The development and implementation of pathological parameters and molecular testing impact prognosis of colorectal adenocarcinoma

Objective:This study aims to analyze how changes in pathological diagnosis practice and molecular detection technology have affected clinical outcomes for colorectal cancer(CRC)patients in Fudan University Shanghai Cancer Center(FUSCC).Methods:This retrospective cohort study analyzed 21,141 pathologically confirmed CRC cases diagnosed at FUSCC from 2008 to 2020.Patients were divided into five groups for different analytical purposes:(1)the before vs.since 2014 groups to analyze the influence of the changes in the classification criteria of pT3 and pT4 staging on the survival of patients;(2)the partial vs.total mesorectal excision(TME)groups to analyze whether evaluation of completeness of the mesorectum have impact on the survival of patients;(3)the tumor deposit(TD)(+)N0 vs.TD(+)N1c groups to analyze the influence of the changes in the pN staging on the survival of patients with positive TD and negative regional lymph node metastasis(LNM);(4)the before vs.since 2013 groups to analyze the influence of the changes in the testing process of deficient mismatch repair on the survival of patients;and(5)the groups with vs.without RAS/BRAF gene mutation testing to analyze the influence of these testing on the survival of patients.Patients’clinicopathological parameters,including age at diagnosis,sex,tumor size,location,differentiation,mucinous subtype,TD,lymphovascular invasion,perineural invasion,tumor depth,LNM and distant metastasis,and tumor-node-metastasis(TNM)stage,were compared between groups.Kaplan-Meier analysis with log rank method was performed for patients’overall survival(OS)and disease-free survival(DFS)analyses.Results:In pathological reports,there were three parameter changes that impacted patient outcomes.Firstly,changes in the pT staging criteria led to a shift of the ratio of patients with stage pT3 to stage pT4 from 1:110.9 to 1:0.26.In comparison to patients admitted before 2014(n=4,754),a significant difference in prognosis between pT3 and pT4 stages was observed since 2014(n=9,965).Secondly,we began to evaluate the completeness of the mesorectum since 2016.As a result,91.0%of patients with low rectal cancer underwent TME(n=4,111)surgery,and patients with TME had significantly better OS compared with partial mesorectal excision(PME,n=409).Thirdly,we began to stage TD(+)LNM(-)as N1c since 2017.The results showed that N1c(n=127)but not N0(n=39)can improve the prognosis of patients without LNM and distal metastasis.In molecular testing,there have been three and five iterations of updates regarding mismatch repair(MMR)/microsatellite instability(MSI)status and RAS/BRAF gene mutation detection,respectively.The standardization of MMR status testing has sharply decreased the proportion of deficient MMR(dMMR)patients(from 32.5%to 7.4%)since 2013.The prognosis of patients underwent MMR status testing since 2013(n=867)were significantly better than patients before 2013(n=1,313).In addition,detection of RAS/BRAF gene mutation status(n=5,041)resulted in better DFS but not OS,for patients with stage I-III disease(n=16,557).Conclusion:Over the past few decades,updates in elements in pathological reports,as well as the development of standardized tests for MMR/MSI status and RAS/BRAF gene mutations have significantly improved patient outcomes.

Comprehensive Assessment of Anxiolytic Properties in 4-HPAA Derivatives: Bridging in Vivo Validation and Molecular Docking Analyses

Anxiety is a significant mental health issue that substantially affects an individual’s quality of life. Feelings of uneasiness, irritability, and sleep disturbances characterize it. 4-Hydroxyphenyl acetic acid (4-HPAA) is identified in brain cells as a physiological byproduct of tyramine. This study hypothesizes that 4-HPAA may regulate anxiety due to its anxiolytic properties, acting as a modulator of the GABAergic system, which plays a crucial role in the pathophysiology of anxiety disorders. Our study aims to enhance the anxiolytic effects of 4-HPAA through chemical modification to improve its pharmacokinetic properties. Three derivatives, namely Isopropyl-4-hydroxy-[phenyl] acetate (IHPA), Isopropyl-4-hydroxy-[phenyl] acetate (MPAA), and 4-methoxyphenyl acetate (MPHA), have been synthesized from 4-HPAA. This assessment will use well-established animal models, specifically the Elevated Plus-Maze (EPM) and Zero Maze (EZM) tests, selected for their validity in replicating anxiety-like symptoms in animals. Chronic caffeine administration via drinking water (0.3 g/l for 14 days) was employed to induce an anxiety state for testing purposes. IHPA and MPAA demonstrated significant anxiolyticactivity when tested in the EPM and EZM experiments. Molecular docking simulations using AutoDock Vina indicated that 4-HPAA derivatives had docking scores ranging from −5.8 to −4.8 kcal/mol, compared to the standard anxiolytic medication Diazepam, which scored −7.1 kcal/mol. These scores suggest a potential for 4-HPAA derivatives to interact effectively with the Gamma-aminobutyric acid (GABA_A) receptor. In conclusion, our in vivo and in silico analyses indicate a promising anxiolytic potential for 4-HPAA derivatives.

