Abstract: One L. hardjo Malaysian isolate and five Leptospira sp. reference strain namely L. pomona, L .icterohaemorrhagae, L. canicola, L. australis and L. hardjo Prajitno were selected in this study. The Leptospira strains were tested for it susceptibility against five types of antibiotics. The antibiotics are tetracycline, oxytetracycline, penicillin G, streptomycin and erythromycin. All the strains were observed under dark-field microscope for their mortality after inoculation of 18 different concentration of antibiotic at 24, 48, 72, 96, 120, 144 and 168 h post inoculation. The study showed that tetracycline is the antibiotic of choice to treat leptospirosis because it is effective against most of the strains used in the study in very short time after inoculation. However, L. canicola was resistant against tetracycline but sensitive to penicillin G.