The effect of treating cultured mouse fibroblasts (L929 cells) with cloned mouse interferon-gamma on the growth of Rickettsia prowazekii within the fibroblasts was studied. Within 48 h after infection, rickettsiae were cleared from a substantial proportion of the initially infected cells and rickettsial growth was inhibited in those cells that remained infected, when L929 cells were treated with cloned mouse interferon-gamma both before and after infection. When L929 cells were treated with cloned mouse interferon-gamma either only before or only after infection with rickettsiae, rickettsial growth was markedly inhibited but rickettsiae were not cleared from many cells. Addition of cycloheximide to L929 cells markedly suppressed the antirickettsial activity of the interferon, and cloned mouse interferon-gamma did not induce antirickettsial activity in human foreskin fibroblasts. The antirickettsial effects of cloned mouse interferon-gamma were similar to those induced by crude mouse lymphokines prepared from concanavalin A-stimulated mouse spleen cells. Equivalent amounts (units) of cloned mouse interferon-gamma produced by Chinese hamster ovary cells or by Escherichia coli caused equivalent inhibition of rickettsial growth in mouse fibroblasts. However, at high concentrations of interferon-gamma, treatment of rickettsia-infected fibroblasts with equivalent amounts (units) of interferon-gamma, as crude mouse lymphokines or cloned mouse interferon-gamma, resulted in slightly greater inhibition of rickettsial growth by the crude lymphokines. Most of the antirickettsial activity of crude mouse lymphokines can be explained by the interferon-gamma that is present in these preparations. Interferon-gamma, by virtue of its ability to inhibit rickettsial growth and effect the clearance of rickettsia from nonprofessional phagocytes, may play a crucial role in the elimination of rickettsiae from the infected host.