Cell-mediated immunity has been reported to play an important role in defense against Encephalitozoon cuniculi infection. Previous studies from our laboratory have underlined the importance of cytotoxic CD8(+) T lymphocytes (CTL) in survival of mice infected with E. cuniculi. In the present study, immune response against E. cuniculi infection in CD4(+) T-cell-deficient mice was evaluated. Similar to resistant wild-type animals, CD4(-/-) mice were able to resolve E. cuniculi infection even at a very high challenge dose (5 x 10(7) spores/mouse). Tissues from infected CD4(-/-) mice did not exhibit higher parasite loads in comparison to the parental wild-type mice. Conversely, at day 21 postinfection, susceptible CD8(-/-) mice had 10(14) times more parasites in the liver compared to control wild-type mice. Induction of the CD8(+) T-cell response in CD4(-/-) mice against E. cuniculi infection was studied. Interestingly, a normal antigen-specific CD8(+) T-cell response to E. cuniculi infection was observed in CD4(-/-) mice (precursor proliferation frequency, 1/2.5 x 10(4) versus 1/10(4) in wild-type controls). Lack of CD4(+) T cells did not alter the magnitude of the antigen-specific CTL response (precursor CTL frequency; 1/1.4 x 10(4) in CD4(-/-) mice versus 1/3 x 10(4) in control mice). Adoptive transfer of immune CD8(+) T cells from both CD4(-/-) and wild-type animals prevented the mortality in CD8(-/-) mice. E. cuniculi infection thus offers an example of an intracellular parasitic infection where CD8(+) T-cell immunity can be induced in the absence of CD4(+) T cells.