To explore the potential of walking alone milestone combined reading-frame rule to improve the early diagnosis of Duchenne muscular dystrophy (DMD).To retrospectively describe the genotype and phenotype of Duchenne and Becker muscular dystrophies (BMD) patients with deletions and duplicates in the dystrophin gene. The sensitivity and specificity of the reading frame rule were calculated and compared to that of the combined reading frame rule and walking alone milestone. The diagnostic coincidence rate of two different methods was analyzed.P < 0.05).One hundred sixty-nine male DMD/BMD patients were enrolled, including 17 cases of BMD and 152 cases of DMD. The diagnostic coincidence rate, diagnostic sensitivity, and specificity of the reading-frame rule for DMD/BMD were 85.2, 86.8, and 70.59%, respectively. The sensitivity and specificity of the reading frame principle combined with the walking alone milestone for DMD/BMD were 96.05 and 70.59%, respectively. The diagnostic coincidence rate increased to 93.49%, significantly different from that predicted by reading- frame rule (The reading-frame rule combined with the walking alone milestone significantly improved the early diagnosis rate of DMD. DMD). The incidence rate did not vary significantly among countries, regions, or races, with one case occurring in every 3,600–6,000 male births  could not walk independently from 10-15 years old. Their average life expectancy under natural conditions was about 20 years . NeverthQuantitative analysis of dystrophy protein in muscle biopsies can help identify DMD/BMD early, but invasive procedures limit its clinical application . An earlThis study aimed to evaluate the early predictive value of the reading-frame rule combined with the walking alone milestone for DMD.A group of unrelated male probands from 208 cases diagnosed in Children's Hospital Affiliated to Zhengzhou University from 2014 to 2021. All the patients had confirmed met the criteria of DMD/BMD.DMD gene with a pathogenic variant.Inclusion criteria: (1) Male, (2) myopathy  genetic testing revealed DMD, (3) at the last follow-up, the cases had not lost ambulation and were under 16.Exclusion criteria: (1) Female, (2) the cases with point mutation in Diagnostic principles of DMD and BMD: Patients who lose ambulation and need wheelchairs before 16 years old (<16 y) were classified as DMD, and patients who did not lose independent walking ability at 16 years old (≥16 y) were diagnosed as BMD.We definite the “criteria for delay of walking alone milestone” as“unable to walk alone until 18 months old.”Two hundred eight cases were registered, 185 (88.9%) cases are deletions/duplications. Twenty-three (11.1%) cases were excluded because of the point mutation. Sixteen cases younger than 16 who could walk independently at the last follow-up were excluded. One hundred sixty-nine cases of DMD/BMD with deletion or duplication were included in our study, T-test was used to compare DMD and BMD's demographic and clinical data. The Chi-square test (χ2-test) was used to compare the diagnostic coincidence rate of different methods. Version 25 of the SPSS software  was used for all statistical analyses, setting the significance at p < 0.05.The study was approved by the ethics committee of the Children's Hospital Affiliated to Zhengzhou University, Zhengzhou, China.P < 0.05; One hundred sixty-nine cases were included in our study. 20 cases had a family history, while the other 149 had no family history or family members refused to provide information. All 169 cases are male. The mean age of onset in DMD and BMD cases was 3.88 ± 2.18 years (range 0.5–9 years) and 6.49 ± 3.06 years (range 2.8–12 years), respectively, but the age of diagnosis delayed, which was 6.80 ± 2.83 (range 1.27–14.34 years) and 7.13 ± 3.45 (2.88 ± 12.12 years), respectively. The mean value of creatine kinase (CK) was 16,197.15 ± 8,750.95U/L  in DMD patients and 8,657.45 ± 5,933.32 U/L  in BMD patients, with the statistical difference (P < 0.05). In BMD group, all cases could walk independently before 18 months, while in DMD cases, 93 cases (61.18%) had an independent walking delay.The age of walking alone in BMD group was between 12 and 14 months, with a mean age of 12.88 ± 0.61 months, while the age of walking alone in DMD group was between 14 and 30 months, with a mean age of 18.03 ± 7.12 months, de novo mutations and 31 with maternal origin . One hundred fifty three cases of deletions were as follows: those in a single exon was the most frequent (28.1%), followed by 5 (13.1%), 2 (9.8%), 3 (8.45%), 6 (7.19%), 8 (7.19%), and 7 (6.54%), 2-test, P = 0.014; In 169 cases, 12 cases of BMD are in the reading frame, and 132 cases of DMD are out of the reading frame, in which 85.2% were in agreement with the rule of reading-frame (86.8% for DMD and 70.59% for BMD). Sensitivity and specificity were 86.8 and 70.59%, respectively, assuming that DMD was the case out of the reading frame and BMD was the case in the reading frame. Sensitivity and specificity were 61.18 and 100%, respectively, assuming that the case was BMD when walking alone milestones were younger than 18 months. The case was DMD when the milestone was older than 18 months. The combined sensitivity and specificity were 96.05 and 70.59%, respectively. The specificity remained unchanged, sensitivity increased by 9.25%, and the diagnostic coincidence rate increased from 85.2 to 93.49%  indicate severe muscle damage . CK is rIn 1988, Monaco et al. proposed the reading-frame rule, and related research supports this hypothesis . In the 2-test, P = 0.06). In the study of scholar Juan Yang, among 118 cases of exon duplication of DMD gene in the Chinese population, there were 9 cases of BMD, indicating that exon duplication is more likely to occur in DMD cases .The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


In multivariate analyses, Hb at T1 was the best predictive biomarker of anemia at T3 with a cut-off value of 120 g/L (specificity 87.5%). Conclusions: The prevalence of anemia at the end of pregnancy remained high in a High-Income Country. Clinical, social, and biochemical parameters did not seem useful to predict anemia at T3 and could not guide iron supplementation. We suggest systematically performing a simple blood count in the first trimester of pregnancy and offering oral iron supplementation for women with Hb < 120 g/L.Background: Systematic iron supplementation may be harmful in pregnant women with non-depleted iron. Our objectives were to estimate the prevalence of anemia at the third trimester of pregnancy (T3) and to identify the parameters at the first trimester (T1), which best predict anemia at T3. Methods: This prospective cohort study in France included pregnant women at T1 without non-iron deficiency anemia. Clinical and social characteristics, health-related quality of life, blood count, and a frozen blood sample were collected at T1 and/or T3. Secondly, a matched nested case–control study was built for women with anemia at T3 but not at T1. Multivariate analyses and ROC curves were used to identify the best predictive parameter(s) of anemia at T3. Results: The prevalence of anemia at T3 in the cohort (629 women) was 21.9% (95% CI 18.7–25.2%). In the matched nested case–control study (256 women), hemoglobin (Hb), serum ferritin (SF) and the SF/soluble transferrin receptor ratio at T1 were predictive of anemia at T3 ( Iron deficiency anemia (IDA) is very common in pregnancy, particularly in the third trimester, due to the increase in iron requirement by the fetus. The consequences of IDA during pregnancy can be serious for the mother: decreased cognitive and working capacities, depression, and susceptibility to infections . Iron deTherefore, we need to identify early in pregnancy women at risk of developing iron deficiency anemia in late pregnancy in order to better target iron supplementation. In clinical practice, serum ferritin (SF) is the most reliable marker to diagnose iron deficiency in pregnancy . Other bA prospective non-interventional single-center cohort study was conducted during one year in France. The study was approved by the French Medical Ethics Committee for the Protection of Individuals Southeast VI  and registered on ClinicalTrials.gov (NCT03176147). All the women provided written consent to participate in the study after being informed in detail about the study procedures. Whole cohort: Pregnant women who consulted the maternity department of Clermont-Ferrand University Hospital for an ultrasonography examination of the fetus between 9 weeks and 11 weeks plus 6 days of gestation, called “first trimester ultrasonography”, and routinely covered by the French public health insurance system were invited to participate in this study. According to French guidelines, women at risk of anemia have a free Hb screening in the first trimester of pregnancy (T1) . Women u2), and the nearest date of inclusion in the study. We did not choose additional matching criteria such as parity, which can be an explanatory criterion of iron deficiency anemia.Matched nested case–control study: A sub-group of the whole cohort was subsequently included in a secondary analysis. Each woman without anemia at T1 but with anemia at T3 was matched to one woman without anemia in T1 and T3, according to age (±5 years), body mass index before pregnancy  according to international recommendations ,8. We suA second blood count was systematically performed during a visit scheduled at T3 between 31 and 34 weeks of gestation. The blood samples collected at T1 were used to determine Hb, mean corpuscular volume (MCV), mean corpuscular hemoglobin concentration (MCHC), red cell distribution width (RDW), and reticulocyte hemoglobin content (RHC). The blood samples frozen at T1 were used to determine serum ferritin (SF), serum hepcidin (Hepc), and serum transferrin receptor (STfR) for the women included in the case–control study. Ferritin was measured by chemiluminescence using a Dimension Vista analyzer from Siemens USA. STfR and Hepc were measured using ELISA methods . As the public financial support of this study was not sufficient to determine the biomarkers of the whole cohort, a matched nested case–control was used. Clinical and social characteristics were collected at T1, including the use of “Universal Medical Coverage”  and health-related quality of life (HRQoL) and social deprivation, using the MOS SF-36  and EPICClinical characteristics at T3 and delivery information were collected: systolic and diastolic blood pressure, hypertension, term of delivery, and birth weight of newborns. The blood samples collected at T3 were used to determine Hb, MCV, and MCHC.The sample size was statistically determined to estimate a prevalence of anemia at T3 of 10%, considering the lowest value found in the literature in High-Income Countries , with a Continuous variables were expressed as medians and interquartile ranges; categorical variables were expressed as numbers and percentages. The prevalence of anemia at T1 and T3 defined as Hb < 110 g/L, the prevalence of hemoconcentration at T3 defined as Hb > 135 d/L and the 95% confidence intervals (95% CIs) were calculated.For the nested case–control study, women with anemia at T3 were compared to matched women without anemia at T3 using Wilcoxon signed-rank tests for continuous variables, McNemar tests for binary variables, and Cochran’s Q tests for categorical variables. p < 0.20. Unadjusted and adjusted odds-ratios (ORs) and their 95% CIs were calculated. Conditional logistic regressions were performed to investigate which sociodemographic and clinical characteristics or biomarkers at T1 were predictive factors of anemia at T3 in the matched nested case–control sample, with a forward selection if needed. We included independent variables significant in the bivariate analyses at For the most predictive biomarker at T1 identified in the conditional logistic regressions, we estimated the area under the receiver operating characteristic (ROC) curve (AUC). The AUC ranges from 0.5 to 1, where higher values indicate better accuracy. Different thresholds were tested for this biomarker to find the optimal positive predictive value using the matched case–control sample. Sensitivity, specificity, and negative predictive value were also estimated. We conducted sensitivity analyses using the whole cohort