2010-2016年唐山市H3N2亚型流感病毒血凝素基因特征分析

Genetic characterization of the hemagglutinin gene of influenza A(H3N2) viruses in Tangshan during influenza seasons from 2010-2016

目的了解2010-2016年唐山市分离的H3N2亚型流感病毒株血凝素HA基因特征和变异规律。方法随机选取2010-2016年唐山市分离的20株H3N2亚流感毒株提取RNA,采用RT-PCR扩增目的片段,测序,采用MEGA 6.0软件构建HA基因核苷酸系统进化树,采用BioEdit v7.2.5软件比较分析其核苷酸序列特征及氨基酸变异情况。结果 0株病毒HA1区核苷酸和氨基酸同源性分别为92.4%和89.1%。系统进化和序列分析显示,2010-2011流行年度分离的5株病毒分别属于HA基因1和5分支,其中1分支毒株在抗原决定簇发生E50K,I260M,R261Q氨基酸替换,5分支毒株发生D53N、Y94H、I230V和E280A替换;2011-2016流行年度分离的15株H3N2亚型病毒属于3C分支,与疫苗株A/Perth/16/2009比较抗原表位发生S45N、T48I、A198S、N278K和N312S位点突变,并逐年进化为3C.1(2010-2011)、3C-2012/13(2012-2013)、3C.3(2013-2014)、3C.3a(2014-2015)和3C.2a(2015-2016)分支。与相应疫苗株相比,3C.3a和3C.2a分支毒株抗原表位A、B及受体结合部位均发生了氨基酸位点突变,其中3C.2a毒株拥有Q311H(C区)和N171K(D区)位点突变。结论 2010-2016年唐山市分离株A/H3N2亚型流感病毒发生抗原漂移,未来应关注3C.2a分支流行株的变化。

Objective To analyze the genetic characteristics and variability of the hemagglutinin （HA） gene of influ- enza H3N2 viruses prevalent in Tangshan from 2010 to 2016. Methods A total of 20 influenza H3N2 strains that were isolated in Tangshan from October 2010 to March 2016 were randomly chosen, and viral RNA was extracted and amplified using RT-PCR. Full length HA sequences were sequenced. A phylogenetic tree of the nucleotide sequences encoding HA was generated by Molecular Evolutionary Genetic Analysis （MEGA） version 6.0, and characteristics of nucleic acid sc quences and amino acid variations were analyzed using the software gioEdit v7.2. 5. Results HA1 of 20 A（H3N2） strains had a nucleotide sequence similarity of 92.4% and an amino acid （aa） sequence similarity of 89.1 %. Phylogenetic analysis and sequencing indicated that A（H3N2） viruses isolated during influenza seasons from 2010 2011 belonged to HA genetic clades 1 and 5. These consisted of clade 1 viruses with aa substitutions E50K, I260M, and R261Q and clade 5 viruses with aa substitutions D53N, Y94H, I230V, and E280A. During influenza seasons from 2011 2016, all 15 A （H3N2） strains belonged to genetic clade 3C and shared aa substitutions at S45N, T48I, A198S, N278K, and N312S in comparison to the vaccine strain A/Perth/16/2009. Strains with those substitutions respectively developed into the suh- clades 3C. 1（2010-2011）, 3（2-2012/13 （2012-2013）, 3C. 3（2013-2014）, 3C. 3a（2014 2015）, and 3C. 2a（2015- 2016）. Compared to the vaccine strains A/Texas/50/2012（2014 2015） and A/Switzerland/ 9715293/2013 （2015- 2016）, strains in two clades （3C. 3a and 3C. 2a） had amino acid mutations at antigenic sites A and B and a receptor binding site. Strains in the elade 3C. 2a also had mutations at epitopes C （Q311H） and D （N171K）. Conclusion These findings confirmed the antigenic drift of influenza A（H3N2） prevalent in Tangshan in recent years. Genetic variations in clade 3C. 2a viruses warrant attention in the future.