We recently showed that class I-restricted CD8+ T cells mediate clearance of primary rotavirus infection in mice: JHD knockout (JHD -/-) (B-cell-deficient) mice depleted of CD8+ T cells become chronically infected with murine rotavirus, and beta2 microglobulin knockout (beta2m -/-) mice have delayed but complete clearance of primary rotavirus infection. In the present work we have analyzed the mechanism used by CD8+ T cells to clear rotavirus infection. We first determined that perforin knockout (perforin -/-) mice and lpr (fas-deficient) mice clear rotavirus infection with the same kinetics as control mice. When perforin -/- or perforin +/+ mice were depleted of CD8+ T cells by administration of an anti-CD8 monoclonal antibody, they showed a delay of 1 to 2 days in the clearance of rotavirus infection compared to the clearance time for untreated control mice, indicating that CD8+ T cells in both groups of mice participate in the resolution of primary rotavirus infection. In addition, passively transferred CD8+ T cells from rotavirus-infected perforin +/+ and perforin -/- mice were able to mediate viral clearance in Rag 2 knockout (Rag 2 -/-) mice chronically infected with rotavirus with similar kinetics, suggesting that CD8+ T cells from perforin -/- mice are as efficient as CD8+ T cells from perforin +/+ mice in clearing a rotavirus infection. Gamma interferon (IFN-gamma) was also shown to be unnecessary for the antirotavirus effect of CD8+ T cells: IFN-gamma knockout (IFN-gamma -/-) mice and JHD -/-, perforin -/-, and perforin +/+ mice depleted of IFN-gamma by administration of an anti-IFN-gamma monoclonal antibody cleared rotavirus infection with the same kinetics as those for control mice. Hence, CD8+ T cells have an antirotaviral effect that is not mediated by perforin and appears to be independent of fas and the release of IFN-gamma.