Abstract
H5N6, the highly pathogenic avian influenza A virus (IAV) of clade 2.3.4.4, causes global outbreaks in poultry. H5N6 has become the dominant IAV subtype in waterfowls and causes human infections with high mortality rates. Here, we isolated two strains of H5N6, XGD and JX, from chickens and ducks, respectively. Growth kinetics were evaluated in duck embryo fibroblasts, chicken embryo fibroblasts, Madin-Darby canine kidney cells, and A549 lung carcinoma cells. Receptor binding specificity was analyzed via sialic acid–binding activity assay. The virulence of each strain was tested in BALB/c mice, and recombinant viruses were constructed via reverse genetics to further analyze the pathogenicity. The two strains showed no significant differences in growth kinetics in vitro; however, JX was more virulent in mice than XGD. We also identified 13 mutations in six viral proteins of the two strains through genetic analysis. Our study showed that the NS1 protein played a crucial role in enhancing the virulence of JX. Specifically, the amino acid 139D in NS1 contributed to the high pathogenicity. Therefore, 139D in NS1 might provide insight into the underlying mechanism of IAV adaptation in mammals.
Highlights
Influenza A virus (IAV) causes acute respiratory distress syndrome in many animal species
We investigated the pathogenicity of two H5N6 strains, A/duck/Hubei/WH18/2015 (JX; NCBI: txid1885579) and A/chicken/Hubei/XG18/2015 (XGD; NCBI: txid1885578), in BALB/c mice to elucidate the underlying mechanism of IAV adaptation in mammals
The two strains were inoculated in Duck embryo fibroblasts (DEF), chicken embryo fibroblasts (CEF), A549, and Madin-Darby canine kidney (MDCK) cells monolayers cultured in 12-well plates, with a multiplicity of infection (MOI) of 0.01
Summary
Influenza A virus (IAV) causes acute respiratory distress syndrome in many animal species. A novel H5N6 IAV has emerged via antigenic drift and reassortment events with other IAVs. A novel H5N6 IAV has emerged via antigenic drift and reassortment events with other IAVs This new highly pathogenic avian influenza (HPAI) A virus (subclade 2.3.4.4) has caused worldwide outbreaks in poultry [1]. H5N6 was first isolated from mallards in 1975 [2]. The novel recombinant H5N6 caused an outbreak in Laos in 2013 and was isolated from waterfowls in southern China [3]. H5N6 has replaced H5N1 as the dominant IAV subtypes in waterfowls that causes human infections in China [4]. The continuous evolution of the virus represents a long-term threat to public health and the poultry industry
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