sample excluding women with anemia at T1 if this biomarker was collected for the whole cohort and not only for the matched nested case–control sample. Statistical analyses were performed with SAS software  and conducted at a two-sided alpha = 0.05 significance level.Among the pregnant women included, 81.0% performed the blood test in T1 . ClinicaThe prevalence of anemia at T3 was 21.9% (95% CI 18.7–25.2%). Among these 138 women with anemia, 10 (7.2%) already had anemia at T1. The prevalence of anemia in the whole cohort at T1 was 3.2% (95% CI 1.8–4.6%). The prevalence of hemoconcentration at T3 was 6.2% (95% CI 4.3–8.1%). Among the 629 pregnant women of the whole cohort, 481 (76.5%) did not have anemia at T1 or T3, 10 (1.6%) had anemia at T1 but not at T3, 10 (1.6%) had anemia at T1 and T3, and 128 (20.3%) did not have anemia at T1 but had anemia at T3. These 128 women without anemia at T1 and with anemia at T3 were defined as the cases of the matched nested case–control study. These cases were matched with 128 women that did not have anemia at T1 or T3 (controls). The social and clinical characteristics and the biomarkers of these 256 women at T1 and at T3 are presented in p < 0.001). MCV, MCHC, and RHC were also lower in women with anemia at T3 than in controls with lower significant p values. Women with anemia at T3 had significantly higher STfR values than controls.All biomarkers at T1 were significantly different between cases and controls, except RDW, Hepc, and the ratio Hepc/SF . HemogloTwo conditional logistic regressions were performed to investigate the predictive factors at T1 of anemia at T3: model 1 with SF and STfR among the independent variables, model 2 with the SF/STfR ratio. The results of the multivariate analyses, adjusted for parity and educational level, are described in n = 256) and for the whole cohort, excluding the 20 women with anemia at T1 (n = 609). The AUC were 0.70 (95% CI 0.63–0.76) and 0.68 (95% CI 0.63–0.73), respectively. In the matched nested case–control sample, the best positive predictive value for anemia at the end of pregnancy was 74.6% (95% CI 63.9–85.4%) for an Hb value < 120 g/L at T1. For this cut-off value, specificity was 87.5% (95% CI 81.8–93.2%), sensitivity 36.7% (95% CI 28.4–45.1%), and negative predictive value 58.0% (95% CI 51.1–65.0%). Using the same cut-off in the whole cohort (Hb < 120 g/L), the positive predictive value was 83.6% (95% CI 80.3–86.8%), specificity 85.7% (95% CI 82.5–88.8%), sensitivity 36.7% (95% CI 28.4–45.1%), and negative predictive value 40.5% (95% CI 31.6–49.5%). For the 81 women with anemia at T3 and Hb of 120 g/L or more at T1, the median SF at T1 was 26 µg/L (interquartile range 13–50). The ROC curves were therefore estimated with Hb for the matched case–control sample  at T1 and similar to that of non-pregnant women of reproductive age in France . The pre% at T1 aA recent guideline  recommenIn our study, the number of subjects needed was not reached. Nevertheless, the prevalence of anemia at T3 (21.9%) was higher than estimated (10%), and our sample size allowed us to estimate the prevalence of anemia at T3 with a precision of 3.2%, which was very close to our objective of 2%. Transferrin saturation (TfS) was not performed because we determined SF, which demonstrated better efficacy for diagnosing IDA in pregnancy ,20. FurtIn this prospective cohort study, we found that an Hb value lower than 120 g/L in the first trimester of pregnancy is a good positive predictive value with very good specificity to predict anemia at the end of pregnancy. In High-Income Countries, we suggest systematically performing a blood count in the first trimester of pregnancy and offering oral iron supplementation for women with an Hb lower than 120 g/L. However, this recommendation should be assessed in a controlled trial in order to confirm the clinical benefit for the mother and the child at birth and during the development of the child.


Through investigation and analysis of the course management of 314 patients with benign prostatic hyperplasia (BPH) during the COVID-19 pandemic, we expected to offer effective recommendations for the management of patients with BPH against the background of the COVID-19 pandemic.  We implemented telephone follow-ups of 314 patients with BPH who were diagnosed at the Urology Clinic of Xiangya Hospital of Central South University before January 24, 2020, and who were admitted to the hospital for reexamination after the epidemic was controlled in China, and we conducted research and analysis of their disease management during the COVID-19 pandemic. Qmax) < 15 ml/s increased from 82.5% to 92.3%, and the proportion with a Qmax < 10 ml/s increased from 8.7% to 15.4%; the individuals with a prostate volume > 30 ml increased from 94.1% to 97.0%; the percentage of men with a bladder residual urine volume > 10 ml was augmented from 81.6% to 89.3%, and patients with prostate nodules on physical examination were elevated from 1.0% to 1.7%. We uncovered no prostate cancer in patients, and the proportion of patients administered the combination drug increased from 78.9% to 91.2%. Compared with patients receiving online or telephone consultations, patients undergoing reexamination at the hospital were better controlled. When we conducted a survey of whether patients chose to go to a public or private hospital for follow-up, we found that 46.6% of the patients chose to go to a private medical institution.  In the follow-up, we found 245 patients (79.3%) over 60 years of age and 187 patients (60.5%) with underlying disease. There were 47 patients (15.2%) who returned for consultation during the COVID-19 pandemic, and of these, 18 were admitted to the hospital for follow-up consultation, and 29 patients underwent consultation via the internet or telephone. Eleven patients underwent surgery during the pandemic, and of these, three experienced emergency surgery. We encountered 65 patients (24.4%) who self-administered medications irregularly and 54 patients (20.3%) who self-medicated and adjusted the dosage and drug types themselves. There were 302 patients (97.7%) who wished to be reexamined during the COVID-19 pandemic. In terms of treatment, the proportion of patients “awaiting observation” declined from 13.9% to 4.4%, and the proportion of patients “awaiting surgery” increased from 4.9% to 16.4%. Using the International Prostate Symptom Score (IPSS) scale, the percentage of patients with moderate-to-severe symptoms increased from 79.9% to 90.1%, and the proportion with a quality of life (QOL) score ≥ 5 rose from 82.5% to 88.9%. The proportions of patients exhibiting storage, voiding, and postmicturition symptoms in lower urinary tract symptoms (LUTS) increased from 77.3%, 21.7%, and 18.8% to 91.9%, 27.5%, and 25.5%, respectively; those manifesting hematuria and urinary retention increased from 0.9% and 0.6% to 2.3% and 1.7%, respectively; those with a prostate specific antigen (PSA) > 4 ng/ml rose from 10.0% to 15.1%; patients with a maximum flow rate ( COVID-19 greatly affected the treatment of patients with BPH. When conditions permit, we recommend that patients first consider going to the hospital for evaluation; however, when this is not possible, medical institutions should provide telephone or online consultation for patients with BPH. Surgical treatment should also be arranged for those in need as soon as possible to avoid delaying the patient's treatment. Coronavirus disease 2019 (COVID-19) is an acute respiratory disease caused by the novel coronavirus ; these sBenign prostatic hyperplasia (BPH) is the most common benign disease that causes urination disorders in middle-aged and older men , 15. TheWe executed telephone follow-up of 314 patients with BPH who were diagnosed at the Urology Clinic of Xiangya Hospital of Central South University prior to January 24, 2020, and who were admitted to the hospital for reexamination after the epidemic was controlled, and we subsequently conducted analyses of their disease management during the COVID-19 outbreak . Since follow-up was carried out during the epidemic stage of the COVID-19 outbreak, no written informed consent was signed physically; however, each patient was informed in detail by telephone regarding the purpose of the follow-up and the details of the present study, and we thereby obtained their consent. We herein assured the patients' personal privacy, and only used the acquired data for clinical scientific research. All data in this project were provided by the patient's own oral narrative.Our inclusion criteria were (1) patients with BPH and normal TPSA and free PSA/total PSA (f/tPSA) ratios; (2) patients with prostate nodules that were observed under color Doppler ultrasonography, CT, MRI, or physical examination, but whose prostate biopsy confirmed BPH; (3) patients with a TPSA > 10 ng/ml or a TPSA at 4–10 ng/ml but abnormal f/tPSA and prostate specific antigen density (PSAD) in whom prostate cancer was excluded by prostate biopsy; and (4) patients with a TPSA at 4–10 ng/ml, but with normal f/tPSA and PSAD, in whom prostate cancer was ruled out combined with objective MRI examination.Exclusion criteria were (1) patients with symptoms of hematuria and patients with BPH in whom malignant tumors were not completely eliminated; (2) patients with prostate nodules as ascertained under color Doppler ultrasonography, CT, MRI, or physical examination, in whom no prostate biopsy was performed to rule out prostate cancer; and (3) BPH patients with a TPSA > 10 ng/ml, or a TPSA at 4–10 ng/ml but with abnormal f/tPSA and PSAD in whom prostate cancer was not eliminated.Although we registered 314 patients with BPH, we were unable to obtain disease information of five men: one patient showed an incorrect phone number and could not be followed up; three patients refused follow-up; and the remaining individual died due to an accident. We therefore ultimately followed up with 309 patients with BPH, and Qmax). A follow-up of 309 patients with BPH showed that 298 patients (96.4%) were reportedly affected by the COVID-19 pandemic and that 302 (97.7%) wished to be evaluated during the pandemic.Our hospital has taken a series of protective measures with respect to staff, as follows. (1) The hospital distributes medical surgical masks to all medical staff daily, has daily online check-in, conducts COVID-19 nucleic acid inspections once per week, is instructed on topics of COVID-19 prevention and control at least once per week, and has intensified self-protection awareness. (2) Those individuals who are required to leave the city on business trips need to file travel applications in advance; prior to returning to work, they are then subjected to a nucleic acid testing, and isolation may be necessitated depending upon their test results. (3) Medical staff with febrile symptoms can immediately return to work after a negative nucleic acid test for SARS-CoV-2. (4) The hospital wards are comprehensively disinfected three times a day. Before medical operation, staff should wash and disinfect hands, wear surgical gloves, take safety precautions, avoid contact with patients' bodily fluids, and immediately after the operation, staff should execute surgical hand-washing and disinfection . (5) TheQmax < 15 ml/s were augmented from 82.5% to 92.3%; those a Qmax < 10 ml/s rose from 8.7% to 15.4%; those with a prostate volume > 30 ml increased from 94.1% to 97.0%; patients with a bladder residual urine volume > 10 ml rose from 81.6% to 89.3; and the percentage of patients with prostate nodules upon physical examination was augmented from 1.0% to 1.7%. We noted no prostate cancer patients. The proportion of patients administered the combination drug increased from 78.9% to 91.2%, and we determined that the relevant indicators for BPH deteriorated to varying degrees—one reason being the patients' failure to modify their lifestyle. In addition, a patient's failure to regularly accept medication was also an important reason. Finally, patient inability to come to the hospital for evaluation and the timely adjustment of treatment plans also caused aggravation of the patient's BPH symptoms, confirming that the restrictions in place during the COVID-19 pandemic affected the treatment of patients with BPH.In addition, we compared some objective indicators of the 309 patients with BPH before COVID-19 