9株甲型肝炎病毒流行株全基因组特征分析

Analysis of the whole genome characteristics of 9 hepatitis A virus strains in China

目的分析四川南充和新疆和田甲型肝炎病毒(hepatitis A virus,HAV)分离株全基因组序列特征。方法收集2006年四川南充同1起暴发的3份和2016年新疆和田6份急性期甲肝患者血清标本,通过核酸提取、逆转录、分段巢式PCR、测序和序列拼接获得9条HAV全基因组序列。利用生物信息学软件进行系统进化、抗原中和位点、重组和选择压力分析。结果VP1-2A连接区基因分型表明9条HAV序列都属于IA亚型,分为4个不同的分支,其中3条南充序列和3条和田序列属于同一分支;全基因组序列表明,3条南充序列核苷酸和氨基酸同源性分别为99.99%~100%和100%,与3条和田序列全基因组核苷酸和氨基酸差异分别为0.50%~0.57%和0.08%~0.17%。9条HAV全基因组核苷酸和氨基酸同源性分别为98.29%~100%和99.62%~100%,与我国hd9和蒙古MNA12-001关系最近(核苷酸和氨基酸同源性分别为:与hd998.60%~99.50%和99.75%~99.92%;与MNA12-00198.45%~99.32%和99.71%~99.87%)。9条HAV序列在已发表的抗原中和位点处未发生改变,未发现重组,选择压力分析表明处于负向选择。结论我国存在多株HAV流行株;9条HAV氨基酸序列在主要抗原中和位点处很保守。

Objective To analyze the characteristics of the whole genome of hepatitis A virus(HAV)strains from Nanchong,Sichuan and Hetian,Xinjiang.Methods Serum samples of 9 patients with acute hepatitis A were selected for HAV whole genome sequencing.Among them,3 were from the same outbreak in Nanchong,Sichuan(2006)and 6 were from Hetian,Xinjiang(2016).Nine full length HAV sequences were obtained by viral RNA extraction,reverse transcription,nested PCR and sequencing.Phylogenetic analysis,neutralizing epitope sites,recombination and natural selection analysis were also performed.Results Genotyping of HAV VP1-2A region indicated that the 9 HAV sequences all belonged to sub-genotype IA and were divided into four different branches,with 3 Nanchong sequences and 3 Hetian sequences in the same branch.While the whole genome sequence showed that the nucleotide and amino acid homology of 3 Nanchong sequences were 99.99%~100%and 100%,respectively.The difference of nucleotides and amino acids were 0.50%~0.57%and 0.08%~0.17%between 3 Nanchong and 3 Hetian sequences,respectively.The nucleotide and amino acid identities of nine HAV genomes were 98.29%~100%and 99.62%~100%,which was closely related to hd9 in China and MNA12-001 in Mongolia(nucleotide and amino acid homology:with hd998.60%~99.50%and 99.75%~99.92%;and with MNA12-00198.45%~99.32%and 99.71%~99.87%,respectively).The 9 HAV sequences did not change at the neutralizing epitope sites,and no recombination events were found.Natural selection analysis indicated that the HAV coding region was subjected to a negative selection.Conclusions There are many HAV epidemic strains in China;the 9 HAV amino acid sequences were conserved at the major neutralizing epitope sites.