Optimization and Testing for PCR System of Rice by Orthogonal Design

[Objective] This study was to screen the economic or stable PCR system of rice and detect the generality of the selected system in different molecular markers based on PCR.[Method] With DNA extracted from rice leaves by CTAB method as the template,PCR system was optimized by L16(45)orthogonal design.[Result] Clear bands were amplified from 16 different combinations,but the amplification effects and yields had difference.The most economic and applicable system was as follows:20 ng DNA template,150 μmol/L dNT...

Antimicrobial susceptibility testing for Helicobacter pylori in times of increasing antibiotic resistance

The gram-negative bacterium Helicobacter pylori(H.pylori)causes chronic gastritis,gastric and duodenal ulcers,gastric cancer and mucosa-associated lymphoid tissue lymphoma.Treatment is recommended in all symptomatic patients.The current treatment options for H.pylori infection are outlined in this review in light of the recent challenges in eradication success,largely due to the rapid emergence of antibiotic resistant strains of H.pylori.Antibiotic resistance is a constantly evolving process and numerous studies have shown that the prevalence of H.pylori antibiotic resistance varies significantly from country to country,and even between regions within the same country.In addition,recent data has shown that previous antibiotic use is associated with harbouring antibiotic resistant H.pylori.Local surveillance of antibiotic resistance is warranted to guide clinicians in their choice of therapy.Antimicrobial resistance is assessed by H.pylori culture and antimicrobial susceptibility testing.Recently developed molecular tests offer an attractive alternative to culture and allow for the rapid molecular genetic identification of H.pylori and resistance-associated mutations directly from biopsy samples or bacterial culture material.Accumulating evidence indicates that surveillance of antimicrobial resistance by susceptibility testing is feasible and necessary to inform clinicians in their choice of therapy for management of H.pylori infection.

Comparison of Histological, Microbiological, and Molecular Methods in Diagnosis of Patients with TBLN Having Different Anti-TB Treatment Background

Objective The influence of anti-tuberculosis （TB） treatment history on tuberculous lymphadenitis （TBLN） diagnosis is unclear. Therefore, this study aims to evaluate the diagnostic methods, including histology, microbiology, and molecular tests, used for TBLN. Methods In this study, suspected patients with TBLN and having different anti-T8 treatment background were enrolled. All the samples were tested simultaneously by histology, ZiehI-Neelsen （ZN） staining, mycobacterial culture （culture）, Xpert MTB/RIF （xpert）, real-time PCR, and high-resolution melting curve PCR （HRM）. Thereafter, the performance of these methods on samples with different anti-TB treatment background was assessed. Results In our study, 89 patients were prospectively included 82 patients with TBLN and 7 with other diseases. The overall sensitivities of Xpert, real-time PCR, histology, ZN staining, and culture were 86.6%, 69.5%, 58.5%, 43.9%, and 22.0%, respectively. The anti-TB treatment history revealed dramatic influences on the sensitivity of culture （P 〈 0.0001）. In fact, the treatment that lasted over 3 months also influenced the sensitivity of Xpert （P 〈 0.05）. However, the treatment history did not affect the performance of remaining tests （P 〉 0.05）. For rifampicin drug susceptibility test （DST）, the anti-T8 treatment showed only significant influence on the success rate of culture DST （P = 0.001）, but not on those of Xpert and HRM tests （P 〉 0.05）. Conclusion Other tests as welt as culture should be considered for patients with TBLN having retreatment history or over 1-month treatment to avoid false negative results.

