禽传染性喉气管炎病毒分离鉴定及gD基因序列分析  

Isolation and Identification of Avian Infectious Laryngotracheitis Virus and Sequence Analysis of Its gD Gene

作　　者：何云凤 顾宇华 匡华丽 李蕴玉[1] 李佩国[1] He Yunfeng;Gu Yuhua;Kuang Huali;Li Yunyu;Li Peiguo(Key Laboratory of Preventive Veterinary Medicine,College of Animal Science and Technology,Hebei Normal University of Science&Technology,Qinhuangdao,Hebei 066604,China)

机构地区：[1]河北科技师范学院动物科技学院预防兽医学重点实验室,河北秦皇岛066604

出　　处：《中国动物检疫》2022年第10期23-27,43,共6页China Animal Health Inspection

基　　金：河北省现代农业产业技术体系蛋鸡产业创新团队岗位项目(HBCT2018150206)。

摘　　要：河北省沧州市某养鸡场饲养的510日龄蛋鸡发生急性上呼吸道疫病。为确定病因,采集发病鸡气管组织样品处理后接种鸡胚,利用特异性引物进行禽传染性喉气管炎病毒(ILTV)、禽传染性支气管炎病毒、新城疫病毒、J型禽白血病病毒等有关病原的PCR扩增,结果从接种病毒的鸡胚绒毛尿囊液中只扩增出ILTV gD基因片段(1133 bp),将分离的毒株命名为Hebei2021株;将分离株扩增产物连接载体构建质粒,测序后进行核苷酸序列同源性分析、氨基酸序列比对,并对分离株及参考株gD基因序列进行遗传进化分析,利用在线预测服务器预测gD蛋白二级结构及糖基化位点。结果显示:分离株与国内外参考株及疫苗株gD基因的核苷酸同源性均在99.0%以上,与参考毒株相比仅极个别氨基酸序列位点发生突变;分离株与江苏省分离株之外的国内其他分离株均在同一进化分支上,与疫苗株相比其糖基化位点并未发生突变。结果表明,此次河北省分离株gD基因较为保守,变异程度较低,当前疫苗对ILTV依然有较好的免疫保护力。本研究为禽传染性喉气管炎的流行病学调查和疫苗选择提供了参考。An outbreak of acute respiratory disease occurred in 510 day-old laying hens in a farm in Cangzhou City,Hebei Province.In order to determine the cause,the trachea tissue samples from infected chicken were collected,treated and inoculated into chicken embryos.The infectious laryngotracheitis virus(ILTV),avian infectious bronchitis virus,Newcastle disease virus,avian leukemia virus type J and other related pathogens were amplified by PCR with specific primers,only ILTV gD gene fragment(1133 bp)was amplified from the chorioallantoic membrane of the inoculated chicken embryos,and the isolated virus was named as Hebei 2021 strain;the amplified products of the isolate were connected to the vector to construct a plasmid,and then sequenced for nucleotide sequence homology analysis and amino acid sequence alignment,and the genetic evolution of gD gene sequences of isolate and the reference strains was analyzed,then the secondary structure of gD protein and glycosylation sites were predicted by use of online prediction serve.The results showed that the nucleotide homology of gD gene between the isolate and the reference strains and vaccine strains at home and abroad was over 99.0%,and only a few amino acid sequence sites were mutated compared with the reference strains;the isolate was located in the same evolutionary branch as other domestic isolates except for those isolated in Jiangsu,and its glycosylation sites did not mutate compared with the vaccine strains.In conclusion,the gD gene of Hebei strain was relatively conservative with lower level of variation,and good immune protection against ILTV was detected.A reference was thus provided for future epidemiological investigation and selection of vaccines against infectious laryngotracheitis.

关 键 词：传染性喉气管炎病毒 GD基因 序列分析 

分 类 号：S831.7[农业科学—畜牧学]