Complete Genome Sequence of an H9N2 Avian Influenza Virus Isolated from Egret in Lake Dongting Wetland

GENOME ANNOUNCEMENT

Avian influenza is an acute infectious disease caused by avian influenza virus (AIV). H9N2 AIV have been isolated from swine and poultry (4, 6), and in particular, H9N2 AIV was isolated from humans in 1999 (7). Therefore, strengthening monitoring for H9N2 AIV has significant implications for public health.

Dongting Lake is a very important habitat and wintering site on the migration route of East Asia's migrating birds. Each year nearly two million waterfowls come here for winter and share the waters with local birds, likely resulting in cross-infection between wild birds and domestic fowl. In January 2012, from fresh feces of egret in the Ardeidae protection region of the Dongting Lake wetland, we isolated an AIV H9N2 strain and it was designated A/Egret/Hunan/1/2012(H9N2). The complete genome of this H9N2 strain consists of eight single-stranded RNA segments: PB2, PB1, PA, HA, NP, NA, M, and NS, with nucleotide lengths of 2,341, 2,341, 2,233, 1,742, 1,565, 1,466, 1,027, and 890, respectively.

The amino acid sequence of the HA cleavage site is PAASDR/G, containing only one basic amino acid which is characteristic of low-pathogenic AIV (1). Analysis of potential glycosylation sites of surface proteins revealed 8 potential N-glycosylation sites in HA (29, 82, 141, 218, 298, 305, 492, and 551), which are similar to those of most H9N2 HA proteins, and 7 potential N-glycosylation sites in NA (61, 69, 86, 146, 200, 234, and 402). The stalk of NA does not have a 3-amino-acid deletion at positions 63 to 65. For polymerase proteins, amino acid substitutions are in sites related to host specificity and virus replication capacity. The PB2 segment of this virus possesses Glu 158, Glu 627, and Asp 701; PB1 has Tyr 436, and PA has Thr 515, indicating the low pathogenicity of this virus in mammals and poultry hosts (2, 5, 9). The amino acids of the M2 protein transmembrane domain such as Leu 26, Val 27, Ala 30, Ser 31, Gly 34, His 37, and Trp 41 present no mutations associated with drug resistance, indicating that the virus is sensitive to M2 ion-channel blockers (3, 8).

The nucleotide homology comparisons revealed that the HA gene of this strain shares 97% homology with the HA gene of a North America wild bird AIV strain, A/Northern Shoveler/Interior Alaska/8BM3470/2008(H9N2). The other gene segments share very high levels of nucleotide homology (≥98%) with corresponding segments from different subtypes, such as H5N2, H5N1, H4N8, H7N3, and H3N6 AIV. The above result indicates that the virus isolated in the current study is a new strain generated through genetic reassortment. Phylogenetic analysis revealed that its HA and NA genes are close to those in the Y439-like (DK/HK/Y439/97) branch, but there are some differences.

In summary, this is a first report of H9N2 AIV isolated from wild birds of Dongting Lake wetland, and the virus is generated through genetic reassortment of genes of different AIV subtypes. These data are of significance for studying AIV epidemiology, reassortment, and evolution in wild birds.