人巨细胞病毒对更昔洛韦耐受性的基因型鉴别方法的建立和应用  

The establishment and application of the method for identifying ganciclovir-resistant genotypes of Human cytomegalovirus

作　　者：曹国君[1] 章莉[1] 华丽[1] 张玥[1] 方风琴[1] 季育华[1] 

机构地区：[1]上海交通大学医学院附属瑞金医院检验科,上海200025

出　　处：《检验医学》2009年第9期642-645,共4页Laboratory Medicine

基　　金：上海市科委重点科技攻关专项基金资助项目(074119521)

摘　　要：目的建立实验室检测人巨细胞病毒(HCMV)更昔洛韦(GCV)耐药的基因型分析的常规方法。方法采用巢式聚合酶链反应(PCR)直接扩增33例HCMV感染患者的外周血白细胞及前期实验同步获得的33例临床分离毒株的HCMV UL97基因片段,对扩增产物进行测序,与HCMV标准毒株AD169序列比对,参照国外报道的耐药相关的热点突变,判定各临床分离毒株的耐药基因型。结果除有1株为HCMV UL97呈M460V突变型外,余32株同标准株AD169的序列一致。同一患者的分离毒株与外周血白细胞中测得HCMV UL97基因序列完全一致。结论建立的实验室检测HCMV耐药的基因型分析完全可以替代传统的耐药表型分析,前者无需分离活病毒,不仅需时短,而且相对安全,其技术要点更适宜临床推广应用。Objective To establish a routine method for analyzing ganciclovir（GCV）-resistant genotypes of Human cytomegalovirus （HCMV）. Methods DNA extracted from peripheral leukocytes of 33 patients with HCMV infection and 33 clinical isolated strains were used to amplify UL97 gene by nested polymerase chain reaction（ PCR）, then they were sequenced. After making a contrast with AD169, the significant mutants were found according to verifying drug-resistant sites. Results After amplifying and sequencing, only one drug-resistant mutant （M460V） was found. The rest of them had sequences identical to AD169. UL97 gene from different samples of one patient was invariable. Conclusions Genotypic assay for drug-resistant detection can completely replace the phenotypic method for clinical routine use due to its advantages, such as rapidity, simplicity, precision, which are more suitable for clinical application.

关 键 词：人巨细胞病毒 UL97基因 更昔洛韦 耐药 基因型 

分 类 号：R373.9[医药卫生—病原生物学]