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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。
Objective: To analyze the variation of hemagglutinin (HA) and neuraminidase (NA) of the influenza A (H1N1) virus of Jiangsu Province in 2013 and analyze its genetic evolution characteristics. Methods: According to the definition of influenza-like cases, samples of influenza-like illness were collected from all sentinel hospitals in Jiangsu Province. The positive samples were identified by virus isolation and culture. 14 strains of positive strains of H1N1 (09pdm) that were representative in different regions of China in different time periods in 2013 were selected. HA and NA genes were amplified by specific primers, and the genetic evolution characteristics were sequenced and analyzed. Results: The nucleotide and amino acid homologies of HA gene of 14 isolates and vaccine strains A / California / 07/2009 (H1N1) were 97.6% -98.4% and 96.5% -98.0%, respectively. NA gene nucleotide and amino acid homologies were 98.4% to 98.9% and 97.0% to 98.5%, respectively. Genetic evolution analysis showed that the 14 isolates HA and NA genes belong to different evolutionary lineages. The molecular characteristics showed that amino acid sequence of HA appeared D114N, K300E, E516K variation, NA molecular characteristics showed N44S locus variation. Influenza A (H1N1) (09pdm) influenza gene diversity increased from about 2013 onwards; a positive pressure was selected by FEL model to select HA amino acid site 310, one positive pressure to select NA amino acid site 4, and 9 positive The HA amino acid sites 179,180,239,301,303,310,311,312,313 (including site 310) were selected under pressure and 5 forward pressure selected NA amino acid sites 4,23,52,287,374 (Including site 4). The HA protein has 9 potential glycosylation sites, 7 on HA1 and 2 on HA2; the NA protein has 9 potential glycosylation sites. No differences were found in the activity of NA protease and the surrounding auxiliary sites among the 14 isolates. CONCLUSION: The genetic variation of HA and NA genes of influenza A (H1N1) (09pdm) in Jiangsu Province has been accelerated and the genetic diversity has been increased in 2013. Future genetic evolution deserves further attention.