Complete Genome Sequencing And Analysis Of Enterovirus 71 Isolated In Kunming City In 2016

Objective To understand the pathogenic features of hand-foot-mouth disease(HFMD)and the genetic characteristics of isolates of enterovirus 71(EV71)in Kunming in 2016.Methods 20 throat swabs were collected from HFMD patients in Kunming City.10 samples(KM01-KM 10)were randomly selected from EV71 positive ones for Vero-E6 cell separation.The whole viral genome clones of 10 samples,excluding the poly-A tail,were obtained by Over-lap extension PCR and then sequenced.The nucleotide and amino acid sequences of KM01-KM 10 were compared with 27 other known EV71 strains.The full-genome sequences of those EV71 strains were analysed systematically by using the softwares of bioinformatics.Results KM01-KM10 had the similarly basic characteristics of Enterovirus.The full length genome of KM01-KM10(not including Poly A tail)were 7405nt and there were only one ORF,which encoding a polyprotein including 2193 amino acids,and no insertion or deletion was included in the coding region.The comparisons of different regions of nucleotide sequences and amino acid of 28 strains showed that KM01-KM10 shared the highest nucleotide and amino acid similarities in VP1,VP4,P1,P2,P3 and complete sequence with the Fuyang strain EU703812.Phylogenetic analysis of 27 strains in VP1,P1 and complete sequence regions of the viral genome indicated that KM01-KM10 strain have the nearest genetic relationship with EU703812 which belongs to C4 subgenotype.Conlusion 1.In this study,complete genome sequence of 10 EV71 samples were obtained.2.The comparison of similarities and the phylogenetic analysis showed that KM01-KM10 strain belong to C4 subgenotype,which corresponded with the subgenotype of EV7 1 outbreaks in other areas of China in 2008.Analysis indicated that the relationship among KM01-KM10 and other EV71 isolated in China 2008 was closed,they may have the same ancestor.3.Based on the bootscan analysis,no recombination pattern were observed in the strain KM01-KM09,but recombinant forms of EV71 are found in KM 10.