The majority of the response of cytotoxic T-lymphocytes (CTL) to lymphocytic choriomeningitis virus (LCMV) in H-2^dmice is directed toward one epitope located on the nucleoprotein (NP, aa 118–126), and usually no primary responses to other epitopes are detectable. Previous studies have shown that thymic expression of lymphocytic choriomeningitis virus-nucleoprotein (LCMV-NP) in H-2^dtransgenic mice (Thy-NP mice) leads to deletion of high-affinity anti-LCMV-NP CTL by negative selection. Selection is incomplete, so that low-affinity NP-specific CTL pass through the thymus and are detectable in the periphery. To analyze the importance of interferon-γ (IFN-γ) in the ability of low-affinity antiviral CTL to clear an acute viral infection, double transgenic mice were generated that are IFN-γ deficient and express the NP of LCMV in the thymus (Thy-NP × IFN-γ −/− mice). When infected with LCMV, these bigenic mice were unable to clear the infection despite generating low-affinity primary antiviral CTL, and they became persistently infected. In contrast, IFN-γ competent Thy-NP mice cleared LCMV within 7–8 days and IFN-γ deficient mice that did not express NP in their thymus generated high-affinity CTL that terminated an acute LCMV infection within 10–12 days post-viral challenge. Persistently infected IFN-γ deficient mice selectively depleted LCMV-specific CTL and displayed reduced levels of antigen-presenting cells in the spleen, and 60% of these mice died at 2–3 months postinfection. Thus, IFN-γ is required for clearing an acute viral infection in the absence of a high-affinity CTL response. In the absence of IFN-γ persistent viral infection results despite the presence of low-affinity CTL.