2019年福建省汉坦病毒分离株全基因组序列测定及分析  

Complete genome sequencing and analysis of hantavirus strains isolated in Fujian,2019

作　　者：刘菁[1] 陈阳[1] 韩腾伟[1] 刘维俊[1] 曾志伟 林文[1] 王加熊[1] 林代华[1] 肖方震[1] Liu Jing;Chen Yang;Han Tengwei;Liu Weijun;Zeng Zhiwei;Lin Wen;Wang Jiaxiong;Lin Daihua;Xiao Fangzhen(Fujian Provincial Center for Disease Control and Prevention,Fujian Provincial Key Laboratory of Zoonosis Research,Fuzhou 350012,Fujian,China)

机构地区：[1]福建省疾病预防控制中心,福建省人兽共患病研究重点实验室,福建福州350012

出　　处：《疾病监测》2022年第9期1165-1169,共5页Disease Surveillance

基　　金：福建省卫生健康科技计划项目(No.2019-1-19,No.2019-ZQN-27);福建省科技创新平台建设项目(No.2019Y2001)。

摘　　要：目的从分子水平分析2018—2020年福建省汉坦病毒(HV)分离株的型别,为疾病的预防控制提供实验依据。方法采集2018—2020年福建省肾综合征出血热(HFRS)急性期患者血清及监测点鼠肺标本。采用胶体金法检测血清样本中HV的IgM、IgG抗体,采用免疫荧光法检测鼠肺样本中HV抗原,阳性样本接种E6细胞分离病毒,用反转录聚合酶链式反应(RT-PCR)方法检测标本培养物上清,免疫荧光检测培养细胞以鉴定HV。采用高通量测序技术对分离株进行全基因组测序,MEGA 11.0软件对其进行序列比对和进化分析。结果捕获鼠形动物879只,阳性率为2.84%。分离得到2株病毒,均来自黑线姬鼠。基因组序列分析显示为汉滩病毒(HTNV)。采用最大似然法构建系统发育树中,所分离病毒株形成独立支系,与LongquanAa-10-101、Z251、LongquanMf-08-92等浙江地区HV分离株基因组核苷酸序列一致性较高,约为95.81%,聚集形成一个分支。氨基酸序列比对分析显示,所分离株与疫苗株Z10核蛋白相比,存在Thr43Ala、Lys95Met突变;与HTNV国际标准株76-118和疫苗株84FLi相比,缺少一个第N766位的糖基化位点。结论本研究提示福建地区流行HV具有独特基因特征,与浙江地区HV分离株具有较近的亲缘关系,可能源自共同进化的祖先,所分离病毒包膜糖蛋白和核衣壳蛋白的氨基酸变异所造成的影响,值得进一步关注和研究。Objective To analyze the gene sequence of Hantavirus(HV)isolates in Fujian province in 2019 at molecular level and provide experimental evidence for the prevention and control hemorrhagic fever with renal syndrome(HFRS).Methods The serum samples of patients with acute HFRS and rat lung specimens were collected from 2018 to 2020.HV IgM IgG antibody in the serum sample were detected by colloidal gold,HV antigen in rat lung specimens were detected by immunofluorescence assay test.Positive samples were inoculated with E6 cell line to isolate virus.The supernatants of the cultures of specimens were detected by reverse transcription-polymerase chain reaction and the cells were detected by immunofluorescence assay test.The genome of HV isolates were sequenced using high-throughput sequencing technique.Finally,the sequence alignment and phylogenetic analysis were conducted with software MEGA 11.0.Results A total of 879 rodents were captured,and the positive rate was 2.84%.Two strains of Hantavirus were isolated from Apodemus agrarius.Genome sequence analysis showed Hantaan virus(HTNV).Phylogenetic analysis using the maximum likelihood method showed that the isolates belonged an distinct lineage and formed a clade with strains isolated in Zhejiang(longquanAA-10-101,Z251 and LongquanMF-08-92),showing 95.81%nt sequence identity.Compared with vaccine strain Z10,the amino variations of Thr43Ala and Lys95Met were observed at nucleoproteins of isolates.Compared with HTNV international reference strain 76-118 and vaccine strain 84FLi,the isolates lacked glycoylation site at N766.Conclusion Hantavirus isolates from Fujian had unique genetic characteristics and exhibited a close evolutionary relationship with strains isolated in Zhejiang which might derive from co-evolving ancestors.The effect of amino variations in envelope glycoproteins and nucleocapsid proteins of Fujian isolates need further research.

关 键 词：肾综合征出血热 汉坦病毒 基因组 序列测定 系统发育树 

分 类 号：R211[医药卫生—中医学] R373.32