新疆油鸡H9N2亚型禽流感病毒分离株HA和NA基因的克隆及进化生物信息分析

Cloning and sequence analysis of HA and NA genes of H9N2 subtype avian influenza viruses isolated from Xinjiang soy sauce chicken

为了从分子水平上掌握新疆油鸡H9亚型禽流感病毒的遗传进化情况,本研究采用RT-PCR技术克隆并测序分离毒株的HA和NA基因,并对其分子特征及全基因序列进行遗传进化分析。结果显示,新疆油鸡分离株的HA和NA基因均属于欧亚分支中的A/CK/BJ/94谱系,HA基因与A/CK/BJ/94在核苷酸和氨基酸水平的同源性依次为90.2%和91.9%,NA基因与A/CK/BJ/94在核苷酸和氨基酸水平的同源性依次为89.3%和90%。分离毒株HA基因的裂解位点为-PSRSSR/G-,符合低致病性AIV的序列特征,HA基因具有人样受体结合的特征(Leu^(234))。HA基因在313位发生突变,比同一谱系的BJ/1/94多1个糖基化位点,NA基因在44位发生突变,比同一谱系的BJ/1/94在该位点多1个糖基化位点,NA基因在3个红细胞吸附区域发生多处突变。试验结果为从分子水平上研究H9N2亚型禽流感病毒跨种传播的机制提供了理论依据。

To evaluate the molecular and evolutional characterization of H9N2 subtype avian influ- enza viruses isolated from Xinjiang soy sauce chicken,the HA and NA genes of the strain were am- plified by RT-PCR. The result indicated that HA and NA genes belonged to the branch of the A/ CK/BJ/94,the nucleotide and amino acids homologies of HA gene between isolated strain and A/ CK/BJ/94 were 90.2% and 91.9% ,the homologies of NA gene were 89.3% and 90%. The se- quence of HA cleavage site was -PSRSSR/G-,which met the characteristic of low-pathogenic avian influenza, HA protein possessed human-like receptor binding specificity （LeuTM ）. The HA gene of isolated strain,compared with BJ/1/94, contains a novel potential glycosylation site at site 313 and the NA gene of isolated strain contains a novel potential glycosylation site at site 44 because of point mutation. There exist several point mutations in the erythrocyte binding sites of NA gene. This study provided a theoretical basis at molecular level for further studying cross-species trans- mission mechanism of H9N2 subtype avian influenza viruses.