OBJECTIVE: Previous studies have demonstrated that mycobacteria can interact with epithelial cells, a property which can be important for establishing infection. In this study we investigated comparatively the ability of Mycobacterium avium, M. tuberculosis and M. smegmatis to invade and multiply within HEp-2 epithelial cells. DESIGN: The ability to invade and to multiply intracellularly in HEp-2 cells was examined using a virulent strain of M. avium, a virulent (H37Rv) and an attenuated (H37Ra) strain of M. tuberculosis and a strain of M. smegmatis. The locus responsible for M. avium invasion was also cloned in Escherichia coli and M. smegmatis. RESULTS: It was observed that M. avium invaded HEp-2 cells with greater efficiency than M. tuberculosis and M. smegmatis, while the H37Rv strain of M. tuberculosis was more efficient in invading HEp-2 than H37Ra and M. smegmatis. Both M. avium and M. tuberculosis were capable of multiplying within HEp-2 cells, while M. smegmatis was not. E. coli K12 and M. smegmatis were transformed with M. avium DNA. The invasive locus of M. avium provided E. coli K12 and M. smegmatis strains S5M101-1 and S5M101-2 with the ability to invade HEp-2 epithelial cells. Transformed M. smegmatis strains were able to grow intracellularly. CONCLUSION: 'Virulent' strains of M. avium and M. tuberculosis were shown to invade and to multiply within HEp-2 epithelial cells. This property was transferred to E. coli K12 and M. smegmatis by transformation with the invasive locus of M. avium. The ability of certain strains of mycobacteria to invade epithelial cells (bronchial, alveolar, intestinal) may represent an important phenotypic characteristic and could be directly related to pathogenicity.