Plasmid pTETnir15, which directs the expression of the non-toxic immunogenic fragment C of tetanus toxin from the anaerobically inducible nirB promoter, was introduced into the Salmonella typhimurium aroA aroD live oral vaccine strain BRD509. The resulting strain, designated BRD847, was used to vaccinate orally BALB/c mice and was tested for plasmid stability and its ability to protect against a lethal tetanus toxin challenge. pTETrar15 was stably inherited by bacteria growing or persisting in the tissues of immunized mice whereas another BRD509 derivative, designated BRD753, harboring plasmid pTET85 which directs fragment C expression from the tac promoter, was highly unstable. Mice immunized with a single oral dose of BRD847 developed high levels of circulating anti-fragment C antibodies and were solidly protected against tetanus toxin challenge. Mice immunized with a single oral dose of BRD753 developed no detectable anti-fragment C antibodies. After boosting, antibodies were detected, but the mice were only partially protected against tetanus toxin challenge. Thus the use of an in vivo inducible promoter such as nirB may be a generally applicable approach to obtaining the stable in vivo expression of heterologous antigens in Salmonella vaccine strains.