Intracellular growth of pathogenic mycobacteria in the continuous murine macrophage cell line J-774: Ultrastructure and drug-susceptibility studies

The comparative action of seven drugs, namely, rifampicin (RIF), ansamycin-LM 427 (ANS), streptomycin (SM), isoniazid (INH), pyrizinamide (PZA), clofazimine (CLF), and pristinamycin (PST), was studied both on extracellularly and intracellularly growing mycobacterial speciesMycobacterium bovis, M. tuberculosis, andM. avium. All the drugs were used at their minimal inhibitory concentrations (MICs) and their obtainable serum levels in man; 10×MICs, when less than the obtainable serum levels, were also tested. The action of drugs on extracellularly growing bacilli was tested with the Middlebrook 7H9-Tween medium, whereas for intracellular growth, an experimental model using a continuous murine macrophage cell line J-774 was developed, and macrophage-mycobacteria interactions by use of cytochemistry, scanning and transmission electron microscopy were investigated. Extracellularly growingM. avium was resistant to all the drugs tested; however, when tested on intracellularly growing bacilli, both RIF and CLF were found to be bactericidal, whereas other drugs only delayed the bacterial growth. In the case of extracellularly growingM. bovis andM. tuberculosis, INH, RIF, and SM were active, whereas PZA was active only onM. tuberculosis. However, on intracellularly growing bacilli, only INH and RIF were found to be bactericidal for both species, CLF to a lesser extent in the case ofM. tuberculosis; ANS and SM were bacteriostatic, whereas both PZA and PST were without any antibacterial effect. This investigation underlined the discrepancies concerning the action of drugs on extra- and intracellularly multiplying mycobacteria and the advantage of using a continuous cell line model while studying drug susceptibility of mycobacteria in relation to their intracellular growth.