摘要
目的研究比较3种念珠菌对氟康唑耐药的易感性及其体外诱导耐药株的耐药机制。方法选取同一患者中同时分离的对氟康唑敏感的白念珠菌、光滑念珠菌和热带念珠菌各1株,在体外氟康唑的作用下诱导其成为耐药株。采用罗丹明6G试验比较敏感株与耐药株的外排泵作用,RT-PCR检测外排相关基因CDR1、CDR2以及靶酶编码基因ERG11的表达,并对ERG11基因进行PCR扩增和测序,同时通过罗丹明123试验对3种念珠菌的线粒体膜电位(ΔΨm)进行检测。结果光滑念珠菌最易被氟康唑诱导为耐药株,其CDR1的过度表达引起外排泵作用增强。白念珠菌和热带念珠菌的诱导耐药株以ERG11表达增加为主,其中白念珠菌的ERG11存在V437I、A430V、S263L和T128K突变位点。此外,白念珠菌和光滑念珠菌的诱导耐药株表现为呼吸缺陷。结论不同念珠菌对氟康唑耐药的易感性不同,耐药机制也存在差异。
Objective To investigate the susceptibility and mechanisms of resistance to fluconazoles in clinical isolates of three Candidaspecies.Methods One strain each of Candida albicans,Candida glabrata and Candida tropicalis was isolated from the same patient simultaneously.These three strains were induced to resist to fluconazole in vitro.Efflux of rhodamine 6G was performed to evaluate the effects of efflux pumps.The expression levels of transporter genes CDR1,CDR2 and azole antifungal target gene ERG11 were examined by real time RT-PCR.The sequence of ERG11 was determined by PCR based DNA sequencing.Meanwhile,rhodamine 123 was used to analyze the respiratory capacity of the three Candida strains.Results Efflux pumps showed stronger effects in fluconazole-resistant C.albicans and C.glabratathan in susceptible isolates,but no obvious change was observed in resistant strain of C.tropicalis.CDR1 was significantly overexpressed in the resistant strain of C.glabrata,while ERG11 was overexpressed in C.albicans and C.tropicalis.ERG11 mutations V437I,A430V,S263L and T128K were identified only in the resistant strain of C.albicans.Fluconazole-resistant C.albicans and C.glabrata were respiratory deficient.Conclusions The three Candida species showed different performance in terms of susceptibility to fluconazole and the mechanisms of fluconazole resistance.
出处
《中国感染与化疗杂志》
CAS
北大核心
2013年第4期296-301,共6页
Chinese Journal of Infection and Chemotherapy
基金
上海交通大学医学院科研基金项目(12XJ10012)
关键词
念珠菌
氟康唑
外排泵
ERG11
呼吸缺陷
Candidaspp.
fluconazole
transporter
ERG11
respiratory deficiency