H9N2 avian influenza virus in Muscovy ducks: a nationwide epidemiological survey and vaccine efficacy assessment.

Avian influenza viruses (AIVs) are recognized as persistent threats to poultry health and global food security. The H9N2 subtype is widely distributed in poultry. A nationwide epidemiological survey was conducted in South, North, Central, and Southwest China, including Muscovy duck farms and live poultry markets, from 2022 to 2024. A total of 1208 Muscovy duck serum samples, 1212 duck-derived oropharyngeal/cloacal swabs and lung/tracheal tissues, and 480 chicken-derived samples were collected. In addition, 31 H9N2 strains were isolated, 2 of which were duck derived and 29 of which were chicken derived. A novel coinfection challenge model combining ocularnasal inoculation of H9N2 with injection of Escherichia coli (E. coli) was established, which induced severe tracheal lesions that closely resembled naturally occurring field pathologies. The coinfection challenge model successfully replicated prevalent field scenarios of viral‒bacterial synergism, resulting in severe lung lesions and a significant increase in viral loads. The results demonstrated that inactivated and live-vector vaccines provided more than 80% protection, with significant reductions in viral shedding and clinical symptoms. This study underscores the urgent need for updated vaccines matching circulating strains and the establishment of a validated framework for evaluating poultry vaccines under real-world coinfection scenarios.