四川省德阳市柯萨奇病毒A组5型基因特征分析

Genetic characteristics of coxsackievirus A5 in Deyang city,Sichuan province

目的分析四川省德阳市手足口病病例中的柯萨奇病毒A组5型(coxsackievirus A5,CVA5)的基因特征,为防控CVA5提供科学依据。方法对德阳市2013—2021年手足口病监测中收集到的疑似病例咽拭子标本,采用实时荧光聚合酶链反应(real-time fluorogenic quantitative polymerase chain reaction,Q-PCR)进行肠道病毒核酸检测,选择Q-PCR检测结果CT值<30的非肠道病毒71型(EV-A71)/柯萨奇病毒A组16型(CVA16)的其他肠道病毒核酸阳性标本,分别进行VP4/VP2区和VP1区基因扩增和核苷酸测序,再用MEGA 7.0.26软件构建CVA5系统进化树,分析其基因特征。结果2013—2021年德阳市共收集手足口病疑似病例标本3115份,经Q-PCR检测非EV-A71/CVA16的其他肠道病毒核酸阳性标本为1314份;选择其中的353份标本进行VP4/VP2区和VP1区核苷酸测序,在280份标本中检测出8种肠道病毒型别,其中10份为CVA5。德阳市10份CVA5标本之间,VP1区核苷酸序列相似性为91.4%~99.8%,推导氨基酸序列同源性96.6%~99.6%;同CVA5原型株比较,核苷酸相似性和推导氨基酸同源性分别为81.3%~82.5%和94.9%~96.6%,有19个位点发生了氨基酸变异。进化树分析显示,德阳市CVA5与绝大多数中国CVA5聚集在一起,均为C1基因亚型3谱系,但分布在3个不同的进化分支中,分支间平均遗传距离为6.2%~6.4%。结论德阳市10株CVA5均为C1基因亚型3谱系,分布在3个不同的进化分支。CVA5不但能引起手足口病和疱疹性咽峡炎,还会引起急性脑病、急性弛缓性脊髓炎、胃肠炎等疾病,今后应加强CVA5的检测,并开展基因组测序,更全面了解其基因特征。

Objective To analyze genetic characteristics of coxsackievirus A5(CVA5)in Deyang city in Sichuan province,so as to provide evidence for its prevention and control.Methods Throat swab specimens were collected from suspected patients with hand,foot and mouth disease(HFMD)from 2013 to 2021 in Deyang and analyzed with commercial real-time fluorogenic quantitative polymerase chain reaction(Q-PCR)assay kit to detect RNA of enteovirus(EV).Specimens with CT values<30 for non-EV-A71/CVA16 enteroviruses were selected for VP4/VP2 and VP1 gene amplification and nucleotide sequencing,respectively.MEGA 7.0.26 software was used to build the phylogenetic tree of CVA5 and to analyze its genetic characteristics.Results A total of 3115 specimens were collected from suspected HFMD cases in Deyang during 2013-2021,of which 1314 were RNA-positive for non-EV-A71/CVA16 enterovirus.Three hundred and fifty-three specimens were selected for nucleotide sequencing of VP4/VP2 and VP1 genes,among which 280 were successfully amplified and sequenced,and 8 types of enterovirus were identified,including CVA5 identified in 10 specimens.Among the 10 CVA5 strains,similarities of nucleotide and deduced amino acid sequences in VP1 gene were 91.4%-99.8%and 96.6%-99.6%,respectively;the similarities compared with CVA5 prototype strain were 81.3%-82.5%and 94.9%-96.6%,respectively,and amino acid variation occurred on 19 sites.Phylogenetic tree analysis of VP1 gene indicated that Deyang strains belong to lineage 3 of C1 subgenotype,clustering with most CVA4 strains detected in other areas in China,but in 3 separate branches with mean genetic distances of 6.2%-6.4%.Conclusions The 10 CVA5 strains detected in Deyang all belong to lineage 3 of C1 subgenotype,distributed in 3 evolutionary branches.CVA5 can cause not only HFMD and herpangina,but also acute encephalopathy,acute flaccid myelitis,and gastroenteritis.Detection of CVA5 should be strengthened in the future,and genome sequencing should be carried out to further understand its genetic characteristics.