpural hemorrhage and deep intra-parenchymal purpural hemorrhage observed predominantly in the hindbrain, the midbrain and the olfactory bulb; the presence of numerous hemorrhagic petechiae in the entire structures of the telencephalon and the diencephalon was determined. The surface of the hemorrhagic lesions constituted 3% to 10% of the total surface of the brain.Level 4—Characterized by the development of deep ecchymotic/confluent intra-parenchymal hemorrhage that occurred in the hindbrain, midbrain, and the entire structures of the telencephalon and diencephalon. Moreover, the presence of confluent hemorrhagic lesions in the subdural and subarachnoid area, pia matter, as well as a presence of intraventricular hemorrhage and hemorrhage to the cranial and the mandibular bones of the skull were observed. The surface of the hemorrhagic lesions constituted over 10% of the total surface of the brain.Representative images of the dorsal projections of the skulls and brains with extra-axial and intra-axial hemorrhage are shown in All the previously published data indicated that ghrelin therapy for CI significantly mitigated hematopoietic injury and decreased the mortality rate of the animals . The purData obtained from the gross pathology and analysis of the daily tally of IHS-ratios for the CI animal cohorts were plotted and are presented in 6 cells·mL−1; sham + vehicle: 1226 ± 59 vs. CI + vehicle: 276 ± 90; p < 0.05). Treatment with ghrelin mitigated this depletion and significantly elevated platelet counts back to 487 ± 45 × 106 cells·mL−1 (p < 0.05).CI is known to induce platelet depletion . As showThe intention of this report is to demonstrate that the acute sickness and moribundity that developed in the B6D2F1/J mice due to CI were associated with intracranial hemorrhage. Administration of ghrelin injected sequentially three times  after CI mitigated sickness, moribundity and incidents of intracranial hemorrhage. To our knowledge, this is the first report indicating the mitigative effect of ghrelin against microvascular injury in brain of B6D2F1/J mice subjected to IR combined with skin burn trauma.It is well documented that damage to the microvascular networks constitutes one of the most important outcomes in the pathogenesis of sickness due to total or partial body IR ,6,21,29.The existing data on CI models indicate that the administration of ghrelin to CI rats and mice can increase their survival seemingly due to the mitigation of bone marrow injury and the suppression of neuronal systemic inflammatory effects ,23. ThusOur observations indicated that the development of intracranial hemorrhage in the B6D2F1/J strain occurred at one or another grade in all cases of moribundity and animal death. The role of hemorrhagic lesions in the pathogenesis of the disease and the mortality rate requires further investigation. However, it would be reasonable to assume that the administration of ghrelin can mitigate the thrombopoietic effects of CI ; therefoTo address the growing demand for R&D of prospective remedies for the management of vascular brain injury, we have proposed a mouse model (B6D2F1/J mice) of CI-induced intracranial hemorrhage and tested the influence of ghrelin on incidents of brain hemorrhagic lesions. The results from our experiments indicate that besides the previously reported systemically beneficial effects and improvement of wound healing ,23,34, tn = 10–11 mice per group.B6D2F1/J female mice were randomly divided into three groups: (1) sham + vehicle, (2) CI + vehicle, and (3) CI + ghrelin. Each group received topical gentamicin cream and were administered with oral levofloxacin. Survival experiments were repeated one time with ad libitum. They were acclimated about eight days before use for experiments. The animal holding rooms had a temperature of 21 °C, 10% relative humidity, and 12 h lighting cycles (6 AM light to 6 PM dark).B6D2F1/J female mice at 12–20 weeks of age were used because male mice are aggressive and will chew each other resulting in unnecessary physical injury that is beyond the wound injury described in the experimental protocol . The female mice were obtained from the Jackson Laboratory . They were sent to AFRRI and kept in plastic microisolator cages bedded with hardwood chip. They received commercial rodent chow and acidified tap water −1), with the field uniformed within ±2%. The mapping data determined each radiation exposure time. The cobalt-60 decay was corrected. Differences in the mass energy absorption coefficients for soft tissues and water were taken into consideration. An ionization chamber adjacent to the mouse rack was used to verify the accuracy of the actual dose delivered to the animals. The verification of dose was calculated aiming the midline soft tissue of animals.The procedure of irradiation has been described previously . Briefly−1 of acetaminophen  and 0.05 mg·kg−1 of buprenorphine immediately after skin injury to alleviate pain. Four hours later, mice were given a second dose of 150 mg·kg−1 of acetaminophen. Skin-wounded only mice received only one dose of 150 mg·kg−1 of acetaminophen immediately after skin injury; sham-operated mice only received one i.p. injection of 0.5 mL sterile 0.9% NaCl.The procedure of skin burn has been described previously . Briefly−1 were administered by lateral tail-vein injections [Ghrelin was purchased from Phoenix Pharmaceutical . Three doses of 113 μg·kgjections  in a vol−1 in 0.2 mL per mouse, was administered per os (p.o.) daily for 14 days on days 3–16. Briefly, a 500 mg tablet was crushed by mortar and pestle. The LVX in the powder was dissolved in a volume of sterile water approximately one-third of the total volume required to prepare the concentration needed for the average body mass of the mice to be treated. The mortar was rinsed with the remaining two-thirds volume of sterile water. The combined suspension was centrifuged to remove the particulate filler and the supernatant solution was passed through a 0.45 μm membrane filter into a sterile amber bottle, which was sealed with a sterile rubber stopper and stored at 4–8 °C.Gentamicin sulfate cream, 0.1% , was applied daily for 10 days to the skin injured area on days 1–10. Levofloxacin (LVX), , 100 mg·kgEntire mouse skulls were collected at the end-point of observations, preserved in 10% neutral buffered formalin, and then the brains were extracted from the cranial cavities and subjected to routine gross pathology. The severity of intracranial hemorrhage  was scored with the provision of the scoring system for hemorrhage assessment previously introduced by Yelverton . In brieThe IHS-ratio for each CI-experimental group at each observation day was then calculated as follows: cumulative IHS per group  was divided by 10 . The calculated data were presented in a histogram of IHS-ratio versus observation days.Intact mouse brains fixed in 10% neutral buffered formalin were obtained as described in the previous paragraph. The brains were subjected coronal sectioning as shown in 6 cells·mL−1.Blood samples collected in EDTA tubes were assessed with the ADVIA 2120 Hematology System . Platelet counts were expressed as mean ± SEM. with units of 10Parametric data were expressed as the mean ± SEM. For each survival experiment, 20–22 mice per group were tested on an individual basis. IHS-ratios for CI-vehicle and CI-ghrelin animal groups over 30 days of observation were compared using the one-way ANOVA and Tukey post hoc test. Platelet data were analyzed with Student’s t-test. A significant level was at or less than 5%.In conclusion, we show that ionizing radiation followed by skin surface burns induces cerebro-vascular impairment and intracranial hemorrhage as well as platelet depletion. The results suggest that platelet depletion probably partly contributes to intracranial hemorrhage, thus this intracranial hemorrhage partly leads to the final mortality. Treatment with ghrelin after irradiation combined with burn trauma significantly decreased platelet depletion and brain hemorrhage. To clarify the role of ghrelin in mitigating the CI-induced brain inflammation process and coagulation systems requires further exploration. However, taken together, the results presented in this report and the data in literature show that ghrelin treatment is an effective therapy for combined radiation injury.