Noninvasive molecular analysis of Helicobacter pylori : Is it time for tailored first-line therapy?

The main problem of Helicobacter pylori(H. pylori) infection management is linked to antibiotic resistances. This phenomenon has grown in the last decade, inducing a dramatic decline in conventional regimen effectiveness. The causes of resistance are point mutations in bacterial DNA, which interfere with antibiotic mechanism of action, especially clarithromycin and levofloxacin. Therefore, international guidelines have recently discouraged their use in areas with a relevant resistance percentage, suggesting first-line schedules with expected high eradication rates, i.e., bismuth containing or non-bismuth quadruple therapies. These regimens require the daily assumption of a large number of tablets. Consequently, a complete adherence is expected only in subjects who may be motivated by the presence of major disorders. However, an incomplete adherence to antibiotic therapies may lead to resistance onset, since sub-inhibitory concentrations could stimulate the selection of resistant mutants. Of note, a recent meta-analysis suggests that susceptibility tests may be more useful for the choice of first than second-line or rescue treatment. Additionally, susceptibility guided therapy has been demonstrated to be highly effective and superior to empiric treatments by both meta-analyses and recent clinical studies. Conventional susceptibility test is represented by culture and antibiogram. However, the method is not available everywhere mainly for methodology-related factors and fails to detect hetero-resistances. Polymerase chain reaction(PCR)-based, culture-free techniques on gastric biopsy samples are accurate in finding even minimal traces of genotypic resistant strains and hetero-resistant status by the identification of specific point mutations. The need for an invasive endoscopic procedure has been the most important limit to their spread. A further step has, moreover, been the detection of point mutations in bacterial DNA fecal samples. Few studies on clarithromycin susceptibility have shown an overall high sensitivity and specificity when compared with culture or PCR on gastric biopsies. On these bases, two commercial tests are now available although they have shown some controversial findings. A novel PCR method showed a full concordance between tissue and stool results in a preliminary experience. In conclusion, despite poor validation, there is increasing evidence of a potential availability of noninvasive investigations able to detect H. pylori resistances to antibiotics. These kinds of analysis are currently at a very early phase of development and caution should be paid about their clinical application. Only further studies aimed to evaluate their sensitivity and specificity will afford novel data for solid considerations. Nevertheless, noninvasive molecular tests may improve patient compliance, time/cost of infection management and therapeutic outcome. Moreover, the potential risk of a future increase of resistance to quadruple regimens as a consequence of their use on large scale and incomplete patient adherence could be avoided.

A Brief Overview of Research on Chemical Molecular Contamination in Cleanrooms

This review proposes associated standards from abroad regarding chemical contamination of the air, and has analyzed the formation of chemical air pollution, the common contaminants and the testing methods of projects. This paper demonstrates the capability of China to begin commercial testing of chemical contamination of the air. In addition, it offers technological references of the feasibility for adopting the international standard ISO14644-8：2006.

Selection and Molecular Biological Identification of a Strain of Bacillus sp. Inhibiting the Growth of Saprolegnia ferax

Based on the theory of biological control of Saprolegnia ferax,antagonism test of nine strains of Bacillus sp. to S. ferax JL was carried out. Bacillus sp.BA1 was screened to have significantly inhibitory effects on the growth of S. ferax JL( P 【 0. 05). Then,the effects of Bacillus sp. BA1 on different sources of S. ferax were carried out. Results showed that BA1 also had significantly inhibitory effects on S. ferax 6#,10# and S2( P 【 0. 05). Sequence of 16 S r DNA of BA1 was analyzed; and homologous alignment analysis showed that BA1 had more than 99% similarity with Bacillus cereus. Therefore,it could be concluded that strain BA1 was B. cereus,which significantly inhibited the growth of S. ferax and could be used as the biological control agent for S. ferax diseases in aquaculture.

