广西地区动物源bla_(NDM)阳性肠杆菌科细菌的流行病学和分子特征研究

Epidemiological and molecular characteristics of bla_(NDM)-positive Enterobacteriaceae from animals in Guangxi

旨在研究广西地区动物源bla_(NDM)阳性肠杆菌科细菌的流行现状和分子特征,并了解bla_(NDM)-5基因的遗传环境和传播特点。2020年6月从广西地区部分养殖场中采集样品130份,通过PCR测序筛选出bla_(NDM)阳性菌株,并进行菌种鉴定和耐药基因的检测;药物敏感性试验和多位点序列分型(MLST)试验分别检测bla_(NDM)阳性分离株的耐药表型和菌株之间的同源性;选取一株代表性bla_(NDM)-5阳性菌株进行全基因组测序、质粒系统发育树的构建和比较基因组学分析,了解bla_(NDM)-5基因的遗传环境和传播情况。结果从样品中共分离出21株bla_(NDM)阳性菌株,分离率为16.15%,包括11株以序列型ST410型为主的bla_(NDM)-5阳性大肠杆菌、3株ST19型bla_(NDM)-1阳性柠檬酸杆菌和7株以ST629型为主的bla_(NDM)-5阳性肺炎克雷伯菌;PCR检出bla_(NDM)、blaCTX、blaCMY-2、tetA、aac(6′)-Ib、qnrB、cfr、fosA等多种耐药基因;药敏结果显示,菌株对美罗培南等13种抗菌药物表现出不同程度的耐药性,且都为多重耐药菌株;全基因组测序结果显示,bla_(NDM)-5定位于可转移的IncFII质粒上,其遗传环境为(IS26-bla_(NDM)-5-bleMBL-trpF-IS91-sul1-qacE-antI-dfrA-IntI1-IS26-Tn3);系统发育分析和比较基因组学分析结果显示,携带bla_(NDM)-5的IncFⅡ质粒可在动物和人类之间进行传播,且具有广泛的宿主。本研究表明bla_(NDM)基因在动物源肠杆科菌中广泛流行,给公众健康安全带来了严重的威胁,应加强对bla_(NDM)基因的监测和防控。

This experiment was to study the epidemic status and molecular characteristics of bla_(NDM)-positive Enterobacteriaceae from animals in the Guangxi area, and to understand the genetic environment and transmission characteristics of the bla_(NDM)-5gene. In June, 2020, we collected a total of 130 samples on the farms in the area. The bla_(NDM)-positive strains were screened by PCR sequencing. At the same time, the resistance genes were detected by PCR. Antimicrobial susceptibility test and MLST typing test were conducted to analyze the resistance phenotype and the homology between different bla_(NDM)-positive isolates. The genetic background and transmission of bla_(NDM)-5were investigated by WGS, phylogenetic tree construction and comparative genomics of plasmids. A total of 21 bla_(NDM)-positive strains were isolated, with a detection rate of 16.15%. Of the strains, there were 11 bla_(NDM)-5-positive E. coli with ST410 as the major phenotype, 3 bla_(NDM)-1positive Citrobacter strains with ST19 as the major phenotype and 7 bla_(NDM)-5positive K. pneumoniae were with ST629 as the major phenotype. BlaNDM, blaCTX, blaCMY-2, tetA, aac(6′)-Ib, qnrB, cfr, fosA and other resistance genes were detected by PCR. The strains showed different degrees of resistance to 13 antimicrobials including meropenem, and all were MDR isolates. WGS revealed that bla_(NDM)-5was located on a transferable IncFII plasmid, and its genetic environment was IS26-bla_(NDM)-5-bleMBL-trpF-IS91-sul1-qacE-antI-dfrA-IntI1-IS26-Tn3. The phylogenetic and comparative analysis showed that bla_(NDM)-5-harboring IncFII plasmids could be transmitted between animals and humans and had extensive hosts. The present study indicated that the bla_(NDM)-5gene was widely prevalent in Enterobacteriaceae from animals, posing a serious threat to public health and safety. Therefore, the monitoring and control of bla_(NDM)gene should be strengthened.