Immune response induced by ppGpp-defective <Emphasis Type="Italic">Salmonella enterica</Emphasis> serovar Gallinarum in chickens

To protect chickens from typhoid caused by Salmonella enterica serovar Gallinarum (S. Gallinarum), the attenuated 9R strain has been used in the field as a vaccine. However, safety concerns have been raised because the mutations in 9R are undefined while its efficacy is still a question under debate. A global regulator, ppGpp, synthesized by RelA and SpoT, has been shown to induce various virulence genes in S. Gallinarum (Jeong et al., 2008). In this study, two mutant strains defective in ppGpp-synthesis were constructed in wild-type S. Gallinarum (ΔppGpp) and 9R strain (9R-ΔppGpp) backgrounds and tested as live vaccines in chickens. After oral inoculation, the LD₅₀ values of ΔppGpp and 9R-ΔppGpp were approximately 5×10¹⁰ colony forming unit (CFU) similarly as 9R strain, which was ∼10⁵-fold higher than that of the wildtype S. Gallinarum strain. Immunological analyses revealed immunization with either of the two attenuated ppGpp-defective strains induced significant antibody responses, the production of antibody-secreting B cells in blood, proliferation of CD4+ and CD8+ T cells in the spleen, and splenic expression of proinflammatory cytokines, such as IFN-γ and TGF-β4, at levels comparable to the 9R strain. Chickens immunized with the mutants (1×10⁸ CFU) were 80% protected against oral challenge with 1×10⁹ wild-type virulent bacteria (4,000-fold LD₅₀ dose), similar to the level of protection achieved by 9R immunization. Based on these data, live attenuated ΔppGpp-defective strains may serve as novel vaccines to control fowl typhoid in chickens.