The mechanism(s) of host resistance against Nocardia asteroides has not been well defined. Since disease due to N. asteroides frequently occurs in patients with impaired cell-mediated immunity, we studied the interaction of N. asteroides with activated and control mouse peritoneal macrophages. Activated macrophages were from mice infected with Toxoplasma gondii or injected with Corynebacterium parvum. N. asteroides in the early stationary phase (>99% in the coccobacillary form) was used for challenge of macrophage monolayers. Growth of two strains of N. asteroides was markedly inhibited in activated macrophages, whereas N. asteroides grew well in control macrophages. Quantitation of macrophage-associated N. asteroides indicated that activated macrophages killed 40 to 50% of N. asteroides within 6 h (P < 0.002). In control macrophage preparations, it appeared as if Nocardia filaments extended from within macrophages to the outside, and many of these filaments appeared to have extended to and then grown through neighboring macrophages. In activated macrophage preparations, Nocardia remained in the coccobacillary form in most macrophages. Control macrophage monolayers were almost completely overgrown with and destroyed by Nocardia 20 h after challenge, whereas activated macrophage monolayers remained intact. Nocardia that grew in control macrophages were not acid-alcohol fast or only weakly so, whereas the few Nocardia that grew in activated macrophages were strongly acid-alcohol fast. Our results indicate that activated macrophages may be important in host defense against N. asteroides.