Typhoid fever is a persistent infection caused by host adapted Salmonella species. Paradoxically, allowing pathogen persistence may also be beneficial for the host when immune-mediated damage outweigh the immediate risk posed by the infection. Our long-term goals are to identify the immune cells and signals activated during Salmonella infection that controls this balance between immune activation required for pathogen eradication with the associated risk of immune-mediated host injury, and immune suppression at the expense of pathogen persistence. The mouse model of S. typhimurium infection reproduces important features of persistent infection characteristic of human typhoid. Together with the abundance of immunological tools and reagents available only in mice, infection in this species is currently the best-available model for identifying the molecular and cellular signals that control this delicate immunological balance. Using this model, our preliminary studies indicate regulatory T cells (Tregs) that comprise a distinct lineage of CD4+ T cells characterized by Foxp3-expression play critical roles in dictating the balance between immune activation and bacterial persistence. Early after infection, Foxp3+Treg-ablation accelerates the kinetics of bacterial eradication, while augmenting Foxp3+Tregs causes reciprocal increases in pathogen burden. Therefore the overall goals of this application are to define the relative importance and identify the suppressive mechanism utilized by Tregs throughout persistent Salmonella infection. Aim 1 will expand on our preliminary findings that demonstrate drastic changes in Treg suppressive potency for cells isolated at early (day 5) and late (day 37) time points, by defining a comprehensive kinetic analysis for changes in Treg suppressive potency during persistent infection. Our specific goals are to identify when Treg suppressive potency peaks, nadirs, and returns to baseline levels, and the absolute magnitude for these changes. These results will establish the frame-work for complementary experiments that verify the importance of Tregs at these critical time points by quantifying the relative impacts of Treg-ablation. Using recently developed techniques for reconstituting Treg- ablated mice with donor Tregs from mice with targeted defects in defined Treg-associated molecules, Aim 2 will further explore the mechanistic basis for these changes in Treg suppression potency. These include experiments that will identify and dissociate the specific Treg-associated molecules that control host defense from those required for sustaining peripheral immune tolerance, and define Salmonella ligands and the corresponding pattern recognition receptors that dictate these changes in Treg suppression potency through cell-intrinsic stimulation. Together these experiments that utilize cutting edge immunological tools will not only fill important gaps in our current knowledge on the importance and the mechanism whereby Foxp3+Tregs control host defense during Salmonella infection, but also establish important paradigms for how Tregs may regulate immunity during other persistent infections. PUBLIC HEALTH RELEVANCE: The pathogenesis of persistent infection is controlled by the balance between opposing immune activation and suppression signals. Using Salmonella infection in mice as a model of human typhoid, this application seeks to define the importance of regulatory T cells, and identify the molecular basis whereby these cells control this delicate immunological balance during persistent bacterial infection.