It has been hypothesized that optimal vaccine immunity against mucosally invasive, intracellular pathogens may require the induction of different types of immune responses in mucosal and systemic lymphoid tissues. Mucosal type 2/3 responses (producing interleukin-4 [IL-4], IL-6 and/or transforming growth factor beta) could be necessary for optimal induction of protective secretory immunoglobulin A responses. On the other hand, systemic type 1 responses (including gamma interferon [IFN-gamma], tumor necrosis factor alpha, and optimal cytotoxic T-cell responses) are likely to be critical for protection against the disseminated intracellular replication that occurs after mucosal invasion. Despite these predictions, we recently found that vaccines inducing highly polarized type 1 immunity in both mucosal and systemic tissues provided optimal mucosal and systemic protection against the protozoan pathogen Trypanosoma cruzi. To further address this important question in a second model system, we now have studied the capacity of knockout mice to develop protective immune memory. T. cruzi infection followed by nifurtimox treatment rescue was used to immunize CD4, CD8, beta2-microglobulin, inducible nitric oxide synthase (iNOS), IL-12, IFN-gamma, and IL-4 knockout mice. Despite the previously demonstrated importance of CD4(+) T cells, CD8(+) T cells, and nitric oxide for T. cruzi immunity, CD4, CD8, and iNOS knockout mice developed mucosal and systemic protective immunity. However, IL-12, IFN-gamma, and beta2-microglobulin-deficient mice failed to develop mucosal or systemic protection. In contrast, IL-4 knockout mice developed maximal levels of both mucosal and systemic immune protection. These results strongly confirm our earlier conclusion from studies with polarizing vaccination protocols that type 1 immunity provides optimal mucosal and systemic protection against a mucosally invasive, intracellular pathogen.