Escherichia coli (n = 12) and Klebsiella pneumoniae (n = 7) tested for quinolone resistance-related genes qnrA, qnrB, qnrC, qnrS and qepA, four each of E. coli and K. pneumoniae possessed qnrS, and another E. coli isolate possessed a new variant of qepA. This is the first detection of qepA in environmentally dwelling bacteria in Bangladesh.Of 19 environmental  Enterobacteriaceae. For years, quinolone resistance was believed to be solely chromosomally mediated through nucleotide substitutions in the quinolone resistance-determining region of DNA gyrase (gyrA and gyrB) and topoisomerase IV (parC and parE), which are the main target molecules of the quinolones [qnrA (currently qnrA1), encoding a protein that protects type II topoisomerases from quinolones, was first reported in 1998 [qepA and oqxAB [Acquisition of resistance towards several classes of antibiotics and the spread of multi-drug resistant bacteria is of immense public health concern globally. Increased use of quinolones, a family of synthetic chemical agents and one of the most commonly prescribed broad-spectrum antimicrobials, in recent years have coincided with the increasing incidence of resistance to this drug among inolones . However in 1998 . Thereaf in 1998 . Another in 1998 . A thirdnd oqxAB , 6.qepA named qepA1, qepA2 and qepA3 have been reported [Enterobacteriaceae. Since aquatic environment serves as a reservoir for diverse microbial populations including bacteria carrying PMQR determinants [qnrA, qnrB, qnrC, qnrS and qepA in members of the family Enterobacteriaceae, namely E. coli and K. pneumoniae, isolated from a lake and a river of Dhaka city, Bangladesh. Both bacteria demonstrated frequent presence of qnrS, and one of the E. coli isolates possessed a new sequence variant of qepA designated as qepA4.QepA was found to be associated with decreasing susceptibility, and 8 to 32-fold increase of minimal inhibitory concentration values of the hydrophilic quinolones such as norfloxacin, ciprofloxacin and enrofloxacin . So far,reported , 7, 8. Areported . All of rminants , selectiE. coli and seven K. pneumoniae were isolated from water samples collected from a lake and a river of Dhaka city. After preliminary identification by standard culturing methods, all isolates were confirmed using API 20 E  strips. Antimicrobial susceptibility was determined by the Kirby-Bauer disk diffusion method according to the Clinical Laboratory Standards Institute recommendations. Commercially available antibiotic discs (Oxoid) of nalidixic acid (30 µg), ciprofloxacin (5 µg) and levofloxacin (5 µg) were used in the antibiotic susceptibility assay. Bacterial DNA was obtained from all isolates by the standard boiling method. Detection of PMQR genes  was performed by polymerase chain reaction (PCR) using the primers as described elsewhere [Twelve lsewhere , 11. In  using overlapping PCR amplicons. The amplified fragments of qepA were then sequenced using an ABI PRISM BigDye Terminator Cycle Sequencing Reaction kit (Applied Biosystems) on an ABI PRISM 310 automated sequencer (Applied Biosystems). The deduced full length sequence of the gene was submitted to GenBank (Accession Number KX580704).F3-TGTTCACCATCGGCAACGA, R3-CACCTTCACCAAGACCACG; all listed 5′–3′) targeting different regions of Enterobacteriaceae.Nearly half of the isolates in the present study were found to be resistant to quinolone antibiotics such as nalidixic acid (52.6%), ciprofloxacin (47.4%), and levofloxacin (47.4%). Among the 19 isolates, ten were resistant to at least one of the tested quinolones. Of the remaining nine isolates, four showed intermediate resistance to nalidixic acid, while one of the ten quinolone-resistant isolates exhibited intermediate resistance to levofloxacin. As quinolones are broad-spectrum antibiotics, people use them for wide ranging infections in Bangladesh. Morgan et al.  reportedE. coli and K. pneumoniae possessed qnrS, while another E. coli isolate possessed qepA. Unlike our study, most of the previous studies were based on clinical isolates and the most frequent PMQR gene hitherto identified has been qnrB [qnrA and qnrC, were found in any of the isolates tested here. Five isolates carrying qnrS were found sensitive to all three quinolone antibiotics tested. Although strains carrying this gene did not exhibit resistance to the tested quinolone drugs, it is possible that qnrS might play a role in the resistance to other quinolones. Nonetheless, an environmental reservoir for the quinolone resistance related genes was demonstrated in the present study, supporting the results of an earlier study [All the isolates n = 19) were screened for PMQR genes by PCR. Of these, four each of  were screr study , which wE. coli isolates resistant to all three quinolone antibiotics tested in the present study. To our knowledge, this is the first time in Bangladesh that a qepA gene was found from environmentally occurring bacteria belonging to the family Enterobacteriaceae. The association of qepA with the rmtB gene contributing to high level resistance to aminoglycosides was reported from Belgium, China and Japan [E. coli showed an association of qepA and rmtB with a composite transposable element flanked by two copies of IS26 [E. coli isolate possessing qepA did not have rmtB, as confirmed by PCR. This result suggests the possibility of independent emergence of E. coli harboring qepA locally in the aquatic environment, and possibly of different ancestry from the ones carrying qepA and rmtB, as reported from other countries [QepA, a recently identified PMQR gene encoding an efflux pump , was detnd Japan , 14, 15.qepA revealed the gene to be different from the three alleles of qepA  that have been reported so far [qepA (Accession Number KX580704) revealed eight nucleotide differences that result in eight amino acid substitutions in the deduced 511 bp amino acid sequence  revealed separation of qepA4 from all qepA alleles reported so far, although it is most closely related to the qepA2 branch (Fig. rmtB gene noted above, suggests a different lineage for E. coli carrying qepA found in Bangladesh. This novel variant of a quinolone resistance determinant and the presence of other PMQR genes in naturally disseminating Enterobacteriaceae call for urgent surveillance to monitor and prevent the spread of these resistant bacteria in vulnerable populations of Bangladesh.In the present study, DNA sequencing of d so far , 7, 8. C


FADS1) shows that it is associated with the FA composition of plasma phospholipids and lipid metabolic indices among several ethnic groups. However, this association requires further confirmation in the Chinese population, and little is known about the effect of polymorphisms in fatty acid-related genes on body fat distribution.Genotyping of the rs174547 polymorphism in the fatty acid desaturase 1 gene (Anthropometric measurements of 951 Chinese adults aged 18–79 were obtained and body fat distribution was estimated using dual-energy X-ray absorptiometry. The FA composition of plasma phospholipids was measured by gas chromatography. Multiple linear regression assessed whether the rs174547 genotype was associated with FA composition, body fat distribution, and metabolic traits in additive, dominant, and recessive models.The rs174547 C minor allele was associated with a higher proportion of linoleic acid, lower arachidonic acid and docosahexaenoic acid, as well as lower delta-6-desaturase and delta-5-desaturase activity. Female C allele carriers had lower android fat percentages and lower levels of low-density lipoprotein-cholesterol, while male C allele carriers had lower gynoid fat percentages and higher triglyceride after adjusting for age, income, BMI, behavioral risk factors, and regional fat percentages.FADS1 rs174547 with the FA composition of plasma phospholipids was identified among this Chinese adult population. The association with body fat distribution and lipid metabolic indices differed between men and women, which might explain sexual differences in body fat distribution and lipid metabolism.An association of  FADS1) and 2 (FADS2) with changes in plasma FA profiles and altered desaturase activity [FADS1 rs174547, a T/C SNP in intron 9 of FADS1, was reported to be associated with arachidonic acid to linoleic acid (AA:LA) FA ratios in both Caucasians and Asians [Previous studies have identified associations between variations in genes encoding fatty acid desaturase 1  using stored blood samples.A total of 1029 community residents aged from 18 to 82 years old were recruited voluntarily between November 2008 and May 2009 from two communities in Hangzhou City, a capital provincial city in eastern China. Two communities were chosen for convenience from Xiacheng and Xiaoshan districts of the city. The subjects were recruited voluntarily by posters, flyers, and community leaders. All subjects completed a standardized questionnaire survey, underwent physical anthropometry and body composition measurements, and provided blood samples at the Obesity and Body Composition Research Center of Zhejiang University School of Public Health. In 2014, 67 participants were sampled with stratification on BMI distribution, and the measurement of plasma FA was made using their stored plasma samples. In 