深圳地区疱疹性咽峡炎疫情的柯萨奇病毒A4型VP1区基因特征分析

Gene characterization of the VP1 region of Coxsackievirus A4 from herpangina cases in Shenzhen of China

目的 分析从2012年和2014年深圳市2起疱疹性咽峡炎疫情中获得的6株柯萨奇病毒A组4型（CVA4）的VP1区遗传进化特征.方法 采用荧光定量反转录（RT）-PCR法对疱疹性咽峡炎患儿咽拭子标本进行总肠道病毒（EV）、人肠道病毒71型（EV71）、柯萨奇病毒A组16型（CVA16）、CVA4、CVA6和CVA10进行检测,对CVA4阳性样本采用RT-PCR方法扩增其VP1区全长序列,并进行核苷酸序列测定和系统进化分析.结果 在2起疫情中共鉴定出6株CVA4病毒,均为GIb基因亚型.其中的2014年深圳CVA4病毒株与2012年深圳CVA4病毒株核苷酸的同源性仅为94.1％（变异率达5.9％）;氨基酸的同源性为98.3％,共发现5个氨基酸突变aa22N-S、aa34T-A、aa63N-S、aa165A-D、aa200T-A.进化分析表明,2012年深圳CVA4病毒株与山东省KF150144株进化关系最近;而2014年深圳CVA4病毒株与吉林省JQ715709株的进化关系最近.结论 2014年深圳CVA4病毒株和2012年病毒株的VP1区存在较大的核苷酸变异,基因内部走势明显不同.

Objective To analyze the phylogeny of the VP1 region of Coxsackie virus A4 （CVA4） from herpangina cases of Shenzhen in 2012 and 2014.Methods Real-time reverse transcription（RT）-PCR method was used to test virus such as human enterovirus71,coxsackievirus A16,coxsackievirus A4,coxsackievirus A6 and coxsackievirus A10.The VP1 gene of CVA4 positive samples were amplified by RTPCR and sequenced.Then the homology and phylogeny analysis of the CVA4 VP1 region was performed.Results The six CVA4 isolates identified in the herpangina cases during 2012 and 2014 were mostly closed with GIb genotypes.The nucleotide and amino acid homology between them were 94.1% （nucleotide mutation rate was 5.9%） and 98.3%,five amino acid mutation were found in CVA4 strain 2014 of Shenzhen：aa22N-S,aa34T-A,aa63N-S,aa165A-D,aa200T-A.The phylogenetic analysis based on VP1 region demonstrates that CVA4 strain of Shenzhen in 2012 had the nearest genetic relationship with CVA4 strain of Shandong isolated in 2010（KF150144）.However,CVA4 strain of Shenzhen in 2014 had the nearest genetic relationship with CVA4 strain of Jilin （JQ715709）isolated in 2006.Conclusions It reveals that all CVA4 strains from the two outbreak of herpangina belong to genotype GIb,the degree of variation in VP1 region of CVA4 strain of Shenzhen in 2014 is obvious compared with that in 2012.There is an obvious difference on internal trend of evolution lineage between the CVA4 strains from 2012 and 2014.