Contribution of H7 haemagglutinin to amantadine resistance and infectivity of influenza virus
Contribution of H7 haemagglutinin to amantadine resistance and infectivity of influenza virus
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DOI：
10.1099/vir.0.82256-0
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发表时间：
2007-04-01
影响因子：
3.8
通讯作者：
Webster, Robert G.
中科院分区：
文献类型：
--
作者：
Ilyushina, Natalia A.;Govorkova, Elena A.;Webster, Robert G.
在本研究中，我们确定了金刚烷胺对流感A/荷兰/219/03（H7 N7）病毒在细胞培养和小鼠模型中的抗病毒作用。浓度< 100 μ M的金刚烷胺不能抑制Madin-Darby犬肾（MDCK）细胞中的病毒复制。当小鼠连续5天口服金刚烷胺15 mg/kg/day（-1）时，不能保护动物免受H7 N7感染的致死性攻击，小鼠器官中的病毒滴度也没有降低。然而，M2蛋白的序列分析显示，没有一个突变先前描述为与金刚烷胺耐药。我们使用反向遗传学产生含有A/Netherlands/219/03病毒的血凝素（HA）或M基因的病毒，以研究这些基因在金刚烷胺敏感性中的作用。所有携带A/Netherlands/219/03（H7 N7）病毒HA片段的重组病毒均对金刚烷胺具有耐药性，无论其其他基因的来源如何。为了研究融合活性在耐药机制中的作用，我们在H7融合肽中引入Gly（23）-> Cys突变。与野生型病毒相比，这种取代导致融合物的pH降低，并且还与MDCK细胞和小鼠中的病毒复制减少有关。我们认为H7 HA蛋白在金刚烷胺耐药中起作用，尽管所有参与耐药的HA氨基酸仍有待鉴定。我们的发现表明，M2蛋白跨膜结构域的序列分析可能无法充分识别所有耐药变异体。
In the present study we determined the antiviral effect of amantadine against influenza A/Netherlands/219/03 (H7N7) virus in cell culture and in a mouse model. Amantadine at concentrations < 100 mu M failed to inhibit virus replication in Madin-Darby canine kidney (MDCK) cells. When orally administered to mice for 5 days, amantadine at 15 mg kg(-1) day(-1) did not protect animals against lethal challenge with H7N7 infection, and virus titres in mouse organs were not reduced. However, sequence analysis of the M2 protein revealed none of the mutations previously described as being associated with amantadine resistance. We used reverse genetics to generate viruses containing the haemagglutinin (HA) or M gene of A/Netherlands/219/03 virus to investigate the role of these genes in amantadine sensitivity. All recombinant viruses carrying the HA segment of A/Netherlands/219/03 (H7N7) virus were amantadine-resistant, regardless of the origin of their other genes. To study the role of fusion activity in the mechanism of drug resistance, we introduced the Gly(23)-> Cys mutation in the H7 fusion peptide. This substitution resulted in a decrease of the pH of fusion and was also associated with reduced virus replication in both MDCK cells and mice, as compared to that of the wild-type virus. We suggest that H7 HA protein plays a role in amantadine resistance, although all HA amino acids that participate in drug resistance still remain to be characterized. Our finding reveals that sequence analysis of the transmembrane domain of M2 protein may not adequately identify all drug-resistant variants.