2015, after excluding the participants aged <18 or >80 years, the The flow of participants included in the analysis is shown in Fig. Weight and height were measured with subjects wearing light clothing and without shoes. Body weight was recorded to the nearest 0.1 kg . Body height was measured with a hypsometer to the nearest 0.1 cm. All values were recorded as the mean of three measures. WC was measured at the approximate midpoint between the lower margin of the last palpable rib and the top of the iliac crest, at minimal respiration. Hip circumference was measured at the maximum circumference of the buttocks with the tape parallel to the floor. Blood pressure was obtained from the right arm of seated participants after a 5-min rest. It was measured twice by the same trained examiners with a mercury sphygmomanometer according to a standard protocol ; the meaDXA  was implemented to measure body fat distribution as previously described . The bonInformation on demographics, socioeconomic status, and lifestyle behavior was collected by face-to-face interview using a structured questionnaire, including questions on age, sex, income, smoking status, alcohol consumption, and physical activity. Participants were classified according to their smoking status as nonsmokers or current smokers, and according to their alcohol consumption as non-drinkers, current drinkers, and abstainers. Physical activity was grouped as low, moderate, or heavy based on its frequency and intensity according to the international physical activity questionnaire , 23.After an overnight fasting, venous blood specimens were collected for biochemical analysis. Fasting plasma glucose (FPG) concentrations were determined using a hexokinase method. Total cholesterol (TC) and TG concentrations were measured using COD-PAP and GPO-PAP methods, and HDL-C and LDL-C were measured using an enzymatic homogeneous assay. The levels of apolipoprotein A1, and apolipoprotein B were determined by immunoturbidimetric assay. All the reagents were obtained from Roche. Then remaining samples were stored at −80 °C for further analysis. Genomic DNA was extracted from 1 to 5 mL whole blood using a commercially available DNA isolation kit  according to the manufacturer’s protocol between April and May 2015.For the analysis of FA composition, total lipids were extracted from 200 μl plasma using chloroform–methanol (2:1 v:v) according to the Folch method . The homt-test for continuous variables or the chi-square test for categorical variables. Pearson’s χ2 test was used to examine whether rs174547 genotype frequencies were in Hardy–Weinberg equilibrium (HWE). The associations between FADS1 rs174547 and plasma phospholipid FAs, body composition indices, and lipid profile indicators were identified by multivariate linear regression models, with each stage of regression analysis performed assuming additive (variant/variant vs. variant/common vs. common/common), dominant (variant/variant + variant/common vs. common/common), or recessive (variant/variant vs. variant/common + common/common) models. The effect value was determined from the model with the lowest P value.The statistical significance of demographic characteristics was analyzed using the FADS1 rs174547 and plasma phospholipid FAs in 67 participants, the first step was to adjust for sex and age, then to adjust for smoking, drinking, and BMI. For the models identifying associations between rs174547 and anthropometric measurements and lipid indicators in 951 participants, we adjusted the same covariates as above. We further adjusted the fat mass in other body regions to determine whether rs174547 independently influenced regional fat distribution. All analyses in the models of anthropometric measurements and lipid indicators were performed separately in men and women because of the interaction of gender. All statistical analyses were conducted using SAS software version 9.3 . A two-tailed value of P < 0.05 was considered statistically significant.For the models identifying associations between n = 67) are shown in Tables n = 951). For whole sample analysis, men showed higher levels of physical activity, alcohol consumption, and smoking prevalence than women, while no sex difference was observed in age, BMI, or household income. The effect allele of rs174547 was the C allele, which had a frequency of 0.343 and conformed to HWE (P = 0.117), with a genotyping success rate of 99.5%.The characteristics and rs174547 genotypes of the FA analysis sample  showed a decreasing trend with increasing numbers of C alleles of the respective genotype after adjusting for age. When further adjusting for covariates and BMI, the association of the C allele with WHpR and android FMP remained significantly negatively correlated, while the association with other indicators of fat distribution diminished. When including gynoid FMP in the model, the android FMP remained significantly associated with the C allele in the dominant model.The associations between the rs174547 genotype and lipid profiles among women and men are shown respectively in Tables FADS1 SNP rs174547 was significantly associated with several lipid metabolic indices and anthropometric indicators. We also confirmed an association between rs174547 and plasma phospholipid FA, and showed that rs174547 was associated with body fat distribution with a significant gender difference.In the present study, we found that the FA metabolism-related FADS gene cluster with plasma phospholipid FA in various ethnicities, and report the correlation of particular alleles with higher levels of LA and lower levels of AA [Several studies have previously explored the association of SNPs in the ls of AA , 7, whicls of AA ; howeverls of AA –28. Howels of AA . It has ls of AA –32, so tGender differences in body fat distribution are well-established , with woA major strength of our study is that it is the first to demonstrate the impact of this FA-related SNP on gender differences in body fat distribution in a Chinese adult population. Although the influence of rs174547 on BMI has been studied previously , fat disFADS1 rs174547 with plasma phospholipid FA profiles, anthropometric measurements, and lipid profiles in this Chinese adult population. The rs174547 C allele was shown to correlate with the level of plasma FA. The association between the rs174547 genotype and body fat distribution and lipid indicators differed between men and women, which might partially explain sexual differences in body fat distribution and lipid metabolism. Future research should investigate the effects of FADS1 and FADS2 on body fat distribution, especially in women.In summary, we investigated the relationship between


Vigna radiata), a widely used legume with promising nutritional and health benefits that was included in the experimental diet as raw or 4 day-germinated seed flour, and aerobic interval training protocol (65–85% VO2 max) has been tested in lean and obese Zucker rats following a 2 × 2 × 2  factorial ANOVA  statistical analysis. Germination of V. radiata over a period of four days originated a significant protein hydrolysis leading to the appearance of low molecular weight peptides. The combination of 4 day-germinated V. radiata and aerobic interval training was more efficient compared to raw V. radiata at improving the aerobic capacity and physical performance, hepatic histology and functionality, and plasma lipid parameters as well as reverting the insulin resistance characteristic of the obese Zucker rat model. In conclusion, the joint intervention with legume sprouts and aerobic interval training protocol is an efficient treatment to improve the alterations of glucose and lipid metabolism as well as hepatic histology and functionality related to the development of NAFLD and the MetS.Metabolic syndrome (MetS) is a group of related metabolic alterations that increase the risk of developing non-alcoholic fatty liver disease (NAFLD). Several lifestyle interventions based on dietary treatment with functional ingredients and physical activity are being studied as alternative or reinforcement treatments to the pharmacological ones actually in use. In the present experiment, the combined treatment with mung bean ( Metabolic syndrome (MetS) is a group of metabolic alterations characterized by central obesity, dyslipidemia, elevated plasma glucose, insulin resistance, elevated blood pressure, pro-thrombotic and pro-inflammatory state . PatientVigna radiata is a widely used legume for human consumption that exhibits an interesting composition of nutrients and bioactive compounds [V. radiata sprouts are usually commercialized to take part in numerous healthy dishes that are becoming increasingly popular for the general consumer.The development of strategies for the prevention and treatment of MetS includes changes in lifestyle such as caloric restriction, low fat and low glycemic index diets, consumption of foods rich in beneficial bioactive ingredients, and regular physical activity ,6. In thompounds ,13. Suchompounds . In factThe effects of different types of exercise on the treatment of MetS have been studied concerning the cardiovascular risk, and high-intensity aerobic interval training has been reported to be more effective at reducing such