利奈唑胺体外诱导粪肠球菌V区突变及耐药相关性研究  

In vitro investigation on linezolid resistance-associated mutation in domain V region of Enterococcus faecalis

作　　者：高睿[1] 张美兰[1] 杜丽[1] 蒲清泉[1] 夏云[1] 

机构地区：[1]重庆医科大学附属第一医院检验科,重庆400016

出　　处：《第三军医大学学报》2016年第8期824-828,共5页Journal of Third Military Medical University

基　　金：国家自然科学基金面上项目(81572055)~~

摘　　要：目的观察利奈唑胺是否能在体外诱导肠球菌产生高水平耐药,同时探索其耐药机制与23S rRNA的V区突变的关系,以及突变的基因拷贝数是否与耐药程度（MIC值）相关。方法 5株耐药机制未明的利奈唑胺低水平耐药粪肠球菌（MIC：4-16μg/m L）持续给予利奈唑胺进行体外诱导,观察其耐药水平变化;采用PCR技术扩增诱导菌和ATCC 29212的23S rRNA的V区片段以及编码核糖体蛋白L3、L4基因片段,分析有无V区和L3、L4突变及V区突变拷贝数。结果通过利奈唑胺持续诱导,5株耐药菌的MIC值较原菌株获得8-32倍增加,最高MIC值超过256 mg/L;其中有3株菌23S rRNA的V583区发生G2576T突变,其余2株菌未发现V区突变;所有诱导菌未发现L3、L4核糖体蛋白氨基酸改变。结论利奈唑胺在体外可诱导粪肠球菌MIC值迅速增加,其V区G2576T突变与高水平耐药密切相关,且突变拷贝数量与MIC值增高呈正相关。Objective To investigate high-level linezolid resistance-associated mutation and copy number of mutation gene in domain V region of 23 S rRNA through inducing Enterococcus faecalis（ E. faecalis）with linezolid in vitro. Methods The linezolid resistance was assessed in 5 strains of E. faecalis that had minimum inhibitory concentration（ MIC） ranging from 4 μg/m L to 16 μg/m L and were induced with linezolid in vitro. After induction with linezolid,PCR and DNA sequencing were used to detect the mutations in 23 S rRNA gene and genes encoding L3 and L4 ribosomal proteins in the induced strains and control strain E. faecalis ATCC29212. The copy number of the mutation gene was determined by DNA sequencing.Results After serial passages in the presence of linezolid in vitro,the MIC of the 5 strains were successfully increased by 8 to 32 times,and the maximum MIC was more than 256 μg/m L. The G2576 T mutation was detected in 3 strains,while all possible mutations were not found in the domain V region of 23 S rRNA gene in the other 2 strains. There were no mutations in the genes encoding L3 and L4 proteins in the strains.Conclusion Linezolid can rapidly induce the MIC increase in E. faecalis in vitro,and the principal mechanism may be associated with G2576 T mutation in the domain V region of 23 S rRNA gene. The copy number of mutation is positively correlated with MIC elevation.

关 键 词：利奈唑胺 粪肠球菌 23S核糖体RNA基因 基因突变 

分 类 号：R378.1[医药卫生—病原生物学] R394.3[医药卫生—基础医学]