Salmonella species causes disease or death in millions of individuals every year who ingest contaminated food or water. Individuals who survive Salmonella infection develop robust immunity that is dependent on antibodies that facilitate bacterial phagocytosis by macrophages, and lymphokines from CD4+ T cells that stimulate macrophages to kill engulfed bacteria. For practical reasons, a subunit vaccine capable of inducing the level of immunity that can be induced by live organisms, is desirable. Unfortunately, immunization with individual Salmonella proteins usually induces only partial immunity. The long-term goal of this project is to overcome this inefficiency and produce an effective subunit vaccine for Salmonella. Our hypothesis is that individual Salmonella proteins are less immunogenic than attenuated organisms because of a lack of inherent adjuvant activity, poor entry into the mucosal lymphoid organs, and poor uptake by the dendritic antigen-presenting cells in these organs. Our specific aim is to produce a single vaccine protein that addresses each of these weaknesses. We propose to identify an immunodominant peptide for CD4+ T cells from a protective antigen; use a peptide:MHC ll-specific monoclonal antibody to identify the dendritic cells that initially generate peptide:MHC II complexes from Salmonella organisms, construct a single chain Fv specific for this dendritic cell. The peptide and single chain Fv will be combined with the Toll-like receptor 5-binding portion of S. typhimurium FliC, which functions an adjuvant, and the beta1 integrin-binding domain of Yersinia pseudotuberculosis, which facilitates M cell translocation of particles into the Peyer s patches. This multi-component vaccine will be tested for its ability to generate protective immunity to Salmonella, and novel methods to track peptide:MHC II complexes and specific CD4+ T cells will be employed to assess the cellular mechanisms responsible for immunity. Completion of these aims will provide a framework for the rational design of effective subunit vaccines for Salmonella, other infectious agents, and cancer.