H3N2亚型猪流感病毒重庆株的分离、鉴定、基因组测定及序列分析
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- 英文论文题名：Isolation,chararcteristics and sequence analysis of the genes of swine influenza viruses H3N2 subtype Chongqing strain
- 论文作者：杨柳 ; 翟少钦 ; 杨睿 ; 张素辉 ; 郑华 ; 张邑帆 ; 付利芝
- 英文论文作者：Yang liu ; Zhai shaoqin ; Yang rui ; Zhang suhui ; Zheng hua ; Zhang yifan ; Fu lizhi ; Chongqing Academy of Animal Sciences ; Key Laboratory of Pig Industry Sciences ; Ministry of Agriculture
- 年：2016
- 作者机构：重庆市畜牧科学院;农业部养猪科学重点实验室;
- 论文关键词：猪流感病毒 ; H3NA亚型 ; HA基因 ; NA基因 ; 序列分析
- 英文论文关键词：swine influenza virus ; H3N2 ; HA gene ; NA gene ; sequence analysis
- 会议召开时间：2016-05-25
- 会议录名称：第七届中国畜牧科技论坛论文集
- 英文会议录名称：Proceeding of the 7th China Animal Husbandry Sci-tech Forum
- 语种：中文
- 分类号：S852.651
- 学会代码：ZGXJ
- 会议名称：第七届中国畜牧科技论坛
- 会议地点：中国重庆
- 主办单位：中国畜牧兽医学会
- 学会名称：中国畜牧兽医学会
- 页数：10
- 文件大小：808k
- 原文格式：O
- 会议级别：全国
摘要
(目的)从猪流感疑似病例中分离鉴定猪流感病毒,并对病毒进行全基因组测序及其序列的分析。(方法)采用鼻棉拭子法从猪流感疑似病猪的鼻中收集病样,制成病毒悬液,经SPF鸡胚增殖培养和纯化。采用红细胞凝集、血凝抑制和神经氨酸酶抑制试验进行病毒亚型的判定,并作电镜观察。运用RT-PCR法扩增病毒的8个基因片段,分别克隆到pMD18-T载体,进行全基因组测序及序列分析。(结果)获得一株猪流感病毒,经血凝和血凝抑制试验,判定为H3N2亚型,命名为A/Swine/Chongqing/CQ/3/2011(H3N2),简称SIV CQ3。基因组序列比较分析显示,SIV CQ3 8个节段与不同时间的猪流感广东分离株(H3N2亚型)相应基因的核苷酸相似性最高,可能来源于H3N2亚型SIV。然而,NP、HA和NA基因编码氨基酸序列与参考毒株A/New York/392/2004(H3N2)的相似性比较低,分别达95%、89%和90%;而血凝素(HA)裂解位点处的氨基酸序列为P-E-K-Q-T-R↓G,与参考毒株的一致,不具有高致病分子特征。此外,HA和NA的糖基化位点、受体结合位点处氨基酸存在一定程度的差异。最后,系统进化树分析显示,NP、HA基因位于SIV群,与A/swine/Guangdong/2006/(H3N2)处于同一分支;而NA基因与禽流感(A/Duck/Ontario/2000/AJ697881)和猪流感位于同一个进化单元,表明这株H3N2亚型流感病毒在不断的发生演变。
aim:isolation,chararcteristics and sequence analysis of the genes of swine influenza virus(SIV) from some SIV-suspected cases in 2011.Method:the viral suspensions were collected from nasal swab material from some SIV-suspected cases,and inoculated into the allantoic cavity of 10-day-old chicken embryos to virus proliferation.The subtype of colleted SIVs can be preliminarily ascertained by using hemagglutinin test(HA),hemagglutination inhibition test(HI) and nueraminidase inhibition test(NI).Allthe eight genes of the SIV were amplified by RT-PCR,cloned into pMD18-T vector and sequenced respectively.Results:A SIV H3 subtype was isolated from SIV-suspected cases according to the results of HI.DNA sequence comparisonrevealed the eight genes from the SIV had sharedthe highest degree of similarity to that of the corresponding genes from the SIV H3N2 subtype from Guangdong isolates at different time,suggested that SIV may be from SIV H3N2 subtype.Comparisons of deduced amino acidsequences of NP,HA and NA of the isolates with that of the reference viruses A/New York/392/2004(H3N2) shared 95%,89%and 90%amino acid similarities,respectively.While the aminoacids sequence of cleavage site of HA is P-E-K-Q-T-R ↓ G,suggesting that SIV did not have the molecular characteristics of high pathogen.In addition,amino acid substitution detection at previously defined glycosylation sites,and receptor binding sites of the HA or NA proteins have some difference.Finally,analysis of NP,HA,NA phylogenetic trees showed the NP,HA might originate from swine influenza virus H3N2 subtype,while NA might originate from aive influenza virus,all that showed this SIV is evolution during the course of its life.
aim:isolation,chararcteristics and sequence analysis of the genes of swine influenza virus(SIV) from some SIV-suspected cases in 2011.Method:the viral suspensions were collected from nasal swab material from some SIV-suspected cases,and inoculated into the allantoic cavity of 10-day-old chicken embryos to virus proliferation.The subtype of colleted SIVs can be preliminarily ascertained by using hemagglutinin test(HA),hemagglutination inhibition test(HI) and nueraminidase inhibition test(NI).Allthe eight genes of the SIV were amplified by RT-PCR,cloned into pMD18-T vector and sequenced respectively.Results:A SIV H3 subtype was isolated from SIV-suspected cases according to the results of HI.DNA sequence comparisonrevealed the eight genes from the SIV had sharedthe highest degree of similarity to that of the corresponding genes from the SIV H3N2 subtype from Guangdong isolates at different time,suggested that SIV may be from SIV H3N2 subtype.Comparisons of deduced amino acidsequences of NP,HA and NA of the isolates with that of the reference viruses A/New York/392/2004(H3N2) shared 95%,89%and 90%amino acid similarities,respectively.While the aminoacids sequence of cleavage site of HA is P-E-K-Q-T-R ↓ G,suggesting that SIV did not have the molecular characteristics of high pathogen.In addition,amino acid substitution detection at previously defined glycosylation sites,and receptor binding sites of the HA or NA proteins have some difference.Finally,analysis of NP,HA,NA phylogenetic trees showed the NP,HA might originate from swine influenza virus H3N2 subtype,while NA might originate from aive influenza virus,all that showed this SIV is evolution during the course of its life.
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