Molecular epidemiologic studies on North American H9 avian influenza virus isolates from waterfowl and shorebirds

Because sequence data on H9 avian influenza virus (AIV) from wild birds are currently limited, we set out to determine the sequence of the hemagglutinin (HA) gene of H9 viruses circulating in North American waterfowl and shorebirds. In this study, we examined the HA gene from H9 AIV isolated from mallards (Anas platyrhynchos) sampled during 1998 and 1999 in Minnesota and ruddy turnstones (Arenaria interpres) sampled during 2003 in Delaware and New Jersey. At these sites, the H9N2 subtype represented 12% and 4% of the avian influenza isolates from mallards in 1998 and 1999, respectively, and 8% of the AIVs isolated from shorebirds between 2000 and 2002. Sequences from these viruses were compared with sequences from H9 AIV isolated from commercial poultry and aquatic birds from North America, Europe, Asia, and the Middle East: four previously reported and three new clades were observed. Sequence data from the HA gene of North American waterfowl and shorebird isolates generated in this study most closely group with the Eurasian H9 viruses in the Y439 clade. In addition, the HA cleavage site (AASNR/G) and receptor binding site was identical to the representative virus of that group (DK/Hong Kong/Y439/97). Viruses in that clade are commonly found in ducks and chickens in Hong Kong and Korea. Positive evolutionary selection (dNonsynonymous > dSynonymous) was observed for the HA gene among the North American waterfowl and shorebird H9N2 viruses, indicating that the H9N2-type viruses are changing in their natural hosts.