break out with those of the 298 patients who were reexamined after effective COVID-19 control . Our survey found that regarding treatment, the proportion of patients “awaiting observation” declined from 13.9% before the COVID-19 outbreak to 4.4% after, and the proportion “awaiting surgery” increased from 4.9% to 16.4%, respectively. With respect to the IPSS scale, patients with moderate-to-severe symptoms increased from 79.9% to 90.1%; those with a QOL score ≥ 5 increased from 82.5% to 88.9%; the proportion of patients with storage, voiding, and postmicturition symptoms in LUTS increased from 77.3%, 21.7%, and 18.8% to 91.9%, 27.5%, and 25.5%, respectively; patients with hematuria and urinary retention increased from 0.9% and 0.6% to 2.3% and 1.7%, respectively; the proportion with a PSA > 4 ng/ml increased from 10.0% to 15.1%; values of those with a Qmax < 15 ml/s were 0% and 5.6%, respectively, while those who consulted medical staff by telephone or online and exhibited a new PSA > 4 ng/ml and a Qmax < 15 ml/s were at 6.9% and 10.3%, respectively. Patients who went to the hospital for reexamination and whose prostate volume and bladder residual urine volume rose again were 38.9% and 27.8%, respectively, while patients who consulted by telephone or internet showed increased prostate volume and bladder residual urine volume, with rates of 65.5% and 51.7%, respectively. There were no men with new prostate nodules who went to the hospital for evaluation, and the proportion of patients who showed new prostate nodules upon telephone or online consultation was 3.4%. The proportion of patients who adjusted their drug treatment plan after going to the hospital for reexamination was 66.7%, and those who adjusted their drug treatment plan after consulting via telephone or the internet was 34.5%. Finally, the percentage of those who traveled to the hospital for reexamination and who manifested improved LUTS was 77.8%, while that for online or telephone consultation was 44.8%. We thus demonstrated that compared with online or telephone consultation, the LUTS of patients who were admitted to the hospital for reexamination improved significantly. Compared with telephone or online consultation, a reexamination in the hospital also achieved specific inspections of related modalities: for example, urinary system ultrasonography, urine flow rate assessment, and PSA quantification. According to these examinations, a patient's condition was accurately assessed and the drug-treatment plan was adjusted in a timely fashion. We therefore posit that compared with telephone or online consultation, it is far better for patients to undergo hospital evaluation in order to control the patient's condition.We also conducted a comparative study of 18 patients with BPH who were admitted to the hospital for reexamination during the COVID-19 pandemic, and 29 patients with BPH who consulted with medical staff via telephone or the internet. We herein found that the proportions of patients with worsened IPSS and QOL scores were 22.2% and 27.8% of patients who went to the hospital for reexamination, respectively, and that the proportions of patients with worsened IPSS and QOL scores were 55.2% and 62.1% of patients who consulted by telephone or online, respectively. The proportions of patients who went to the hospital for reexamination with a new PSA > 4 ng/ml and It was reported that patients with COVID-19 were more likely to manifest deterioration of their symptoms toward more serious disease and should therefore visit the hospital for treatment in a timely fashion when the disease deteriorates—particularly when pathologic deterioration occurs . During COVID-19 pandemic greatly affected the treatment of patients with BPH. We recommend that if conditions permit, patients should first consider going to the hospital for evaluation; when this is not feasible, medical institutions need to provide telephone or online consultation for patients with BPH. Surgical intervention should also be arranged as soon as possible for those in need so as to avoid delaying patient treatment.


The prevalence of hepatocellular carcinoma (HCC) is increasing worldwide and it is now the third most common cause of cancer-related death. HCC is becoming a major health burden with steadily increasing incidence globally.This is an observational study over a 3-year period in a tertiary care center in India. Three hundred and thirty-nine patients diagnosed to have HCC were included in this study. Patients’ clinical, etiological, radiological and cytohistological data and therapy offered were recorded and analyzed.Cirrhosis of the liver was seen in 73.2% of the patients. 16.8% of patients were asymptomatic at the time of presentation. Ascites (57.2%) and jaundice (22.4%) were the most common signs of hepatic decompensation. The most common etiology of HCC was cryptogenic/non-alcoholic fatty liver disease (NAFLD) in 51% of the patients, while hepatitis B and C were seen in 17.4% and 5.8% of the patients, respectively. Advanced and end-stage disease with Barcelona Clinic Liver Cancer (BCLC) stages C and D were seen in 62.4% of patients. 56.6% had Albumin-bilirubin (ALBI) score of 2, while 62.8% had Okuda stage II disease. High alpha-fetoprotein (AFP) levels (>400 ng/mL) were seen in 48.9% of patients. Macrovascular invasion and metastases were seen in 45.9% and 22.2% of the patients, respectively. 17.6% of patients had evidence of tumor thrombus. 14.5% of biopsy specimens showed associated steatosis/steatohepatitis along with confirmation of HCC. Only 26.6% of the cirrhotic HCC patients were diagnosed during surveillance.HCC due to unknown cause/NAFLD appears to be overtaking hepatitis B as the commonest cause for HCC. Despite the advances in diagnostic methods and surveillance, most cases of HCC tend to be diagnosed at advanced stages. The incidence and prevalence of hepatocellular carcinoma (HCC) is rising, mainly due to the epidemic of non-alcoholic fatty liver disease (NAFLD), and is poised to become the leading cause of liver cancer . The 201This is a retrospective cross-sectional study of prospectively maintained data, conducted at a tertiary care center in India. Institutional Ethical Committee (IEC)   clearance was taken for the study (IEC: 410/2020). The clinical records of all patients admitted with  HCC at our center between March 2017 and March 2020 were reviewed. All the patients who fulfilled  the diagnostic criteria of  HCC according to European Association for the Study of the Liver (EASL) guidelines were taken into study . Data weAll patients’ details of clinical presentation, history of alcohol consumption, presence of cirrhosis, duration of cirrhosis (if known cirrhotic), blood investigations including complete blood counts, liver function tests, hepatitis B surface antigen (HBsAg) by chemiluminescence method, anti-hepatitis C virus (HCV) antibody by chemiluminescence method, alpha-fetoprotein (AFP) by enzyme chemiluminescence immunoassay (ECLIA) method, radiological features including tumor size, number, and location were recorded. Details of biopsy specimen and therapy offered were also recorded. Diagnosis of cirrhosis was made on the basis of clinical, biochemical, endoscopic, and radiological findings. Hepatitis B virus (HBV)-related cirrhosis was diagnosed when HBsAg was detectable in serum. HCV-related cirrhosis was diagnosed when detectable anti-HCV, HCV ribonucleic acid (RNA) , or both were present in serum.Etiology of cirrhosis was considered to be alcohol, if the alcohol consumption was more than 40–80 g/day for males and 20–40 g/day for females for more than10 years . SeveritDiagnosis of HCC was based on non-invasive criteria with typical findings of HCC in triple phase contrast multidetector computed tomography (MDCT)/magnetic resonance imaging (MRI) and/or histopathology. Image characteristics of liver lesion of arterial phase hyperenhancement according to LI-RADS (Liver Imaging Reporting and Data System) classification and washout on portal venous and/or delayed phases were diagnosed as HCC . Complett test and Mann-Whitney non-parametric U test. For all tests, p values < 0.05 were considered statistically significant. The analysis was performed using Statistical Package for the Social Sciences (SPSS) software .Categorical variables were presented as percentages, while continuous variables were presented as mean (standard deviation) or as median (interquartile range). Comparisons between proportions were performed using the Chi-square test and continuous variables using the student During the study period of 3 years, a total of 3038 patients with diagnosis of chronic liver disease (CLD) were admitted. Three hundred and forty-nine patients of HCC with 541 admission events were noted. A total of 339 patients with HCC  were included.n=91) of HCC patients were non-cirrhotic. One hundred and fifty-eight patients (63.7% of cirrhotic HCC ) were detected to have HCC and cirrhosis on the first presentation. In the remaining 90 patients, HCC was detected during the follow-up, either due to surveillance  or due to symptoms during follow-up . Among the surveillance group, the median duration from diagnosis of CLD to detection of HCC was 35.5 months  of the patients were male. 73.2% (248/339) of the patients with HCC had a background of cirrhosis on imaging (ultrasonography [USG]/computed tomography [CT]/MRI) at the time of diagnosis, while the remaining 26.8% (n=55) were asymptomatic or were diagnosed incidentally, when being evaluated for unrelated illness, of whom 40 patients were cirrhotic, while the remaining were non-cirrhotic. The most common clinical presentations were abdominal pain (24.4%), ascites (24.1%), and anorexia (20%) followed by weight loss (17.9%) and fatigue (10.9%)  bleed, and hepatic encephalopathy were noted in 194 (57.2%), 76 (22.4%), 22 (6.4%), and 3 (0.9%) of patients, respectively. Eleven patients (3.2%) had spontaneous rupture of HCC at first presentation.The most common cause of HCC was unknown/cryptogenic in 51% of the patients, while HBV and HCV constituted 17.4% and 5.8 % of the causes, respectively. Significant alcohol intake was seen in 19.4% of the patients. Rest of the etiology were constituted by combination of factors: alcohol + HBV, alcohol + HCV and HBV + HCV in 3.2%, 1.1% and 0.5% of patients, respectively  had no obvious precipitating factor, while HBV accounted for 8.9% and significant alcohol was seen in 7.6%.Among cirrhotic HCC patients, the most common cause was unknown in 39.9%. Significant alcohol was seen in 23.7%, while viral etiology of HBV and HCVwas seen in 20.5% and 7.6% of patients. A combination of factors  was seen in 6.85% of cirrhotic patients.Diabetes mellitus (DM) was present in 44.2% of the patients, while hypertension (HTN) was seen in 32.4%, ischemic heart disease in 8.5%, hypothyroidism in 2.9%, cerebrovascular accident (CVA) in 3.8%, chronic kidney disease (CKD) in 3.2%, and chronic obstructive pulmonary disease (COPD) in 3.5%. Among the cryptogenic group, 51.1% and 41.3% of the patients had DM and HTN, respectively. Three patients had human immunodeficiency virus (HIV) infection, two among the HBV group and one among the unknown group.Normal AFP levels up to 20 ng/mL were seen in 27.4% of patients, while AFP levels more than 400 ng/mL were seen in 48.9% of patients. Coagulopathy with international normalized ratio (INR) > 1.5 was seen in 14.5% of patients. Elevated aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels more than twice the upper normal limit were seen in 45.1% and 12.7% of patients. Elevated ALP levels more than two times the upper normal limit were seen in 49.3% of patients (Table 5 IU/mL. Majority of patients had hepatitis B e antigen (HBeAg) negative (24 out of 26 patients).HBV viral load was available for 32 patients, among whom 53.1% of patients had levels more than 10Most of the patients were having Child-Pugh category B followed by A in 45.4% and 41%, respectively. Majority belonged to BCLC C (48.8%) followed by B (25.4%). Very-early/early-stage disease (BCLC-0 and A) was seen in 12.1% of patients, and advanced stage/end-stage disease (BCLC-C and D) was seen in 62.4% of all patients. 56.6% had ALBI score of 2, while 62.8% had Okuda stage II disease , while the rest 6.4% had involvement of supradiaphragmatic IVC and/or extension to the right atrium. 