Primary viral infections, including primary HIV infection, trigger intense activation of the immune system, with marked expansion of CD38+ CD8+ T cells. Whether this expansion involves only viral-specific cells or includes a degree of bystander activation remains a matter of debate. We therefore examined the activation status of EBV-, CMV-, and influenza virus (FLU)-specific CD8+ T cells during primary HIV infection, in comparison to HIV-specific CD8+ T cells. The activation markers CD38 and HLA-DR were strongly expressed on HIV-specific CD8+ T cells. Surprisingly, CD38 expression was also up-regulated on CD8+ T cells specific for other viruses, albeit to a lesser extent. Activation marker expression returned to normal or near-normal values after 1 year of highly active antiretroviral therapy. HIV viral load correlated with CD38 expression on HIV-specific CD8+ T cells but also on EBV-, CMV-, and FLU-specific CD8+ T cells. In primary HIV infection, EBV-specific CD8+ T cells also showed increased Ki67 expression and decreased Bcl-2 expression, compared with values observed in HIV-seronegative control subjects. These results show that bystander activation occurs during primary HIV infection, even though HIV-specific CD8+ T cells express the highest level of activation. The role of this bystander activation in lymphocyte homeostasis and HIV pathogenesis remains to be determined.