The ex vivo antiviral CD8⁺ repertoires of 34 human immunodeficiency virus (HIV)-seropositive patients with various CD4⁺ T-cell counts and virus loads were analyzed by gamma interferon enzyme-linked immunospot assay, using peptides derived from HIV type 1 and Epstein-Barr virus (EBV). Most patients recognized many HIV peptides, with markedly high frequencies, in association with all the HLA class I molecules tested. We found no correlation between the intensity of anti-HIV CD8⁺ responses and the CD4⁺ counts or virus load. In contrast, the polyclonality of anti-HIV CD8⁺ responses was positively correlated with the CD4⁺ counts. The anti-EBV responses were significantly less intense than the anti-HIV responses and were positively correlated with the CD4⁺ counts. Longitudinal follow-up of several patients revealed the remarkable stability of the anti-HIV and anti-EBV CD8⁺ responses in two patients with stable CD4⁺ counts, while both antiviral responses decreased in two patients with obvious progression toward disease. Last, highly active antiretroviral therapy induced marked decreases in the number of anti-HIV CD8⁺ T cells, while the anti-EBV responses increased. These findings emphasize the magnitude of the ex vivo HIV-specific CD8⁺ responses at all stages of HIV infection and suggest that the CD8⁺ hyperlymphocytosis commonly observed in HIV infection is driven mainly by virus replication, through intense, continuous activation of HIV-specific CD8⁺ T cells until ultimate progression toward disease. Nevertheless, highly polyclonal anti-HIV CD8⁺responses may be associated with a better clinical status. Our data also suggest that a decrease of anti-EBV CD8⁺ responses may occur with depletion of CD4⁺ T cells, but this could be restored by highly active antiretroviral treatment.