白假丝酵母菌耐吡咯类药物的ERG11基因分析

Genetic Analysis of ERG11 in Candida Albicans Resistance to Azole Agents

目的 研究白假丝酵母菌耐吡咯类药物的ERG11的变异情况.方法 将93例诊断为真菌性阴道炎的患者的阴道分泌物标本进行真菌培养,筛选白假丝酵母菌菌株,利用纸片扩散法进行氟康唑、酮康唑、咪康唑药敏试验,用加热裂解法提取菌株DNA,扩增ERG11基因,扩增后的PCR产物进行双向测序,测序结果与GenBank中的标准序列（SC5314）比较分析.结果 93例均培养出假丝酵母菌,包括60株白假丝酵母菌,19株热带假丝酵母菌,9株克柔假丝酵母菌和5株光滑假丝酵母菌;白假丝酵母菌对氟康唑、酮康唑、咪康唑耐药率分别为13.33%,20.00％和51.67％;对白假丝酵母菌的ERG11基因测序发现存在25个碱基突变位点,其中13个同义突变,12个错义突变,其中有6个是新变异：V36F,V51L,T123I,E194K,Y257H和K344N.结论 耐吡咯类药物白假丝酵母菌ERG11基因有多个错义突变位点,其中某些位点突变导致的氨基酸变异可能与其耐药性产生有关.

Objective To investigate the mutations of ERG11 gene in azole resistant Candida albicans.Methods The vaginal secretion specimens of 93 fungal vaginitis patients were given fungal culture.Disk diffusion method （K B method） was performed for preliminary culture of strains resistant to fluconazole,ketoconazole and miconazole.Total DNA was extracted by heating pyrolysis method.And the target genes were amplified.The amplified PCR product underwent DNA sequencing,and the sequencing results were compared with the standard sequence in GenBank （SC5314）.Results 60 strains of Candida albicans,19 strains of Candida tropicalis,9 strains of Candida krusei and 5 strains of Candida glabrata were cultured by fungal culture;of Candida albicans,the resistant rate to fluconazole,ketoconazole,and miconazole were 13.33 ％,20.00 ％ and 51.67 ％,respectively.A total of 25 mutation sites were existed in Candida albicans,in which there were 13 synonymous mutations and 12 missense mutations.There were 6 new mutations included V36F,V51L,T123I,E194K,Y257H and K344N.Conclusion Multiple sites of missense mutations are existed in ERG11 gene of azole-resistant Candida albicans,the amino acid mutation caused by some of which may be related to the resistance to drugs.