产酸克雷伯菌新生儿分离株对碳青霉烯类耐药的分子机制  

Molecular mechanism of the carbapenem-resistant Klebsiella oxytocastraines recovered from neonates in a tertiary hospital

作　　者：李笃军 冯渐焘 刘真真 刘入华[2] 贾楠[2] 陈海霞 赵辉[2] 朱元祺[2,3] LI Du-jun;FENG Jian-tao;LIU Zhen-zhen(Department of Clinical Laboratory,the Yeda Hospital of Yantai City,Yantai 264006,China)

机构地区：[1]烟台业达医院检验科,山东烟台264006 [2]青岛大学附属医院检验科,山东青岛266555 [3]青岛大学医学院,山东青岛266071

出　　处：《中国实验诊断学》2020年第1期115-118,共4页Chinese Journal of Laboratory Diagnosis

基　　金：烟台市科技计划项目（2019MSGY132）;烟台业达医院科技发展项目（2018-1-82）

摘　　要：目的探讨碳青霉烯类耐药产酸克雷伯菌新生儿分离株的耐药机制。方法 3株耐碳青霉烯类产酸克雷伯菌是2013年9月-2013年12月分离于新生儿患者。菌株的鉴定和药敏采用梅里埃Vitek-2,经PCR扩增及产物测序确认其携带的耐药基因。脉冲场凝胶电泳和多位点序列检测菌株间的同源性。基于PCR的质粒分型、质粒液相接合和Southern杂交分析质粒的特性。结果 3株产酸克雷伯菌除了对氨曲南敏感外,对目前常用的β-内酰胺类抗生素都耐药。这3株供体菌(产酸克雷伯菌TJ11-TJ13)与受体菌(Escherichia coli J53Azi R)经液相接合获得接合子(TJC11-TJC13)。经检测,供体菌与其接合子均携带blaNDM-1、blaOXA-1、blaDHA-1、qnrB4和aac(6′)-Ib-cr耐药基因。3株产酸克雷伯菌脉冲场凝胶电泳显示克隆聚集性,且都属于ST2型。质粒的S1核酸酶切、Southern杂交和PCR分型表明产酸克雷伯菌携带大小约100kb和275kb两个质粒,而接合子仅携带1个大小约275kb的质粒,且二者携带的blaNDM-1基因都位于275kb的IncHIB型质粒上。结论研究表明ST2型产酸克雷伯菌对碳青酶烯类耐药的分子机制是由于携带了blaNDM-1、blaOXA-1、blaDHA-1、qnrB4和aac(6′)-Ib-cr耐药基因,而且在新生儿中存在该克隆株的播散。经检索,这属于国内外文献首次报道。Objective To explore the molecular mechanism of the clinical carbapenem-resistant Klebsiella oxytocas isolated from neonates in a tertiary hospital.Methods Three isolates of carbapenem-resistant Klebsiella oxytocas,collected from Sep 2013 to Dec 2013 from three neonates,were determined by VITEK-2 system for identification and antimicrobial susceptibility testing.Carbapenemase and other resistance determinants were identified by PCR and sequencing.Genetic relatedness was verified by pulsed-field gel electrophoresis(PFGE)and multilocus sequence typing(MLST).Plasmid localization of blaNDM-1 in isolates and transconjugants was investigated by S1-PFGE and Southern hybridization.Plasmid typing was also performed using PCR-based replicon typing(PBRT)method.Results The three isolates were highly resistant to most availableβ-Lactams except for aztreonam.The carbapenemase and other resistance genes could be transferred into an Escherichia coli J53 AziR recipient by conjugation experiments.And the coproduction of blaNDM-1、blaOXA-1、blaDHA-1、qnrB4 and aac(6′)-Ib-cr were observed in both Klebsiella oxytoca isolates and their transconjugants.The PFGE demonistrated that the three NDM-1-producing Klebsiella oxytocas were clonal relatedness,and belong to ST2.The PBRT,S1-PFGE and Southern blotting revealed that the Klebsiella oxytoca carried two plasmids,100 kb,275 kb,respectively,and that the blaNDM-1 gene was located on a 275 kb IncHIB plasmid in all isolates.Conclusion Our results demonstrated that the molecular mechanism of carbapenem resistance in three ST2 Klebsiella oxytocas is duo to these isolates harboring blaNDM-1、blaOXA-1、blaDHA-1、qnrB4 and aac(6′)-Ib-cr genes and that there was a clonal spread caused by them.To the best of our knowledge,it is firstly documented in the worldwide.

关 键 词：产酸克雷伯菌 blaNDM-1 新生儿 基因 

分 类 号：R446.5[医药卫生—诊断学]