In vitro cultivation of Mycobacterium X from Mycobacterium leprae infected tissues in propane-tetradecane medium (a preliminary communication).

Host grown Mycobacterium leprae and cultures of Mycobacterium X, cultivated from M. leprae infected armadillo and human specimens, were inoculated into propane and propane-tetradecane media. The media contained in one litre distilled water KH2PO4, 7 g; Na2HPO4, 0.5 g; (NH4)2SO4, 2 g; MgSO4, 0.1 g; ferric ammonium citrate, 20 mg and yeast extract (Difco), 0.1 g. Twenty ml media, distributed into each of 50 ml screw cap tubes, were inoculated with the bacilli and bubbled aseptically for 10 s with 99% purity propane gas. Tetradecane-propane media were prepared by adding 0.1 ml tetradecane to each of the tubes containing 20 ml propane medium. When incubated at 32 degrees C a logarithmic growth rate was counted in the propane-tetradecane media following a one to two week latency period. The time of division was estimated at seven days. In the propane-tetradecane medium, growth occurred at the interface of the tetradecane oil and water as a thin veil developing into a 1 to 3 mm thick emulsion in two to three months. No growth occurred in the propane medium and growth was extremely slow in the tetradecane medium. When added to the tetradecane medium, propane considerably shortened the latency period and the generation time, resulting in increased bacterial yield. Bacilli were strongly acid-fast; the culture did not grow on Löwenstein-Jensen or in Dubos media, but produced the localized disease typical of M. leprae in the foot pads of mice.