Isolation and analysis of genetic characteristics of a new strain of enterovirus EV-C99 from a healthy child in Yunnan Province / 中华微生物学和免疫学杂志

ABSTRACT

Objective To isolate and analyze the genetic characteristics of a new strain of entero-virus EV-C99 from a healthy child in Yunnan Province in 2016. Methods Virus isolation was performed ac-cording to the World Health Organization ( WHO) recommended procedures. Viral RNA was extracted from the supernatant of cell culture. RT-PCR and sequencing analysis of VP1 gene were used for virus identifica-tion. VP1 sequence was edited and stitched by Sequencher 5. 0 software. The edited sequence was BLAST searched in GenBank and the preliminary result indicated that it was an EV-C99 virus. Nucleotide (nt) and amino acid (aa) identities were calculated by MEGA5. 2 software. Serotype of the virus was identified ac-cording to the standard for enterovirus sequence typing. Results The virus could be amplified by 494/496 and 495/497 primer pairs and the edited sequence was about 1 200 bp. Result of the " BLAST" search showed that it was a new strain of enterovirus EV-C99. Comparative analysis with the prototype strain BAN00-10461 (GenBank IDEF015008) by MEGA5. 2 software showed that the VP1 gene of that virus was 909 bp and the identities between them were 77. 05% in nt sequence and 90. 04% in aa sequence. According to the standard for enterovirus sequence typing,the virus was a new strain of enterovirus EV-C99 belonging to hu-man enterovirus species C (EV-C). Conclusion A new strain of enterovirus EV-C99 was isolated during an investigation of enteroviruses among healthy children in Yunnan Province in 2016. To our knowledge,this is the third report of EV-C99 in China. Phylogenetic analysis reveals that the EV-C99 Yunnan strain,Xinjiang strains and Shandong strains all belong to genotype B,but group into different clusters,indicating that Chinese strains have diverse genetic characteristics.