鲍曼不动杆菌分子特征及其在替加环素压力下外排泵表达的变化  

Molecular characteristics of Acinetobacter baumannii and the changes of efflux pump expression under tigecycline pressure

作　　者：程建军 Kesavan Dinesh Kumar 阴晴[2] 王慧璇 蔡伟 陈建国[3] 苏兆亮 许化溪 CHENG Jian-jun;Kesavan Dinesh Kumar;YIN Qing;WANG Hui-xuan;CAI Wei;CHEN Jian-guo;SUZhao-liang;XU Hua-xi(InstituteofLaboratoryMedicine,JiangsuUniversity,Zhenjiang 212013,Jiangsu,China;LaboratoryDepartmentof Affiliated Hospital,Jiangsu University,Zhenjiang 212001,Jiangsu,China;Laboratory Department of Affiliated People's Hospital,Jiangsu University,Zhenjiang 212002,Jiangsu,China)

机构地区：[1]江苏大学医学院检验医学研究所,镇江212013 [2]江苏大学附属医院检验科镇江212001 [3]江苏大学附属人民医院检验科,镇江212002

出　　处：《医学研究生学报》2020年第1期18-24,共7页Journal of Medical Postgraduates

基　　金：国家自然科学基金（81771756）;江苏省博士后基金（1601002C）

摘　　要：目的鲍曼不动杆菌是医院内感染常见的细菌。文中旨在分析其分子流行特征,检测外排泵和调节蛋白基因的携带率,并研究替加环素对外排泵和调节蛋白基因表达的影响。方法选取2017年5月至2019年3月江苏大学附属医院住院患者183株鲍曼不动杆菌。分为耐药组(一类及以上抗菌药物耐药的菌株,139株),敏感组(药敏试验中的药物均为非耐药的菌株,44株)。应用重复序列PCR作菌株的同源性分析,并以脉冲场凝胶电泳(PFGE)为同源性分析的金标准对部分菌株进行验证和比较。PCR检测耐药相关基因。依据同源性分析、外排泵携带率检测及药敏试验结果,选取6株外排泵基因全携带但耐药表型不同的临床菌株作为实验株,包括敏感株(SAB)、多药耐药株(MDRAB)、广泛耐药株(XDRAB),进行替加环素体外诱导,诱导后的菌株为诱导组,以不含替加环素LB培养的相同菌株为对照组。检测AdeB、TetB等外排泵基因及AdeS、BaeS等调节蛋白的基因表达变化。结果同源性分析显示,183株临床分离株包含有45个克隆群。耐药组AdeB、TetB基因(97.84%、98.56%)检出率均高于敏感组(63.64%、20.45%),CraA、AbeM等基因亦高于敏感组(P<0.05)。替加环素诱导前MDRAB、XDRAB中AdeB(4.17±0.94、6.86±3.23)、TetB(13.06±0.38、13.96±0.63)表达较SAB(0.00±3.70、0.00±0.70)升高(P<0.05),MDRAB菌株中MacB、BaeS表达较SAB菌株亦升高(P<0.05)。MDRAB、XDRAB菌株中AbaQ表达较SAB菌株降低(P<0.05)。诱导组AdeB、TetB在各菌株的表达较对照组明显升高(P<0.01);除SAB1菌株外,AbaQ、AbeM在其余菌株中的表达较对照组明显升高(P<0.05);AdeS在SAB1、SAB2、MDRAB1、XDRAB2菌株中的表达较对照组升高(P<0.05);负调节蛋白SoxR在SAB1、XDRAB1菌株中表达较对照组降低(P<0.05)。结论AdeB、TetB、AbeS、AdeS、BaeS在临床鲍曼不动杆菌多药耐药过程中发挥着重要的作用,其中AdeB和TetB在菌株对替加环素耐药过程�Objective Acinetobacter baumannii(A.baumannii)is a commonly infective bacterium in the hospital.This study aims to analyze its molecular epidemiological characteristics,detect the carrying rate of efflux pump and regulatory protein genes,and investigate the effects of tigecycline on the efflux pump and expression of regulatory protein genes.Methods A total of 183 A.baumannii strains were collected from inpatients of the affiliated hospital of Jiangsu University from May 2017 to March 2019.They were divided into an antimicrobial-resistant group(one or more antimicrobial-resistant strains,139 strains)and a sensitive group(the drugs in the drug sensitivity test were all non-resistant strains,44 strains).Repeated sequence PCR was used for homology analysis of the strains,and pulse-field gel electrophoresis(PFGE)was used as the gold standard for homology analysis to verify and compare some strains.PCR was used to detect the occurrence of drug resistance-related genes.Based on homology analysis,efflux pump carrying rate detection and antibiotics sensitivity test results,6 clinical strains carrying all efflux pump genes but different resistance phenotypes were selected as experimental strains,including sensitive strains(SAB),the multidrug resistance strain(MDRAB)and the extensively drug-resistant strain(XDRAB).All strains were induced in vitro with the minimum inhibitory concentration(MIC)of tigecycline.The induced strains were categorized as induction group,and the same strains cultured in LB agar without tigecycline was used as a control group.MIC was used to analyze the tigecycline susceptibility,and RT-qPCR was used to detect the gene expression of efflux pumps,such as TetB,AbaQ and regulatory proteins(AdeS and BaeS),in drug-resistant strains.Results Homology analysis showed that there were 45 clonal groups in the detected clinical isolates,with no obvious outbreak of epidemic clonal groups.Efflux pumps and regulatory proteins were widely distributed in the clinical isolates,and the expression of AdeB,TetB,AbeS,AdeS

关 键 词：鲍曼不动杆菌 外排泵基因 同源性分析 脉冲场凝胶电泳 替加环素 耐药 

分 类 号：R378[医药卫生—病原生物学]