risk in rats with MetS than moderate-intensity continuous training did . With reDifferent in vivo experimental models have been used to test the beneficial effects of diet and exercise on several parameters of MetS. Among them, the obese Zucker rat is a widely accepted animal experimental model to study the multifactorial effects of diet and exercise on MetS associated conditions. This rat strain is known to present a genetic defect in leptin receptor that causes the development of hyperphagia leading to obese phenotype , and shaV. radiata as dietary source of bioactive compounds and hydrolyzed protein obtained using natural methods, together with an AIT protocol. Such joint intervention (diet plus exercise) in the above mentioned experimental animal model of the obese Zucker rat was an efficient strategy to improve the NAFLD condition related to the development of MetS.The aim of this study was to assess the effects on glucose and lipid metabolism parameters, as well as on liver histology and functionality, exhibited by a novel joint intervention combining the consumption of 4 day-germinated Vigna radiata L.  seeds were obtained from a commercial establishment and powdered (0.18 mm sieve) for sample analysis. Sprouting was done in seeds previously sterilized by immersion in sodium hypochlorite for 3 min and washed out with sterilized, type-2 water to eliminate any traces of sodium hypochlorite. The seeds were then left soaking in water during 8 h and distributed in trays over sheets of filter paper where they were left covered in darkness, at 30 °C, during 4 days. After this period, sprouted V. radiata seeds were collected, milled and lyophilized. Once lyophilized, the sprouts were milled again to a fine powder (0.18 mm sieve) for sample analysis and diet preparation.V. radiata flours and experimental diets was determined by drying to constant weight in an oven at 105 ± 1 °C. Total N was determined according to Kjeldahl’s method. Crude protein was calculated as N × 6.25. Insoluble N and soluble protein and non-protein N of the legume flours were measured as described by Kapravelou et al. [Moisture content of the raw (RVR) and 4 day-germinated (GVR) u et al. . SamplesTM, Sigma, St. Louis, MO, USA).Was done according to the method of Laemmli . The finv/v) of B at 15 min. Total run time was 25 min and post-delay time 5 min. Mobile phase flow rate was 0.4 mL/min.An Ultra Performance Liquid Chromatography (UPLC)  coupled by quadrupole-time-of-flight mass spectrometry  was employed for all high-resolution mass spectrometry analysis. Prior to mass spectrometry analysis, all powdered samples (100 mg) were extracted with 10 mL of acidified water (pH 2), and filtered through 0.22 μm nylon disk filters . 10 µL of the final solution were injected into the chromatograph. Analytical separation of peptides and proteins was performed on an Acquity BEH 300 C4 analytical column . A mobile phase consisting in a gradient program combining deionized water with 0.1% of acid formic as solvent A and acetonitrile with 0.1% of acid formic as solvent B was used. The initial conditions were 95% A and 5% B. A linear gradient was then established to reach 95% (m/z) range 50–1200.After chromatographic separation, a high-resolution mass spectrometry analysis was carried out in positive electro spray ionization (ESI+ve). The gas used for desolvation (800 L/h) and Cone (25 L/h) was high-purity nitrogen. Spectra were recorded over the mass/charge (fa/fa) (O) and 40 lean heterozygous (fa/+) (L) Zucker rats (aged 5 weeks) with an initial mean body weight of 137 ± 2.5 and 120 ± 1.6 g, respectively, were allocated to eight different experimental groups  within 2 different experiments RVR and GVR (raw vs. 4 day-germinated V. radiata dietary interventions). Two of the experimental groups in each experiment (an obese and a lean one) performed aerobic interval exercise (E) according to an established training protocol  while the two remaining were considered as sedentary (S) groups . The experiment lasted for 8 weeks, during which the animals were housed in a well ventilated, thermostatically controlled room (21 ± 2 °C). A reversed 12:12 light/dark cycle was implemented so the animals would perform the training protocol in darkness. Throughout the trial, animals had free access to type 2 water and consumed the experimental diet  and centrifuged at 1458× g for 15 min to separate plasma that was subsequently frozen in liquid nitrogen and stored at −80 °C. The liver was extracted, weighed, photographed for macroscopic studies, divided into various portions and immediately frozen in liquid nitrogen and stored at −80 °C. All experiments were undertaken according to Directional Guides Related to Animal Housing and Care [Forty male obese homozygous  , in orde2 max, and consisted of successive 4 min exercise periods at 65–80% of VO2 max, followed by 3 min recovery periods at 50–65% of VO2 max. The intensities and length of the training were gradually incremented every week , and the area under the curve (AUC) was determined. Biochemical parameters of lipid metabolism, and liver functionality were measured in plasma using a Shenzhen Mindray BS-200 Chemistry Analyzer  at the Bioanalysis Unit of the Scientific Instrumentation Centre . A portion of liver was lyophilized in order to determine the percentage of water. Hepatic lipids were extracted from the freeze-dried liver portion using the method described by Folch et al. [Fasting blood glucose and glucose tolerance test were performed by oral glucose overload following the protocol described by Prieto et al. . Blood gh et al.  with slih et al. . The extn = 40 samples per group). Histological alterations were evaluated according to the following grading score: -, non-existent; +, mild; ++, mild/moderate; +++, moderate; ++++, abundant; +++++, severe.Liver area of the macroscopic image was estimated in all the liver images of the eight experimental groups assayed by morphometric study using the software Image Pro Plus 6.0 . A portion of liver was fixed in 10% phosphate-buffered formalin, dehydrated in ethanol, embedded in paraffin, and sectioned for histological examination using hematoxylin-eosin (HE), and Masson’s trichrome (MT) staining for general microscopy morphology and fibrosis development, respectively. Four different preparations of each staining were analyzed for each animal, and 10 animals were evaluated in each experimental group . Liver homogenates were centrifuged at 13,000× g for 45 min, at 4 °C and supernatants were aliquoted and stored at −80 °C, until further use for western blot analysis. Protein concentration was measured by the method of Lowry et al. [Liver samples were homogenized in 20 mM Tris HCl buffer containing 0.1% Igepal, 100 mM Ethylene glycol-bis(2-aminoethylether)y et al. . Equal afa/+ or fa/fa genotype), AIT protocol, and dietary intervention with RVR or GVR on final body weight, aerobic capacity and physical performance, plasma and liver biochemical parameters, hepatic antioxidant enzyme activity, and protein expression was analyzed by 2 × 2 × 2 factorial ANOVA with phenotype (lean vs. obese), AIT protocol (sedentary vs. exercise) and dietary intervention (RVR vs. GVR) as main treatments. The use of a 2 × 2 × 2 factorial ANOVA is based on the potential interactions among the three interventions assayed  being significant in our statistical model in addition to single effects of exercise or dietary treatment in each of the two phenotypes tested. To reinforce the potential integrative strength of the statistical model implemented, the R2 statistic has been included in the tables as a measure of goodness of fit of the model, given that the coefficient of determination indicates the proportion of variability in a data set that can be accounted by the statistical model. Results are given as mean values and pooled standard error of the mean. Bonferroni’s test was used to detect differences between treatment means. The analyses were performed with SAS, version 9.0 , and the level of significance was set at p < 0.05.Time-repeated measurement analysis was applied to weekly food intake and body weight gain data as well as to blood glucose content after an oral glucose overload in order to analyze within subject effects (time) or within group effects (phenotype or AIT protocol) on the above parameters. The effect of phenotype  of 93–893.p = 0.025 and p < 0.0001, respectively).The effects of phenotype, AIT protocol and dietary treatment with RVR or GVR on weekly food intake and weekly body weight gain are presented in p = 0.001).With regard to the dietary treatment, GVR led to lower food intake and body weight gain when compared to RVR diet, although the effects on weight gain were more pronounced in the obese when compared to the lean animals, thus giving rise to a significant phenotype × diet interaction . Significant effects of phenotype and exercise were also observed in running time, maximal speed and distance run that were lower in obese compared to lean animals and improved substantially in both groups as a result of the AIT protocol  Furthermore, a significant effect of exercise was observed on VO2 max that increased in both phenotypes (p < 0.001), although the effects were more pronounced in lean compared