17.6% (60 patients) of the patients had radiological evidence of tumor thrombosis.The most common site of metastases were lungs in 62.6%, followed by liver and bone in 16% and 13.3%, respectively. Other organs included peritoneum (6.6%), adrenal gland (6.6%), brain 4%), spleen (2.6%), and pancreas/kidney (1.3% each)  of patients. One patient underwent transarterial radioembolization (TARE) and another patient underwent liver transplantation. Palliative stereotactic body radiotherapy (SBRT) was offered to 3 patients and one patient received palliative radiotherapy to vertebral metastasis for symptom relief. Systemic chemotherapy with sorafenib was offered to 54.1% (n=143) of patients. Palliative therapy was given to 21.9% (n=58) of patients.Treatment details were available for 264 patients. Therapy with curative intent was offered to 13.6% of patients. Surgical resection was offered to 16 (6%) patients, of whom 4 underwent transarterial chemoembolization (TACE) prior to resection. Radiofrequency ablation was offered to 19 patients (7.1%). TACE was offered to 13.2% . Four patients among the FNA group had findings suggestive of HCC, while one with inadequate sample required repeat biopsy for characterization of the lesion. Sixty-one patients underwent USG-guided percutaneous liver biopsy, while one underwent intraoperative liver biopsy and two underwent biopsy from skeletal metastatic lesions. Thirty-five biopsy specimen were processed with additional immunohistochemistry markers like arginase-1, HepPar-1, and others like CK7 and CK20. All biopsy results except for one patient (who underwent repeat biopsy) yielded positive report. Among these biopsy specimen, 7 had evidence of macrosteatosis and 3 had steatohepatitis, while four patients showed evidence of liver fibrosis along with findings of HCC.p=0.0001), ALBI (p<0.001), Okuda scoring (p=0.036), and CTP class (p<0.001). Platelet-to-lymphocyte ratio (PLR) was found to be significantly lower in the group of cirrhotic HCC. AFP levels were comparable in both the groups. Patients in non-cirrhotic HCC group were having higher age, larger size of the lesion, and lower MELD score compared to the cirrhotic HCC group , presence of thrombosis (p<0.0001), and across BCLC stages (p<0.0001) and Okuda stages (p=0.03069).AFP levels were found to correlate significantly with size of lesion (p=0.026), size of lesion , MELD , CTP score , BCLC stages (p=0.0002), Okuda (p<0.0001), and ALBI staging (p=0.005). However, it did not correlate with AFP levels  and with respect to presence of thrombus (p= 0.1356). Similarly, PLR was found to be significantly different with presence of metastases (p=0.001), size of lesion , BCLC, and ALBI staging. However, there was no correlation of PLR with respect to AFP (p= 0.2189), CTP (p= 0.2809), and MELD score (p= 0.3141).There was statistically significant correlation of neutrophil-to-lymphocyte ratio (NLR) with presence of metastases  and HTN (41.3%). The high prevalence of DM and HTN among patients with unknown probably points towards NAFLD as an etiology. The lack of biopsy from  non-tumorous liver parenchyma for all patients was a barrier in identifying the true prevalence of NAFLD among our patients. Our study being retrospective in nature, the risk factors for NAFLD could not be assessed. NAFLD-associated HCC is more likely to arise even in the absence of cirrhosis. It is estimated that half of the cases of non-alcoholic steatohepatitis (NASH)-induced HCC arise in non-cirrhotic patients –29.Viral infection with HBV and HCV was seen in 17.4% and 5.8% of all HCC patients. Viral hepatitis was seen in 9.8% of non-cirrhotic patients, while it was seen in 28.2% of cirrhotic HCC patients. Significant alcohol intake was seen in 19.4% of all patients and accounting for 7.6% of non-cirrhotic HCC and 23.7% of cirrhotic HCC patients. Combination of factors were seen in 3.2% , 1% , and 0.5% (HBV + HCV) of all patients, and all were having cirrhosis.Our study findings were in contrast to the findings from other reported series from India, which showed viral hepatitis (HBV and HCV) to be commonest etiology for HCC , 23–25. Macrovascular invasion and metastases were seen in 45.7% and 22.1% of patients, while tumor thrombus was noted in 17.6% of patients. AFP > 400 ng/mL was seen in 48.9% of patients. Only 12.4% of patients had very early and early stage disease (BCLC- 0 and A) amenable for curative treatment. The nature of lesions including macrovascular invasion and metastases was similar to other studies from India , 23–25. Our study shows that both NLR and PLR correlate significantly with the presence of metastases and size of lesion and with BCLC and ALBI staging, however not correlating with AFP levels. AFP levels were found to correlate with size of the lesion, presence of thrombosis, and across BCLC and Okuda stages. Both the NLR and PLR were identified as predictors of overall survival and recurrence-free survival .Only 26.6% (24/90) of known cirrhotic HCC patients were diagnosed during surveillance and had median period of 35.5 months of cirrhosis prior to detection of liver lesion. In an American study, only 22% of cases known to have cirrhosis had undergone HCC screening prior to diagnosis . In a stSpontaneous rupture of HCC was seen in 3.2% of patients in our study. To our knowledge, this is the first study which noted prevalence of rupture of HCC among Indian patients as there are only few case reports and series. In recent reports from Italy and China, the reported incidence of rupture of HCC varies from 3% to 4.8% , 32.In conclusion, the majority of patients with HCC present at an advanced-stage limiting the therapeutic options that can be offered. NAFLD probably is becoming the most common etiological factor among both cirrhotic and non-cirrhotic patients of HCC. These findings must be interpreted in light of the limitations of the present study being a cross-sectional study with no follow-up. Current surveillance protocol and its compliance need to be assessed in a prospective study to validate the benefit of surveillance in these patients.


Lead is naturally available toxic heavy metal which is one of the main causes of environmental pollution and produces detrimental effects on health, particularly young children. Lead toxicity has become an emerging global burden of disease varying with the age, socio-economic status, occupation, industrialization, social customs and behaviours. The objective of this study was to find out the baseline blood lead level among children between 8-18 years.A descriptive study was conducted in Gokarneshwor Municipality, Kathmandu with a total of 160 children between 8-18 years of age from 2018 to 2019 after taking ethical approval from Research and Institutional Review Committee (Reference number: 17-074/075). Informed written consent was taken from the principal along with their respective parents and semi structured questionnaires were asked to students. Sample size was calculated and simple random sampling was done. The data was analyzed using Statistical Package for Social Science version 16. Point estimate at 95% Confidence Interval was calculated along with frequency and descriptive statistics.The mean blood lead level of the children was 4.39+7.35 μg/dl. Mean hemoglobin being 12.63g/dl; out of 160 children 30 (18.75%) children had blood lead levels. Children with elevated blood lead level had mean haemoglobin level within normal range (13.05gm/dl), however out of 30 children, 8 (27%) had haemoglobin level below normal.Lead exposure in the children of urban area of Nepal is considerably high as compared to similar studies coducted in similar settings. Data were collected from 2018 to 2019 after taking ethical approval from the Research and Institutional Review Committee, NMCTH (Reference number: 17-074/075). Children who were willing to participate were included while children suffering from any chronic illness were excluded. Simple random sampling was carried out and the sample size was using the formula:2 × (σ)2 / e2n = Z2 × (5.3)2 / (1)2  = (1.96)  = 108where,n = sample sizeZ = 1.96 at 95% confidence level11σ = Standard Deviation (5.3) according to a study done among older children from urban areas of Chennaie = margin of error, 1%.12 procedure i.e. 1+9 dilution with solution composed of 0.2% (NH4)2HPO4, 0.5% Triton X-100 and 0.2% HNO3 at Department of Biotechnology, Tribhuvan University, Kirtipur. A semi structured questionnaire was asked from students & values of interest were recorded. The informed written consent was taken. Data was then analyzed using Statistical Package for Social Sciences (SPSS) version 16. Analysis included standard descriptive statistics. Normally distributed variables were expressed in terms of mean±SD (standard deviation), whereas non normally distributed variables were presented as medians and ranges. Continuous data (BLLs) were expressed as mean±SD.Adding a non-response rate of 10% the required sample was 119. However, 160 children were included in the study. Total 3ml of blood from each participant was drawn from ante-cubital vein under strict aseptic conditions. Haemoglobin estimation by Drabkin's method in whole blood by using Drabkin reagent (Potassium ferricyanide / Potassium cyanide) at laboratory of Biochemistry Department of Nepal Medical College while BLLs estimation done in serum by Graphite Furnace AAS & Flame AAS applying lysing method at National Academy of Science and Technology (NAST), Lalitpur. Sample preparation for this method was done following Miller et al2. Hb was found to be ranged from 6.0-22.80g/dl, mean±SD of Hb being 12.63± 2.91g/dl. . Majority of the children were within the age between 14-16 years 37 (42.5%) male and 50 (57.5%) females. Least of children were below 10 years 6 (60%) males and 4 (40%) females. The mean age of children was 13.9±2.18 years. The mean BMI was found to be 20.06±3.77kg/m.91g/dl. .Elevated BLL was seen in 14-16 years of age group 15 (9.4%) followed by 11-13 years of age group 8.96 (5.6%) and ≤ 10 years of age 6 (3.75%).Children with elevated BLL had mean Hb level within normal range (13.05gm/dl), however out of 30 children 8 (27%) had Hb level below normal .BLL of 105 (65.62%) children was measured by GFAAS method with mean value of 3.72 μg/dl (range 0.10-49.56 μg/dl) while BLL of 55 (34.37%) children was measured by FAAS with mean 5.65 μg/dl (range 1.0-25.0μg/dl) .BLL among samples of20 childrenwere measuredby both GFAAS and FAAS method with mean value of 6.26 μg/dl (range 1.03-11.73μg/dl) and 3.55 vg/dl (range 2.0-4.0μg/dl) respectively. By GFAAS method, a wider range of BLL can be measured than by FAAS method .Among the sociodemographic and environmental variables studied, lead exposure had been related to the age group, frequent consumption of instant noodles, children exposed to renovation work, and children playing with batteries .1 which was 64.4%, exceeding the cut off points of CDC i.e. ≥5μg/dl. However, in this study so much high exposure is not found but still it is significant (18.75%).Very high prevalence of the lead exposure has been found in the study conducted by Dhimal, et al.4 concluded that the prevalence of BLL in children from the industrial city of Nepal was alarmingly high. Children exposed with chipped paints were related to BLL. Though not so high, the significant prevalence of the lead exposure in this study warrants need for screening programmes.Gautam K et al,5 found more than half of the children (57% of 251) were anemic at 15 months of age, and elevated BLLs were not significantly associated with anemia.In this current study those children with elevated BLL had mean Hb level within normal range, however 8 out of 30 children had Hb level below normal. Mohan VR et al,9 found the mean BLLs of Chinese children living in industrial and urban areas were significantly higher than those of children in suburbs and rural areas. The children's BLLs in China are higher than those of their counterparts in other countries due to its heavy lead pollution. China is considered heavily polluted with lead, especially urban areas. Large scale study is needed in our country to calculate the load of lead pollution.Meta-analysis by Wang S and Zhang J10 showed significant association of BLL with the environmental factors such as consumption of tap water in homes with lead service