[Molecular characterization genetic diversity of extended-spectrum β-lactamases-harboring conjugative plasmids identified in multi-drug resistant Escherichia coli isolated from food in China].

Objective: The purpose of this study was to investigate the molecular characteristics of ESBL-encoding conjugative plasmid identified in muti-drug resistant Escherichia coli isolated from food. Methods: 465 Escherichia coli isolates were collected from national foodborne disease surveillance net from 2013 to 2014 (salad, n=159; meat, n=102; processed meat, n=95; cakes/rice, n=46; cooked dish, n=63). ESBLs strain was detected by Mueller-Hinton agar plate, and then its drug resistance was tested by agar dilution method. Polymerase chain reaction (PCR) and DNA sequencing were performed to identify the corresponding ESBL genes. Plasmids were typed by PCR-based replicon typing and their characteristics were determined by S1-nuclease pulsed-field gel electrophoresis method. Broth mating assays were carried out for all isolates to determine whether the ESBL marker could be transferred by conjugation. Results: 12 E. coli were found to be resistant to cefotaxime, and all of which were confirmed as ESBLs. The 12 isolates all carried different types of CTX-M genes resistant to drug, and 7 of which carried TEM type as well. All 12 isolates contained at least one plasmid and some had four plasmids, with size ranging from 47-to 220-kb by S1-PFGE anaylsis. Seven isolates demonstrated the ability to transfer their cefotaxime resistance marker to the recotper strain J53 by only one plasmid. Conclusion: This study highlights the diversity of the multi-drug resistant E. coli and also the diversity of ESBL genes in China. Plasmids carrying these genes poses a serious threat to food safety in China.目的： 研究食品中分离的多重耐药大肠埃希菌携带ESBLs可转移质粒分子特征。 方法： 465株大肠埃希菌株来自2013—2014年全国污染物监测网食源性致病菌（凉拌菜，n=159；生肉，n=102；预制肉制品，n=95；蛋糕米面，n=46；热菜，n=63）。采用Mueller-Hinton琼脂平板进行ESBLs筛选后，使用琼脂稀释法对菌株进行耐药性检测；采用PCR和序列分析对ESBLs菌株进行基因、质粒分型研究；采用S1-PFGE法分析菌株携带质粒的特征；运用肉汤接合实验确定编码ESBLs基因质粒的可移动性。 结果： 465株大肠埃希菌株中有12株对头孢噻肟耐药，并且均为产ESBLs菌株。12株头孢噻肟耐药菌株均携带CTX-M型耐药基因，其中7株还同时携带TEM型。通过S1-PFGE的分析，12株头孢噻肟耐药菌株中，每株菌至少携带了1个质粒，最多携带有4个质粒，质粒大小范围为47~220 kb。共有7株菌可以将携带CTX-M质粒转移到受体菌株J53，且每个成功发生转移的供体菌仅转移1个质粒。 结论： 食品中多重耐药菌株及ESBLs基因型别均具有多样性；ESBLs耐药基因质粒的可转移性威胁我国食品安全。.