A simple, robust assay system which can be used to screen for inhibitors of mycobacterial growth has been developed. A strain of the rapidly growing saprophyte Mycobacterium aurum is used as the test organism. Inhibition of its growth is highly predictive of activity against Mycobacterium tuberculosis, which cannot itself be used in screening because of its growth characteristics and highly infectious nature. The viability of M. aurum in the presence of a test sample is monitored by measuring the uptake of radiolabelled uracil into the cells. In a microtiter plate format, the screen has the potential for testing several thousand samples per day.Tuberculosis has reemerged in recent years as a serious public health problem (3,11,14). The resurgence has been linked to coinfection with the human immunodeficiency virus (2) and to the emergence of strains resistant to available therapies (10). Multidrug-resistant Mycobacterium tuberculosis is refractory to treatment with any of the commonly available drugs, so new classes of compound are needed to counter the threat posed by these strains. In addition, new drugs are required. These drugs will need to have improved pharmacokinetic properties which reduce the frequency and/or duration of therapy and a broader spectrum of antimycobacterial activity to include emerging pathogens such as Mycobacterium avium.The first proportional methods used to determine mycobacterial susceptibility were developed in the 1950s (4, 13). Conventional antibiotic susceptibility testing protocols were developed from these methods and include testing on both solid and liquid media. Solid media have included Lowenstein-Jensen (4) and Middlebrook 7H11 (12) agars. Several problems are inherent in this type of assay, including drug instability (long incubation period and protein binding of drug affect potency), binding of drug to some medium components, and contamination by extraneous microorganisms, and because of the slow growth rate of M. tuberculosis, it may take up to 3 weeks to obtain a result. A microdilution MIC method with Middlebrook 7H9 broth has been developed (22), but it still takes 2 weeks to produce macroscopic growth.BACTEC is a rapid, radiometric, broth susceptibility system which is now routinely used to measure drug susceptibilities (7)(8)(9)(17)(18)(19) We are seeking to identify novel molecules which possess antimycobacterial activity. The slow growth rate and, more importantly, the highly infectious nature of M. tuberculosis preclude its use in a high-throughput screen. Thus, we have developed a rapid and sensitive broth susceptibility system using a strain of the rapidly growing, nonpathogenic organism Mycobacterium aurum. This assay offers the possibility of screening large numbers of compounds in a system which is highly predictive of activity against M. tuberculosis. MATERIALS AND METHODSMedia and supplements. Middlebrook 7H11 agar and Middlebrook 7H9 broth (5) (Difco, West Molesey, United Kingdom) were prepared as described in the Difco Manual (6). The supplements OADC (...