New fecal test for non-invasive Helicobacter pylori detection:A diagnostic accuracy study

AIM To assess the diagnostic accuracy of a new fecal test for detecting Helicobacter pylori(H. pylori), using ^(13)Curea breath test as the reference standard, and explore bacterial antibiotic resistance. METHODS We conducted a prospective two-center diagnostic test accuracy study. We enrolled consecutive people≥ 18 years without previous diagnosis of H. pylori infection, referred for dyspepsia between February and October 2017. At enrollment, all participants underwent 13 C-urea breath test. Participants aged over 50 years were scheduled to undergo upper endoscopy with histology. Participants collected stool samples 1-3 d after enrollment for a new fecal investigation(THD fecal test). The detection of bacterial 23 S rRNA subunit gene indicated H. pylori infection. We also used the index diagnostic test to examine mutations conferring resistance to clarithromycin and levofloxacin. Independent investigators analyzed index test and reference test standard results blinded to the other test findings. We estimated sensitivity, specificity, positive(PPV) and negative(NPV) predictive value, diagnostic accuracy, positive and negative likelihood ratio(LR), together with 95% confidence intervals(CI).RESULTS We enrolled 294 consecutive participants(age: Median 37.0 years, IQR: 29.0-46.0 years; men: 39.8%). Ninetyfive(32.3%) participants had a positive ^(13)C-urea breath test. Twenty-three(7.8%) participants underwent upper endoscopy with histology, with a full concordance between ^(13)C-urea breath test and histology in detecting H. pylori infection. Four(1.4%) out of the 294 participants withdrew from the study after the enrollment visit and did not undergo THD fecal testing. In the 290 participants who completed the study, the THD fecal test sensitivity was 90.2%(CI: 84.2%-96.3%), specificity 98.5%(CI:96.8%-100%), PPV 96.5%(CI: 92.6%-100%), NPV 95.6%(CI: 92.8%-98.4%), accuracy 95.9%(CI: 93.6%-98.2%), positive LR 59.5(CI: 19.3-183.4), negative LR 0.10(CI: 0.05-0.18). Out of 83 infected participants identified with the THD fecal test, 34(41.0%) had bacterial genotypic changes consistent with antibiotic-resistant H. pylori infection. Of these, 27(32.5%) had bacterial strains resistant to clarithromycin, 3(3.6%) to levofloxacin, and 4(4.8%) to both antibiotics. CONCLUSION The THD fecal test has high performance for the non-invasive diagnosis of H. pylori infection while additionally enabling the assessment of bacterial antibiotic resistances.

Cancer Chemosensitivity Testing: Review

Since chemotherapy started in 1940s, chemosensitivity testing has been both a very attractive field and one fraught with potential pitfalls. Many methods were developed and brought initial promises, yet later ending in disappointment and were eliminated. For example, in the 1970s clonogenic assay was generally believed to be the best testing method for predicting clinical outcome. However, technical problems including low evaluation rate limited its use. Currently, MTT, ATP, DISC and Kern’s assay hold better promises. Since the 90s, the study of molecular biology has been progressing rapidly. It accelerated the understanding of molecular mechanisms of drug response. Numerous papers were published, but only few techniques can be applied in clinical practice. This review summarizes the controversies and current progress of chemosensitivity assays based on available online information, and makes a suggestion about their future routine practice.

Genetic testing vs microforceps biopsy in pancreatic cysts:Systematic review and meta-analysis

BACKGROUND Carcinoembryonic antigen(CEA)and cytology in pancreatic cystic fluid are suboptimal for evaluation of pancreatic cystic neoplasms.Genetic testing and microforceps biopsy are promising tools for pre-operative diagnostic improvement but comparative performance of both methods is unknown.AIM To compare the accuracy of genetic testing and microforceps biopsy in pancreatic cysts referred for surgery.METHODS We performed a literature search in Medline,Scopus,and Web of Science for studies evaluating genetic testing of cystic fluid and microforceps biopsy of pancreatic cysts,with endoscopic ultrasound with fine-needle aspiration(EUSFNA)prior to surgery and surgical pathology as reference standard for diagnosis.We evaluated the diagnostic accuracy for:1-benign cysts;2-mucinous low-risk cysts;3-high-risk cysts,and the diagnostic yield and rate of correctly identified cysts with microforceps biopsy and molecular analysis.We also assessed publication bias,heterogeneity,and study quality.RESULTS Eight studies,including 1206 patients,of which 203(17%)referred for surgery who met the inclusion criteria were analyzed in the systematic review,and seven studies were included in the meta-analysis.Genetic testing and microforceps biopsies were identical for diagnosis of benign cysts.Molecular analysis was superior for diagnosis of both low and high-risk mucinous cysts,with sensitivities of 0.89(95%CI:0.79-0.95)and 0.57(95%CI:0.42-0.71),specificities of 0.88(95%CI:0.75-0.95)and 0.88(95%CI:0.80-0.93)and AUC of 0.9555 and 0.92,respectively.The diagnostic yield was higher in microforceps biopsies than in genetic analysis(0.73 vs 0.54,respectively)but the rates of correctly identified cysts were identical(0.73 with 95%CI:0.62-0.82 vs 0.71 with 95%CI:0.49-0.86,respectively).CONCLUSION Genetic testing and microforceps biopsies are useful second tests,with identical results in benign pancreatic cysts.Genetic analysis performs better for low-and high-risk cysts but has lower diagnostic yield.

