Modification of neutralizing epitopes of hemagglutinin for the development of broadly protective H9N2 vaccine

•Modification of major neutralizing epitopes of Hemagglutinin (HA) for broad protection against distinct H9N2 viruses.•We displayed modified HA on the baculovirus envelope (BacHAm).•Evaluated Bac-HAm vaccine constructs efficacy against H9N2 viruses in a mouse model.•Cross-protective efficacy of BacHAm vaccine against distinct influenza H9 viruses. The H9N2 avian influenza viruses cause significant economic losses in poultry worldwide and could potentially cause human pandemic. Currently, the available vaccines have limited efficacy due to antigenic drift of H9N2. To improve vaccine efficacy, we developed monovalent vaccine strain via the modification of neutralizing epitopes on hemagglutinin (HA) to broaden the protection against H9N2 viruses. In this study, single and multiple mutation were introduced to amino acid at position 148, 150 (site I) and 183, 186, 188 (site II) on the full-length HA gene of H9N2 strain (A/Hong Kong/33982/2009). These mutant HA constructs were displayed on the baculovirus surface (BacH9), and evaluated for their cross-protective efficacy against H9N2 viruses in a mouse model. Our findings indicate that mice immunized with multiple BacH9 mutant constructs (148–150 183 and 186) induced cross-protective immunity against circulating H9N2 in the viral challenge study and prove to be a promising vaccine candidate for H9N2.