connections, peeling paint or recent renovations in old housing, hand-mouth behavior, passive smoking and having a mother born in a country where lead is often used while in this current study among many environmental factors, only children frequently consuming instant noodles, children playing with batteries, children exposed to recent renovation work at home and age group respectively were related with BLL.Etchevers et al.13 which showed the mean lead level for the entire group was 7.9μg/dl (SD= 4.5μg/dl). CDC has recommended intervention if a child's BLL is more than 15μg/dl. In this study also those children who had elevated BLL should undergo detailed medical evaluation.In this study BLL was found to be highly scattered with minimum BLL value, maximum BLL value, mean BLL, median, standard deviation. It may be due to the fact that the subject's BLLs were measured by two methods (FAAS and GFAAS) of different sensitivity. This was found to be lower compared to a similar study done in the urban area of Karachi, conducted by Khan et al.14 found high BLLs were significantly correlated with low BMI and low Hb status. However, in this current study the mean Hb of the children of the elevated BLL was found to be normal. BMI didn't show relation with BLL.Mitra, et al.15 However, detailed clinical history, IQ were not evaluated even those children who had high BLL were not followed up. Sherchand O et al,16 had conducted the first cross sectional study in primary school children in Nepal which revealed that the median BLL of school children was 5.5μg/dl. In this study however the level of toxicity was not seen high to appear clinical symptoms.Lead pollution has created health problems worldwide. Even relatively low levels of exposure can cause serious health conditions including neurological damage. To appear clinical symptoms BLL should be >70μg/dl.17 determined BLLs of the Japanese children by Inductively Coupled Plasma Mass Spectrometry (ICP-MS) and found the mean to be 0.96μg/dl and concluded lead exposure was generally low among Japanese children. In contrast, 18.75% children with elevated BLL beyond CDC normal reference range predict that children are alarmingly at high risk of lead poisoning, therefore public awareness and preventive programs are to be recommended.Ilmiawati et al.The present study had few unavoidable limitations. This study could not include children of lower age group due to many difficulties. We had to limit the sample size because of financial limitations. This study could not assess other risk factors including socioeconomic status, residency near industries and did not measure the presence of lead from the other sources of contamination e.g. lead level in soil, paint, dust, water supply, fashion accessories, folk remedies and so on. Also, the study did not investigate cognitive function or intelligence quotient (IQ) in children. As Hb was calculated by Drabkin method by colorimeter accurate Hb measurement could not be done. It warrants the large volume study of lead level measurement in children and its effect in Hb values measured by standard method. Children with elevated BLLs were not followed up or retested. This study did not establish any causal pathway between suspected sources and causes of elevated BLLs. It had a cross-sectional design, and lead exposure throughout the entire childhood development could not be assessed. Other hazardous environmental pollutants  were not evaluated.Elevated BLLs were more common among school children living in urban area. The GFAAS was a better method to measure BLL than FAAS. Children frequently consuming instant noodles, children playing with batteries, children exposed to recent renovation work at home and the age group related to the elevated BLL. Lead exposure in school children is becoming an alarming health problem in cities of Nepal. Study in large population is recommended for further evaluation of the current situation regarding the lead exposure to children. Screening programmes for BLL at least among school children living in urban areas should be emphasized.


Tenebrio molitor) can masticate nylon 11 film at the rate of 0.25 ± 0.07 mg per fifty larvae per day. The body mass of larvae did not differ from that of starvation control while feeding on nylon 11. Comparison of gut microbiota in nylon-fed and starving larvae showed a shift in composition. There was a significant variation in community composition among the nylon 11-fed experimental groups, suggesting that many organisms are capable of metabolizing nylon 11 fragments and/or possess a growth advantage in a nylon-fed gut environment. We also discovered that a significant fraction of gut microbiome of control larvae is capable of metabolizing nylon 11 monomer (11-aminoundecanoic acid) even in the absence of prior exposure to nylon 11. This is the first study demonstrating ingestion of nylon polymers by invertebrates, and our results suggest the potential of mealworm larvae for nylon 11 biodegradation applications.Nylon 11, which can be found in many commercial products, is a synthetic plastic that has previously been considered non-biodegradable. Increasing nylon 11 and other plastics in landfills and in the environment pose an environmental concern. Recent studies on plastic biodegradation revealed that initial mechanical fragmentations increase the rate of degradation. In this study, we discovered that the larvae of mealworm ( Two earlier biodegradation studies were performed with nylon 11/chitosan mixture or nylon 11/L-alanine co-polymers but not with pure nylon 11 Our previous study of nylon 11 enrichment culture with soil microorganisms led to the identification of 11-aminoundecanoic acid-degrading bacteria 2.2.1.−1 tryptone, 0.75 g·L−1 soytone, 0.625 g·L−1 dextrose, 1.25 g·L−1 sodium chloride and 0.625 g·L−1 dipotassium phosphate. For solid media, 15 g·L−1 agar was added. Nylon 11 was heated then stretched to approximately 0.1 mm thickness Tenebrio molitor were purchased from Ward's Science .Nylon 11 and other chemicals were purchased from Sigma-Aldrich . Deionized water was used for creating growth media. DEPC-treated water was used for molecular experiments. Bacterial strains were grown on defined media 2.2.Larvae from the single initial supply were separated into six sets of 50 individuals. On day 1, 5 grams of nylon 11 was added to three of the sets. No additional food or water source was added. The other three larvae sets were left under starvation (no food or water) and served as a negative control. Another set of three boxes, each containing 100 larvae, were also incubated with nylon 11. Larvae were incubated at room temperature. The masses of nylon 11 and the larvae were monitored for the span of 32 days. The microscopic images of the nylon 11 pieces were recorded using an Olympus BX60 microscope coupled with DP70 , or by Olympus CKX41 and recorded via Mic-Fi digital microscope .2.3.−1, 10−2, 10−3, 10−4 and 10−5 concentrations and spot plated as previously described 50 mg of fecal matter collected at the end of the study was resuspended in 1 mL 50 mM phosphate buffer (pH 8). The solutions were diluted to 102.4.The DNA from the mealworm feces was extracted using E.Z.N.A. stool DNA kit  according to the instructions. RNA was removed by incubation with RNase at 37 °C. The quality and the quantity of total RNA was confirmed by gel electrophoresis and absorbance at 260 and 280 nm. The sequences of the V3-V4 region of the 16S rRNA gene were obtained by Illumina MiSeq sequencing. Sequencing was performed by Psomagen Inc. .2.5.miccaphyloseq, vegan, biostrings, reshape, ape, scales, picante, ggpubr and gridExtravegan, ggplot2 and ggbiplot in R The Illumina reads were assembled and processed using 3.3.1.Since larvae did not ingest intact nylon 11 pellets (3 mm in diameter), nylon 11 was melted and stretched to a thin film. Ingestion by larvae at a measurable rate was observed in the areas of reduced thickness . The thiTo examine the fate of ingested nylon 11, we first observed fecal matter under the microscope. Microscopic investigation did not identify fragments distinctive from what was found in the starvation control . Additio−1 and 0.66 mg·day−1 for incubation with 50 and 100 worms, respectively. The rate nearly doubled when the number of worms was increased two-fold. This suggests that increasing the number of worms per mass of nylon 11 and/or increasing the accessible edges could further increase the rate of fragmentation.Despite the fact that no other food or water was added, a vast majority of larvae lived and continued to ingest nylon 11 for the 32-day study period . The rat3.2.To study whether ingested nylon 11 supports larvae growth and/or exerts negative impacts, the change in body mass and survival rate of larvae were analyzed. The body weight declined continuously during the study period . The rat3.3.To study whether fragmented nylon 11 impacted gut bacterial composition, we investigated the gut microbiota via high-throughput sequencing of 16S rRNA gene fragments. Sequences with 97% identity were grouped into OTUs. A total of 266 OTUs were identified, and taxonomic information was assigned. As shown in Spiroplasma), OTU2 (Enterobacter), OTU5 (Enterococcus), OTU6 (Lactobacillus), OTU7 (Lactococcus) and OTU8 (Enterobacteriaceae). One of the control samples (C2) and two nylon-fed samples (N1 and N3) contained many unique OTUs with low to medium abundance . We did not find any sequences that were significantly more abundant in all nylon-fed samples than in control with p < 0.05. However, we identified sequences that were uniquely higher in one of the nylon-fed sample than controls. OTU22  composed 2.3% of the N1 sequences while it was 0.02% of controls (400-fold enrichment). OTU13 (unknown), OTU17 (Streptophyta), OTU32 (unknown) and OTU12 (Clostridiaceae) were also uniquely enriched in the N1 sample . OTU5 (Enterococcus) was enriched 2.6-fold in the N2 sample. In the N3 sample, OTU15 (Staphylococcus), OTU2 (Enterobacter) and OTU14 (Enterobacteriaceae) were enriched by 6.3, 5.2 and 2.6-fold, respectively.There were sequences that were more abundant in controls (OTU3 and OTU8) than in nylon-fed groups with weak statistical significance . Next, to see if there were any changes in community composition that were linked to nylon 11 metabolism, we analyzed the C.F.U. on minimal media containing 11-AUA as a sole source of carbon and nitrogen. We reasoned that if ingested nylon 11 were fragmented into oligomers and/or monomers and available for bacterial metabolism, then bacteria capable of utilizing nylon 11 monomer (11-AUA) would be enriched. To our surprise, the number of 11-AUA degrading bacteria was high even in control samples  The ingested nylon 11 fragments were not available for metabolism by the gut bacteria; 2) nylon 11 was metabolized via a pathway that does not involve 11-AUA as an intermediate; or 3) resident bacteria were metabolizing 11-AUA, but their relative abundance did not change. We would like to note that this method does not allow us to detect a change in non-culturable populations. It is possible that non-culturable populations  play a significant role in nylon fragment metabolism in the gut.The presence of 11-AUA degrading bacteria in the gut of control larvae is unique, compared to our separate experiment with the adult mealworm beetle. In the adult beetles study, the starvation control did not harbor 11-AUA degrading bacteria. Ingestion of nylon 11 increased the 11-AUA degrading bacteria by 10,000-fold. Differences in the gut microbiome of larvae and adult, despite that they both ingest nylon 11, is of interest. We plan to study further to decipher the optimum strategies for nylon 11 biodegradation using mealworms at different stages of life.4.In this study, we have shown that mealworm larvae can ingest nylon 11. 0.25 mg of nylon 11 films was ingested by fifty worms per day for a period of 32 days. Absence of visible fragments in their fecal matter suggests that nylon 11 pieces were fragmented to a very small size or completely mineralized. The shift of gut microbial community composition was observed. Interestingly, mealworms devoid of previous exposure to the nylon 11 carried a significant number of monomer-degrading bacteria. Our results illuminate a potential use of mealworm larvae for nylon 11 bioremediation applications.


