Shiga-toxin producing Escherichia coli O157:H7 (O157)-based vaccines can provide a potential intervention strategy to limit foodborne zoonotic transmission of O157. While the peripheral antibody response to O157 vaccination has been characterized, O157-specific cellular immunity at the rectoanal junction (RAJ), a preferred site for O157 colonization, remains poorly described. Vaccine induced mucosal O157-specific antibodies likely provide some protection, cellular immune responses at the RAJ may also play a role in protection. Distinct lymphoid follicles were increased in the RAJ of vaccinated/challenged animals. Additionally, increased numbers of interferon (IFN)γ-producing cells and γδ + T cells were detected in the follicular region of the RAJ of vaccinated/challenged animals. Likewise, adjuvanted-vaccine formulation is critical in immunogenicity of the O157 parenteral vaccine. Local T cell produced IFNγ may impact epithelial cells, subsequently limiting O157 adherence, which was demonstrated using in vitro attachment assays with bovine epithelial cells. Thus, distinct immune changes induced at the mucosa of vaccinated and challenged animals provide insight of mechanisms associated with limiting O157 fecal shedding. Enhancing mucosal immunity may be critical in the further development of efficacious vaccines for controlling O157 in ruminants and thus limiting O157 transmission to humans.