to obese animals, thus resulting in significant phenotype × exercise interaction (p < 0.001). Regarding the inclusion of RVR or GVR in the experimental diets, running time, maximal speed and distance run were enhanced by the GVR compared to RVR diet, with more pronounced effects in lean animals. All these, gave rise to significant diet effect (p < 0.001) and diet × phenotype interactions . In addition, the parameters related to physical performance were differentially affected by exercise depending on the type of diet consumed. In this regard, the enhancing effect of exercise in both lean and obese animals was greatest in rats fed GVR, thus resulting in significant diet × exercise interaction . The AIT protocol assayed tended to increase all the hematic parameters related to aerobic capacity . Likewise, diet significantly affected the influence of exercise on AUC, with a less evident effect in GVR compared to RVR, resulting in a significant diet × exercise interaction. All these effects were reflected in different trends of evolution in blood glucose values after an oral glucose overload in GVR compared to RVR , Alanine Transferase (ALT), and Alkaline Phosphatase (ALP) activities were used as markers of liver function. A significant phenotype effect was observed on these parameters leading to higher values in the obese groups when compared to the lean ones. Exercise managed to decrease the activities of AST, ALT, and ALP among the obese groups, but a significant increase in ALP was observed for the lean animals. There was also a significant effect of diet in AST and ALT activities with lower values found in GVR than in RVR animals. The effects of diet were more obvious in obese than in lean animals, thus giving rise to a significant phenotype × diet interaction. With regard to the hepatic antioxidant enzyme activities, the influence of the different interventions assayed on the outcomes studied varied widely and did not reach significant biological importance.There was a clear phenotype effect on liver histopathological parameters related to the development of NAFLD , Table 6p < 0.001). GVR diet tended to increase AMPK phosphorylation compared to RVR, although the results did not reach statistical significance (p = 0.129). With regard to the AIT protocol, it decreased the degree of AMPK phosphorylation in all the experimental groups with the exception of lean animals fed RVR, thus resulting in significant exercise effect (p < 0.001) and exercise × diet interaction (p = 0.001).The effects of phenotype, AIT protocol, and dietary treatment on protein expression of AMPK and pAMPK are shown in V. radiata in combination with the AIT protocol was more efficient than the raw legume at reversing the alterations in aerobic capacity, glucose and lipid metabolism, and liver histology and functionality.In the present study, we aimed to study the beneficial effects of a mixed intervention with aerobic interval training protocol and RVR or GVR diets on plasma and liver parameters of hepatic functionality using the obese Zucker rat experimental animal model. Obese rats exhibited higher food intake and body weight, and suffered significant alterations in plasma lipid profile, glucose tolerance test, liver tests and histology. Exercise increased the aerobic capacity of both rat phenotypes and diminished the severity of MetS alterations, especially those related to glucose and lipid metabolism. Consumption of germinated V. radiata was effective at breaking down proteins in small molecular weight peptides such as shown by both chemical and electrophoretic methods. The results were further confirmed by mass spectrometry that showed the appearance of newly formed low molecular weight peptides with low retention times. Our results are in agreement with those reported by Urbano et al. [The germination process selected to obtain a natural protein hydrolyzate from o et al.  in pea, o et al.  due to bObese Zucker rats are known to present a genetic defect in leptin receptor that causes the development of hyperphagia as well as other metabolic disturbances . A decreV. radiata [Physical performance was always lower in obese when compared to lean Zucker rats due to the severe metabolic disturbances, impaired skeletal muscle perfusion, and muscular atrophy inherent to this experimental model. Moreover, low intrinsic aerobic capacity in rats has been related to lower energy expenditure and reduced whole body and hepatic mitochondrial lipid oxidation, which can induce a higher susceptibility to dietary-induced hepatic steatosis . As a re radiata  resultedV. radiata L. are mainly assigned to different non-nutritional components like d-chiro-inositol and the synergistic effects of its two major phenolics: vitexin and isovitexin [V. radiata are thought to arise from its phytosterol and dietary fiber content that act in the prevention of cholesterol biosynthesis and absorption and cecal fermentation [The development of MetS in obese Zucker rats was associated to alterations in glucose and lipid metabolism as reported by other authors ,32. Our ovitexin , as wellovitexin . The strovitexin . The hypentation . NeverthIn the present study, using the Non Alcoholic Steatohepatitis grading score described by Brunt et al. , a valueV. radiata sprouts [In addition, the administration of GVR diet led to a greater improvement in hepatic status compared to RVR as demonstrated by the lower liver weight and area of the obese animals that consumed this specific diet. Such increased potential of the GVR diet may be due to the compositional changes that take place during germination such as the increase in polyphenol, antioxidant capacity and vitamin C levels of  sprouts ,41. CholActivation of AMPK depends on the ADP:ATP ratio, and it is reduced in the liver of obese Zucker rat due to an excess of energetic substrates entering this tissue . In addiV. radiata and physical exercise) represented an efficient and feasible complement for the treatment of MetS.The beneficial effects on insulin resistance, hyperlipidemia, and liver tests and histology exerted by the combination of the two lifestyle strategies assayed (protein hydrolyzates from germinated Since patients affected with MetS have increased risk of developing NAFLD, we put forward the hypothesis that lifestyle interventions based on dietary treatment with functional ingredients in combination with a specific training protocol are an alternative or complementary treatment to improve the metabolic alterations caused by the disease. Therefore, liver histological and functionality analyses may contribute to a better knowledge about the pathophysiology of NAFLD and to establish an efficient treatment based on nutritional and healthy lifestyle strategies to improve metabolic syndrome and derived hepatic dysfunction.


Adequate fluid management is an important component of patient care following cardiac surgery. Our aim in this study was to determine the benefits of tolvaptan, an oral selective vasopressin-2 receptor antagonist that causes electrolyte-free water diuresis, in postoperative fluid management. We prospectively examined the effect of tolvaptan on renal excretion of electrolytes and urea nitrogen in cardiac surgery patients.n = 30) or conventional loop diuretic therapy plus tolvaptan . Fractional excretions of sodium (FENA), potassium (FEK) and urea nitrogen (FEUN) were measured in both groups during post-surgical hospitalization.Patients undergoing coronary artery bypass surgery were randomized to receive conventional loop diuretics . However, postoperative FENA values in Group C did not decrease, and the values were similar in both groups. Serum potassium levels remained lower and FEK values remained higher than the preoperative values, but only in Group C . BUN increased postoperatively in both groups, but it remained higher than its preoperative value only in Group C . Group T showed an initial increase in BUN, which peaked and then returned to its preoperative value within a week. The FEUN increased postoperatively in both groups, but the change was more pronounced in Group T .Urine output was greater with tolvaptan (Group T) than without it (Group C), and some patients in Group C required intravenous as well as oral loop diuretics. Serum sodium concentrations decreased after surgery in Group C, but were unchanged in Group T The present study is registered with the UMIN Clinical Trials Registry (ID:  Fluid management following cardiac surgery is one of the most important parts of postoperative cardiac patient care , 2. LoopOne promising candidate is tolvaptan, an oral selective vasopressin V2-receptor antagonist that causes electrolyte-free water diuresis , 7. WateTo the best of our knowledge, no detailed study has yet examined the effect of tolvaptan on renal electrolyte excretion following cardiac surgery. Therefore, we prospectively compared the fractional excretion of electrolytes and urea nitrogen in postoperative cardiac surgery patients following treatment with loop diuretics with or without tolvaptan.Data on the effects of tolvaptan on changes in body weight and other clinical parameters after bypass surgery were collected at Juntendo University Hospital. Seventy patients undergoing off-pump coronary artery bypass surgery