Objective: To understand the origin of HA and NA genes of swine influenza A (H9N2) viruses isolated from pigs in the mainland of China and on basis of these to reveal the pathogenicity of them in pigs.
Methods: The target gene was amplified by PCR, the PCR product was ligated with PGEM-T Easy Vector (Promega company, USA) at 4 degrees, the recombined plasmid was transferred into DH-10-beta bacteria; positive colonies were selected and identified then digested with restriction enzyme. Afterwards,the nucleotide sequence was determined. Finally,phylogenetic analysis of the sequencing data was performed with MegAlign (Version 1.03) and Editseg (Version 3.69) softwares.
Results: Two strains of swine influenza A(H9N2) virus isolated in the mainland had an amino acid residue, leucine (L) at position 226 (H3 numbering) on HA protein molecule found in H9N2 viruses isolated either in pigs or humans previously; the amino acid sequence at HA connecting peptide of isolates possessed R-L-S-R, whereas the other H9N2 viruses with virulence in poultry had R-S-S-R at HA connecting peptide. The two pig H9N2 isolates shared the same three-amino-acids deletion in the NA stalk at 62.64 position found in A/Shaoguan/408/98 and A/Swine/Hong Kong/9/98, as well as A/Duck/Hong Kong /Y280/97(H9N2) viruses. The analysis of the phylogenetic tree indicated that the HA and NA genes of new isolates were closely related to those of A/Chicken/Hong Kong/G23/97 and A/Chicken/Hong Kong/G9/97 and A/Shaoguan/408/98 viruses, respectively.
Conclusion: The HA and NA genes of swine influenza A(H9N2) viruses isolated in the mainland of China probably were derived from those of avian influenza A(H9N2) virus. The occurrence of substitution of amino acid sequence at HA connecting peptide, could result in the H9N2 virus from non pathogenic to pathogenic in pigs. However, avian influenza A(H9N2) virus had deletion in the stalk of the NA that resulted in host range transmission. Therefore they could infect pigs directly.