耐亚胺培南肺炎克雷伯菌分子流行病学及耐药传播机制研究

Molecular epidemiology and drug-resistance transfer mechanism of imipenem resistant Klebsiella pneumonia

目的探讨临床收集亚胺培南耐药肺炎克雷伯菌的耐药分子机制和克隆流行情况。方法采用K-B纸片法进行药敏实验,多位点序列分型(MLST)进行临床菌株克隆分型,滤膜接合法进行质粒接合实验,PCR扩增筛选菌株的常见耐药基因和其周围序列结构。结果亚胺培南耐药肺炎克雷伯菌对青霉素类、头孢菌素类、含抑制剂β-内酰胺类、氨基糖苷类等药物均显示出很强的多重耐药性,MLST分型显示耐药克隆具有明显的流行特征,13株携带blaKPC-2基因的流行克隆均为ST11型。这13株细菌亚胺培南耐药性均由可接合性质粒介导,并且同时介导和转移了编码CTX-M型、TEM型或DHA型的β-内酰胺酶基因。Tn4401-Tn3复合转座子或其变体2介导了blaKPC-2基因的转移。结论临床肺炎克雷伯菌亚胺培南耐药的主要原因是产KPC-2酶菌株的克隆播散,同时可接合质粒造成的耐药基因快速水平转移也起到了重要作用。

Objective To characterize the clone dissemination of imipenem resistant Klebsiella pneumoniae and its re-sistance mechanism. Methods Antimicrobial agents susceptibility testing was determined by K-B method. Multi Locus Sequence Typing （MLST） was used to investigate the clonality of clinical isolates. Plasmid conjugation assay was used by filter mating. PCR amplification and sequencing were used to screen common antimicrobial resistance genes and their adja-cent genetic environment. Results Imipenem resistant Klebsiella pneumoniae isolates exhibit multidrug resistance to peni-cillin, cephalosporins, β-lactam-β-lactamase inhibitor combinations, and aminoglycosides. MLST result supported the evidence of clone dissemination. Thirteen isolates that harbored KPC-2 type carbapenemase gene were all ST11 type clone. Resistances of these 13 isolates were all mediated by conjugant plasmid, which encoded CTX-M, TEM, KPC type β-lac-tamase genes. The composite transposon Tn4401-Tn3 or its variant 2 mediated the transfer of carbapenemase gene blaKPC-2. Conclusion The main resistance mechanism of clinical Klebsiella pneumoniae isolates was the dissemination of KPC-2-carbapenemase-producing ST11 type clone. Plasmid conjugation or horizontal gene transfer also promotes the rapid preva-lence of resistance.