Molecular Detection and Sequencing for S1 Glycoprotein Gene of Bronchitis Virus of 2016 Epidemic from Sindh and Punjab

Infectious Bronchitis (IB) is highly contagious disease of commercial poultry causing substantial economic loses by producing poor quality meat in broilers and effecting production in breeder birds. The causative agent has been reported as most hazardous pathogen among other infectious agent even after being immunized with multi-variant strain vaccine. Currently, different strain such as H-120, 4/91 and D274 have been used extensively for immunoprophylaxis against velogenic strain across Pakistan with minimal protection reported. In current study PCR analysis was used to investigate the molecular nature of IB isolates from Punjab and Sind province of Pakistan in 2016 epidemics. Total of 100 tracheal samples were considered for virus inoculation in 10 days old chicken embryonated eggs. The IBV infected amniotic fluid was neutralized with monoclonal antisera of H-120, 4/91 and D274 strains. The IBV screened samples were subjected for RNA extraction and subsequent to PCR using type specific primer of each strain. The amplified product of 840 bp was sequenced through Sanger sequencing. On the basis of PCR results, four similar amplified products from both regions were obtained showing similarities in agarose gel electrophoresis, but they differ from each other on the basis of nucleotides sequence. Phylogenetic analysis revealed that nucleotide sequences of isolates from Karachi were similar to the IBV H-120, Mass-41 and Connecticut 46 reference strains. Whereas, isolates from the Punjab province are analogous to the Mans-2, Mans-3, 9/41(UK) but did not show significant similarity with other reference strain. Therefore, it is recommended that use of M-41 and H-120 in vaccine production could be effective measure against velogenic infectious agent in Sindh particularly in Karachi, whereas, it would be better to incorporate either of the variant GQ281656.1, AY279533.1 in vaccine because of their highest level of resemblance with genetically sequenced isolates from Lahore and its surroundings.

梅毒实验室诊断的研究现状与展望

[背景]近年来,梅毒发病数呈逐年上升趋势,引起广泛关注,已成为全球性公共卫生问题.鉴于梅毒病程的隐匿性、复杂性和多样性,迫切需要高灵敏度的诊断方法和有效的疗效监测指标,以实现梅毒的早诊断和早治疗.[进展]本文对梅毒实验室检测方法及诊断程序的研究现状进行综述.病原学检查可以直接确诊感染,但易受取材部位及取材方法等因素影响,灵敏度较低.血清学检测是梅毒诊断的主要检测方法,但由于抗体至少在感染2周后才产生,难以筛查窗口期的患者.由于血清学诊断程序的行业标准尚未统一,各实验室对3种血清学诊断程序的选择也不尽相同.聚合酶链式反应(polymerase chain reaction,PCR)对于一期或二期梅毒患者皮损组织具有较高灵敏度和特异度,但对于较易获得的血液样本,其梅毒螺旋体(Treponema pallidum)DNA检出率低,国内至今尚无获得医疗器械注册证的梅毒PCR检测试剂,无法应用于临床检测.[展望]首先,研发梅毒螺旋体抗体和非特异性抗体联合检测试剂,避免单一试验假阴性造成的漏诊;其次,开展血液、脑脊液、尿液等标本的梅毒螺旋体抗原谱研究,缩短检测的窗口期;最后,引入质量管理体系,制定梅毒分子诊断的统一标准.

