高毒力肺炎克雷伯菌中CRISPR-Cas系统的分布及其与毒力和耐药的关系  

Distribution of CRISPR-Cas systems and its potential relationship with virulence and antibiotic resistance in clinical hypervirulent Klebsiella pneumoniae strains

作　　者：廖文建 祝兰兰[1] 杜芳玲 龙丹[1] 张伟[1] 刘洋[1] LIAO Wenjian;ZHU Lanlan;DU Fangling;LONG Dan;ZHANG Wei;LIU Yang(Department of Respiratory Medicine,the First Affiliated Hospital of Nanchang University,Nanchang 330006,China)

机构地区：[1]南昌大学第一附属医院呼吸科,南昌330006

出　　处：《中国感染与化疗杂志》2020年第2期163-168,共6页Chinese Journal of Infection and Chemotherapy

基　　金：国家自然科学基金地区项目(81860368)。

摘　　要：目的了解高毒力肺炎克雷伯菌中CRISPR-Cas系统的分布特征,并探究其与毒力和耐药的关系。方法收集非重复高毒力肺炎克雷伯菌61株,使用VITEK 2-Compact全自动微生物分析系统进行菌株鉴定及药物敏感性分析,PCR检测CRISPR-Cas系统4个相关基因(Cas1、Cas3、CRISPR1和CRISPR2)、筛查荚膜血清分型基因K1/K2、14种毒力基因及14种耐药基因,用χ2检验比较CRISPR-Cas系统阳性菌株与阴性菌株毒力基因及耐药差异。大蜡螟毒力实验分析比较两组细菌的毒力差异,并且采用多位点序列分型(MLST)分析所有高毒力肺炎克雷伯菌分子分型与CRISPR-Cas系统的关系。结果CRISPR-Cas系统阳性检出率为34.4%(21/61),所有CRISPR-Cas系统阳性的菌株均为ST23型K1菌株,除毒力基因kpn、repA外,CRISPR-Cas系统阳性的高毒力肺炎克雷伯菌携带毒力基因阳性率均大于对应的阴性菌,其中5种毒力基因差异具有显著统计学意义。除耐药基因blaNDM-1、qnrB外,CRISPR-Cas系统阳性高毒力肺炎克雷伯菌的8种耐药基因阳性率均小于对应的阴性菌株,差异具有显著统计学意义。CRISPR-Cas系统阳性高毒力肺炎克雷伯菌感染大蜡螟平均半数生存时间少于对应的阴性菌感染,差异具有统计学意义。MLST结果表明高毒力肺炎克雷伯菌CRISPR-Cas系统分布与分型密切相关。结论CRISPR-Cas系统阳性高毒力肺炎克雷伯菌主要为ST23型K1菌株,CRISPR-Cas系统阳性肺炎克雷伯菌相对于阴性菌株的毒力强,毒力基因阳性率高,耐药率低,耐药基因的阳性率低。临床上应警惕CRISPR-Cas系统阴性ST11型“碳青霉烯类耐药高毒力肺炎克雷伯菌”的暴发,此类细菌进化获得毒力质粒pLVPK毒力强、高度耐药、传播性强,可能成为危害公共健康并难以控制的超级细菌。Objective We aimed to characterize the CRISPR-Cas systems in clinical hypervirulent K.pneumoniae isolates and to investigate the potential association of CRISPR-Cas systems with virulence and antibiotic resistance.Methods A total of 61 hypervirulent K.pneumoniae strains were collected from inpatients in a teaching hospital in Jiangxi Province.The strains were identified and tested for antimicrobial susceptibility by using VITEK 2-Compact automatic microbiology analyzer.All the isolates were assessed for four CRISPR-Cas system related genes,hypervirulent capsular serotypes K1/K2,14 virulence genes,and 14 resistant genes by PCR.The virulence and antibiotic resistance were compared between CRISPR-Cas strains and CRISPR-Casstrains by Chi-square test.The potential virulence of all the clinical hypervirulent strains was tested in a Galleria mellonella infection model.Multilocus sequence typing(MLST)was used to analyze the genotypes of the hypervirulent K.pneumoniae strains.Results The prevalence of CRISPR-Cas system was 34.4%(21/61)in the strains tested.All the K.pneumoniae strains carrying CRISPR-Cas system belonged to ST23 type K1.The prevalence of virulence genes(except kpn and repA)was higher in the strains carrying CRISPR-Cas system than in the strains without CRISPR-Cas system.Five virulence genes showed significantly different prevalence in terms of CRISPR-Cas system.The prevalence of antibiotic resistance genes(except blaNDM-1 and qnrB gene)in the strains carrying CRISPR-Cas system was higher than in the strains without CRISPR-Cas system.Eight antibiotic resistance genes showed significantly different prevalence in terms of CRISPR-Cas system.The average median survival time of Galleria mellonella larvae was significantly shorter after infection with CRISPR-Cas-positive hypervirulent K.pneumoniae strains than after infection with CRISPR-Cas-negative strain.MLST analysis showed the distribution of CRISPR-Cas systems was closely related to MLST types in hypervirulent K.pneumoniae strains.Conclusions All the hyperv

关 键 词：高毒力肺炎克雷伯菌 CRISPR-Cas 细菌毒力 细菌耐药性 

分 类 号：R378.99[医药卫生—病原生物学]