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目的对7例临床水痘患者的疱疹液样本,进行水痘带状疱疹病毒(varicella-herpes zoster virus,VZV)的分离及鉴定分析。方法对7例临床水痘患者的疱疹液,用2BS细胞进行病毒分离及病毒滴度检测;用特异性引物分别对病毒分离株及疫苗株(Oka株)的ORF68、ORF54、ORF38进行聚合酶链反应(Polymerase Chain Reaction,PCR)扩增及测序,用DNAMAN软件对病毒分离株的ORF68序列与Dumas株序列进行比对鉴定,并对病毒分离株和Oka株的ORF54、ORF38进行限制性片段长度多态性(restriction fragment length polymorphism,RFLP)分析。结果从7例临床水痘患者的疱疹液样本中,分离到4株VZV病毒株;病毒分离株的细胞病变(cytopathic effect,CPE)及病毒滴度随传代次数的增加而增强;4株病毒分离株的ORF68序列与Dumas株完全一致;ORF54、ORF38的RFLP结果显示,4株病毒分离株均为BglⅠ+PstⅠ+型,Oka株为BglⅠ+PstⅠ-型。结论成功分离的4株病毒分离株均为VZV野生毒株。
Objective To isolate and identify varicella-herpes zoster virus (VZV) from herpes simplex of seven patients with clinical chickenpox. Methods The herpes zoster fluid of 7 patients with clinical chickenpox were tested for virus isolation and virus titer with 2BS cells. The specific primers were used to detect the ORF68, ORF54 and ORF38 of the virus isolates and vaccine strains (Oka strain) (Polymerase Chain Reaction, PCR) amplification and sequencing, DNAMAN software was used to identify the ORF68 sequence of the virus isolates and Dumas strain sequence, and the ORF54 and ORF38 of the virus isolates and Oka strains were subjected to restriction fragment length polymorphism Restriction fragment length polymorphism (RFLP) analysis. Results Four strains of VZV were isolated from herpes fluid samples of seven patients with clinical chickenpox. Cytopathic effect (CPE) and virus titer of the virus isolates were increased with the increase of passage number. Four strains of virus isolates ORF68 sequence was identical with that of Dumas. RFLP results of ORF54 and ORF38 showed that all four isolates were BglⅠ + PstⅠ + and Oka was BglⅠ + PstⅠ-. Conclusion All the 4 isolates that were successfully isolated were wild type VZV strains.