To assess the involvement of bacterial microflora in the development of host defenses, we compared in vitro LPS-induced cytokine production by macrophages in germ-free and E. coli monoxenic mice. E. coli implantation significantly increased IL-1 and IL-6 and, to a lesser extent, TNF activities of peritoneal and bone marrow-derived macrophages. These results suggest that exposure to microflora primes macrophages for an enhanced cytokine production, which may contribute to the activation of the antiinfectious defense. The priming was not restricted to peritoneal macrophages but was associated with a more general effect of the flora since the enhanced response of bone marrow-derived macrophages indicates an effect on macrophage precursors. Furthermore, a higher ability of peritoneal macrophages to produce IL-1 in axenic and monoxenic mice was observed as compared to bone marrow-derived macrophages. In contrast, bone marrow-derived macrophages demonstrated a higher ability to produce IL-6 and TNF but only 3 weeks after bacterial administration.