A systematic review and meta-analysis was performed to evaluate the prevalence of suicide ideation among HIV/AIDS patients in China.Systematic search of CNKI, Wanfang, China biology medicine database, Weipu, EMBASE, Web of science and PubMed for studies related to the suicide ideation of HIV/AIDS patients. The incidence of suicide ideation of HIV / AIDS patients in China was investigated by meta-analysis.n = 6,174). The incidence of suicidal ideation in HIV/AIDS patients was 30.6% (95%CI: 21.4–39.9%). The results of subgroup analysis showed that the incidence of suicidal ideation in male was 36.1%, which was higher than that in female (32.8%), homosexual patients (39.7%) higher than heterosexual patients (27.1%), 2013–2021 survey (35.2%) higher than 2003–2012 survey (26.5%), the unmarried patients (39.6%) were higher than the married patients (34.5%), the patients diagnosed >1 year (28.4%) were higher than the patients diagnosed <1 year (27.6%), and the depression patients (34.3%) were higher than patients without depression (20.5%) and CD4 cell counts ≤200 cells/ul group (20.6%) were higher than those in >400 cells/ul group (19.8%).A total of 16 studies were included (The incidence of suicide ideation in HIV/AIDS patients in China is relatively high. Acquired immunodeficiency syndrome (AIDS) is an infectious disease caused by Human Immunodeficiency Virus (HIV). As of December 2020, 1.053 million AIDS patients were registered and reported in China , and more than 100,000 new HIV-infected patients were reported that year . StudiesRetrieval databases include Chinese databases: CNKI, China Biology Medicine Database (CBM), Wanfang and Weipu; English databases: Web of Science, PubMed and Embase. The database was searched for “AIDS” or “acquired immunodeficiency syndrome” or “HIV”, “suicide” or “suicidal ideation”, “suicide” or “suicidal ideation”, “incidence” or “prevalence” and “China” by subject headings, abstracts, titles (or article titles) and keywords for Chinese databases and MeSH vocabulary and free words for English databases, and the retrieval time was set from 2003 to 2021. Besides, the study traceability method was applied to further search.Inclusion criteria: cross-sectional study from China; age ≥ 18 years old; with clear number of samples (≥25); the basis of suicidal ideation is clearly recorded.Exclusion criteria: repeated publication or incomplete information; unreasonable research design and statistical methods; review or expert commentary.Studying retrieval was carried out according to the retrieval strategy, and two researchers conducted preliminary screening of the retrieved studies, and then read the full text for secondary screening to determine the included studies according to the inclusion and exclusion criteria. Studies with different opinions were judged by a third researcher. Data were extracted according to a unified table, including the first author, publication year, study area, sample source, age range, sample size, incidence of suicidal ideation, etc. Suicidal ideation was defined as the presence of suicidal thoughts or suicidal intent.The quality was scored using the AHRQ  cross-sectional study quality evaluation list  . 0–3 poi2 < 50%, it was considered that there was moderate or low degree of heterogeneity, so the fixed effect model was used for analysis; if I2 ≥ 50%, the random effect model was applied for analysis. Subgroup analysis was used to evaluate differences in gender, sexual orientation, year of investigation, marital status, time of diagnosis, history of depression and CD4 cell count. Sensitivity analysis was conducted by excluding individual studies to observe changes in the results. If the results do not change much, it indicated that the results were relatively stable. Publication bias analysis was assessed using Egger's test and funnel plots. For all statistical tests, P < 0.05 was considered statistically significant.Stata12.0 software was used for heterogeneity test. If the heterogeneity test IM = 278). Among the studies included in this study, 5 are of high quality, 10 are of medium quality and 1 is of low quality , which were screened according to the inclusion and exclusion criteria. A total of 16 studies were finally selected, including 12 in Chinese and 4 in English, as shown in lity see .2 = 99.3%, P < 0.05). Therefore, the random effects model was used in this study. The incidence of suicidal ideation in Chinese HIV/AIDS patients was 30.6% (95%CI: 21.4~39.9%)  see .The incidence of suicidal ideation in male was 36.1% (95%CI: 29.1–43.2%), which was higher than that in females (32.8% (95%CI: 22.0–43.6%); the incidence of suicidal ideation in homosexual patients was 39.7% (95%) CI: 32.3–47.1%), higher than that in heterosexual patients 27.1% (95% CI: 22.1–32.1%); 35.2% (95%CI: 26.8–43.5%) in 2013–2021 and 26.5% (95%CI: 18.0–35.0%) in 2003–2012, indicating the incidence of suicidal ideation with an upward trend. The incidence of suicidal ideation in unmarried patients  was higher than that in married patients ; the incidence in patients diagnosed >1 year was 28.4% (95%CI: 23.9% ~33.0%), slightly higher than that in patients <1 year (27.6% (95%CI: 95%CI: 9.3% ~45.9%). Compared with non-depressive patients , depressive patients  was more likely to have suicidal ideation. The incidence of suicidal ideation in the CD4 cell count ≤ 200 cells/ul group was 27.3% (95%CI:6.3–48.4%), which was slightly higher than that in the CD4 cell count >400 cells/ul group  see .t = 1.08, P = 0.19, indicating that there was no publication bias. After excluding any studies in the sensitivity analysis, the overall results did not change significantly, confirmed the stability of the serious results.It was found that the symmetry was general through funnel plot, and Egger's test showed Suicidality and HIV/AIDS are currently two major public health issues of widespread concern, which are more prominent in developing countries . There aThe subgroup analysis found that the incidence of suicidal ideation in males was higher than in females, and the results were consistent with the studies by Kelly B et al.  in SwitzThe advantage of this study is that it systematically assessed the occurrence of suicidal ideation among HIV/AIDS patients in China, and provided new thinking and scientific basis for the formulation of psychological intervention policies and prevention and control priorities for AIDS patients. There were still several deficiencies in this study: (1) Due to the lack of a unified measurement scale for suicide in AIDS patients, a unified standard cannot be used to assess suicidal ideation, and there is information bias and selection bias; (2) This study also did not explore the effects of other co-morbid mental disorders and the impact of major emergencies in life on patients.The incidence of suicide ideation in HIV/AIDS patients in China is relatively high. In order to effectively reduce the suicide risk of HIV/AIDS patients, relevant government departments should strengthen the level of AIDS-related knowledge and further enhance the level of social support for people with HIV/AIDS. Policy makers should strengthen the cooperation between the Centers for Disease Control and Prevention and specialized mental health medical institutions, and highlight mental health education and psychological crisis intervention services on the basis of “four exemptions and one assistance”, so as to improve the quality of life of HIVAIDS patients and reduce the risk of suicide.The dataset generated and analyzed during the current study could be available from the corresponding authors on reasonable request.Study design: SL and HZ. Analysis and interpretation of data: YJia, QY, JY, and YJi. Drafting of the manuscript: SL and YJia. Critical revision of the manuscript: SY and HZ. Approval of the final version for publication: All authors. All authors contributed to the article and approved the submitted version.


Unilateral spinal anesthesia (USpA) is a technique used to restrict the effect of the spinal block on the operative side. 10–15 degrees reverse Trendelenburg position has been used to control the height of the spinal block using hyperbaric drugs. We aimed to study the effect of the 10-degree reverse Trendelenburg position on the quality of block and hemodynamic stability in unilateral spinal anesthesia in this hospital-based, double-blind, randomized clinical trial. 60 patients of both sexes between 20–60 years of age, undergoing below-knee orthopedic surgeries, were randomized into 2 groups. In both groups, spinal anesthesia was given with 2 mL bupivacaine heavy (0.5%), and the lateral position was maintained for 10 mins. Group 2 patients were kept in a 10-degree reverse Trendelenburg position throughout the surgery. The hemodynamic parameters and block characteristics of the two groups were compared using Epi Info statistical software. The onset of sensory block was faster in Group 1 (recumbent) compared to Group 2 (reverse Trendelenburg). The two-segment regression time was longer in the second group. In group 2, 73.3% of patients reached a level at T8 or below T8, compared to 46.7% in Group 1. The duration of sensory block and anesthesia was longer in Group 2. We conclude that reverse Trendelenburg of 10 degrees immediately after spinal anesthesia significantly limits the level of sensory block and prolongs the duration of unilateral spinal anesthesia. Spinal anesthesia (SA) has long enjoyed a history of "anesthesia of choice" for below umbilical surgeries, being used by humanity for over a century . Spinal There are relatively few investigations in which the reverse Trendelenburg position was employed to control the height of the spinal block with hyperbaric medications , 10. FurThis research was carried out in a "tertiary hospital" in northern India using a double-blind, randomized clinical trial including patients who underwent knee or sub-knee surgery. This analysis was carried out in line with the ethical standards of the 1975 Helsinki Declaration, as amended in 2000. Patients presenting for knee or below knee surgeries in a supine position under the subarachnoid block, aged 20–65, either sex, ASA physical status 1 or 2, and hemodynamically stable were involved in the research. Patients with ASA physical status of more than 2, any history of cardiovascular, renal, hepatic, respiratory, endocrine, and neuromuscular disorders, patients with epilepsy, neurological or psychiatric disorders, bleeding or coagulation disorders, local spinal deformity or local illness, and people with known allergy to local anesthetic agents were omitted from the research. Patients not consenting to regional anesthesia or patients who requiered surgery in a lateral position were also excluded from the study.2 (oxygen saturation) were applied. Vital signs  were recorded before surgery.Randomization of participants was done using a computer-generated sequence with the help of sequentially numbered opaque envelopes to be opened by the concerned anesthetist. All observations were performed by the same observer to eliminate subjective errors. All participants gave informed consent in their native language during the preoperative visit. Ranitidine 150 mg and alprazolam 0.25 mg were administered on the night of surgery and 2 hours before surgery. The patient was transferred to the operating table, and a venous catheter was secured with an 18G cannula on the forearm. Loading was performed with 10 ml/kg of Ringer's lactate liquid. 