南宁市2007～2009年H3N2亚型流感病毒NA基因变异分析

目的 对2007～2009年度南宁市流行性感冒病毒NA序列分析,阐明NA基因变异和进化特征.方法 提取2007 ～ 2009年30株(每年10株)H3N2亚型病毒RNA,采用RT-PCR扩增病毒NA基因后进行序列测定,利用VectorNTI 9.0对测序结果分析处理,并与WHO推荐疫苗株进行比对,用Mage 4.1构建系统进化树.结果 南宁市2007 ～ 2009年H3N2亚型流感病毒被分为3个分枝,其中疫苗株A/Wisconsin/67/2005独立形成一个分支(类群Ⅰ),A/Brisbane/59/2007与2007年毒株(除339外)形成类群Ⅱ,2008和2009年毒株聚集成类群Ⅲ.NA基因在质尾区域、极性跨膜区、柄部区域有个别毒株个别位点发生了替换.抗原区域328～ 336替换频率较高,2008年毒株抗原位点更活跃.A/nanning/242/2007在酶活性位点发生了D151E替换,可能导致NA对扎纳米韦和奥斯他韦的轻微耐药性；A/nanning/242/2007、A/nanning/561/2008分别在非活性位点N234D,在G286D的替换则可能影响NA介导的病毒粒子的释放从而影响NA蛋白的活性.结论 南宁市2007～2009年毒株NA均发生了较大变异,当年疫苗株均滞后南宁流行株.

Genetic variations of NA gene of influenza H3N2 virus in Nanning in 2007-2009

Objective To analyze the sequence of influenza and understand the genetic variation and evolution characteris tics of NA gene of influenza H3N2 viruses circulating in Nanning city in 2007～2009.Methods 30 H3N2 strains（10 each year） isolated from influenza surveillance during years of 2007～2009 were selected.RNAs were extracted,NA genes were am plified by RT-PCR and then sequenced.The data obtained were compared with the vaccine strains recommended by WHO with the software VectorNTI 9.0.Genetic distance and phylogenetic tree of these strains were constructed by using MEGA 4.1.Results The NA phylogenetic tree showed that the strains were divided into three branches,vaccine strain A/Wisconsin/67/ 2005 was located in branchⅠseparately,all strains isolated in 2007（in addition to the strain 339）were located in branch Ⅱ,and strains isolated in 2008-2009 were located in branch Ⅲ.There were individual substitutions in the cytoplasmic tail,trans membrane domain,and stalk region of NA in some strains.Higher variation frequency was showed in antigenic region 328 ~ 336,and antigenic sites were more active in 2008 strains.E 151 amino acids that located in enzyme active site of A/nanning/ 242/2007 NA gene were replaced by D,which might lead to a slight resistance to zanamivir and oseltamivir.The sites N234D in A/nanning/242/2007 and G286D in A/nanning/561/2008,mapped away from the active site of NA,affected the NA-mediat ed viral particle release and NA enzymatic activity.Coclusion Relatively genetic variations occurred to the strains isolated during 2007~2009.The vaccine strains lagged behind the circulating strains in Nanning.