were prospectively enrolled in the study and randomized to either the group receiving conventional loop diuretic treatment only (Group C) or the group receiving conventional loop diuretic treatment plus tolvaptan (Group T). Patients in Group T received tolvaptan 7.5 mg once daily on postoperative days 1 and 2, in addition to conventional oral loop diuretics; intravenous loop diuretics were given only if needed. Tolvaptan was administered as needed from postoperative day 3 in Group T patients. Patients in Group C received only conventional loop diuretic therapy. Patients with renal insufficiency, such as those with chronic kidney disease stage 4 or greater, were excluded from the study. Patients requiring emergency surgery, those undergoing cardiopulmonary bypass, and those undergoing concomitant extracardiac/vascular and/or valvular surgery were also excluded. Laboratory values were serially measured after surgery for up to 7 days during hospitalization, and the fractional excretion of sodium (FENA), potassium (FEK), and urea nitrogen (FEUN) were calculated for both groups of patients.The study was approved by the institutional review board of Juntendo University Hospital (UMIN000011039). All participants received detailed information about the study and provided written informed consent.Data are presented as mean ± SD. Normality was evaluated for each variable from normal distribution plots and histograms. Data obtained at the same time-point were compared between groups using Student’s unpaired two-tailed t-test for continuous variables and the chi-square test for categorical variables. Intragroup changes in preoperative vs. postoperative values were assessed with Student’s paired t-test. All data were analyzed using the Statistical Analysis Systems software JMP 11.0 .A total of 70 patients scheduled to undergo off-pump coronary artery bypass surgery were initially reviewed. Of those, 2 patients were excluded as they required another procedure in addition to bypass surgery. After randomization of patients into Groups T and C, incomplete datasets were collected from 7 patients in Group T and 4 patients in Group C. Therefore, analyzable data were obtained from 27 patients in Group T and 30 patients in Group C Fig. .Fig. 1FlPatient demographics, laboratory values prior to the surgery, and operative information for both groups are summarized in Table Both groups received loop diuretics after surgery in a routine manner in order to rectify postoperative hypervolemia. In our institution, Lasix® 40 mg per day orally is generally given to patients undergoing coronary bypass surgery. Additional Lasix® is administered intravenously if a patient’s hourly urine output is approximately less than 50 mL. In the present study, only some patients in Group C required additional intravenous loop diuretics Fig. . Even sop = 0.046, 0.027, and 0.029, respectively), but they remained unchanged in Group T. Comparison of the values between the groups revealed that postoperative serum sodium concentrations at postoperative days 2, 3, 5, and 7 were higher in Group T than in Group C. However, even though the serum sodium concentrations decreased postoperatively in Group C, the renal sodium excretion in that group remained constant. In other words, FENA did not decrease in Group C.Figure p < 0.001). After postoperative day 3, serum potassium levels in Group T returned to the preoperative range, whereas the levels stayed lower in Group C. The FEK values increased from their preoperative value on postoperative day 1 in both groups (both p < 0.001). In Group C, the FEK values further increased until postoperative day 3, and they remained higher than the preoperative value . In Group T, the FEK value, which was higher on postoperative day 1 than on the preoperative value, decreased and returned to the preoperative range after postoperative day 2.Figure p = 0.0003). In Group T, the increase in BUN peaked on postoperative day 2 and then gradually decreased to its preoperative value.Figure p = 0.049 and 0.025, respectively).In both groups, FEUN increased postoperatively and remained higher than the preoperative values. The increase in postoperative FEUN was more pronounced in Group T. Intergroup comparison showed that the FEUN values at postoperative days 2 and 7 were statistically higher in Group T than in Group C  the amount of daily urine output was well maintained and somewhat greater with tolvaptan (Group T) than without it (Group C), although some patients in Group C required intravenous loop diuretic administration; (ii) serum sodium concentrations decreased after surgery only in Group C, but the postoperative FENA values were not significantly different with or without tolvaptan; (iii) in the absence of tolvaptan (Group C), postoperative serum potassium concentrations decreased and stayed lower than the preoperative level, with an associated increase in FEK. By contrast, the addition of tolvaptan (Group T) maintained the postoperative serum potassium levels at essentially the preoperative levels; and (iv) both groups showed an increase in BUN and FEUN after surgery, but tolvaptan treatment led to a higher FEUN and a rapid decrease in BUN as time progressed.We speculate that the similar FENA values with and without tolvaptan after surgery may reflect an enhancement of the effectiveness of loop diuretics by tolvaptan. Previous papers have also noted that concomitant administration of tolvaptan enhances the effects of loop diuretics . Nishizap = 0.024). Probably because the number of studied patients was too small, we failed to show a tolvaptan effect on eGFR values (Fig. In the current study, both groups of patients were placed on a salt-restricted diet soon after surgery, and the amount of peroral sodium intake did not likely differ between the groups. Therefore, the higher serum sodium concentration seen in Group T, compared to Group C, might have resulted from the effective free-water diuresis induced by tolvaptan in Group T. Indeed, the amount of daily urine volume was larger in Group T soon after surgery Fig. . In addiThe decrease in serum potassium concentrations and the slight increase in FEK after surgery in Group C are reasonable responses to loop diuretic therapy. Group T patients also received loop diuretics, although the dosage was smaller than in Group C, and the changes in their serum potassium and FEK values soon after surgery showed similar trends to those in Group C.The increase in BUN just after surgery most likely reflects surgery-induced protein catabolism. Although the operative duration was about 4 h and the peri-operative fasting period was approximately 1 day in both groups, maintenance fluid replacement, together with the fasting, would elicit protein degradation. However, the sustainment of this increase in BUN after surgery may also reflect neurohormonal activation and renal hypoperfusion , 17, 18.Operative stress induces neurohormonal activation and increases serum AVP levels , 9. The We admit that tolvaptan is a relatively expensive medication; however, it effectively enables diuresis without causing electrolyte imbalance, which is helpful in the management of postsurgical patients who are at high risk for cardiac arrhythmia and for progression to acute kidney injury. Therefore, the high cost of this treatment may be offset by reductions in the lengths of stays in the intensive care unit and further costly postsurgical treatment. Additional studies are needed to analyze the efficacy of tolvaptan on a medical-care cost basis in patients undergoing cardiac surgery. Moreover, elevated BUN and low sodium concentration have well known associations with poor patient prognosis , 11, 16,The present study had several limitations. First, this was a single-center, retrospective, observational analysis with a small number of patients. According to the protocol, tolvaptan was given only on postoperative days 1 and 2 in Group T, and the decision to continue tolvaptan was up to the attending doctor. Therefore, among the Group T patients, some received tolvaptan throughout their hospital stay and some received it for only 2 days. This may have affected the data obtained after postoperative day 3, which complicates the interpretation of our results. In addition, because most of the patients were able to consume fluids orally within a day after surgery, we were unable to estimate the effect of intravenous fluid administration on serum and/or urinary electrolytes. We also did not measure plasma AVP levels or urine Aquaporin-2, which have been reported to be predictive markers of the response to tolvaptan , 25. OthIn conclusion, loop diuretics can cause continuous excretion of sodium and potassium, irrespective of their serum concentrations, which may result in electrolyte imbalance. The addition of tolvaptan in the present study prevented the generation of an electrolyte imbalance in the cardiac surgery patients investigated here. Furthermore, tolvaptan seems to enhance the effectiveness of loop diuretics. Tolvaptan also showed potential to promote the excretion of urea nitrogen produced by protein catabolism and in response to operative stress, and it does so more effectively than loop diuretics in patients undergoing coronary artery bypass surgery.


