A previous study indicated that spleens from reovirus-infected chickens contained macrophages that were primed to produce nitric oxide (NO). The presence of these primed macrophages correlated with depressed in vitro T cell mitogenesis. The current studies indicated that splenic adherent macrophages from virus-exposed chickens inhibited concanavalin A (ConA) induced proliferation of normal spleen cells. ConA-stimulated spleen cells from uninfected chickens, but not virus-exposed chickens, produced large quantities of interleukin-2 (IL-2) and a factor that induced NO production. This factor was tentatively named NO inducing factor (NOIF). The removal of macrophages from the spleens of virus-exposed chickens by plastic adherence resulted in partial recovery of ConA-induced proliferation and the production of normal levels of IL-2 and increased levels of NOIF, although these remained below normal. However, nonadherent spleen cells produced substantial quantities of NO, which indicated an incomplete removal of macrophages. Because removal by plastic adherence did not result in the depletion of all macrophages, spleen cells were panned with anti-CD3 antibody to obtain an almost pure population of T cells. Fractionated T cells from virus-exposed chickens proliferated vigorously to ConA and produced normal levels of IL-2 and NOIF. When splenic adherent cells from virus-exposed chickens were added to purified T cells, the T cells failed to respond to ConA. Addition of splenic adherent cells from virus-free chickens did not induce mitogenic inhibition. Further, the addition of purified T cells from the spleens of reovirus-infected chickens to T cells from virus-free birds did not adversely affect T cell mitogenesis. These data indicated that reovirus infection in chickens does not compromise the functional capabilities of T cells but induces suppressor macrophages that inhibit T cell functions.