During acute cytomegaloviral (CMV) mononucleosis, mononuclear leukocytes displayed diminished responsiveness to certain mitogens. Responses to phytohemagglutinin were normal in patients with community-acquired CMV illness but were reduced in patients with transfusion-associated infection. Both groups were hyporesponsive to pokeweek mitogen and concanavalin A. Cells from convalescent CMV mononucleosis patients responded as well as cells from normal donors to the three mitogens. Preculture of cells from patients with acute CMV mononucleosis for up to seven days before addition of concanavalin A greatly enhanced the blastogenic response to that mitogen. The effect was increased further by depletion of adherent cells after preculture for seven days and could be markedly reduced by the addition of fresh, autologous adherent cells to the precultured nonadherent cells. Suppressor activity was not observed in the serum of patients with acute CMV mononucleosis. These studies suggest that mitogen hyporesponsiveness in CMV mononucleosis may be mediated by suppressor cells included among the plastic-adherent mononuclear leukocyte population.