Deficiency of macro and micro-minerals in the ration of dairy cows adversely affects growth, milk production and reproduction efficiency. It is essential to examine mineral concentrations in feeds offered to dairy cows in practical farms.Two villages from each taluka were selected at random for taking representative samples of feeds, forages and hair. Within the village, help was sought from village milk producers and district animal husbandry officer for identification of 4 to 5 farmers and collection of representative samples. All the samples were processed and analyzed for chemical composition as well as major macro and micro-minerals, using Inductively Coupled Plasma-Optical Emission Spectrometer.Ca content in wheat straw (0.29%), crushed maize (0.02%) and wheat bran (0.12%) was found to be below the critical level (0.30%). The P content in concentrate ingredients was high (0.26–0.96%), but low in dry roughages (0.06–0.12%). Cereal straws (0.14%) and grains (0.12%) were deficient in Mg. Feeds and forages were found to be adequate in K (1.50%). Cereals straws were found to be deficient in S (0.11%). Greens were good source of Cu (12.02 ppm). Wheat straw was found to be low in Zn (18 ppm), but high in Mn (225 ppm) and Fe (509 ppm). Local grasses and azolla green were found to be rich source of Co (>1.00 ppm). Se (0.63 ppm) was present in appreciable quantities in most of the feedstuffs.From the present study, it was apparent that the feeds and forages available in the state of Jharkhand may not meet the requirements for Ca, P, Mg, Cu, Zn and Co in order to sustain a milk production of ~10 kg/day. Therefore, it is necessary to supplement these deficient minerals through area specific mineral mixture in the ration of dairy cows for improving productivity and reproduction efficiency. Dietary nutrition plays a significant role in any livestock development programme and the optimum expression of genetic potential for milk production in dairy cows depend on adequate supply of nutrients. Micronutrients, particularly the mineral elements are considered to be inevitable for the normal metabolic and physiological processes of animal systems. The importance of minerals in regulating biological systems, growth, production and reproduction is well documented , however035’ N and longitude of 85033’ E. Atmospheric temperature ranges from 5 to 45 °C during different seasons. In this study, two dimensional survey methods was adopted to map relevant mineral elements, by collecting feeds and forage samples from the representative villages, according to random sampling design based on conceptual landscape units [Information Network for Animal Productivity and Health, a windows based Internet linked application, developed by National Dairy Development Board of India to assess the prevailing status of the nutrient provision to the animal against the animal’s nutrient requirements [Two villages from each taluka were selected at random for taking representative samples of feeds, forage and hair. Total area of Jharkhand state is 79,710 sq km distributed in to 24 districts, 211 blocks and 32,620 villages. The district is having annual rainfall of 1,400 mm, latitude of 23Weekly basis composite samples of wet forages, cereal straws/dry forages, concentrate feed ingredients and the compound cattle feed (concentrate mixture) were collected from all over the surveyed area. Surveyed area used for the places/villages in the district from where representative samples of feed, wet forage, dry roughage and hair samples were collected. Wet forage samples were dried in hot air oven at 100 °C for 24 h and subsequently ground (1 mm). Ground samples of concentrate and forages were stored in airtight bags until analysis. The samples of feeds, wet forages and dry roughages were analyzed for crude protein (CP), ether extract (EE), crude fibre (CF) and acid insoluble ash (AIA) as per AOAC  and for The word “critical” is used in this article to note a concentration in feedstuffs below (or above with excesses) what is considered the requirement for dairy cow . This asThe data were analyzed statistically as per the Snedecor and Cochran , with thZea mays), jowar green (Sorghum bicolor) etc. Some milk producers offered crushed maize, crushed wheat alone or their mixture. Feeding mustard oil cake, wheat bran, linseed cake and maize germ cake was also observed in some parts of state. Those milk producers, who didn’t feed concentrate ingredients, were feeding compound cattle feed depending on the level of milk production. The use of common salt and mineral mixture supplementation was not a common practice in the surveyed area, except for therapeutic purpose on prescription by veterinary officer.Crop residues were found to be the main source of dry roughages in the ration of dairy cows. It was noticed that some of the milk producers fed cultivated forages like, maize (Cicer arietinum), arhar/pigeon pea (Cajanus cajan) and urd (Phaseolus mungo). Neutral detergent fibre (NDF) and acid detergent fibre (ADF) content was found to be highest in dry roughages. The data on proximate composition of the feedstuffs are in agreement [The feed and forage samples collected from surveyed area were analyzed for chemical composition Table . Mustardgreement –19.TableIn order of priority, available good quality feed resources are first allocated to lactating dairy cows followed by dry pregnant, dry, heifers, growing calves and non-productive cows. In the surveyed area our observation indicates that metabolizable energy were in excess in the ration of more than 65% of cows, crude protein was deficient in the diet of more than 85% of dairy cows, Ca and P were deficient in the ration of about 70% of the cows because of inadequate mineral mixture supplementation. Milk producers in most of the developing countries often do not feed adequate quantities of mineral mixture to their dairy cows due to non-availability, lack of knowledge on the benefits of feeding mineral mixtures . In viewThe straws of wheat and paddy were the main roughage sources in the surveyed area Tables  and 3. TCu content was found below the critical level (<8 ppm) in wheat straw, paddy straw and crushed grains Tables  and 5. ZHair samples collected during survey were analyzed for the same minerals as in feeds and forages. Mineral levels in hair must reflect the concentration and/or activity of the certain minerals in other parts of the body and reflect dietary mineral status of dairy cows . The aveThe daily intake of different minerals by a HF crossbred cow (400 kg body weight) yielding 10 kg milk (4% fat), with the prevailing feeding system in the surveyed area is presented in Table Information on the actual intake of each type of feeds and forage for a particular level of milk production was collected from each of the individual dairy farmer, to calculate intake of various mineral elements against the requirement. Total mineral intake from feeds and forages was compared against the requirements on dry matter basis Table , to idenIt was evident from the present study that majority of the dairy cows in Jharkhand state were deficient in Ca, P, Mg, S, Cu, Zn and Co. Therefore, it is necessary to supplement these minerals in the ration of dairy cows by formulating area specific mineral mixture, having highly bio-available mineral salts. Deficient trace minerals, except Co, may be supplemented in the form of chelates, for better bio-availability and improving productivity, reproductive efficiency and productive life of dairy cows.


Nanoscale materials have been produced with unprecedented speed due to their widespread use, and they may eventually be released into the environment. As effective adsorbents for heavy metals, carbon nanomaterials can be used to immobilize metals in contaminated soil, but little information is available regarding their effects on soil microarthropods. This study was designed to investigate the influence of three types of carbon nanomaterials, graphene (G), graphene oxide (GO) and carbon nanotubes (CNTs) on soil microarthropod communities under turfgrass growth conditions. The application of carbon nanomaterials resulted in increased abundance of all soil microarthropods, especially in the GO and CNT treatments. GO also significantly increased the abundances of multiple trophic functional groups, including predators, detritivores, herbivores and fungivores. Further, the dominant genera varied among the treatments. Herbivorous microarthropods predominated in the control, whereas predatory species predominated in the carbon nanomaterial treatments. Carbon nanomaterials also increased the total taxonomic richness, Shannon diversity index, and dominance index of the microarthropod community, but they decreased the evenness index. Higher diversity of soil microarthropods indicates an environment suitable for soil mesofauna and for enhanced decomposition and nutrient cycling in the soil food web. Therefore, these organisms are good indicators reflecting the changes in ecosystems4. Changes in the community structure of microarthropods can be used to characterize soil conditions, such as the soil organic matter content, soil disturbances and pollution levels6. Microarthropods include the Collembola (springtails), Acari (mites), Diptera and Coleoptera. In particular, Collembola and Acari are the most important and abundant microarthropod fauna in soils, and are the most valuable groups for soil quality evaluations7.Soil microarthropods represent a class of soil fauna that is widespread in the soil ecosystem. These organisms play important roles in soil organic matter decomposition, nutrient mineralization, microbial activity and soil aggregation8. Oxidized carbon nanomaterials include a large amount of chemically bonded oxygen on the surface. Due to their large specific surface areas and porous structures with many functional groups, carbon nanomaterials have shown good adsorption abilities for metal ions in environmental applications10. Tan et al.11 used graphene oxide (GO) membranes as