Abstract
Bacillus subtilis, a probiotic bacterium with engineering potential, is widely used for the expression of exogenous proteins. In this study, we utilized the integrative plasmid pDG364 to integrate the hemagglutinin-neuraminidase (HN) gene from Newcastle disease virus (NDV) into the genome of the B. subtilis 168 model strain. We successfully constructed a recombinant B. subtilis strain (designated B. subtilis RH) that displays a truncated HN antigen fragment on the surface of its spores and further evaluated its immunogenic effects in mice. Using ELISA, we quantified the levels of IgG in serum and secretory IgA (sIgA) in intestinal contents. The results revealed that the recombinant B. subtilis RH elicited robust specific mucosal and humoral immune responses in mice. Furthermore, B. subtilis RH demonstrated potential mucosal immune adjuvant properties by fostering the development of immune organs and augmenting the number of lymphocytes in the small intestinal villi. Additionally, the strain significantly upregulated the relative expression of inflammatory cytokines such as IL-1β, IL-6, IL-10, TNF-α, and IFN-γ in the small intestinal mucosa. In conclusion, the B. subtilis RH strain developed in this study exhibits promising mucosal immunogenic effects. It holds potential as a candidate for an anti-NDV mucosal subunit vaccine and offers a novel preventive strategy for the poultry industry against this disease.