应用于食品检测领域的分子即时检测技术研究进展

食品安全问题关系人类民生。为有效预防和控制食品安全风险,减少食品安全经济损失,迫切需要开发出针对食源性病原体、食物掺假、转基因作物的高特异性、高敏感性、快速的核酸检测技术。即时检测(point-of-care testing,POCT)作为一种新兴的快速检测手段,由于其检测速度快、操作简单、现场检查等特点,在食品检测领域应用广泛。本文首先介绍了POCT技术的发展历史,而后根据食品安全事件的特点,从食源性致病菌、食物掺假、转基因作物3个方向分析了食品检测的主要对象,阐述了分子POCT技术在食品检测领域的应用现状,最后对应用于食品检测领域的分子POCT技术存在的问题和解决办法进行了分析总结,对食品检测分子POCT技术的发展方向进行了展望。本文有助于进一步了解分子POCT技术在食品检测中的应用,并为分子POCT技术在食品快速检测中的研究和应用提供参考。

2022年结核分枝杆菌分子检测能力验证结果分析

目的通过组织实施能力验证了解我国实验室在结核分子检测能力方面的现状。方法在2022年组织相关实验室报名参加本次能力验证,并制备能力验证样本并发放给参加实验室。通过对参加实验室的所在地区、实验室类型、上报结果和错误类型等方面进行统计和分析,评估其结核分子检测能力和管理水平。结果共有来自于11个省份的20家实验室参加本次能力验证活动,总体满意率为89.5%。不满意的原因主要为将阴性样本错误地检测为阳性。部分实验室出现填报结果错误等实验室管理问题。结论大多数参加本次能力验证的实验室具有较好的结核分子检测能力,但部分实验室应当关注实验室核酸交叉污染和实验室质量管理问题。本能力验证有助于进一步提升我国实验室的结核分子检测能力。

PCR荧光探针技术对结核分枝杆菌复合群及利福平耐药性检测性能的评价

目的:评价PCR荧光探针技术检测痰液样本中结核分枝杆菌复合群(Mycobacterium tuberculosis complex,MTBC)和利福平耐药性效果,为进一步开展“结核分枝杆菌复合群及利福平耐药核酸检测试剂盒”多中心临床试验提供可行性研究数据。方法:采用前瞻性研究方法,参照入组标准收集2021年12月至2022年8月上海市3家结核病定点医院肺科门诊就诊的205例初诊疑似肺结核患者的3份痰液样本(即时痰、夜间痰、晨痰),分别进行痰涂片、BACTEC MGIT 960(MGIT 960)、GeneXpert MTB/RIF(Xpert)和PCR荧光探针法检测,评价PCR荧光探针法对痰液样本中MTBC及利福平耐药性的检测效能。另对随机选取同期上海市疾病预防控制中心结核病实验室菌株库中保存的96株MTBC临床菌株(包括利福平表型耐药菌株47株和利福平表型敏感菌株49株)分别进行PCR荧光探针法和Xpert检测利福平耐药性,分析两种检测方法对利福平耐药相关基因突变的检出情况。结果:205例疑似肺结核患者中,PCR荧光探针法对MTBC的检出率为22.44%(46/205),与MGIT 960[21.95%(45/205)]、Xpert[20.98%(43/205)]和痰涂片[16.10%(33/205)]的差异均无统计学意义(χ^(2)=0.014,P=0.904;χ^(2)=0.129,P=0.719;χ^(2)=2.650,P=0.104)。以临床诊断为参照标准,PCR荧光探针法、Xpert、MGIT 960和痰涂片检测MTBC的敏感度(95%CI)分别为48.10%(36.93%~59.56%)、48.10%(36.93%~59.56%)、50.63%(39.23%~61.97%)和40.51%(29.79%~52.15%),特异度(95%CI)分别为93.65%(87.47%~97.12%)、96.03%(90.52%~98.53%)、96.03%(90.52%~98.53%)和99.21%(95.01%~99.96%),一致率分别为76.10%(156/205)、77.56%(159/205)、78.54%(161/205)和76.59%(157/205),Kappa值分别为0.453、0.482、0.507和0.446。PCR荧光探针法检测痰液样本中MTBC的涂阴和极低级别阳性样本的检出率[10.73%(19/177)]与Xpert检测结果[8.47%(15/177)]的差异无统计学意义(χ^(2)=0.793,P=0.373)。PCR荧光探针法在检出MTBC的46份样本中同时检出6份样本发生利福平耐药基因突变,突变率为13.04%(6/46);而Xpert在检出MTBC的43份样本中仅同时检出1份样本发生利福平耐药基因突变,突变率为2.33%(1/43)。在96株临床菌株中,两种分子检测方法均检测到56株菌株发生利福平耐药基因突变,除2株探针突变类型不一致外,其他探针突变类型均一致,且多发生在probe E(529~533)和FAM通道(531/533突变)。结论:以临床诊断为参照标准,PCR荧光探针法检测MTBC的效能与MGIT 960、Xpert和痰涂片基本相当,且与Xpert检测MTBC临床菌株的利福平耐药性效果一致,认为PCR荧光探针法不仅是一种等同于Xpert检测功能的新技术,还具有操作简单、价格低于Xpert、真正实现一体化和全封闭等优势。建议在临床推广过程中进一步优化PCR荧光探针法以提高MTBC检出率,并开展多中心临床试验验证研究。

