鸡胚中H9N2亚型禽流感病毒的分离鉴定

Isolation and Identification of H9N2 Subtype Avian Influenza Virus in Chicken Embryos

将来自有产蛋下降的肉用型父母代种鸡群的种蛋孵化,每日观察鸡胚死亡情况。在孵化的50个鸡胚中9日龄时开始死亡1个,尿囊液无血凝性,盲传一代后,鸡胚尿囊液有血凝性。在HI试验中,对H9亚型禽流感病毒(AIV)单因子血清在1∶26稀释度时呈现血凝抑制活性,对新城疫病毒(NDV)和H5亚型AIV单因子血清不呈血凝抑制活性,由此确定为低致病性禽流感病毒,命名为ZKH90901。对该毒株进行HA和NA基因扩增克隆测序,把获得的HA和NA基因与已经发表的H9N2毒株的相应序列比较,结果表明该毒株确实为H9N2亚型禽流感病毒,HA基因的裂解位点序列为KSGR↓GLF,符合低致病性禽流感病毒H9N2蛋白裂解位点的分子特征,仍属于低致病力毒株。从鸡胚中分离到H9N2亚型禽流感病毒在国内尚未见报道。

Among 50 eggs from a meat-type parent breeder chicken flock with egg dropping, 1 embryo was dead at 9 d after incubation,which of allantoic fluid had＇t hemagglutinatiou activity,but after one passage in embryo,the allantoic fluid showed hemagglutination activity. By hemagglutination inhibition （HI）test against Newcastle disease virus（NDV）,H5 and H9 subtype avian influenza virus （AIV）,the agent for HA activity in allantoie fluid of dead embryos were identified as H9 subtype AIV and designated as strains ZKH90901. HA and NA genes were amplified by RT-PCR,then were cloned into pMD18-T vector and sequenced. The results showed that ZKH90901 was the H9N2 strain of avian influenza virus,The cleavage site of HA gene was KSGR ↓ GLF,it still belonged to low pathogenicity strain. This is the first report of isolation of H9N2 subtype avian influenza virus from eggs in China.