Phylogenetic and Pathogenic Analysis of H5N1 and H5N6 High Pathogenicity Avian Influenza Virus Isolated from Poultry Farms (Layer and Broiler Chickens) in Japan in the 2023/2024 Season

During the 2023-2024 winter, 11 high pathogenicity avian influenza (HPAI) outbreaks caused by clade 2.3.4.4b H5N1 and H5N6 HPAI viruses were confirmed in Japanese domestic poultry among 10 prefectures (<i>n</i> = 10 and 1, respectively). In this study, we aimed to genetically and pathologically characterize these viruses. Phylogenetic analysis revealed that H5N1 viruses were classified into the G2d-0 genotype, whereas the H5N6 virus was a novel genotype in Japan, designated as G2c-12. The G2c-12 virus shared PB2, PB1, PA, HA, and M genes with previous G2c viruses, but had NP and NS genes originating from avian influenza viruses in wild birds abroad. The N6 NA gene was derived from an H5N6 HPAI virus that was different from the viruses responsible for the outbreaks in Japan in 2016-2017 and 2017-2018. Experimental infections in chickens infected with H5N1(G2d-0) and H5N6(G2c-12) HPAI viruses showed no significant differences in the 50% chicken lethal dose, mean death time, or virus shedding from the trachea and cloaca, or in the histopathological findings. Different genotypes of the viruses worldwide, their introduction into the country, and their stable lethality in chickens may have triggered the four consecutive seasons of HPAI outbreaks in Japan.