耐碳青霉烯类肺炎克雷伯菌药物敏感性和耐药基因研究  

Susceptibility and characterization of drug resistance genes in carbapenem-resistant Klebsiella pneumoniae

作　　者：王健[1] 潘亚萍[1] 徐元宏[1] 沈继录[1] 王中新[1] Wang Jian;Pan Yaping;Xu Yuanhong(Dept of Clinical Laboratory,The First Affiliated Hospital of Anhui Medical University,Hefei 23002)

机构地区：[1]安徽医科大学第一附属医院检验科,合肥230022

出　　处：《安徽医科大学学报》2018年第8期1231-1235,共5页Acta Universitatis Medicinalis Anhui

基　　金：国家自然科学基金(编号:81171606)

摘　　要：目的研究碳青霉烯类抗生素耐药的肺炎克雷伯菌(CRKP)抗生素敏感性和耐药基因分析,以及通过表型检测分析外排泵抑制剂对CRKP的亚胺培南、美罗培南和替加环素耐药性影响。方法收集并筛选医院临床分离非重复的CRKP,采用纸片扩散法和Vitek2法测定临床常见抗菌药物的敏感性;采用Carba-NP试验和改良Hodge试验对碳青霉烯酶进行筛选,并同对碳青霉烯酶耐药基因、ESBL和Amp C酶基因进行PCR扩增和序列分析;采用琼脂稀释法检测筛选菌株外排泵的表型;ERIC-PCR进行同源性分析。结果共筛选出31株CRKP,Carba-NP试验和改良Hodge试验显示有1株细菌不产碳青霉烯酶;KPC酶基因的PCR检测结果显示有27株KPC酶检测阳性,与碳青霉烯酶表型检测基本一致,部分菌株同时还产生了ESBL和Amp C酶;外排泵的抑制试验结果显示有3株对亚胺培南和美罗培南的最低抑菌浓度(MIC)分别下降4倍和8倍,而替加环素却未下降;ERIC-PCR分型显示有9种类型,其中1型13株,为主要流行株。结论 CRKP存在耐药基因积聚共存现象,导致CRKP对大多数临床常用抗菌药物呈高度耐药;CRKP产碳青霉烯酶以KPC-2型酶为主,是CRKP对碳青霉烯类耐药的主要机制;外排泵系统可能参与CRKP对碳青霉烯类耐药。Objective To investigate the antimicrobial susceptibility and analysis of antibiotic resistant gene in carbapenem-resistant Klebsiella pneumoniae（ CRKP） strains. Efflux pumps inhibitor phenotype analysis were used to study the role of efflux pump inhibitor in resistance to meropenem, imipenem,and tigecycline among clinical isolates of CRKP. Methods The CRKP strains were collected and screened. The drug sensitivity tests were performed by the Kirby-Bauer method and Vitek2 compact system. Carbapenemase production were confirmed by modified Hodgetes test and Carbapenemase Nordmann-Poirel（ Carba NP） test. The genotypes of carbapenem resistant genes,ESBLs genes,and Amp C were detected by PCR and DNA sequence analysis. The contribution of efflux mechanisms was screened by agar dilution method. Enterobacterial repetitive intergenic consensus（ ERIC-PCR）was used to type 31 clinical CRKP isolates. Results A total of 31 CRKP strains were isolated,the Carba-NP test and the modified Hodge test showed that only 1 strains of CRKP did not produce carbapenems. PCR results showed that 27 strains were positive for KPC enzyme gene,which was basically consistent with the detection of carbapenemases phenotype. In the inhibition experiment,the minimum inhibitory concentration of meropenem,imipenem decreased by 4 times and 8 times respectively after treated with PAβN in 3 isolate. 31 Klebsiella pneumoniae strains were classified into 9 types based on ERIC-PCR profiles,including 13 genotyping 1,which was the main epidemic strains. Conclusion The coexistence of drug resistance genes in CRKP shows high levels of resistance to routine antibiotic. Production of KPC-2-type carbapenemase is the common resistant mechanism to carbapenems for Klebsiella pneumoniae in our hospital. The efflux pump system might be involved in the carbapenem resistance of Klebsiella pneumoniae.

关 键 词：肺炎克雷伯菌 碳青霉烯酶 外排泵 耐药 

分 类 号：R446.5[医药卫生—诊断学]