The Isolation of Vibrio cholera and Other Enteric Bacteria with Molecular Characterization of Vibrio cholera during the Outbreak of Baghdad/Iraq in 2015

Vibrio cholera, causing acute watery diarrhea known as cholera disease, affects all ages and both genders. Cholera infection outbreaks in Iraq have been reported for several years. The recent cholera outbreak, emerged throughout 2015, was investigated using bacteriological laboratory tests, singleplex and multiplex PCR technique for the detection of V. cholera from stool samples. Furthermore the toxigenic potential coupled with the antibiotic susceptibility test for cholera and other bacteria were also investigated. The stool samples were collected from 5698 patients admitted to Al-Yarmouk Teaching hospital and health care centers in Baghdad/Al-Karkh, Iraq, from the 1st of August to the 30th of December 2015. The V. cholera was isolated from 194 cases (3.4% of the cases age between 21 - 50 years). In addition, other enteric infections: Salmonellosis and Shigellosis 7 and 21 respectively, protozoan parasite Giardia lamblia and Entamoeba histolytica 2 and 43 cases respectively were also reported. High percentage of V. cholera infection was detected in October (122 cases, 62.8%), compared with other enteric infections that show high percentage of diarrheal disease in September and November. The results have confirmed that the cholera outbreak was caused by V. cholera O1, biotype El Tor, and serotype Inaba. Seven virulence genes were identified ctxA, toxR, zot, ace, rfbO1, tcpA and ompW. Moreover, the cholera isolated strains were found sensitive to most antibiotic but resistant to nalidixic acid.

低分子肝素及胚胎植入前非整倍体检测对不明原因复发性流产患者妊娠结局的影响

目的:比较行胚胎植入前非整倍体检测(PGT-A)助孕和未行PGT-A助孕的不明原因复发性流产(URSA)患者应用低分子肝素(LMWH)后临床结局的差异。方法:回顾分析2016年1月1日至2022年5月17日URSA患者于郑州大学第三附属医院生殖医学科首次移植冷冻囊胚共1158个周期的临床资料,按移植前是否行PGT-A分为PGT-A组(行PGT-A+ICSI,315例)和非PGT-A组(行IVF/ICSI,843例),再根据移植日是否使用LMWH分为肝素组和非肝素组,为调整混杂因素进行1∶5倾向性评分匹配(PSM),对PSM后患者数据进行分析,比较两组患者种植率、临床妊娠率、流产率、早期流产率、活产率及单胎活产围产结局的差异。对非PGT-A组和PGT-A组PSM前数据、肝素PGT-A组和肝素非PGT-A组临床妊娠结局进行多因素logistic回归分析。结果:非PGT-A组中,肝素组和非肝素组的不孕年限差异有统计学意义(P<0.05),种植率(33.8%vs 32.1%,P=0.610)、临床妊娠率(45.4%vs 43.8%,P=0.733)、流产率(28.0%vs 28.4%,P=0.958)、早期流产率(23.8%vs 25.4%,P=0.797)、活产率(31.9%vs 30.1%,P=0.671)比较差异无统计学意义(P>0.05)。PGT-A组中,肝素组和非肝素组的女方体质量指数(BMI)比较差异有统计学意义(P<0.05),种植率(59.8%vs 57.6%,P=0.763)、临床妊娠率(59.4%vs 57.6%,P=0.806)、流产率(20.4%vs 14.7%,P=0.448)、早期流产率(16.4%vs 14.7%,P=0.803)、活产率(46.9%vs 49.2%,P=0.752)比较差异无统计学意义(P>0.05)。非PGT-A组中,肝素组和非肝素组内获得单胎活产患者的围产期结局比较差异均无统计学意义(P>0.05);PGT-A组中,肝素组较非肝素组小于胎龄儿(10.3%vs 0.8%,P=0.023),差异有统计学意义(P<0.05),余获得单胎活产患者的围产期结局差异均无统计学意义(P>0.05)。结论:行PGT-A助孕和未行PGT-A助孕的URSA患者应用LMWH并不能改善其妊娠结局,且行PGT-A助孕应用LMWH患者的围产期小于胎龄儿发生风险显著升高。

