Generation of a H9N2 Avian Influenza Virus Whose Ha Proteins Fused C3d-P29 and Its Application in Vaccine Development

Maternal-derived antibodies (MDAs) interfere with immune responses, leading to the failure of H9N2 avian influenza vaccination in poultry. So far, none of commercial H9N2 avian influenza vaccines used in poultry can overcome MDAs interference. To develop a vaccine overcoming MDAs interference, different copies of minimum-binding domain (P29) of complement protein C3d were inserted between the signal peptide and the head domain of hemagglutinin (HA) of a H9N2 avian influenza virus (A/Chicken/Shanghai/H514/2017, named H514). The HA proteins containing P29 stimulated stronger typeⅠinterfere than pure HA proteins in vitro. The modified viruses with HA containing one copy of P29 (rH514-P29.1) and two copies of P29.2 (rH514-P29.2) were successfully rescued by using reverse genetics. The inactivated H9N2 vaccines developed with rH514-P29.1 or rH514-P29.2 induced higher and faster adaptive immune responses than the vaccine developed with rH514 in chickens without MDAs. Importantly, in the chickens with MDAs, the inactivated rH514-P29.2 vaccine induced significantly higher HI titers than the vaccine without P29, and reduced viral shedding after challenged with H514, which suggests that vaccine antigens fused two copies of P29 can decrease the interference of MDAs to immunity in chickens. Overall, our results provide a new strategy to overcome MDAs interference.