Heterosubtypic protection against avian influenza virus by live attenuated and chimeric norovirus P-particle-M2e vaccines in chickens

Avian influenza in poultry continues to be a great concern worldwide, and the currently licensed inactivated influenza vaccines are not effective against the novel strains of influenza virus that continue to emerge in the field. This warrants the development of more broadly protective influenza vaccines or vaccination regimens. Live attenuated influenza vaccines (LAIVs) and subunit vaccines derived from viral peptides, such as the highly conserved ectodomain of influenza virus matrix protein 2 (M2e), can offer a more broadly reactive immune response. In chickens, we previously showed that a chimeric norovirus P particle containing M2e (M2eP) could provide partial but broad immunity, when administered as a standalone vaccine, and also enhanced the protective efficacy of inactivated vaccine when used in a combination regimen. We also demonstrated that a naturally-selected NS1-truncated H7N3 LAIV (pc4-LAIV) was highly efficacious against antigenically distant heterologous H7N2 low pathogenicity avian influenza virus challenge, especially when used as the priming vaccine in a prime-boost vaccination regimen. In this study, we investigated the cross-subtype protective efficacy of pc4-LAIV in conjunction with M2eP using single vaccination, combined treatment, and prime-boost approaches. Chickens vaccinated with pc4-LAIV showed significant reduction of tracheal shedding of a low pathogenicity H5N2 challenge virus. This cross-subtype protective efficacy was further enhanced, during the initial stages of challenge virus replication, in chickens that received a vaccination regimen consisting of priming with pc4-LAIV at 1?day of age and boosting with M2eP. Further, H5N2-specific serum IgG and pc4-LAIV-specific hemagglutination-inhibition antibody titers were enhanced in LAIV-primed and M2eP boost-vaccinated chickens. Taken together, our data point to the need of further investigation into the benefits of combined and prime-boost vaccination schemes utilizing LAIV and epitope-based vaccines, to develop more broadly protective vaccination regimens.