Fifty mycobacterial strains were isolated from freshly imported tuberculin-negative Macacus rhesus and Cercopithecus ethiops monkeys. Of these strains, 14 were identified as Mycobacterium simiae and 4, as Mycobacterium asiaticum. These two species are slow growing with a delayed photochromogenicity. M. Simiae is niacin-positive. Both species are resistant to the antituberculous chemotherapeutic compounds streptomycin, isoniazid, p-aminosalicylate, and rifampin but are sensitive to cycloserine. The two species are virulent for white mice. Infection is contagious; 25% of noninfected cage mates become infected during 12--60 days of exposure to infected animals. Intrauterine transmission of infection also occurs. Utilizing the gel-precipitation method, we have observed up to 16 antigens in each species. Four to six antigens are shared with Mycobacterium tuberculosis. Mycobacterium species strain 52 is antigenically distinct. The 14 strains of M. simiae belong to two serotypes. Mycobacterium habana belongs to M. simiae serotype 1.