To define the immune correlates of protection against cytomegalovirus (CMV) end-organ disease, CMV-specific interferon (IFN)- gamma enzyme-linked immunospot (ELISPOT) and CD8(+) and CD4(+) intracellular IFN- gamma synthesis assays were performed for subjects with CD4(+) cell counts of < or =50 cells/ microL who were enrolled in a prospective observational study of CMV infection in the era of highly active antiretroviral therapy. Of 87 subjects, 46 developed viremia, 14 developed end-organ disease, and 20 died. Positive ELISPOT assay results, but not positive results for CD4(+) or CD8(+) intracellular IFN- gamma synthesis, were associated with delayed development of viremia and CMV end-organ disease or death. CMV viremia did not appear to boost CMV-specific immunity. ELISPOT assays may be used to identify HIV-infected patients who might benefit from anti-CMV prophylactic interventions.