Genetic evolution and pathogenicity analysis of two natural recombinant isolates of avian influenza

Avian influenza virus (AIV) reassortment plays a crucial role in viral evolution, contributing to genetic diversity and expanding host range. Among low pathogenic avian influenza viruses (LPAIVs), H9 and H6 subtypes are widely prevalent and have significant potential for gene reassortment. In this study, two H6 subtype and seven H9 subtype LPAIVs were isolated and characterized from poultry in Fujian, China. Notably, the two viruses H6N2-C5 and H9N2-C1 shared identical seven gene segments except for the hemagglutinin (HA) gene, with H6N2-C5 carrying the HA gene from an H6N6 virus. Phylogenetic analysis revealed distinct lineages for H6 and H9 viruses, with divergence from commercial vaccine strains. Both H6N2-C5 and H9N2-C1 strains exhibited the ability to replicate efficiently in multiple mammalian and avian cell lines. Animal infection experiments demonstrated that both viruses caused mild to moderate respiratory symptoms in chickens, Muscovy ducks, and mice. H6N2-C5 showed broader tissue tropism and higher viral loads, including neurotropism in mice, while co-infection intensified pathological damage. These findings highlight the active reassortment and cross-species adaptability of H9 and H6 LPAIVs, emphasizing the necessity for continuous surveillance to mitigate their potential threat to animal and public health.