Emerged HA and NA Mutants of the Pandemic Influenza H1N1 Viruses with Increasing Epidemiological Significance in Taipei and Kaohsiung, Taiwan, 2009–10

The 2009 influenza pandemic provided an opportunity to observe dynamic changes of the hemagglutinin (HA) and neuraminidase (NA) of pH1N1 strains that spread in two metropolitan areas -Taipei and Kaohsiung. We observed cumulative increases of amino acid substitutions of both HA and NA that were higher in the post–peak than in the pre-peak period of the epidemic. About 14.94% and 3.44% of 174 isolates had one and two amino acids changes, respective, in the four antigenic sites. One unique adaptive mutation of HA2 (E374K) was first detected three weeks before the epidemic peak. This mutation evolved through the epidemic, and finally emerged as the major circulated strain, with significantly higher frequency in the post-peak period than in the pre-peak (64.65% vs 9.28%, p<0.0001). E374K persisted until ten months post-nationwide vaccination without further antigenic changes (e.g. prior to the highest selective pressure). In public health measures, the epidemic peaked at seven weeks after oseltamivir treatment was initiated. The emerging E374K mutants spread before the first peak of school class suspension, extended their survival in high-density population areas before vaccination, dominated in the second wave of class suspension, and were fixed as herd immunity developed. The tempo-spatial spreading of E374K mutants was more concentrated during the post–peak (p = 0.000004) in seven districts with higher spatial clusters (p<0.001). This is the first study examining viral changes during the naïve phase of a pandemic of influenza through integrated virological/serological/clinical surveillance, tempo-spatial analysis, and intervention policies. The vaccination increased the percentage of E374K mutants (22.86% vs 72.34%, p<0.001) and significantly elevated the frequency of mutations in Sa antigenic site (2.36% vs 23.40%, p<0.001). Future pre-vaccination public health efforts should monitor amino acids of HA and NA of pandemic influenza viruses isolated at exponential and peak phases in areas with high cluster cases.

2009年流感大流行为观察在台北与高雄这两个都会区传播的pH1N1毒株的血凝素（hemagglutinin, HA）与神经氨酸酶（neuraminidase, NA）的动态变化提供了研究契机。本研究观察到，HA与NA的氨基酸替换累积增幅在疫情流行峰后期显著高于峰前期。在174株病毒分离株中，约14.94%与3.44%的毒株分别在4个抗原位点上出现1个和2个氨基酸突变。血凝素2亚基（HA2）的一处独特适应性突变E374K最早于疫情峰前三周被检出。该突变伴随疫情进程不断演化，最终成为主流流行毒株，其在峰后期的检出频率显著高于峰前期（64.65% vs 9.28%，p<0.0001）。E374K突变株在全国范围内启动疫苗接种后的十个月内持续存在，未发生进一步抗原变异（例如在选择压力达到峰值之前）。在公共卫生干预措施层面，疫情于奥司他韦治疗启动后的第七周达到流行高峰。E374K突变株的传播早于首次班级停课高峰，在疫苗接种前于高密度人口区域持续扩散，在第二次班级停课浪潮中占据主导地位，并随着群体免疫的建立成为固定优势毒株。E374K突变株的时空传播在峰后期更为集中（p=0.000004），在7个存在显著空间聚集性的行政区中分布更为密集（p<0.001）。本研究首次通过整合病毒学、血清学与临床监测手段，结合时空分析与干预政策评估，针对流感大流行的免疫 naive（naïve）阶段的病毒变异情况展开了探究。疫苗接种提升了E374K突变株的占比（22.86% vs 72.34%，p<0.001），并显著提高了Sa抗原位点的突变频率（2.36% vs 23.40%，p<0.001）。未来的疫苗接种前公共卫生防控工作中，应在聚集性病例高发区域，针对分离自病毒指数增长期与流行峰期的大流行性流感病毒，监测其HA与NA的氨基酸序列变异情况。