分离自AID患者的白念珠菌唑类耐药株中.pdf

摘 要 分离自 AIDS 患者的白念珠菌唑类耐药株中 Erg11 基因表达研究 研究生：张丽娟 导师：惠艳 教授 摘 要 目的：探讨分离自艾滋病人的白念珠菌唑类耐药株中 Erg11 基因的突变及其与 唑类耐药的关系。方法：将分离自艾滋患者的 100 株真菌标本，行科玛嘉显色培养 鉴定为白念珠菌 85 株，根据 CLSIM27-A 方案的液体微量稀释法行体外药敏试验， 鉴定出 27 株唑类耐药株（耐氟康唑和或伊曲康唑和或伏立康唑）。对27 株耐药株和 3 株敏感株用碱裂解方法提取基因组 DNA ，然后对目的基因进行分段PCR 扩增，将 所得产物测序并与 genebank 中敏感株进行比对分析。结果：在 27 株唑类耐药株中共 发现 25 处点突变，其中 5 处引起氨基酸替换, 分别为 D116E 、E266D 、H485T 、V447I 和 V488I 。30 株实验菌株均发生 H485T 突变，7 株耐药株发生 E266D 突变, 3 株耐药 株发生 V488I 突变，其中 1 株同时发生 E266D 和 V488I ，1 株发生 V447I 突变；2 株敏感株也发生了有意义碱基突变，1 株敏感株发生了有意义基因突变。结论：本研 究共发现 5 个有意义的碱基突变，引起其编码的氨基酸的替换 ,其中 V447I 突变和 H485T 突变国内外文献无报道。 关键词: 白念珠菌；耐药机制；Erg11 基因；突变 1 新疆医科大学医学硕士学位论文 Study on Erg11gene of candida anbicans azole-resistanceisolate from AIDS Postgraduate:ZHANG Li-juan Supervisor: HUI Yan professor Abstract Objective: To investigate the piont mutation of cytochrome P450 lanosterol 14-a demethylase gene Erg11 and its relationship with azole-resistance candida albicans isolated from AIDS. Methods: The azole-resistance and sensitive isolate (fluconazol, Itraconazole, voriconazole) were selected in vitro according to CLSI M27-A protocols.the DNA was extracted by enzymatic lysis method, the objective gene were divided two fragment for PCR reaction respectively, PCR product were sequenced and then contrast with the standarded sensitive strain in Genebank. Results:The 25 piont mutation were found in the 27 azole-resistance strains,and there are 5 significant site which had made amino acids replacement ：D11