adsorbents for removing heavy metals from water and found that the maximum adsorption capacities of the GO membranes for Cu2+, Cd2+ and Ni2+ were approximately 72.6, 83.8 and 62.3 mg/g, respectively, whereas Lu and Chiu12 used single-walled and multi-walled oxidized carbon nanotubes to adsorb Zn2+ in water and found that their adsorption capacities at pH = 7 and 25 °C were 43.66 and 32.68 mg/g, respectively. Adsorbents based on graphene are employed for removing pollutants from the environment13. Carbon nanomaterials, however, will inevitably be released into the soil environment after application, with mostly unknown consequences. Previous studies have evaluated the toxicological effects and the potential mechanisms of toxicity of carbon-based nanomaterials in bacteria, mammalian cells, and animal models15, but very little work has explored the impacts of carbon nanomaterials on soil fauna, especially on the community of soil microarthropods.Carbon nanomaterials are composed of approximately 97–99% of carbon atoms in the form nearly spherical particles with diameters between 10 and 100 nmThe present study aimed to determine the impacts of three different carbon nanomaterials, graphene (G), graphene oxide (GO) and carbon nanotubes (CNTs) on the composition and diversity of the soil microarthropod community in a turfgrass system. We characterized the responses of soil microarthropods to carbon nanomaterials in order to provide a scientific basis for the application of carbon nanomaterials for remediating soil contaminated with heavy metals.Palaeacarus (Oribatida), Spinibdella (Prostigmata) and genera belong to Tarsonemidae (Prostigmata), accounting for 47.1% of the individuals. In the GO treatments, Mahunkania and Cheylostigmaeus, belonging to Prostigmata, were the dominant genera and presented 17.6% and 34.3%, respectively, of the total individuals. In the CNT treatments, there were four dominant taxa that together accounted for 76.5% of the individuals: Brennandania 28.1%, Petalomium 15.6%, Trombidiidae 17.2% and Rhodacarellus 15.6%.In total, 24 genera of Acari and 8 genera of Collembola were recorded in this study Table . The ideThe abundance of total microarthropods was strongly affected by the treatments Table . The aveNeognathus were grouped into fungivorous, detritivorous, herbivorous and predatory guilds according to their feeding habits and the results for their abundances are shown in Fig. All of the collected soil microarthropods except for P < 0.05) in the Shannon–Wiener index and dominance relative to control values were observed in all of the carbon nanomaterial treatments. Moreover, the GO and G treatments showed significantly greater species richness than did the CNT and control treatments. Although the GO treatment showed lower evenness, the evenness indices were not significantly different among the G, CNT and control treatments.The diversity indices were significantly influenced by all the carbon nanomaterials Table . Signifi4. Organic fertilizers may promote increases in faunal populations by increasing the quality and quantity of food needed by soil microarthropods17. Greater microarthropod densities with increased soil fertility were also observed by Cole et al.18. However, Cao et al.19 found that applications of chemical fertilizer decreased the number of soil mites, perhaps because of the direct toxicity of metal contaminants in the chemical fertilizers20. In the present study, the abundance of soil microarthropods significantly increased in the turfgrass plantation under the application of G, GO and CNT. This may be attributable to an increase in the supply of soil nutrients  through the addition of carbon nanomaterials. Previous studies demonstrated that the growth of soil microarthropods abundance depends on increases in the soil C and N concentrations5.Soil microarthropods are important components of the soil ecosystem. The species composition and abundance of soil microarthropods, the trophic functional groups, and the community indices can be used as sensitive indicators for evaluating ecosystem processes21. In general, application of a low level of fertilizer tends to result in increased faunal diversity due to its positive effect on detrital inputs17. However, graphene oxide showed the most significant effect on the abundance and diversity of soil microarthropods. Previous studies demonstrated that GO can enhance cell adhesion and proliferation and promote biological growth and reproduction, thereby affecting the populations and diversity of total microarthropods22.Both graphene and graphene oxide increased the diversity and richness of soil microarthropods, effects that were mainly attributable to the marked increases in Oribatida and Prostigmata. In a previous study, an increase in the diversity of soil fauna was correlated to an increase in the diversity of available food resources23. Based on the presence of Oribatida in soils treated with carbon nanomaterials, these soils tend to contain nearly all functional groups of microarthropods24. In the present study, Oribatida was mainly represented by detritivores, which are sensitive to the soil environments25. The greater abundances of Oribatida in carbon nanomaterial treatments suggest that the nutrient level and soil quality were improved. Furthermore, detritivorous Oribatida are involved in organic matter decomposition, nutrient cycling and soil formation in the soil ecosystem25. In the GO treatment, the number of microarthropods in predatory, herbivorous and fungivorous groups increased at a significance level of 51-fold, 10-fold and 16-fold, respectively, in comparison with the control, whereas no significant differences in abundance of these trophic groups were found among the control, G and CNT treatments. This result indicates that different trophic communities have different adaptive mechanisms to the treatments. Predatory microarthropods were found to be dominant in carbon nanomaterial-treated soils, whereas herbivorous species were dominant in the control treatment. The increase in predatory abundance was likely caused by an increase in the availability of prey species, e.g., Enchytraeidae and nematodes26. Nematodes are a preferred food source for most predatory species28. As suggested in previous studies, predators may more directly regulate the abundances of detritivorous and fungivorous microarthropods by limiting their population expansion in the natural soil30. Further studies should be conducted to identify the mechanisms by which carbon nanomaterials affect soil microarthropods, perhaps through effects on the soil properties.The function of soil and the maintenance of soil quality are closely linked to the micro-fauna community composition; therefore, understanding the responses of soil microarthropod functional groups is useful for revealing the ecological effects of carbon nanomaterialsIn conclusion, the application of carbon nanomaterials  in turfgrass soil significantly increased the abundance and diversity of soil microarthropods. The GO treatment produced significant increases in the abundances of four trophic microarthropods. Predatory arthropods predominated in the carbon nanomaterial treatments, whereas herbivorous species predominated in the control. The increased microarthropod populations support enhanced decomposition and nutrient cycling in the soil food web by strengthening both bottom-up and top-down processes.−1, and organic matter, total phosphorus, and total nitrogen contents of 52.3, 3.75 and 2.15 g kg−1, respectively. Festuca arundinacea Schreb. was selected as the tested turfgrass. Municipal solid waste (MSW) compost was obtained from Xiaodian composting plants in Tianjin, China. The properties and origins of the carbon nanomaterials, graphene (G), graphene oxide (GO) and carbon nanotubes (CNTs), are reported in Table The tested soil was gathered from the top 20 cm of an experimental site at the Tianjin Normal University campus. It is a sandy loam with a pH (in water) of 7.3, water content of 19.4%, conductivity of 2250 μS cmFestuca arundinacea was sown in each pot. Cultivation was performed in a greenhouse under natural light conditions (646-27090 LX). During the experiment, the night and day average temperatures were 15 °C and 26 °C, respectively, and the relative humidity was between 35% and 55%. Water was supplied daily to maintain adequate substrate moisture for turfgrass growth. Turfgrasses were mowed once, on day 65.For this study 1500 g of tested soil, 50 g of MSW compost and 15 g of carbon nanomaterials were mixed thoroughly and transferred to plastic pots . Treatments without the nanomaterials were used as the controls, and each treatment was replicated three times. Then, 5 g of seeds of 31 and then preserved in 70% alcohol. All extracted faunal samples were sorted and counted under a dissection microscope and identified to the genus level32. The soil microarthropods were grouped into predatory, herbivorous, detritivorous and fungivorous guilds38.Soil samples were taken from the experimental pots on day 130. Soil microarthropods were extracted from 100 g of fresh soil using the Tullgren methodH′), Margalef richness index (SR), Pielou evenness index (J) and Simpson dominance index (C) were used to describe the diversity of the soil microarthropod community39. H′ = −∑PilnPi; SR = (S − 1)/lnN; J = H′/lnS; C = 1 − ∑(Pi)2, where S is the number of species, N is the total number of individuals and Pi = Ni/N is the ratio between the individual number in a genus and the total number of individuals.The Shannon-Wiener index . Data presented in the Tables and Figure are means ± standard deviations (SD) of three replicates for each treatment. SigmaPlot 12.5 was used to plot the graph40.The responses of the microarthropod community structure to carbon nanomaterials were examined by using analysis of variance (ANOVA). Differences between means were evaluated by using the least significant difference (LSD) test. All statistical tests were conducted at a significance level of Dataset 1