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A mycobacterial growth factor was present in the conditioned media of Escherichia coli and Enterobacter cloacae cultures, but not in Pseudomonas aeruginosa culture. This factor potentiated the growth of Mycobacterium avium-M. intracellulare complex (MAC), a patient isolate, and well-established strains of M. avium and M. intracellulare. The growth factor was not a polypeptide; it was heat-stable and possessed a molecular weight < 500 D. Acetate production by enterobacteria was responsible for the biological activities observed. Acetate promoted mycobacterial growth at concentrations up to 3 mM; higher levels were toxic. The effects of acetate on MAC growth were not influenced by the pH of the media. Our data suggest that production of acetate by enterobacteria may regulate mycobacterial growth, and therefore, intestinal acetate might be a cofactor in the pathogenicity of MAC.