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Insertion mutations were constructed in cloned pmi and rfc genes of Salmonella typhimurium, and these mutations were recombined (singly) into the chromosome of mouse-virulent S. typhimurium C5, displacing the wild-type alleles. Phage sensitivity profiles, lipopolysaccharide analysis, and DNA blotting all confirmed that the replacement events had occurred. The mutations were complemented by plasmid-borne wild-type alleles, as judged by the restoration of wild-type phage plaquing profiles and lipopolysaccharide production (both mutants) and the restoration of pmi-encoded enzyme production (pmi mutant). The virulence, persistence, and immunizing capacities of the mutants fed to mice were compared with those of the wild-type strain and complemented mutants. Both mutants were much reduced in virulence, with the rfc mutant being avirulent even at 10(9) bacteria per mouse. This mutant was also avirulent at up to 10(6) bacteria per mouse when administered intraperitoneally. Both the rfc and pmi mutant strains persisted in the Peyer's patches of the gut after feeding and were capable of colonizing the deeper tissues of the mice from such initial infective foci. Both mutant strains were effective as live oral vaccines (10(7) bacteria or more) against oral S. typhimurium challenge (10(4) 50% lethal doses; 6 x 10(8) bacteria) in mice.

Mutations at rfc or pmi attenuate Salmonella typhimurium virulence for mice