We genetically modify Salmonella serovars Typhimurium, Paratyphi A and Typhi as vaccines to exhibit regulated delayed attenuation in vivo, regulated delayed in vivo synthesis of protective antigens specified by codon-optimized DNA sequences and regulated delayed lysis in vivo. These vaccines are grown under conditions that enable the orally administered vaccines to display the capabilities of a wild-type strain to survive host defense stresses and more efficiently colonize lymphoid effector tissues before manifesting attenuation to preclude causing disease symptoms and to synthesize protein antigens to induce protective immune responses. We have engineered strains to eliminate or decrease synthesis of serotype-specific LPS O-antigen and flagellar antigens to enable reuse of the vaccine vector and to expose conserved LPS core and over-express immunologically cross-reactive surface outer membrane protein antigens needed for the acquisition of essential iron and manganese ions. This results in induction of cross-protective immunity to enteric bacterial pathogens. We often include other mutations to preclude biofilm formation and diminish induction of gastroenteritis symptoms while retaining abilities to recruit innate immunity and exhibit biological containment. Recent work focuses on further refinement of the regulated delayed lysis phenotype to develop a superior means to induce cellular immunity, delivery of DNA vaccines to prevent viral infections and to target and destroy solid tumors. These vaccine strains exhibit complete biological containment to preclude persistence in vivo and survival if excreted. Strains are totally safe at high doses to newborn, pregnant, protein malnourished and immunocompromised mice. We are using these technologies to develop vaccines to prevent infections of newborns with Streptococcus pneumoniae, Mycobacterium tuberculosis, a diversity of enteric bacterial pathogens causing diarrheal disease and viral influenza. Vaccines are also being developed against pathogens of agriculturally important animals, especially to enhance food safety by preventing infections by Salmonella, E. coli pathovars and Campylobacter jejuni that can be passed through the food chain to humans. Research supported by grants from the Bill & Melinda Gates Foundation, the National Institutes of Health, the United States Department of Agriculture and the Ellison Medical Foundation.