Murine infection with Salmonella typhimurium provides models for typhoid fever and long-lasting protective immunity conferred by oral vaccination with viable attenuated bacteria. To further understand the role of T cells in these systems, we identified a bacterial Ag recognized by murine T cells responding to a Salmonella infection. From orally infected mice, we derived a CD4+ Ak-restricted T cell clone (7.4.8) the stimulatory Ag of which was provided by S. typhimurium or its flagella, but not by other salmonellae or S. typhimurium mutants unable to synthesize the flagellar filament protein FliC. We mapped antigenic activity to FliC hypervariable region VI using a generally applicable method of sequential C-terminal truncation of recombinant MalE-FliC fusion proteins. Residues 339-350 are the minimal FliC structure capable of stimulating 7.4.8 and represent the first reported Salmonella-specific epitope recognized by T cells from infected mice. T cells with this specificity are generated by oral immunization, reactivity can be recovered for at least 5 mo afterwards, and FliC is the dominant recall Ag for CD4+ T cells from protectively immunized C3H/HeJ mice. FliC 339-350 is presented by macrophages infected with viable S. typhimurium, and presentation, but not bacterial uptake, is greatly enhanced by pretreatment of macrophages with IFN-gamma. These data point to the importance of IFN-gamma-activated macrophages in the stimulation of T cells responding to facultative intracellular pathogens like S. typhimurium and provide a model system for studying Ag-specific T cell responses in murine salmonellosis.