A multimeric S1-nucleocapsid fusion antigen elicits cellular and humoral immune responses against porcine epidemic diarrhea virus in mice.
Porcine epidemic diarrhea virus (PEDV) causes severe enteric disease in piglets, with mortality approaching 100% in naïve herds and extensive global economic losses. Protection relies on maternal vaccination to confer lactogenic immunity; however, current vaccines provide incomplete protection. To improve the immunogenicity of spike-based subunit vaccines, we developed a recombinant S1-nucleocapsid (N) fusion antigen, combining the S1 domain, which contains multiple neutralizing epitopes, with highly conserved N protein within a single immunogen. The antigen formed heterogeneous multimeric assemblies when expressed in mammalian cells and was formulated with Montanide™ Gel 02 PR or ISA 61 VG adjuvants. Immunogenicity was evaluated in BALB/c mice using a prime-boost intramuscular vaccination regimen by assessing antigen-binding and neutralizing antibodies, as well as IFN-γ- and IL-5-secreting cells. Vaccination induced robust IgG, IgG1, IgG2a, and IgA responses against both S1 and N, with higher neutralizing antibody titers in the Gel 02 PR group than the ISA 61 VG group. Adjuvant formulation influenced cytokine response patterns. Both Gel 02 PR and ISA 61 formulations showed higher IFN-γ and IL-5 responses than the control group with Gel 02 inducing highest IFN-γ levels amongst the groups. These pre-clinical findings demonstrate that the S1-N-T4f fusion antigen is a promising vaccine candidate for further evaluation of its protective potential against PEDV infection.