Studies On The Molecular Biological Characteristics And Phylogenetics Of H6N1 Subtype AIV Strains Isolated From Mallard

In our study, five H6N1 subtype of influenza A viruses were collected in 2007 in Heilongjiang Sanjiang Nature Reserve. A genome was carried out, including four partial gene sequencing, an important amino acid sites, and genetic homology comparison evolutionary was researched.Viruses collected in the field were subcultured in SPF chicken embryos for three generations. And then total viral RNA were extracted from the allantoic fluid.Reference to a published whole influenza virus subtype H6N1 gene sequences, eight full-length gene specific primers were designed, The total viral RNA was amplified by RT-PCR gene sequence. Gene amplification was cloned into PMD18-T vector, and sequenced. Biological software such as DNAStar, MEGA, DNASIS, etc were used to join the nucleotide sequence together. After ucleotide splicing,homology comparison, finding a special amino acid, and the mapping of the genetic relationship between the phylogenetic tree, the analysis of the obtained results showed that all of the gene sequences contain complete gene sequence of ORF; Non-continuous were found in A/mallard/Sanjiang/380/2007 (H6N1) HA cleavage site amino acid sequence of multiple basic amino acids, with a low pathogenic influenza typical characteristics of the virus. There were eight glycosylation sites in HA amino acid sequence. Compared with other referred strains of subtype H6N1, amino acid substitution did not exist; five amino acid sequence of the influenza virus NA in the eight glycosylation sites are conservative, while replacement phenomenon exists in another three and common replacement exists in 4 of the 17 potential antigenic sites; the eight gene segments of A/mallard/Sanjiang/380/2007 (H6N1) were probably from Eurasian formation 7 other than HA, a high homology and close evolutionary relationship existed between the Eurasian strain and the virus;this virus may be recombined by H6N2, H5N1 and H6N1 subtypes of influenza virus;There were close genetic relationship between NA genes of H6N1 and H1N1 subtypes isolated from duck,so the phenomenon of gene flow may exist in the two NA genes.Come to a conclusion that AIVs of wildbirds have widespread rearrangement between different subtypes,and have genetic exchange with AIVs isolated from avian.