Protection and immune responses induced by attenuated Salmonella typhimurium UK-1 strains

We previously reported that Salmonella typhimurium SR-11 mutants with deletion mutations in the genes encoding adenylate cyclase (cya) and the cAMP receptor protein (crp) are avirulent and protective in mice. Salmonella typhimurium UK-1 is highly virulent for chicks (oral LD₅₀ of 3 x 10³ CFU) and mice (oral LD₅₀ of 8.5 x 10³ CFU) and is capable of lethal infections in pigs, calves and horses. We postulated that attenuated derivatives of this lethal strain would probably induce a higher level of protective immunity than achieved with attenuated derivatives of less virulent S. typhimurium strains such as SR11. To test this hypothesis, we have constructed S. typhimurium UK-1 Δcya-12 Δcrp-11 mutant strain (χ)3985 and its virulence plasmid cured derivative (χ)4095 to investigate their avirulence and immunogenicity in mice. We found that the mutants are avirulent and able to induce protective immune responses in BALB/c mice. These mutant strains retained wild-type ability to colonize the gut associated lymphoid tissue but reach and persist in spleen and liver at a significantly lower level than the wild-type parent strain. Mice survived oral infection with > 1 x 10⁹ CFU of (χ)3985 (the equivalent to 10⁵ 50% lethal doses of wild-type S. typhimurium UK-1) and were fully protected against challenge with 10⁵ times the LD₅₀ of the wild-type parent. Immunized mice developed a high level of serum IgG titre to Salmonella LPS and delayed-type hypersensitivity (DTH) response to S. typhimurium outer membrane proteins. Compared to the virulence plasmid-containing strain (χ)3985, the virulence plasmid cured Δcya Δcrp mutant strain (χ)4095 was more attenuated and less protective, as some mice immunized with (χ)4095 died when challenged with the wild-type UK-1 strain. This work demonstrates that S. typhimurium UK-1 Δcrp Δcya mutant strain may be a potential live vaccine to induce protective immunity against Salmonella infection or to deliver foreign antigens to the immune system.