浙江衢州地区儿童病毒性脑炎埃可病毒优势血清型与VP1基因分析  

Predominant serotypes and VP1 gene analysis of Echovirus from viral encephalitis children in Quzhou area of Zhejiang province

作　　者：邱晓枫[1] 曹国平[2] 潘劲草[1] 张国忠[1] 李钧[1] 占炳东 陈树昶[1] 吕华坤[3] Qiu Xiaofeng;Cao Guoping;Pan Jincao;Zhang Guozhong;Li Jun;Zhan Bingdong;Chen Shuchang;Lyu Huakun(Hangzhou Center for Disease Control and Prevention,Hangzhou 310021,China;Quzhou Center for Disease Control and Prevention,Quzhou 324000,China;Zhejiang Provincial Center for Disease Control and Prevention,Hangzhou 310051,China)

机构地区：[1]杭州市疾病预防控制中心,310021 [2]衢州市疾病预防控制中心,324000 [3]浙江省疾病预防控制中心,杭州310051

出　　处：《中华实验和临床病毒学杂志》2020年第2期169-174,共6页Chinese Journal of Experimental and Clinical Virology

基　　金：浙江省公益技术应用基金(2012C33063);杭州市农业与社会发展科研计划项目(20170533B76)。

摘　　要：目的了解浙江省衢州地区儿童病毒性脑炎埃可病毒(Echovirus,ECHOV)优势血清型与变迁并分析ECHOV VP1基因分子特征。方法采集53例病毒性脑炎患儿脑脊液标本进行病毒分离培养。采用荧光RT-PCR和PCR分别检测脑脊液标本中人肠道病毒(human enterovirus,HEV)包括ECHOV、柯萨奇病毒(coxsackie virus,CV)和新型肠道病毒(enterovirus,EV),以及流行性乙型脑炎病毒(japanese encephalitis virus,JEV)、腮腺炎病毒(mumps virus,MuV)、西尼罗病毒(west nile virus,WNV)、基孔肯雅病毒(chikungunya virus,CHIKV)、单纯疱疹病毒(herpes simplex virus,HSV)和巨细胞病毒(cytomegalovirus,CMV)。HEV阳性脑脊液标本采用RT-PCR法扩增HEV-B组全长VP1基因序列,测序后用多种生物信息学软件分析ECHOV VP1基因型、基因重组和遗传进化特征。结果RD细胞分离培养出6株毒株,但Hep-2细胞分离培养结果均阴性。11份标本HEV通用荧光RT-PCR结果阳性,但其他病毒检测结果均阴性。11份HEV阳性标本中6份检出ECHOV,与分离培养结果一致,分别为4株ECHO6、1株为ECHO7和1株ECHO30血清型,柯萨奇病毒和EV检测结果均阴性。4株ECHO6病毒属于ECHO6-C2基因亚型,但分为ECHO6-41/46和ECHO6-45/48两个流行克隆,ECHO7和ECHO30分别属于ECHO7-C和ECHO30-C基因型。ECHO6与ECHO30病毒在进化过程中存在VP1基因重组。结论ECHOV是该地区儿童病毒性脑炎主要病原体,且可能存在局部暴发流行。ECHO6与ECHO30病毒VP1基因存在重组可能性,ECHO6有成为ECHOV优势血清型的可能。Objective To investigate the predominant serotypes and diversification of Echovirus(ECHOV)from viral encephalitis children in Quzhou area of Zhejiang province and the molecular characteristics of the ECHOV VP1 genes.Methods Cerebrospinal fluid samples from 53 children with viral encephalitis were collected for viral isolation/culture.Fluorescent RT-PCR or PCR was used to detect human enteroviruses(HEV)including ECHOV,coxsackievirus(CoV)and new enterovirus(EV),and japanese encephalitis virus(JEV),mumps virus(MuV),west Nile virus(WNV)and chikungunya virus(CHLKV)or herpes simplex virus(HSV)and cytomegalovirus(CMV)in the cerebrospinal fluid samples.The complete VP1 gene sequence of HEV-B group in the HEV-positive cerebrospinal fluid samples was amplified by RT-PCR and sequenced,and then the typing of Echovirus isolates was performed.The VP1 genotypes of Echovirus isolates,gene recombination,inheritance and evolution characteristics were analyzed using multiple bioinformatic software.Results Six viral strains were isolated by cell culture using rhabdomyosarcoma(RD)cells but not human epithelial-2(Hep-2)cells.Eleven cerebrospinal fluid samples were positive for HEV by RT-PCR but the detection result of all the other viruses were negative.In the 11 HEV-positive samples,6 samples were positive for ECHOV(4 for ECHO6,1 for ECHO7 and 1 for ECHO30 serotype),which was coincident with the isolation result,but CoV and EV were undetectable.The 4 ECHO6 isolates belonged to ECHO6-C2 subgenotype but can be divided into two epidemic clones(ECHO6-41/46 and ECHO6-45/48).The ECHO7 and ECHO30 isolates belonged to ECHO7-C and ECHO30-C genotypes.The VP1 gene recombination between the ECHO6 and ECHO30 isolates were found during their evolutionary process.Conclusions ECHOV is the major pathogen of viral encephalitis children in the area,and there is a possibility of local outbreak or epidemic.Because of the possibility of recombination of the VP1 gene of ECHO6 and ECHO30 virus,ECHO6 may become the dominant ECHOV serotype.

关 键 词：病毒性脑炎 埃可病毒 VP1基因 血清型/基因型 基因重组 

分 类 号：R72[医药卫生—儿科] R4[医药卫生—临床医学]