Bayesian inference towards the resolution of molecular evolution: application to the “Trichoderma harzianum sensu lato” clade

The Hypocrea lixii/Trichoderma harzianum species aggregate contains a group of taxa (H. lixii /T. harzianum, T. aggressivum, T. tomentosum, T. cerinum, T. velutinum, H. tawa) of which some (e.g. T. harzianum) are important for biocontrol of plant pathogenic fungi in agriculture, whereas others are aggressive pathogens of Agaricus spp. and Pleurotus spp. in mushroom farms (T. aggressivum), or opportunistic pathogens of immunocompromised mammals including humans (T. harzianum). We characterized the evolutionary properties of three genomic regions in Hypocrea/Trichoderma: the internal transcribed spacer regions ITS1 and 2 of rDNA, the large intron of translation elongation factor 1-alpha (tef1a), and a portion of the large exon of the endochitinase 42 gene (ech42), selected the best model which describes the evolution of every fragment, tested the molecular clock hypothesis and made an estimation of the usability of the combined three fragments data matrix for the phylogenetic analysis of the genus as a whole as well as on the level of the holomorphic H. lixii/T. harzianum species clade and separate clonal lineages. To this end, we applied Bayesian phylogenetic inferences to 124 sequences of ITS1 and 2 and of the large tef1a intron, and to 64 ech42 gene sequences to resolve the evolution of H. lixii/T. harzianum with respect to the position of other taxa with closely related phenotypes. The resulting phylogram clearly identified T. aggressivum, T. velutinum, H. tawa, T. cerinum and T. tomentosum as phylogenetic species, and in addition identified three new unknown phylogenetic species as members of this clade. The clear distinction between T. tomentosum and T. cerinum was not recognized in all trees, but was supported by multivariate analysis of phenotype micro arrays. In contrast, H. lixii/T. harzianum did not form a single phylogenetic species in this study, as its monophyly was not supported in any analysis. Strains morphologically identified as H. lixii or T. harzianum formed either a proliferating megaclade (three loci combined tree) or appeared as several independent branches on the star-like phylogram (ITS1 and 2 and large tef1a intron) . However most terminal clades had a high posterior probability and thus fulfilled the criterion to consider their member isolates as a phylogenetic species. The clade containing the ex-neotype strain of T. harzianum included exclusively clonally propagating isolates from temperate climates, and was clearly distinguished from all other “H. lixii/T. harzianum” lineages by multivariate analysis of phenotype microarray data. We propose that only this lineage should be regarded as T. harzianum sensu stricto, and that the other lineages be regarded as T. harzianum sensu lato until phenetic characters have been defined which allow their description as separate taxa.

耐药结核病实验室诊断技术研究进展

结核病(TB)是严重危害公众健康的全球性公共卫生问题。耐药结核病(DR-TB)是TB防治的重点和难点。目前,常用的体外药物敏感性试验方法包括表型药物敏感性检测技术和分子药物敏感性检测技术。表型药物敏感性检测技术包括固体药物敏感性试验、液体药物敏感性试验、最低抑菌浓度法,分子药物敏感性检测技术包括GeneXpert MTB/RIF、线性探针技术、基因芯片技术、荧光聚合酶链反应(PCR)熔解曲线法、FluoroType MTBDR、Xpert MTB/XDR、核酸质谱分析、全基因组测序技术。文章就表型药物敏感性检测和分子药物敏感性检测对DR-TB实验室诊断技术的研究进展进行综述,旨在为DR-TB的早期诊断提供参考。