摘要
本实验室于2016年在天津分离到一株猪流感病毒(SIV),命名为A/swine/Tianjin/312/2016(H1N1)(简称为TJ312株)。为了解该株病毒的生物学特性及遗传演化特征,本研究对经PCR分段扩增该病毒全基因组并测序,采用SeqMan软件拼接成全基因组序列经BLAST比对,得到与该SIV各基因节段序列同源性最高的病毒株,采用mafft分析其各基因节段编码氨基酸序列的分子特征,利用MEGA5.1软件分别构建该病毒8个基因节段的进化树。结果显示,SIV TJ312株各基因节段与2016年分离自天津的H1N1 SIV相应各基因节段的同源性在99%~100%。进化树结果显示该病毒株HA、NA和M基因均来自欧亚类禽H1N1(EA H1N1)SIV谱系;PB2、PB1、PA和NP基因来自2009/H1N1流感病毒谱系;NS基因来自北美三重配H1N2 SIV谱系。蛋白分子特征分析结果显示,HA蛋白碱性氨基酸裂解位点序列为PSIQSR↓G,符合低致病性流感病毒分子特征;受体结合位点符合人源唾液酸受体(α-2,6唾液酸受体)的分子特性;NA蛋白aa275为H、aa295为N,且该蛋白头部区域未缺失,表明该病毒对奥司他韦类药物敏感;PA蛋白^(356)R为病毒复制增强和对哺乳动物致病性增强的分子特征;NP蛋白存在^(41)I以及^(210)E病毒聚合酶活性增强的分子特征;M2蛋白的^(31)N表示该SIV对金刚烷胺类药物耐药。将该病毒以10^(6) EID_(50)/只经鼻腔感染2只雪貂,感染后96 h剖杀,采集各脏器组织观察其剖检病变并对出现病变的肺脏制备病理切片观察其组织病变,采用免疫组化试验检测肺脏组织中的病毒抗原。将采集的所有脏器处理后接种鸡胚检测脏器内的病毒滴度评估该病毒在雪貂体内的复制能力;将该株病毒以上述剂量经鼻腔感染3只雪貂分别放入3个铁笼内作为感染组,24 h后在相邻3个铁笼内分别放入1只未感染雪貂作为传播组。所有雪貂自感染组接种病毒后2 d起,隔天采集鼻洗液至第14 d,通过检测雪貂鼻洗液内�
In order to investigate the genetic evolution background and biological characteristics of the swine influenza virus(SIV)A/swine/Tianjin/312/2016(H1N1)isolated from Tianjin Province in 2016,the whole genome of the virus was amplified by PCR and sequenced in this study,and the complete genome sequence was spliced by SeqMan software and compared by BLAST to obtain the virus strain with the highest homology with each gene segment sequence.Mafft was used to analyze the molecular characteristics of amino acid sequences encoded by each gene segment and MEGA5.1 software was used to construct the evolutionary tree of 8 gene segments of the virus.The results showed that all gene segments of SIV TJ312 strain share 99%-100%homology to the corresponding gene segments of H1N1 SIVs isolated from Tianjin in 2016.The results of the evolutionary tree showed that the HA,NA and M genes of this strain came from the Eurasian avian H1N1(EA H1N1)SIV lineage.PB2,PB1,PA and NP genes were derived from 2009/H1N1 influenza virus lineage.The NS gene was derived from the North American triple reassortment H1N2 SIV lineage.The protein molecular characteristics analysis results showed that the sequence of the basic amino acid cleavage site of HA protein was PSIQSR↓G,which was consistent with the molecular characteristics of low pathogenic influenza virus.The receptor binding sites correspond to the molecular properties of human sialic acid receptors(α-2,6 sialic acid receptors).The NA protein aa275 is H and aa295 is N,and the head region of this protein is not missing,indicating that the strain is sensitive to oseltamivir drugs.PA ^(356)R was a molecular characteristic of increased viral replication and pathogenicity in mammals.NP protein ^(41)I and ^(210)E could enhance the activity of viral polymerase.^(31)N of M2 protein is a molecular characteristic of resistance to amantadines.In this study,the ability of TJ312 strain to replicate and spread by droplets in ferrets was further evaluated.Two ferrets were infected through the nasal cavity
作者
张奕杰
孟飞
杨焕良
宋祖晨
陈艳
陈化兰
ZHANG Yi-jie;MENG Fei;YANG Huan-liang;SONG Zu-chen;CHEN Yan;CHEN Hua-lan(Animal Influenza Key Laboratory of the Ministry of Agriculture and Rural Affairs,State Key Laboratory of Animal disease control and prevention,Harbin Veterinary Research Institute,Chinese Academy of Agricultural Science,Harbin 150069,China)
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2023年第10期987-993,共7页
Chinese Journal of Preventive Veterinary Medicine
基金
十四五国家重点研发计划(2021YFD1800200)。
关键词
猪流感病毒
欧亚类禽H1N1
遗传演化
飞沫传播
swine influenza virus
Eurasian avian-like H1N1
genetic and evolution
droplet transsmission
