Numerous studies have shown the potential of Salmonella typhimurium as a vector for delivery of heterologous proteins for vaccination against other pathogens. Earlier studies showed that the inefficient elicitation of MHC class I-restricted responses could limit the use of S. typhimurium as a heterologous antigen delivery vector for vaccination. We recently developed an approach to overcome this limitation by using a bacterial-encoded specialized protein secretion system, termed type III, to deliver proteins into the class I antigen presenting pathways. Thus, peptides of interest fused to proteins bearing the type III secretion signal, which can elicit protective CTL responses. Because protective immunity is usually assessed a few weeks after vaccination, there is a paucity of information regarding duration of protective immunity induced by this system. We show here that mice immunized orally with S. typhimurium vectors expressing a MHC class I-restricted epitope of the lymphocytic choriomeningitis virus (LCMV) nucleoprotein developed specific antiviral CTL responses. CD8+ T cells were found to be necessary for this CTL activity against targets presenting the LCMV epitope. The survival of mice challenged with lethal doses of LCMV 60 or 135 days after vaccination was as complete as the survival of mice challenged 2 weeks after immunization with the same vectors. By demonstrating their ability to induce prolonged protective immunity after oral delivery, S. typhimurium vectors have met an essential requirement in support of their development as vectors for heterologous vaccination.