Combined Salmonella and Pasteurella multocida Antigens Single-Dose and Adsorbed in Aluminum Hydroxide–Chitosan Protect Swiss Mice Against Lethal Challenge

Background/Objectives: Avian cholera and typhoid are major bacterial diseases in the poultry industry, causing reduced productivity and economic losses. Here, we developed a single-dose, bivalent, inactivated vaccine. Methods: Different concentrations of the experimental vaccine containing Pasteurella multocida (P. multocida) and Salmonella Gallinarum (S. Gallinarum) antigens adsorbed to an aqueous matrix based on chitosan, aluminum hydroxide, and saponin were evaluated in mice to determine the optimal single-dose concentration. Results: Vaccine efficacy was assessed by survival rates; the experimental vaccine provided complete protection, whereas 67% of animals immunized with a commercial vaccine survived a challenge with 1 × 1010 CFU/mL (2 × 109 CFU/mice) of P. multocida and 5 × 1010 CFU/mL (1 × 1010 CFU/mice) of S. Gallinarum. Subsequently, experiments were conducted to determine the cytokine profiles of animals immunized with the experimental vaccine (single-dose vs. double-dose regimens) followed by a challenge. Pre-challenge cytokine analysis revealed similar profiles for interleukin (IL)-2, IL-4, IL-10, IL-17, interferon gamma (IFN-γ), and Tumor necrosis factor (TNF-α) across groups; however, IL-6 levels were significantly higher in the single-dose experimental vaccine group (G1) than in the commercial vaccine group (G2). In the post-challenge comparative study, the cytokine response induced by the single-dose vaccine was robust and stable, showing no negative interference following the S. Gallinarum challenge, although the double-dose regimen did boost cellular and regulatory cytokines (IFN-γ, TNF-α, and IL-10) specifically against S. Gallinarum. Conversely, no significant differences in cytokine profiles were observed among the treatments for groups challenged with P. multocida or for unchallenged groups. Conclusions: The single-dose regimen proved sufficient to confer complete protective immunity against both pathogens, suggesting it is a promising strategy for simultaneously preventing avian typhoid and cholera.