Genetic evolution, phylodynamics, geographic spread of H9N2 avian influenza viruses in China from 2014 to 2025: an increasing potential zoonotic risk

Subtype H9N2 of the avian influenza virus (AIV) poses a growing threat to the poultry industry and public health. However, since 2014, there has been no systematic study of its genetic evolution, genotype, spatial dynamics, and pathogenicity in China. Therefore, we performed a large-scale sequence analysis of the genome of Chinese H9N2 viruses from 2014 to 2025 using public databases and laboratory isolates. We identified 52 different genotypes in 1,591 H9N2 viruses, including 4 previously recognized genotypes (G6, G57, G58, and G68) and 48 newly defined genotypes (G118–G163) in this study. G57 and G118 were the main epidemic genotypes in China from 2014 to 2025. Bayesian phylogeographic analysis showed that there were 12 obvious migration paths for the spread of H9N2 AIVs in China from 2017 to 2022. In particular, the South China region was the main transmission centre. H9N2 AIV continues to circulate in chickens and ducks in China and spreads to other hosts. The H9N2 AIVs with the G57 and G118 genotypes could effectively replicate in MDCK, CEF, A549, and HBE cells with titres of 0.97–8.5 lgTCID50/mL. The G57 and G118 genotypes H9N2 viruses preferentially bound to α–2,6–linked sialic acid glycopolymers (human receptors), and effectively replicate in multiple organs of mice and chickens and cause pathological changes in the lungs. Thus, it is necessary to strengthen the monitoring and prevention of H9N2 AIVs in China.