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Molecular Evolution of HA1 in Influenza A (H3N2) Viruses Isolated in Japan from 1989 to 2006

Objectives: To evaluate the trend of phylogenetic evolution among influenza A (H3N2) viruses isolated in Gunma and A (H3N2) vaccine strains, we studied the transition of gene mutations and amino acid substitution of the sites A and B in HA1 during long-term seasons. Methods: A total of 15 A (H3N2) strains were obtained from patients in Gunma, Japan, during the 1989–2006 seasons. A partial HA1 gene was amplified by reverse transcription-polymerase chain reaction (RT-PCR). The phylogenetic tree and amino acid sequences were determined with nucleotide sequences. Results: We found that the nucleotide sequences of the isolates in the 1989–1990 and 1997–1998 seasons were similar to those of the A/Beijing/352/89 and the A/Sydney/5/97 strains, respectively, indicating a lag between the circulated virus and the vaccine strains. Phylogenetic analysis made possible to classify into two clusters in genetic evolutions among these strains, which showed large antigenic conversion. Moreover, we confirmed that basic amino acids tended to substitute uncharged polar amino acids, like K135T (lysine-to-threonine substitution at position 135), K145N, H155T, K156Q, R189S and R197Q, as confirmed by multiple alignment of amino acid sequences. Conclusions: We showed a new understanding of genetic evolution and amino acid substitutions of HA1 to recur pandemic.