Mycobacterial growth and sensitivity to H₂O₂ killing in human monocytes in vitro

The intracellular growth and susceptibilities to killing by H₂O₂ in cultured human monocytes of a number of mycobacterial species including laboratory strains and clinical isolates of Mycobacterium tuberculosis, and Mycobacterium bovis bacillus Calmette-Guerin (BCG) and a clinical isolate of Mycobacterium avium-M. intracellulare were examined. The clinical isolate of M. avium-M. intracellulare did not replicate in freshly explanted monocytes (generation time of >400 h); BCG replicated with a generation time of 95 h, and M. tuberculosis strains CDC551, H37Rv, and H37Ra replicated with generation times of 24, 35, and 37 h, respectively, during the 4-day growth assay. When cultured in monocytes for 4 days, the mycobacteria were variably sensitive to H₂O₂-induced killing. A positive correlation between the generation time and percent killing of intracellular bacilli was observed. By comparison, mycobacterial strains were similarly sensitive to H₂O₂ treatment in cell-free culture media and in sonicated cell suspensions. Using a number of inhibitors of reactive oxygen intermediates we determined that other than catalase the inhibitors tested did not affect H₂O₂-induced killing of intracellular mycobacteria. Our studies suggest that the killing of mycobacteria growing in human monocytes in vitro by the addition of exogenous H₂O₂ is dependent on the susceptibility to a peroxide-induced killing pathway as well as on the intracellular growth rate of the mycobacteria.