Orally Dissolving Film-Based Influenza Vaccines Confer Superior Protection Compared to the Oral Administration of Inactivated Influenza Virus

<b>Background</b>: Self-administered orally dissolving films (ODFs) encapsulating inactivated influenza vaccines represent an effective strategy for stimulating mucosal immunity. While this vaccination method offers several advantages over conventional influenza vaccines, a comparative efficacy study remains lacking. <b>Methods</b>: Female BALB/c mice were immunized with inactivated A/PR/8/34 (H1N1) either via orogastric inoculation or through the oral mucosal delivery using pullulan and trehalose-based ODF vaccines. Each group received equivalent antigen doses across three immunizations. Humoral responses and antibody functionality were assessed using sera collected post-immunization. After lethal viral challenge, other immunological and virological parameters were determined in corresponding tissues. Body weight and survival were monitored over a 14-day period after challenge. <b>Results</b>: ODF vaccination elicited significantly higher virus-specific IgA levels, HAI titers, and neutralizing antibody activity than oral gavage. After the viral challenge, ODF-immunized mice exhibited stronger IgG and IgA responses in respiratory tissues, increased antibody-secreting cells in lungs and spleen, and elevated germinal center B cells and CD8⁺ T cell responses. Both vaccination methods reduced lung pro-inflammatory cytokines and provided full protection against lethal challenge; however, the ODF group showed lower cytokine levels, better weight maintenance, and reduced viral loads. <b>Conclusions</b>: ODF vaccination elicits more robust systemic and mucosal immune responses than oral vaccination and may serve as a promising alternative method of influenza vaccine delivery.