Retamal Miguel, Abed Yacine, Rhéaume Chantal, Baz Mariana, Boivin Guy
Research Center in Infectious Diseases of the CHU of Québec and Laval University, Québec City, Quebec, Canada.
J Gen Virol. 2017 Jun;98(6):1224-1231. doi: 10.1099/jgv.0.000800. Epub 2017 Jun 20.
Influenza A(H1N1)pdm09 virus continues to circulate worldwide without evidence of significant antigenic drift between 2009 and 2016. By using escape mutants, we previously identified six haemagglutinin (HA) changes (T80R, G143E, G158E, N159D, K166E and A198E) that were located within antigenic sites. Combinations of these mutations were introduced into the A(H1N1)pdm09 HA plasmid by mutagenesis. Reassortant 6 : 2 viruses containing both the HA and NA genes of the A(H1N1)pdm09 and the six internal gene segments of A/PR/8/34 were rescued by reverse genetics. In vitro, HA inhibition and microneutralization assays showed that the HA hexa-mutant reassortant virus (RG1) escaped A(H1N1)pdm09 hyper-immune ferret antiserum recognition. C57Black/6 mice that received the vaccine formulated with A/California/07/09 were challenged with 2×104 p.f.u. of either the 6 : 2 wild-type (WT) or RG1 viruses. Reductions in body weight loss, mortality rate and lung viral titre were observed in immunized animals challenged with the 6 : 2 WT virus compared to non-immunized mice. However, immunization did not protect mice challenged with RG1 virus. To further characterize the mutations causing this antigenic change, 11 additional RG viruses whose HA gene contained single or combinations of mutations were evaluated in vitro. Although the RG1 virus was still the least reactive against hyper-immune serum by HAI testing, mutations G158E and N159D within the Sa antigenic site appeared to play the major role in the altered antigenicity of the A(H1N1)pdm09 virus. These results show that the Sa antigenic site contains the most prominent epitopes susceptible to cause an antigenic drift, escaping actual vaccine protection.
甲型H1N1pdm09流感病毒继续在全球传播，在2009年至2016年期间没有明显抗原漂移的证据。通过使用逃逸突变体，我们之前鉴定出位于抗原位点内的六个血凝素（HA）变化（T80R、G143E、G158E、N159D、K166E和A198E）。通过诱变将这些突变的组合引入A（H1N1）pdm09 HA质粒。通过反向遗传学拯救了包含A（H1N1）pdm09的HA和NA基因以及A/PR/8/34的六个内部基因片段的重配6:2病毒。在体外，HA抑制和微量中和试验表明，HA六突变体重配病毒（RG1）逃避了A（H1N1）pdm09超免疫雪貂抗血清的识别。接受用A/加利福尼亚/07/09配制疫苗的C57Black/6小鼠用2×104个菌斑形成单位（p.f.u.）的6:2野生型（WT）或RG1病毒进行攻毒。与未免疫的小鼠相比，在用6:2 WT病毒攻毒的免疫动物中观察到体重减轻、死亡率和肺病毒滴度的降低。然而，免疫并未保护用RG1病毒攻毒的小鼠。为了进一步表征导致这种抗原变化的突变，在体外评估了另外11种HA基因包含单个或组合突变的RG病毒。尽管通过HAI检测RG1病毒对超免疫血清的反应性仍然最低，但Sa抗原位点内的G158E和N159D突变似乎在A（H1N1）pdm09病毒抗原性改变中起主要作用。这些结果表明，Sa抗原位点包含最易导致抗原漂移、逃避实际疫苗保护的突出表位。
