The frequencies of human immunodeficiency virus type 1 (HIV-1) Gag- and Epstein-Barr virus (EBV)-specific cytotoxic T lymphocyte precursors (CTLp) were studied longitudinally in peripheral blood mononuclear cells from 9 HIV-1-infected persons. By antigen-specific stimulation, HIV-1 Gag-specific CTLp were detected in vitro throughout the course of HIV-1 infection, even after the onset of overt disease. In 4 patients, however, HIV-1 Gag-specific CTLp frequencies declined over time in the presence of maintained EBV-specific CTLp. This decline was correlated with decreasing CD4 (r = .38; P < .05) and CD8 (r = .75; P < .001) cell numbers. The maintenance of EBV-specific CTLp in patients with low CD4 cell numbers indicated that EBV-specific CTL-mediated immunity may remain longer unaffected by HIV-1-induced immune dysfunction. Consistent with this observation, the growth of EBV-specific CTL could be supported in vitro by EBV-infected lymphoblastoid B cell lines, independent of both CD4 cells and exogenous cytokines.