5-lead ECG (electrocardiography), NIBP (non-invasive blood pressure monitoring), and SpO2 were monitored. Monitoring was started after completion of the injection, every 5 minutes until the first 60 minutes, and after that, every 10 mins until the surgery was over. Sensory analgesia was tested bilaterally along the midclavicular line every 30s until sensory blockage appeared  and then every 5 min until both segments regressed below the maximal level.An epidural catheter (Romson EPI KIT[GS-2006]) was placed into the L2-3 intervertebral space with an 18G Tuohy needle inside the lateral position using all aseptic precautions. The catheter was fixed after being inserted 4 cm into the epidural space. A subarachnoid puncture was performed at the patient's midline in a lateral position using a Quincke BD needle 26G×3.5-inch spinal needle that cuts the dura mater (0.45 mm×90 mm) in the L3-4 intervertebral space. Then, 10 mg heavy 0.5 percent bupivacaine was slowly inserted intrathecally at a 0.2 ml per sec rate. The beveled end of the spinal needle was kept facing down in the lateral position. There were 2 groups of patients. In Group 1, the patient stood in the lateral position for 10 min after spinal blockade without reverse Trendelenburg position and served as a control. In Group 2, patients stood in a lateral position for 10 min after the spinal block, with a reverse Trendelenburg of 10 degrees throughout the procedure. None of the patients was sedated, as this could interfere with our sensory blockade evaluation. Patients with incomplete block/partial effect or patients unable to perforate the L2-3 or L3-4 space due to epidural block and spinal block were excluded from the study. Blood pressure , respiratory rate, heart rate, and SpOA maximum degree of sensory block was seen 20 minutes after the intrathecal injection. The 2-segment regression interval was computed as the block regression time two segments down from the maximal sensory block. The sensory analgesia duration was determined as the time required for the block level to return to T12.Requests for the first analgesic based on analgesia duration were noted. Motor blocks were tested using an adjusted Bromage scale . Grade 0GPower 3.1.9.7 was used to determine the sample size. Given a type I error of 5% and a study power of 80% to detect differences in one sensory segment, a sample size of 46 for two groups was calculated. Statistical analysis was conducted using Epi Info statistical software. Quantitative data were assessed with independent t-tests, and paired data were examined with paired t-tests. Qualitative data were assessed with chi-square and Fisher's exact test. A finding was deemed statistically significant if the value of p was below 0.05.60 patients completed our study, as shown in The two groups were similar in intraoperative hemodynamic profiles, namely heart rate, mean arterial pressure (MAP), systolic blood pressure (SBP), and diastolic blood pressure (DBP) . The senThe maximum level of sensory block achieved was higher in Group 1 than in 2 (P=0.0001). The number of intermediate segments blocked at T12 or higher was higher in Group 1 than in Group 2 (p=0.0001). In Group 2, 73.3% of patients reached levels below T8 (or 26.7% of patients had sensory levels exceeding T8). In Group 1, 46.7% of patients reached levels below T8 (or 53.3% of patients with sensory levels above T8). Two-segment regression time was longer in Group 2 (86.16±12.50 min) than in Group 1 (72±7.94 min). The sensory block time was longer in Group 2 (144±18.21 minutes) than in Group 1 (123.33±17.6 min) (p=0.0001). The motor block time was longer within Group 2 (85.66±10.8 min) than in Group 1(80.33±11.0 min). Total pain relief duration was significantly longer in Group 2 (180.13±8.47 min) than in Group 1 (165.4±6.79) (P=0.0001) . In GrouLocalized  spinal analgesia in surgery was described in 1909 by Jonnesco . Since tThe orthopedic anesthesia plan requires customization per the patient's need for a safe outcome. Low-dose bilateral and USpA with epidural anesthesia has advantages over conventional spinal anesthesia. It delivers more stable hemodynamics by providing intraoperative activation of epidural anesthesia and postoperative analgesia , 13. TheLow-dose spinal anesthesia and unilateral spinal anesthesia (USpA) with epidural anesthesia have advantages for orthopedic anesthesia. This ensures stable hemodynamics with a sufficient height of the spinal block. Spinal anesthesia has disadvantages such as hypotension and bradycardia because the block height cannot be predicted , 11. TheIn our work, the onset of sensory blockade was quicker in Group 1 than 2. Ciceki et al.  found siThe analgesia duration was also substantially prolonged in the reverse Trendelenburg group. This research was assisted by a similar work . Group 2The difference in regression time found in our study was only 10–15 min. but was similar to that of Lee et al. . Our conOne of the important limitations of our study is the use of 10 mg of the drug, which may be too high to produce a true hemispinal. Also, results of the reverse Trendelenburg position cannot be extrapolated to other population groups like obstetric patients due to physiological changes in pregnancy. Further larger sample-sized clinical examinations are needed to confirm our hypothesis.We conclude that keeping patients in reverse Trendelenburg of 10 degrees immediately after providing spinal anesthesia considerably reduces the level of sensory block. It can be clinically advantageous in controlling block height in high-risk patients involving lower limb surgeries where levels above T8 are rarely required. At the same time, low sensory block levels led to prolonged analgesia duration. Therefore, we recommend further studies using the reverse Trendelenburg position in geriatric and other high-risk groups. It can be a ray of hope to find a way that has not been achieved perfectly until today since the inception of spinal anesthesia a century ago.


Trachypithecus leucocephalus) is exclusively distributed in the karst forests and is critically endangered owing to habitat fragmentation. Gut microbiota can provide physiological data for a comprehensive study of the langur’s response to human disturbance in the limestone forest; to date, data on spatial variations in the langurs’ gut microbiota are limited. In this study, we examined intersite variations in the gut microbiota of white-headed black langurs in the Guangxi Chongzuo White-headed Langur National Nature Reserve, China. Our results showed that langurs in the Bapen area with a better habitat had higher gut microbiota diversity. In the Bapen group, the Bacteroidetes (13.65% ± 9.73% vs. 4.75% ± 4.70%) and its representative family, Prevotellaceae, were significantly enriched. In the Banli group, higher relative abundance of Firmicutes (86.30% ± 8.60% vs. 78.85% ± 10.35%) than the Bapen group was observed. Oscillospiraceae (16.93% ± 5.39% vs. 16.13% ± 3.16%), Christensenellaceae (15.80% ± 4.59% vs. 11.61% ± 3.60%), and norank_o__Clostridia_UCG-014 (17.43% ± 6.64% vs. 9.78% ± 3.83%) were increased in comparison with the Bapen group. These intersite variations in microbiota diversity and composition could be accounted for by differences in food resources caused by fragmentation. Furthermore, compared with the Banli group, the community assembly of gut microbiota in the Bapen group was influenced by more deterministic factors and had a higher migration rate, but the difference between the two groups was not significant. This might be attributed to the serious fragmentation of the habitats for both groups. Our findings highlight the importance of gut microbiota response for the integrity of wildlife habitats and the need in using physiological indicators to study the mechanisms by which wildlife responds to human disturbances or ecological variations.The white-headed black langur ( Of the Alouatta pigra; Procolobus gordonorum; Macaca mulatta) distributed in different geographical populations being grouped by altitude conditions, which is manifested by the production of new and unique microbiota (Microcebus griseorufus), which may be associated with varying degrees of human invasion of each habitat, occurring in Bale monkeys (Chlorocebus djamdjamesis) as well  are exclusively distributed in the limestone forest in Southwest Guangxi, China, and are listed as critically endangered on the IUCN Red List , which is covered by limestone forest with an altitude ranging from 400 to 600 m . Fecal s2.2.We collected langur fecal samples with the permission of the Administration Center of Guangxi Chongzuo White-headed Langur National Nature Reserve. This study did not involve any invasive animal tissue procedures. Furthermore, we collected the samples after the langurs left their sleeping sites to avoid any stress reactions caused by the collection.2.3.® Soil DNA Kit . A NanoDrop 2000  was used to detect the concentration and purity of the DNA, and 1% agarose gel electrophoresis was used to detect the extraction quality of the DNA. The TransGen AP221-02 reaction system  comprised 4 μL of 5× FastPfu buffer, 4 μL of 2.5 mM dNTPs, 0.8 μL of forward primer (5 μm) and reverse primer (5 μm), 0.4 μL of FastPfu polymerase, 0.2 μL of BSA, 10 ng of template DNA, and enough ddH2O to bring the total amount of reagent to 20 μL. In this system, highly specific amplification primers were used . Three replicates of each sample were run, and products from the same sample were mixed and detected using 2% agarose gel electrophoresis and recovered using an AxyPrep DNA Gel Extraction Kit . The PCR products were detected and quantified using a Quanti Fluor™-ST Blue Fluorescence Quantification System , after which the corresponding proportion was mixed as per the sequencing volume requirement of each sample. According to the fluorescence quantitative measurement results, a NEXTFLEX Rapid DNA-Seq kit  was used to construct a sequencing library. The sequencing platform used was an Illumina Miseq PE 300 .Total DNA was extracted from fecal samples of white-headed black langurs using an E.Z.N.A.2.4.The following procedures were performed on Majorbio Cloud PlatforminvSimpson represent the reciprocal of Simpson index, and the variations in values with the Shannon index are proportional to species diversity. The ACE and Chao indexes indicate the species richness of the community. The principal co-ordinate analysis (PCoA) reflects the similarity or variability of the community composition in the samples. The distance between different sample points was calculated using Qiime 1.9.1. Significant differences were analyzed for sample species between two sites using a Wilcoxon rank–sum test with a confidence interval of 95%, and the results were expressed as corrected p-values. Based on the Kyoto Encyclopedia of Genes and Genomes (KEGG) database, the functional prediction of gut microbiota in fecal samples was performed by PICRUSt 2   . They wem value) and the impact of random effects on the construction process (indicated by R2). The value of m is inversely proportional to the dispersal limitation, which shows that abundant taxa are generally higher than rare taxa because the latter are more likely to disappear from a single host owing to ecological drift. Within the range of 0–1, R2 represents the stochastic effect and the remaining part represents the deterministic effect (1 − R2).We then constructed the NCM based on an OTU table to demonstrate the relative importance of stochastic processes on gut microbiota community assembly . This moUsing the randomForest package, massive decision trees were constructed after processing the data according to the random and put-back sampling principle . Objects3.3.1.After the quality control, a total of 11,291,593 optimized sequences were obtained from 203 fecal samples, with an average of 55,624 ± 13,334 sequences per sample . The OTUThe langurs from the Banli and Bapen groups shared 1878 OTUs, 31 phyla, and 284 families. The Banli group possessed 1,024 unique OTUs and 79 unique families, and the Bapen group exhibited 189 unique OTUs and 21 unique families. Moreover, all groups had two unique phyla .At the phylum level, all samples were occupied by Firmicutes (Banli group: 86.30% ± 8.60% vs. Bapen group: 78.85% ± 10.35%), Bacteroidetes (4.75% ± 4.70% vs. 13.65% ± 9.73%), and Actinobacteria (3.88% ± 3.56% vs. 1.82% ± 2.04%), which accounted for >94% of the total relative abundance . At the Dominant phyla and families in the samples were analyzed for intergroup differences using the Wilcoxon rank–sum test. The results showed that the relative abundance of Firmicutes and Actinobacteria in the Banli group