2017-2019年呼和浩特市甲型A(H1N1)pdm09流感病毒基因特征分析

Genetic characterization of the influenza A(H1N1)pdm09 virus in Hohhot,2017-2019

目的了解呼和浩特市甲型A(H1N1)pdm09流感病毒遗传进化特征及抗原位点、糖基化位点和耐药位点的变异情况,为流感防控提供科学依据。方法采集2017-2019年呼和浩特市3家哨点医院的流感样病例(ILI)咽拭子标本进行核酸检测,阳性样本通过MDCK细胞和鸡胚进行病毒分离。随机抽取15株甲型A(H1N1)pdm09流感毒株进行基因测序分析。采用MEGA7.0.14软件、DNASTAR7.0.1软件和NetNGlyc1.0软件进行基因进化树分析、核苷酸同源性分析和糖基化位点分析。结果呼和浩特市2017-2019年15株甲型A(H1N1)pdm09分离株与疫苗株A/Brisbane/02/2018和A/Michigan/45/2015共同属于6B.1分支。各年度分离株与疫苗株A/Brisbane/02/2018的组间遗传距离分别为0.004、0.011和0.013。相较于疫苗株A/California/07/2009和A/Michigan/45/2015,2017年起甲型A(H1N1)pdm09病毒抗原位点变异较大,但与疫苗株A/Brisbane/02/2018相比并未发生抗原漂移现象,与疫苗株匹配效果较好。分离株与疫苗株A/California/07/2009相比多了糖基化位点162NQS/T。分离株与神经氨酸酶抑制剂耐药相关的9个关键氨基酸位点没有发生变异。结论随着时间推移,呼和浩特市甲型A(H1N1)pdm09流感病毒与疫苗株的基因相似性逐渐降低,提示呼和浩特市病毒在不断进化中,应持续监测。本次研究中的分离株与WHO推荐的2019-2020年北半球疫苗株A/Brisbane/02/2018相比未发生抗原漂移现象,WHO推荐的疫苗株对目前流行的甲型A(H1N1)pdm09流感仍有较好的保护效果。所有分离株NA基因均未发生H275Y耐药突变,提示呼和浩特市流行的甲型A(H1N1)pdm09流感仍然对奥司他韦等NA抑制剂药物敏感。

This study analyzed the genetic evolutionary characteristics of the influenza A(H1N1)pdm09 virus in Hohhot,including variations in antigenic loci,glycosylation loci and drug resistance loci,to provide a scientific basis for influenza prevention and control.Pharyngeal swab specimens were collected from influenza-like cases(ILI)at three sentinel hospitals in Hohhot from 2017 to 2019,and subjected to nucleic acid testing.Positive samples were isolated with MDCK cells and chicken embryos.Fifteen strains of influenza A(H1N1)pdm09 were randomly selected for gene sequencing analysis.MEGA7.0.14,DNASTAR7.0.1 and NetNGlyc1.0 software were used for gene evolutionary tree analysis,nucleotide homology analysis and glycosylation site analysis.The Hohhot 2017-201915 A(H1N1)pdm09 isolates belonged to branch 6B.1,together with vaccine strains A/Brisbane/02/2018 and A/Michigan/45/2015.The intergroup genetic distances between the isolates and vaccine strain A/Brisbane/02/2018 were 0.004,0.011 and 0.013 for each year,respectively.Compared with those of vaccine strains A/California/07/2009 and A/Michigan/45/2015,the antigenic locus of A(H1N1)pdm09 virus was more variable from 2017,but no antigenic drift occurred with respect to vaccine strain A/Brisbane/02/2018,which matched better with the vaccine strain.The isolate had an additional glycosylation site 162NQS/T with respect to vaccine strain A/California/07/2009.The isolate had an additional glycosylation site 162NQS/T with respect to vaccine strain A/California/07/2009.The isolate showed no variations in nine key amino acid sites associated with neuraminidase inhibitor resistance.The genetic similarity between Hohhot A(H1N1)pdm09 influenza virus and the vaccine strain gradually decreased over time,thus suggesting that the Hohhot virus is evolving and should be continually monitored.The isolates in this study did not show antigenic drift with respect to the WHO-recommended 2019-2020 Northern Hemisphere vaccine strain A/Brisbane/02/2018.The WHO-recommended vaccine strain still provides better protection against the currently prevalent influenza A(H1N1)pdm09.No isolates had H275Y drug resistance mutations in the NA gene,thus suggesting that the influenza A(H1N1)pdm09 prevalent in Hohhot is still sensitive to NA inhibitor drugs such as oseltamivir.