A Combination of Recombinant HA1-and Nucleoprotein-Based Chitosan Nanoparticles Induces Early and Potent Immune Responses Against the H9N2 Influenza Virus

The effectiveness of inactivated H9N2 influenza vaccines is doubtful due to changes in antigenic regions of the virus hemagglutinin (HA) protein. One strategy for the development of the efficacious vaccine is the use of nanoparticles that display more immunogenic regions of the influenza virus. In this study, chitosan (CS)-based nanoparticles were developed as a delivery system for intranasal immunization using recombinant H9N2 virus HA1 and nucleoprotein (NP), for the induction of humoral and cellular responses. CS-HA1 and CS-NP nanoparticles were prepared by the ionic gelation method and characterized for their physicochemical properties and shape. The immunogenicity and the protective efficacy were evaluated by measuring antibody titers, T cell proliferation response, CD4 + /CD8 + ratio, and quantitative real-time RT-PCR following intranasal administration of the prepared nanoparticles alone or in combination in chickens compared to an inactivated H9N2 vaccine. The average size, surface charge, and spherical structure of the synthesized nanoparticles showed high quality. Serologic analysis revealed that the immunization of inactivated vaccine groups resulted in strong influenza antibodies, which were significantly ( p