2015—2019年上海市青浦区腹泻门诊致泻性大肠埃希菌的耐药情况及分子分型

Drug resistance and molecular typing of diarrheagenic escherichia coli from the diarrhea clinics in Qingpu District,Shanghai City from 2015 to 2019

目的分析2015—2019年上海市青浦区腹泻门诊病例中5种致泻性大肠埃希菌(DEC)分离株的耐药情况和多位点序列分型。方法于2015年1月至2019年12月,采集上海市复旦大学附属中山医院青浦分院腹泻门诊病例的肛拭子样本,将经分离培养鉴定为DEC的菌株以微量肉汤稀释法测定菌株的最小抑菌浓度。根据药敏试验结果,选取对第三代头孢菌素类或碳青霉烯类等抗生素耐药,或产超广谱β-内酰胺酶的菌株进行全基因组测序。基于测序获得的MLST分型,通过BioNumerics 7.6软件构建最小生成树,分析当地优势菌群。结果从4494份肛拭样本中共分离出513株DEC,检出率为11.42%;完成其中500株对4类9种抗生素的抗生素药物敏感性试验,分别为330株产毒性大肠埃希菌(ETEC)、72株黏附性大肠埃希菌(EAEC)、95株致病性大肠埃希菌(EPEC)、1株出血性大肠埃希菌(EHEC)和2株侵袭性大肠埃希菌(EIEC)。2015—2019年不同年份间对头孢噻肟-克拉维酸的耐药检出率差异有统计学意义(P<0.05);各毒力分型DEC对萘啶酸的耐药检出率差异有统计学意义(P<0.05)。71株DEC的全基因组测序结果显示,共检测到77种耐药基因,并被归类为32个ST分型,优势基因型为ST-1491(29.6%,21/71)和ST-10 Complex(23.9%,17/71)。其中,ST-1491全部产ESBLs,均为bla_(CTX-M)耐药基因突变株,ST-10 Complex优势基因型为ST-218(35.3%,6/17)。8株EAEC、14株EPEC和49株ETEC分别被归类为7个、14个和18个ST分型。结论上海市青浦区腹泻门诊病例的DEC对重点药物的耐药情况较严重;EAEC和EPEC的ST分型呈高度多样性,当地优势ST型与国内东南地区的常见基因型基本一致。

Objective To analyze the drug resistance and multilocus sequence typing of five types of diarrheagenic Escherichia coli(DEC)isolated from diarrhea outpatients of diarrhea comprehensive monitoring designated hospital in Qingpu District,Shanghai City from 2015 to 2019.Methods From January 2015 to December 2019,five types of DEC,isolated and identified from diarrhea outpatient cases'anal swabs of the Qingpu branch of Zhongshan Hospital were collected to determine the minimal inhibitory concentration by using the micro broth dilution susceptibility test.The strains,resistant to the third-generation cephalosporins or carbapenems,or producing ESBLs,were selected based on the results of sensitivity tests and determined by WGS.The MLST typing of DEC was analyzed based on the WGS technology and the minimum spanning tree was constructed by BioNumerics 7.6 software to analyze the local dominant flora.Results A total of 513 strains of DEC were detected and isolated from 4494 anal swabs,with a detection rate of 11.42%.About 500 strains were tested for drug sensitivity to nine antibiotics in four classes,including 330 strains of enterotoxigenic E.coli(ETEC),72 strains of enteroaggregative E.coli(EAEC),95 strains of enteropathogenic E.coli(EPEC),1 strain of enterohemorrhagic E.coli(EHEC),and 2 strains of enteroinvasive E.coli(EIEC).From 2015 to 2019,the resistance rate of cefotaxime-clavulanic acid was significantly different(P<0.05).The resistance rate of virulence types of DEC to nalixic acid was significantly different(P<0.05).About 71 strains of DEC were determined by WGS,and 77 drug-resistant genes were detected.Strains were classified into 32 ST subtypes,with the dominant genotypes being ST-1491(29.6%,21/71)and ST-10 Complex(23.9%,17/71).All ST-1491 produced ESBLs,which were bla_(CTX-M) gene mutant strains.The dominant type of ST-10 complex was ST-218(35.3%,6/17).In addition,8 strains of EAEC,14 strains of EPEC and 49 strains of ETEC were classified into 7,14 and 18 ST subtypes,respectively.Conclusion The drug resistance of DEC strains from the diarrhea outpatient case of Qingpu District is serious.The ST types of EAEC and EPEC are highly polymorphic.The dominant ST types of DEC are basically consistent with the common genotypes in southeast China.