[Evolutionary changes in the NA and HA genes of H3N2 influenza virus in the Moscow Region during 2003-2009].

During the winter 2009 outbreak in the Moscow Region, H3N2 influenza viruses were isolated from the nasopharyngeal washes of patients via their propagation in the human intestinal (Caco-2) and bronchial (Calu-3) epithelial cell cultures maintaining the proteolytic cleavage of HA0--> HA1+HA2 and multicycle virus replication. Analysis of the nucleotide sequences of virus RNA indicated that the 2009 viruses differed from those isolated in 2003 in 14 and 21 amino acids of the neuraminidase (NA) and hemagglutinin (HA) genes, respectively. The NA gene was 1762 nucleotides long whereas the 2003 isolates had a deletion of 66 nucleotides (22 amino acids) in the stalk region (short-stalk NA genotype) of viruses. The NA gene of the 2009 and 2003 isolates possessed an amino acid profile characterized for oseltamivir- and zanamivir-susceptible viral strains. The HA gene of the 2009 viruses contained an N-glycosylation site at Asn181 (an analog to Asn 65 numbering from a signal peptide), which correlated with the long-stalk NA gene. The 2009 viruses had Phe209 in the HA receptor binding center whereas the 2003 isolates possessed Ser209, which correlated with their differences in HA activity. Phylogenetic analysis showed that the NA genes of the 2003 and 2009 Moscow strains were located in the same genetic clade with a single common precursor while their HA genes were diverged in more genetic distance and located in different clades. Viral distribution in the phylogenetic tree indicated that the Moscow strains isolated in 2009 were not direct ancestors of those isolated in 2003; and during the period of 2003 to 2009, H3N2 influenza virus with a short-stalk NA genotype was substituted for a migrant virus possessing a long-stalk NA gene.