H9亚型禽流感病毒山西株的分离鉴定及其HA基因的遗传进化分析

Identification of avian influence virus H9 subtype isolated from Shanxi province and genetic variation analysis of HA gene

从山西省某些鸡场采集疑似病料,接种SPF鸡胚后分离病毒,采用A型流感病毒通用引物进行HA基因的扩增,并连接T载体克隆后进行序列测序和遗传进化分析。结果显示：获得4株H9亚型禽流感病毒完整HA序列,开放阅读框1 683nt,编码560个氨基酸,各自之间核苷酸同源性较高,达95.5%~99.6%,且推导的氨基酸同源性介于92.0%~99.6%之间;遗传进化分析结果显示该4株属4.2.5分支,与国内常用疫苗株4.2.3分支存在一定差异,但仍属于4.2大分支;HA蛋白的裂解位点氨基酸顺序为RSSR↓GLF,为低致病性禽流感病毒,并且含有7个潜在的N-糖基化位点以及8个受体结合位点。结果表明：本试验分离的H9亚型禽流感病毒与近年来流行毒株HA基因同源性及蛋白关键位点相似性较高。

Samples were collected from a poultry farm of Shanxi province.Four strains of AIV were isolated by inoculation of SPF chicken embryos and screened by RT-PCR with H9 subtyping primers,then the cDNA of these four HA genes was cloned into T-vector and sequenced,and analysed the genetic variation.The results showed that the full length of HA gene consisted of 1 683 nucleotides encoding 560 amino acid,nucleotide and amino acid sequence homology of four HA genes was 95.5%-99.6% and 92.0%-99.6%,respectively.Genetic analysis revealed that the strains belonged to clade 4.2.5,were similar to prevalent strains,and differed from the common vaccine strains belonged to clade 4.2.3,and the strains had the same RSSR↓GLF motif at the cleavage site of HA,representing low pathogenicity in chicken.In addition,four strains had 7potential N-glycosylation sites and 8receptor binding sites.In conclusion,there was high similarity in HA gene homology and protein key sites between four H9 subtype AIV strains and pandemic strains in recent years.