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目的阐明体外诱导利奈唑胺耐药粪肠球菌的核糖体23SrRNA V区基因位点变异特征。方法收集1株血流感染的粪肠球菌和1株粪肠球菌质控菌ATCC29212(编号分别为F3和F4菌株,均为利奈唑胺敏感株),通过体外浓度倍增法诱导利奈唑胺耐药;挑取单克隆,经E-test条测定MIC值,获得各菌株的耐药浓度梯度;提取耐药菌株基因组DNA,PCR扩增23SrRNA V区基因,扩增产物经测序后与野生株比较,获得V区的突变位点。结果经体外多步法诱导利奈唑胺耐药的不同MIC值粪肠球菌共13株。PCR测序分析2株母株均无变异位点,23SrRNA V区的突变位点主要是G2576U,此外还有T2504A、G2505A、C2610A、C2424U。结论体外多步法可诱导粪肠球菌利奈唑胺耐药,耐药机制与23SrRNA V区位点突变密切相关,突变位点随着MIC值的增高而增多。
Objective To elucidate the variation of ribosomal 23S rRNA V-region gene mutation induced by Enterobacter oximaillin-resistant Enterococcus faecalis in vitro. Methods One strain of Enterococcus faecalis and one strain of Enterococcus faecalis ATCC29212 (numbered F3 and F4, respectively, linezolid susceptible strains) were collected and inactivated by Linezolid in vitro The monoclonal antibody (McAb) was selected and the MIC value was determined by E-test. The drug resistance concentration gradient of each strain was obtained. The genomic DNA of resistant strain was extracted and the 23SrRNA V region was amplified by PCR. The amplified product was compared with the wild- Obtain the mutation site of V region. Results In vitro multi-step method of linezolid resistance induced by different MIC values of Enterococcus faecal 13 strains. PCR analysis of the two strains showed no mutation sites, 23SrRNA V region mutation sites are mainly G2576U, in addition to T2504A, G2505A, C2610A, C2424U. Conclusion Multi-step in vitro method can induce genistein-resistant linezolid. The mechanism of drug resistance is closely related to the mutation of 23SrRNA V-site, and the number of mutation sites increases with the increase of MIC value.