Role of the amino acid mutations in the HA gene of H9N2 avian influenza virus under selective pressure in escape vaccine antibodies

It has been well-documented that some amino acid mutations in hemagglutinin (HA) of H9N2 avian influenza virus (H9N2 virus) alter the viral antigenicity, but little is reported about the role of antibody escape mutations in escape vaccine antibodies. In this study, we found that the evolution of F/98 strain in chicken embryos or chickens resulted in significant differences in immune escape, and identify the contribution of HA mutations to the antigenic variation and immune escape of H9N2 virus. Among amino acid mutations in the HA of the antigen variant viruses occurring in embryonated chicken eggs and/or chickens with or without the selection pressure of vaccine antibodies, the mutations, S145N, Q164L, A168T, A198V, M224K and Q234L, affect the antigen drift of H9N2 virus. Specially, the A198V mutation, located at the receptor-binding site on the head domain of HA, significantly contributed the antigenic variation of H9N2 virus. The mutation A198V or Q234L significantly improved the receptor binding activity, while S145N mutation decreased the receptor binding activity. Single S145N mutation could promote viral escape from polyclonal antibodies (pAbs) by preventing Ab binding physically, and single A198V mutation could promote viral escape from pAbs by enhancing the receptor binding activity. Additionally, either the mutation S145N or A198V did interfere with the immunogenicity of the inactivated vaccine, resulting in reduction of the protective efficiency of H9N2 inactivated vaccine, which contributed escape from the antibody-based immunity. Our findings provided an important reference for the accurate evaluation of the role of the amino acids mutation in HA affecting the antigenicity of H9N2 virus on immune escape, and delivered a new perspective for monitoring the adaptive evolution of H9N2 virus. ImportanceIn this study, the role of the HA mutations of H9N2 virus occurring with and without antibody selective pressure on escaping from the antibody-based immune response in host was analyzed. The results demonstrated that (i) the HA mutations S145N, Q164L, A168T, A198V, M224K, and Q234L occurring in the process of the adaptive evolution of H9N2 virus in embryonated chicken eggs and/or chickens could affect the antigenic variation of H9N2 virus. Among these mutations, the HA mutation A198V had the most significant effect on the antigenic variation; (ii) S145N mutation promoted viral escape from pAbs by preventing Abs binding physically; (iii) A198V mutation did promote viral escape from pAbs by enhancing the receptor binding activity; (iv) neither the HA mutation S145N or A198V interfered with the immunogenicity of the inactivated vaccine, resulting in reduction of the protective efficiency of H9N2 inactivated vaccine.