质粒介导的产NDM-1肠杆菌科细菌分子水平研究

Molecular Study of plasmid-mediated NDM-1-producing Enterobacteriaceae

目的对分离的29株产新德里金属-β-内酰胺酶1(NDM-1)肠杆菌科细菌进行同源性分析,了解浙江省NDM-1的流行情况;通过blaNDM-1基因周围序列分析,阐明该基因所处的周围环境。方法PCR扩增浙江医院分离的29株产NDM-1肠杆菌科细菌耐药基因;接合试验分析质粒传播转移能力;以PCR为基础的质粒复制子分型方法(PBRT)分析质粒型别;PCR扩增blaNDM-1基因周围序列,分析其周围序列异同点。结果29株携带blaNDM-1基因菌株中,86.9%(26/29)合并超广谱β-内酰胺酶或其他碳青霉烯酶耐药基因。PFGE结果显示6株产气肠杆菌,5株阴沟肠杆菌,2株大肠埃希菌以及2株产酸克雷伯菌分别具有同源性。多位点序列分型发现3株大肠埃希菌为ST167,5株阴沟肠杆菌为ST114。接合实验发现,27株菌株接合成功。PBRT质粒分型发现16株为IncX3型质粒;对菌株进行blaNDM-1周围环境分析发现,16株存在blaNDM-1-bleMBL-trpF序列。结论NDM-1已成为浙江省碳青霉烯类抗生素耐药的重要原因。不同菌种细菌间存在IncX3型质粒广泛传播,blaNDM-1-bleMBL-trpF基因片段广泛存在于产NDM-1菌株中。

Objective To analyze the homology of 29 New Delhi Meta-β-lactamase 1(NDM-1)-producing Enterobacteriaceae strains,and to understand the prevalence of NDM-1 in Zhejiang Province,and the surrounding environment of blaNDM-1 gene was elucidated by sequence analysis.Methods The drug resistance genes of 29 NDM-1-producing Enterobacteriaceae strains isolated from Zhejiang Hospital were amplified by PCR;the plasmid types were analyzed based on PCR-based plasmid replicon typing(PBRT),and the surrounding sequence of blaNDM-1 gene was amplified by PCR to analyze the similarities and differences of the surrounding sequence.Results Among the 29 blaNDM-1-carrying strains,86.9%(26/29)were associated with extended spectrum beta-lactamases or other carbapenemase resistance genes;PFGE results showed that six strains of Enterobacter aerogenes,five strains of Enterobacter cloacae,two strains of Escherichia coli and two strains of Klebsiella oxytoca had homology,respectively.Multilocus sequence typing found that three strains of Escherichia coli were ST167 and five strains of Enterobacter cloacae were ST114.The conjugation experiments showed that 27 strains were successfully conjugated;16 strains were found to be IncX3 type by PBRT plasmid typing,and 16 strains were found to carry blaNDM-1-bleMBL-trpF gene fragment by analyzing the surrounding environment of blaNDM-1.Conclusion NDM-1 has become an important cause of carbapenem antibiotic resistance in Zhejiang Province.IncX3 plasmids are widely spread among different bacterial species,and blaNDM-1-bleMBL-trpF gene fragments are widely disseminated in NDM-1-producing strains.