不同抗原性甲3(H3N2)亚型流感病毒的鉴定及分子生物学研究

The identification and molecular biology of influenza A(H3N2) strains with distinctively different antigenicity

目的 对不能用当年标准血清进行鉴定的甲 3 (H3N2 )亚型流感病毒进行抗原性分析及分子生物学研究。方法 用交互血凝抑制试验对毒株进行抗原性分析 ,提取病毒RNA ,用一步法逆转录聚合酶链反应 (RT PCR)扩增HA1 (血凝素重链区 )基因片段 ,产物纯化后测序并分析结果。结果发现两株从鸡胚分离到的甲 3 (H3N2 )亚型“D”相毒株与 2株从MDCK细胞分离到的“O”相毒株抗原性已明显不同 ;其中 1 999年分离的“D”相与“O”相 2毒株的抗原比大于 2 56,氨基酸序列同源性为 93 % ,抗原决定簇A区和B区氨基酸位点相差分别为 50 %及 3 5% ;受体结合部位 (RBS)有 6个氨基酸位点发生变异 (左侧壁 1个、前壁 3个、右壁 2个 )。结论 人群中存在没有发生“O”相变异的甲 3 (H3N2 )亚型“D”相毒株 ,其抗原性与当前流行的“O”相变异株已明显不同 ,提示今后在鉴定甲 3 (H3N2 )亚型毒株时 ,需根据毒株的来源和相特性选择适合的标准血清进行鉴定。

Objective Antigenic analysis and molecular biology studies on those strains of influenza A(H3N2) virus which could not be identified by standard serum of the current year. Methods The cross hemagglutination inhibition test was used for antigenic analysis, RNAs were extracted by one step RT PCR to amplify HA1 gene and then the PCR products were sequenced. Results A distinctive antigenic difference was found between two strains of 'D' phase influenza A(H3N2) virus isolated from embryonated chicken eggs and two strains of 'O' phase influenza A(H3N2) virus isolated by MDCK cell culture. The antigenic distribution ratio of 'D' phase and 'O' phase strains isolated in 1999 is above 256 and the homology of amino acid sequence is 93%. The amino acid difference in antigenic determinant A and B were 50% and 35% respectively; 6 amino acids were changed in receptor binding site(RBS)(1 in left side, 3 in front side and 2 in right side). Conclusion 'D' phase strains of influenza A(H3N2) viruses which have not undergone 'O' phase variation still existed in the population and the antigenicity was distinctively different from that of 'O' phase viruses. The results indicated that the origination and phase characteristics were essential points in the selection of standardized antisera for identification of influenza A(H3N2) virus.