江苏省2013年甲型H1N1(09pdm)流感病毒血凝素和神经氨酸酶分子特征分析

目的:分析江苏省2013年甲型H1N1(09pdm)流感病毒血凝素(hemagglutinin,HA)和神经氨酸酶(neuraminidase,NA)变异情况,分析其遗传进化特征?方法:按照流感样病例定义,收集江苏省各哨点医院流感样病例标本,阳性样本经病毒分离培养及亚型鉴定?选取2013年不同时间段?不同地区具有代表性的14株甲型H1N1(09pdm)阳性毒株,利用特异性引物扩增HA?NA基因,测序并分析其遗传进化特征?结果:14株分离毒株与疫苗株A/California/07/2009(H1N1)的HA基因核苷酸和氨基酸同源性分别为97.6%~98.4%和96.5%~98.0%?NA基因核苷酸和氨基酸同源性分别为98.4%~98.9%和97.0%~98.5%?遗传进化分析表明,14株分离株HA和NA基因分属于不同的进化谱系?分子特征表现为HA氨基酸序列出现D114N?K300E?E516K的变异,NA分子特征表现为N44S位点变异?从2013年左右开始甲型H1N1(09pdm)流感基因多样性增加;通过FEL模型得到一个正向压力选择HA氨基酸位点310,一个正向压力选择NA氨基酸位点4,通过REL模型得到9个正向压力选择HA氨基酸位点179?180?239?301?303?310?311?312?313(含位点310),5个正向压力选择NA氨基酸位点4?23?52?287?374(含位点4)?HA蛋白具有9个潜在糖基化位点,7个位于HA1上,2个位于HA2上;NA蛋白共有9个潜在糖基化位点?14株分离毒株在NA蛋白酶活性中心及周围辅助位点上均未发现变异?结论:2013年江苏省甲型H1N1(09pdm)流感HA?NA基因变异加快,遗传多样性增加,未来的遗传进化值得进一步关注?

Molecular characteristics of hemagglutinin and neuraminidase genes of influenza A/H1N1(09pdm) viruses, Jiangsu province, 2013

Objective:To analyze the genetic characterization of hemagglutinin and neuraminidase genes of influenza A/H1N1 (09pdm) viruses in Jiangsu province, 2013. Methods: Specimens from influenza-like cases in Sentinel hospitals from Jiangsu province were isolated by cell cultivation and performed by subtype identification. Fourteen representative positive strains of A/H1N1 (09pdm) viruses were selected within different regions in 2013. HA and NA genes of 14 isolates were sequenced and the genetic characterization was analyzed. Results: Compared with vaccine strain A/California/07/2009(H1N1), the HA and NA genes of 14 isolates shared the highest nucleic acid sequence similarity (97.6%~98.4% and 98.4%~98.9%, respectively), and amino acid sequence similarity (96.5%~98.0%, 97.0%~98.5%, respectively). The phylogenetic analysis showed that the HA and NA genes of all 14 strains were divided into two clusters. The amino acid substitutions of the HA proteins (D114N,K300E and E516K) and NA proteins (N44S) of 14 isolates were observed. The evolution dynamics revealed that the genetic diversity of influenza A/H1N1(09pdm) viruses increased. The positive pressure sites were observed in HA proteins site 179, 180, 239, 301, 303, 310, 311, 312, 313 (including site 310) and NA proteins 4, 23, 52, 287, 374 (including site 4) by FEL and REL model. Of 14 isolates, the HA protein contained 9 potential glycosylation sites(7 in HA1 and 2 in HA2), whereas NA protein contained 9 potential glycosylation sites. Conclusion: In 2013, the influenza A/H1N1 (09pdm) viruses had undergone molecular evolution to generate genetic diversity and this emphasize the importance of reinforcing virus surveillance.