血清4型禽腺病毒W株的分离鉴定和全基因组序列分析

doi:10.16431/j.cnki.1671-7236.2019.07.022

摘要/Abstract

摘要：

为了解血清4型禽腺病毒（fowl adenovirus genotype 4，FAdV-4） W株全基因序列、结构特征及遗传变异情况，本试验采集发病鸡肝脏和肾脏进行PCR鉴定、SPF鸡胚和鸡胚肝细胞分离、电镜观察证明该病毒为FAdV-4，TCID₅₀为10^-7.2/0.1 mL；电镜观察可见70 nm左右的二十面体无囊膜病毒颗粒；经绒毛尿囊膜途径接种10日龄SPF鸡胚后能明显抑制鸡胚发育出现侏儒胚，接种10 d内死亡率达100%；进一步对W株进行全基因组扩增及序列分析。病毒序列分析结果显示，其全基因组长为43 591 bp，共有60个开放阅读框，W株与GenBank中公布的FAdV-4参考毒株全基因组核苷酸序列同源性为98.4%~100%，其中与标准株非致病性毒株ON1（登录号：GU188428.1）的同源性为98.4%，主要缺失ORF19（脂肪酶基因）和ORF27；通过对病毒主要结构蛋白Hexon、Fiber-1和Fiber-2基因分析发现，W株与近几年国内分离株亲缘关系较近，与ON1株及其他国外分离株差异较大。综上说明，该分离株具有高致病性，与国外分离株存在较大差异，为进一步分析FAdV-4的毒力增强机制提供了基础依据。

关键词: 禽腺病毒; 分离; 鉴定; Fiber基因

Abstract:

In order to understand the complete gene sequence,structural characteristics and genetic variation of fowl adenovirus genotype 4 (FAdV-4) W strain,the liver and kidney of infected chicken were identified by PCR.The isolation of SPF chicken embryo and electron microscope observation showed that the virus was FAdV-4 and TCID₅₀ was 10^-7.2/0.1 mL.The virus was about 70 nm in diameter and without envelope under electron microscope.Chorioallantoic membrane inoculation of 10-day-old SPF chicken embryos could significantly inhibit the development of chicken embryos and produce dwarf embryos,and the death rate was 100% within 10 days after inoculation with chorioallantoic membrane.The complete genome of W strain was amplified and sequenced.The virus was 43 591 bp,which had 60 open reading frames.The nucleotide sequence homology was 98.4% to 100% between W strain and referenced FAdV-4 strains published in GenBank.The homology was 98.4% with standard non-pathogenic virus strain ON1 (accession No.:GU188428.1).The ORF19 (lipase gene) and ORF27 were not found.It was found that W strain was closely related to the domestic isolates by analyzing the Hexon,Fiber-1 and Fiber-2 genes of the main structural proteins in recent years.It had a great difference with the ON1 strain and other foreign strains.The results showed that the strain had high pathogenicity and was different from foreign isolates,which provided the basis data for further analysis of the virulence enhancement mechanism of FAdV-4.

Key words: fowl adenovirus; isolation; identification; Fiber gene

中图分类号:

S852.65⁺9.1

赵玉杰, 刘建勋, 陈田田, 王赛楠, 何春辉, 陈盼盼, 刘琳, 李新生. 血清4型禽腺病毒W株的分离鉴定和全基因组序列分析[J]. 《中国畜牧兽医》, 2019, 46(7): 2059-2068.

ZHAO Yujie, LIU Jianxun, CHEN Tiantian, WANG Sainan, HE Chunhui, CHEN Panpan, LIU Lin, LI Xinsheng. Isolation, Identification and Complete Genome Sequence Analysis of Fowl Adenovirus Genotype 4 W Strain[J]. , 2019, 46(7): 2059-2068.

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