Genomic events contributing to the high prevalence of amantadine-resistant influenza A/H3N2.

Background: The prevalence of amantadine-resistant influenza A/H3N2 viruses (belonging to the N-lineage), possessing an S31N mutation in the M2 protein and S193F and D225N substitutions in their HA1 subunit, has significantly increased worldwide since 2005. The aim of this study was to clarify the genomic events contributing to the evolution and continuity of the N-lineage amantadine-resistant viruses. Methods: The full genome sequence of A/H3N2 isolates, including both amantadine-resistant and amantadine-sensitive viruses, collected in Japan between 2006 and 2008, was determined and phylogenetically compared with isolates obtained from the database. Results: On the basis of the full genome sequence analysis, the N-lineage could be further divided into three genetically related clades: Ni (A/Wisconsin/67/2005-like amantadine-resistant viruses from years 2005-2007), N2 (amantadine-sensitive viruses from 2007) and N3 (A/Brisbane/10/2007-like amantadine-resistant viruses from 2007 and 2008). The 2006/2007 season showed cocirculation of antigenic variants of amantadine-resistant viruses of clades Ni and N3 in addition to the N2-sensitive viruses. In the 2007/2008 season, the clade N3 amantadine-resistant lineage dominated and replaced other strains. Phylogenetic analysis of each individual segment suggested that N2 and N3 were generated from two independent reassortment events involving clade Ni viruses and pre-N-lineage strains. Conclusions: Our data show that several reassortment events have contributed to the evolution of amantadine-resistant A/H3N2 strains and, consequently, to the successful spread of this lineage. Although amantadine resistance is caused by single amino acid mutations in the M2 protein, genome-wide adjustment involving multiple genes appears to be necessary to obtain efficient replication and transmission of resistant viruses. Such adjustments are attainable through reassortment of segments among different virus lineages.