[Construction of recombinant Lactococcus lactis strains expressing classical swine fever virus E2 antigen and evaluation of their immunogenicity in murine models].

Classical swine fever virus (CSFV) infects pigs primarily via the respiratory and digestive tracts and inflicts enormous economic losses on the global swine industry. To address the challenge of early CSFV invasion, this study developed an oral vaccine based on a Lactococcus lactis vector to induce sIgA-mediated mucosal immunity. This approach aims to provide robust early protection and a scalable, stress-free vaccination strategy for modern pig farming. In this study, we first assessed the colonization capacity of L. lactis NZ9000 in mice by in vivo luciferase imaging and fecal bacterial counting. We then employed homologous recombination to construct two recombinant L. lactis strains: NZ9000/pNZ8148-pgsA-E2 for surface display of the CSFV E2 protein and NZ9000/pNZ8148-E2 for intracellular expression of E2. The expression of E2 protein was confirmed by Western blot and indirect immunofluorescence assay. Immunogenicity of the recombinant strains was evaluated in mice. The results showed that L. lactis NZ9000 achieved short-term colonization in mice. NZ9000/pNZ8148-pgsA-E2 and NZ9000/pNZ8148-E2 achieved surface display and intracellular expression of E2, respectively. Both recombinant strains induced specific immune responses following gavage. Notably, NZ9000/pNZ8148-pgsA-E2 showed higher capacity in promoting Th1-type cytokine secretion and enhancing CD4+ T cell activation than NZ9000/pNZ8148-E2. This study successfully constructs recombinant L. lactis strains expressing the CSFV E2 protein and supports the further development of oral mucosal vaccines against classical swine fever.