Genotyping Of Varicella-Zoster Virus Clinical Strains Isolated In Fujian, China

Objectives:To isolate varicella-zoster virus (VZV) clinical strains from patients with varicella or herpeszoster in Fujian province, China, to differentiate vaccine-strains from wild-strains and identifygenotype(s) of the wild-strains.Methods:1. Vesicular fluids from patients with varicella or herpes zoster were directly inoculated ontomonolayer of human fetal lung fibroblast cell line MRC-5, and passaged for one to three times.When characteristic cytopathic effects (CPEs) of the virus-infected cells were visible, cell smearswere stained immunohistochemically with a monoclonal antibody against immediate early62(IE62) antigens to verify the isolated viruses.2. Viral genomic DNAs were then extracted and their ORF21, ORF22and ORF62wereamplified respectively by polymerase chain reaction (PCR) with a high-definity DNA polymerase.The ORF62PCR products were then digested with restricted endonuclease Sma, BssH II andNae I, respectively, and then compared with those of wild strain pOka and vaccine strain vOka todifferentiate vaccine-strains from wild-strains.3. The ORF21and ORF22PCR products of wild-strains were then sequenced and comparedwith those of Japanese reference strains vOka, as well as those of Dumas,11.8, DR, CA123strains in GENEBANK to identify their genotype(s).Results:1. Forty-four VZV clinical strains were successfully isolated from69patients with varicellaor herpes zoster with a successful isolation rate of63.8%. 2. The ORF62PCR products were not digested by Sma, BssH II, or Nae I. Their restrictionmaps were Sma-BssH II-Nae I-, which were consistent with those of wild strain pOka.3. Among these44VZVclinical isolates, the bases at single neucleoside polymorphism(SNPs) sites,33725and33728in ORF21,37902,38055,38081and38177in ORF22, wereconsistent with those of Japanese reference strains vOka.Conclusions:1. All the44VZV clinical strains isolated from patients with varicella or herpes zoster inFujian province, China, were wild strains.2. Their genotypes were type J, which is identical to clade2according to the internationalVZV genotyping programme. Their genotypes were similar to those of VZV clinical strains fromJapan, South Korea, Mongolia, and other parts of China.