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Inability to cultivate Mycobacterium leprae in vitro has been regarded as the bottleneck which obstructs research on modes of transmission, pathogenesis, immunization and the treatment of leprosy . The bottleneck in the cultivation problem always has been that of gaining significant information from an organism that has given no useful response to intensive inquiries by the disciplines of biochemistry, metabolism or biology. The two mycobacterial pathogens have I been regarded as 'obligate intracellular parasites' , because the efforts of several investigators have failed to induce their growth in bacteriologic media . The work on M. leprae has been continuous since shortly after its discovery by Hansen in 1 873 , a period of 1 1 2 years . Mycobacterium lepraemurium, agent of murine leprosy, was noncultivable since its discovery by Stefansky in 1 903 until the convincing report of Nakamura in 1 972, a total period of 69 years . Table 1 offers a comparison of pertinent properties of M. leprae and M. lepraemurium. Both species fail to grow in pulmonary tissues of natural and experimental hosts (occasionally, M. leprae grows slowly in lungs of infected armadillos) . This property is interpreted to mean that the oxygen sensitivity of M. lepraemurium will also be encountered in M. leprae. The table also reveals why M. lepraemurium was used earlier as an interium model for M. leprae. Table 2 describes the significance of adenosine triphosphate (ATP) in such studies. In order to investigate obligate intracellular bacteria one needs a method that measures immediately and directly the physiologic status of unwashed cells . Bioluminescent determinations of A TP qualify for several reasons. A TP measures energy levels that are fundamental to biosynthesis and growth. During growth the increase in A TP per culture should coincide with microscopic counts. ' Figure 1 demonstrates the utility of A TP data while cells are growing and, even more importantly, while their metabolic systems are deteriorating. Curves A and M show that during growth of M. lepraemurium in Nakamura's medium the A TP per culture and microscopic counts are equivalent. Curve B, with 67% air

Limitedin vitromultiplication ofMycobacterium leprae:application to screening potential antileprosy compounds