The T-cell-mediated resolution of herpes simplex virus type 2 (HSV-2) genital infections is not fully understood. In these studies, the mechanisms by which CD8+ T cells clear virus from the genital epithelium were examined. Ovalbumin (OVA)-specific CD8+ T cells from OT-I transgenic mice cleared a thymidine kinase-deficient, ovalbumin-expressing HSV-2 virus (HSV-2 tk- OVA) from the genital epithelium of recipient mice, and clearance was abrogated by in vivo neutralization of gamma interferon (IFN-gamma). Further, CD8+ OT-I T cells deficient in IFN-gamma were unable to clear HSV-2 tk- OVA from the vaginal epithelium. The requirement for cytolytic mechanisms in HSV-2 tk- OVA clearance was tested in radiation chimeras by adoptive transfer of wild-type or perforin-deficient OT-I T cells to irradiated Fas-defective or wild-type recipients. Although a dramatic decrease in viral load was observed early after challenge with HSV-2 tk- OVA, full resolution of the infection was not achieved in recipients lacking both perforin- and Fas-mediated cytolytic pathways. These results suggest that IFN-gamma was responsible for an early rapid decrease in HSV-2 virus titer. However, either perforin- or Fas-mediated cytolytic mechanisms were required to achieve complete clearance of HSV-2 from the genital epithelium.