Mycobacterium ulcerans causes Buruli ulcer, an emerging infectious disease for which antimicrobial therapy has only recently proven to be beneficial. The discovery and development of new drugs against M. ulcerans are severely impeded by its very slow growth. Recombinant bioluminescent strains have proven useful in drug development for other mycobacterial infections, but the ability of such strains to discriminate bacteriostatic from bactericidal activity has not been well demonstrated. We engineered recombinant M. ulcerans strains to express luxAB from Vibrio harveyi. In drug susceptibility tests employing a wide range of antimicrobial agents and concentrations, the relative light unit (RLU) count measured in real time was a reliable surrogate marker for CFU counts available 3 months later, indicating utility for the rapid determination of drug susceptibility and discrimination of bacteriostatic and bactericidal effects. A second important finding of this study is that the addition of subinhibitory concentrations of the ATP-binding cassette transporter inhibitor reserpine increases the susceptibility of M. ulcerans to tetracycline and erythromycin, indicating that drug efflux may explain at least part of the intrinsic resistance of M. ulcerans to these agents.