无菌性脑膜炎暴发中病原学检测及埃柯30型肠道病毒的基因特征和分子进化分析

Detection of echovirus 30 causing aseptic meningitis outbreak in Quanzhou and analysis on its genetic characteristics and molecular evolution

目的了解引起福建省泉州地区2011年无菌性脑膜炎暴发疫情的埃柯30型肠道病毒(echovirus 30,ECHO30)病原及基因特征,分析基因变异情况及进化来源。方法对泉州市2011年无菌性脑膜炎暴发疫情中的临床样本采用实时荧光定量反转录-聚合酶链反应(rRT-PCR)方法进行肠道病毒通用型、肠道病毒71型(EV71)和柯萨奇病毒A组16型(Cox A16)核酸检测,随机选5份阳性样本进行肠道病毒VP1区核苷酸序列测定,将测序所得VP1序列用BLAST程序在GenBank上序列搜寻比对,确定其基因型,并通过构建进化树分析其遗传进化规律。结果 47份样本经rRT-PCR检测,有39份非EV71和Cox A16的其他肠道病毒核酸阳性,5份阳性样本的肠道病毒VP1区核苷酸序列均为876 bp,且同源性最高,均大于98.9%,最高仅相差一个核苷酸,与它们具有较高同源性的均是ECHO30。与本次分离株亲缘关系最近的一组ECHO30分别是2008年河南省和浙江省的分离株,与中国台湾2001年无菌性脑炎分离株以及印度2011年分离株则距离较远;原型株Bastianni和20世纪90年代美国分离株和本次分离到的ECHO30毒株距离最远。结论 2011年ECHO30在泉州地区发生一定程度的传播流行,并导致无菌性脑膜炎疫情的局部暴发流行,进化树分析表明本次分离株与国内近年ECHO30分离株亲缘关系较近,而与国外分离株则相对较远。

Objective To understand the pathogen causing an aseptic meningitis outbreak in Quanzhou in 2011 and pathogen＇ s genetic characteristics and molecular evolution. Methods Clinical specimens were collected from the aseptic meningitis cases to detect enterovirus 71 （EV71）, coxsackievirus A16 （Cox A16） and universal RNA by using real-time RT- PCR. The complete nucleotide sequencing of VP1 region of 5 random positive specimens were conducted, BLAST program was used to identify the genotype by nudeotide pairwise-alignment with all available sequences in GenBank database, and the phylogenetic tree was drawn to compare with other enteroviralsequences. Results Real-time RT-HR indicated that 39 of 47 clinical specimens were universal enterovirus positive but not EV71 or Cox A16. The complete nudeotide sequence of VP1 region of 5 enterovirus isolates was 876nt with homology of 〉98.9%, and had closer relationships with echovirus 30. These strains had close distance with the strains isolated in Henan and Zhejiang in 2008 and far distance with those isolated in Taiwan of China （2001） and India （2011）. When compared with echovirus 30 prototype strain Bastianni, the distance was farthest. Conclusion Echovirus 30 circulated in Quanzhou in 2011 and caused aseptic meningitis outbreak in some areas. Phylogenetic trees analyses showed that the strains isolated from this outbreak had close distance with these isolated in China in recent years and far distance with those isolated abroad.