was significantly higher than that in the Bapen group at the phylum level, whereas Bacteroidetes and Proteobacteria in the Bapen group were significantly increased compared with the Banli group . At the We used a random forest model to construct classifiers to rank the importance of gut microbiota, which promotes the formation of intersite variations. At the phylum level, the most important taxa contributing to intersite variations in the gut microbiota of white-headed black langurs were Bacteroidetes, followed by Proteobacteria and Campylobacterota. At the family level, these variations were accounted for by Butyricoccaceae, Peptococcaceae, and Rikenellaceae. The AUC values of the phylum and family levels were 0.963 and 1.000, respectively, which indicate an exceptional classification .3.2.invSimpson index (57.575 ± 22.396 vs. 73.890 ± 23.367) of the gut microbiota of langurs in the Banli and Bapen group differed significantly . However, the ACE index and Chao index did not show significant intersite variations  and riations . The detR2 = 0.093, p = 0.001) between geographic groups  than in the Banli group (m = 0.124). Of the gut microbiota of the Banli group, 59.7% were within the theoretical prediction range of the random effects calculated by the model, whereas 23.6% were above this value and 16.7% were below it. The proportion of gut microbiota in the Bapen group that fell within the interval of theoretical predictions as measured by the model was 78.8%, with 13.2% above this limit and 8.0% below it  and 48.1% (for the Bapen group) of the community assembly of the white-headed black langurs’ gut microbiota. The migration rate of gut microbiota was higher in the Bapen group  (0.98) (Eulemur fulvus) (0.5) (Macaca thibetana) (2.65)  (0.98)  and brows) (0.5) , as well) (2.65) . In this) (2.65) . The higBifidobacteria, the most widely recognized, is extensively used as a probiotic and can also assist in the hydrolysis of plant polysaccharides by producing glycosyl hydrolases , which indicates that the community assembly of the gut microbiota of langurs is jointly built on a combination of stochastic and deterministic processes. Our results were similar to those of a study on latitudinal phytoplankton distribution  can be found at: The animal study was reviewed and approved by Administration Center of Guangxi Chongzuo White-headed Langur National Nature Reserve.ZH designed the study. YiL and YC analyzed the data. YiL wrote the manuscript. JZ, ZL, DN, and JL collected samples. YoL and ZH revised the manuscript. All authors read and approved the submitted manuscript.This work was supported by the National Natural Science Foundation of China .The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


To investigate the feasibility and dosimetric index features of dose painting guided by perfusion heterogeneity for brain metastasis (BMs) patients.H) with less than 25% of the maximum CBF value were defined as the dose escalation region. The planning target volume (PTV) and PTVH were calculated from GTV and GTVH respectively. The PTVN was obtained by subtracting PTVH from PTV, and conventional dose was given. Three kinds of radiotherapy plans were designed based on the CBF values. Plan 1 was defined as the conventional plan with an arbitrary prescription dose of 60 Gy for PTV. For dose painting, Plan 2 and Plan 3 escalated the prescription dose for PTVH to 72 Gy based on Plan 1, but Plan 3 removed the maximum dose constraint. Dosimetric indices were compared among the three plans.A total of 50 patients with single BMs were selected for this study. CT and MR simulation images were obtained, including contrast-enhanced T1-weighted images (T1WI+C) and cerebral blood flow (CBF) maps from 3D-arterial spin labeling (ASL). The gross tumor volume (GTV) was determined by fusion of CT and T1WI+C images. Hypoperfused subvolumes  cm3, accounting for 49.3% of GTV. Both conventional plan and dose painting plans achieved 98% target coverage. The conformity index of PTVH were 0.44 (Plan1), 0.64 and 0.72 . Compared to Plan 1, the D2%, D98% and Dmean values of the PTVH escalated by 20.50%, 19.32%, and 19.60% in Plan 2 and by 24.88%, 17.22% and 19.22% in Plan 3 respectively (P<0.05). In the three plans, the index of achievement value for PTVH was between 1.01 and 1.03 (P<0.05). The dose increment rates of Plan 2 and Plan 3 for each organs at risk (OARs) was controlled at 2.19% - 5.61% compared with Plan 1. The doses received by OARs did not significantly differ among the three plans (P >0.05).The mean GTV volume was 34.5 (8.4-118.0) cmBMs are associated with significant heterogeneity, and effective escalation of the dose delivered to target subvolumes can be achieved with dose painting guided by 3D-ASL without extra doses to OARs. Brain metastases (BMs) are the most common intracranial malignant tumors, and approximately 8-10% of tumor patients will develop BMs during their disease course . And larAt present, radiotherapy (RT) is considered a curative-intent treatment for BMs patients primarily consisting of whole-brain radiotherapy (WBRT) and stereotactic radiosurgery (SRS) , 7. WBRT3D-arterial spin labeling (3D-ASL) perfusion imaging analyzes CBF parameters noninvasively and quantitatively and is independent of blood-brain barrier (BBB) damage . In addiIncreasing the doses to BMs can significantly improve the curative effect. Therefore, safe dose escalation for target subvolumes of BMs is crucial. This study therefore explored the feasibility of 3D-ASL-guided subvolume segmentation of BMs based on CBF map variation. In addition, we further studied the dosimetric indices of dose painting plans to provide additional reference for the formulation and modification of individualized RT to BMs patients.Fifty patients, namely, 29 males and 21 females , with a single BMs who received RT were selected from July 2018 to September 2020. There were 33 cases of lung cancer, 9 of breast cancer, 4 of renal carcinoma, 2 of esophageal cancer and 2 of colon cancer. All patients were diagnosed with BMs with imaging and the maximum tumor cross-sectional diameter was more than 2 cm. The retrospective analysis of the medical records was approved by the Institutional Review Board of Shandong Cancer Hospital.CT simulation images  were obtained by Brilliance Big Bore CT scanner . MR imaging was performed on 3.0 T Discovery 750 W MR scanner  with the same head position as CT simulation. 3D-ASL and contrast-enhanced T1-weighted (T1WI+C) images were acquired using 3D volume scanning . For 3D-ASL images, the special acquisition conditions were as follows: repetition time (TR) = 5160ms; echo time (TE) = 11.5 ms; and postlabeling delay (PLD) = 2025 ms. David et al. pointed out that it is more reasonable to set the PLD of healthy people older than 70 years old and adult clinical patients to around 2000ms . For T1WH), hyperperfused and nonperfused subvolumes based on CBF values , and gross tumor volume (GTV) was defined as the region with high signal. The GTV was divided into hypoperfused  constraint of 66 Gy (110% of prescription dose). For dose painting, Plan 2 escalated the PTVH to 72 Gy with a Dmax constraint of 79 Gy (110% of prescription dose) based on Plan 1. Moreover, Plan 3 was designed depending on Plan 2 without Dmax constraint.Conventional and dose painting plans were implemented by intensity-modulated radiotherapy (IMRT) and simultaneous integrated boost (SIB) IMRT. These plans were designed using Eclipse . For larger tumors, with the most common doses being 27 Gy in 3 fractions and 30 Gy in 5 fractions applying with SRS based on NCCN . Prelimimax<50 Gy for eye, Dmax<54 Gy for optic nerve, Dmax< 8 Gy for lens, and Dmax<54 Gy for brainstem.The optimized parameters and dose constraints for organs at risk (OARs) among three plans were unified. OARs were restricted to the following doses: DDose calculation was performed in anisotropic analytical algorithm optimization mode (version 15.512) using 6 MV X-rays. The calculated grid was 2.5 mm×2.5 mm, and the prescription dose was acquired to cover 95% of the target volume.2%, D98% and Dmean  were the maximum, minimum and mean dose, respectively. The Dmax values of eyeballs, optic nerves, lenses and brainstem were compared. Furthermore, the target coverage, conformity index (CI) and index of achievement (IOA) were calculated. The proposed IOA is formulated as the volume-weighted average of the deviation between prescription dose and planned dose  is the volume of the kth PTV, where j is the total number of volumes, Dj is the jth volumes dose value, and dDVHPTV(K)(Dj) is the absolute volume (cc) of the jth dose volume in the kth PTV.where CI near 1 indicates that region receiving the reference dose closely matches the shape of the target region . IOA havF value is used to evaluate the difference between groups. The larger the F is, the more significant the equation is, and the better the fitting degree is. The least significant difference was used in pairwise comparisons. All data are expressed as mean ± standard deviation P<0.05 indicates a significant difference.Statistical analysis was performed using SPSS Statistics Version 22.0 . Analysis of variance was used to evaluate the differences among the three plans. 3, with an average volume of 34.5 cm3. And mean GTVH was 17.0 (4.5-58.3) cm3,accounting for 49.3% of the GTV. The PTV, PTVH and PTVN were 72.0 cm3, 41.5 cm3 and 30.5 cm3, respectively. The ratios of PTVH to PTV and PTVN to PTV were 57.6% and 42.4%, respectively, as shown in GTV had a volume range of 8.4-118.0 cmH up to 99.90%. Compared with Plan 2, Plan 3 significantly increased the CI levels of PTVH by 12.50% (P< 0.05). Owing to the use of targeted gradient doses instead of a group-based uniform dose, CI values of the PTV and PTVN were lower in the dose painting plans than in the conventional plans. For PTVH, the IOA values of the three plans were between 1.01 and 1.03 (P< 0.05). Meanwhile, the IOA values of PTV and PTVN were between 1.01 and 1.10 (P< 0.05).As shown in 2% were statistically significant (P< 0.05). Compared to those of Plan 1, the D2%, D98% and Dmean of the PTV were increased by 20.18%, 8.34% and 18.38% in Plan2 and by 24.05%, 6.77% and 17.00%, respectively, in Plan 3. The dosimetric indices above for PTVH were increased by 20.50%, 19.32% and 19.60% in Plan 2, while those of Plan 3 were increased by 24.88%, 17.22% and 19.22%. Similarly, the dosimetric indices of PTVN were increased in Plan 2 and Plan 3 . Additionally, the D2% values of PTVs showed increasing trends among three plans, while the increment rates of D98% and Dmean for Plan 2 were higher than those for Plan 3, as shown in The differences in all dosimetric indices of PTVs except Dmax received by OARs between conventional and dose painting plans (P > 0.05). Nevertheless, the dose painting plans slightly increased the Dmax of all OARs compared to Plan 1, while right eye increased by 5.61% to 0.50 Gy in Plan 3. The other OARs increased by less than 5.40% (2.19% - 5.30%), as shown in There was no signifificant difference in DThis study demonstrated that BMs can be divided into high and low perfusion subvolumes based on 3D-ASL. We therefore presented a new method for assessing the clinical feasibility of dose painting plans directed by 3D-ASL for BMs. The results showed that the technique above can be completed feasibly without increasing the dose delivered to OARs which holds the potential to achieve better control of BMs than traditional RT treatment.BMs are significant health problems whose incidence is increasing, and the median survival time is only 1-2 months without treatment . NumerouSome studies have shown that BMs are highly heterogeneous according to their pathological sources or even sites of pathological origin . Brown e18F-fluoromisonidazole , the Taishan Scholars Project of Shandong Province (Grant No. ts201712098) and National Key Research and Development Program of China (Grant No. 2017YFC0113202).We gratefully acknowledge Qingtao Qiu for his support with text proofing.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.