规模化鸡场粪样和苍蝇产CMY-2大肠杆菌中耐药基因与毒力基因调查及其共转移研究

Investigation and co-transfer of resistance genes and virulence genes among Escherichia coli producing CMY-2 isolated from feces and flies among large-scale poultry farms

摘要:

为了解全国20家规模化鸡场粪样和苍蝇产CMY-2大肠杆菌中耐药基因与毒力基因的流行现状及其共转移情况，对2013-2015年间分离自粪样和苍蝇中的549株大肠杆菌，采用聚合酶链反应（PCR）方法筛选出blaCMY-2阳性菌株；脉冲场凝胶电泳（PFGE）分析blaCMY-2阳性菌株的遗传进化关系；K-B纸片法检测阳性菌株对13种抗菌药物的耐药性；利用PCR技术检测19种相关耐药基因以及14种毒力基因；接合转移试验和质粒复制子分型研究耐药基因及毒力基因的传播方式。结果，33株大肠杆菌呈CMY-2阳性，阳性率为6.0 %，且均表现为多重耐药；检测到15种耐药基因（blaTEM-1、blaCTX-M-55、blaOXA-1、qnrBDS、aac(6’)-Ib-cr、oqxA、tetA、tetC、sul1、sul2、sul3、rmtB和flor）和4种毒力基因（iutA、traT、fyuA和VagC）；33株携带blaCMY-2基因菌株中有21株接合转移成功，blaCMY-2基因可通过lncA/C或lncI1质粒与耐药基因（blaTEM-1、blaCTX-M-55、tetA或sul2）和（或）毒力基因（traT或VagC）共同转移。本研究阐明了规模化鸡场产CMY-2大肠杆菌携带耐药基因与毒力基因的情况，证实了细菌中质粒介导的耐药与毒力的共转移，为耐药性传播及疾病防控提供了参考。

Abstract:

The prevalence and co-transfer of resistance genes and virulence genes among Escherichia coli producing CMY-2 from feces and flies among twenty large-scale poultry farms in China were investigated. Polymerase chain reaction (PCR) was used to examine for the presence of E.coli producing CMY-2. The genetic relationship between isolates carrying blaCMY-2 gene was determined by pulsed-field gel electrophoresis (PFGE). Antimicrobial susceptibilities to 13 antimicrobials agents of isolates producing CMY-2 were tested by diffusion methods. Characterization of isolates producing CMY-2, including other resistance genes, and virulence genes was detected by PCR. The transmission of resistance and virulence genes was demonstrated by conjugation experiment, PCR-based replicon typing (PBRT). The results showed that 33 isolates were confirmed to carry blaCMY-2 gene with a positive rate of 6.0%, which showed resistance to multiple antimicrobial agents. Fifteen resistance genes conferring resistance to other antimicrobial agents were identified, including blaTEM-1, blaCTX-M-55, blaOXA-1, qnrBDS, aac(6’)-Ib-cr, oqxA, tetA, tetC, sul1, sul2, sul3, rmtB, and flor. Furthermore, four virulence genes including iutA, traT, VagC, and fyuA gene were also detected. Twenty-one transconjuncgants were detected in 33 E.coli Producing CMY-2. The co-transfer of between resistance genes and virulence genes was observed on transferred plasmids of lncA/C or lncI1. This study confirmed the prevalence and co-transfer of resistance genes and virulence genes in isolates of E.coli producing CMY-2, providing a reference for disease prevention and transmission in poultry farms.