Virus isolation and gene evolution analysis on Jiangsu local strain of coxsackievirus B4 type

Abstract

Objective To established the isolation method of coxsackievirus B4 (CVB4), and to perform genetic evolution analysis of the isolated Jiangsu local strain. Methods Fluorescence quantitative PCR was used to detect enteroviruses in the throat swabs from patients with respiratory infections admitted to the sentinel hospitals of Jiangsu province. The positive samples were used for virus isolation with HeLa cell line, followed by VP4 gene sequencing and phylogenetic analysis. Results Twelve samples were positive of enterovirus by fluorescence quantitative PCR. Cytopathic effect was observed in one sample during virus isolation. The VP4 gene were amplified by specific RT-PCR. The sequence analysis showed that the similarities of the VP4 gene sequence with the other 20 CVB4 strains ranged from 86.56% to 93.59%, and the strain was identified as a Jiangsu local strain of CVB4. The phylogenetic tree showed that the strain only clustered with CVB4 strain KY369904. Conclusions HeLa cell line was used to isolate and identify one strain of enterovirus in the present study. The virus strain was identified as Jiangsu local strain of CVB4 using evolution analysis of VP4 gene. The work presented important value for the prevention and control of local enterovirus infections. Key words: Coxsackievirus B4; Enterovirus; Virus isolation; Cytopathic effect