In analyzing mechanisms of protection against intracellular infections, a series of human CD1-restricted T cell lines of two distinct phenotypes were derived. Both CD4⁻CD8⁻ (double-negative) T cells and CD8⁺ T cells efficiently lysed macrophages infected withMycobacterium tuberculosis. The cytotoxicity of CD4⁻CD8⁻ T cells was mediated by Fas-FasL interaction and had no effect on the viability of the mycobacteria. The CD8⁺ T cells lysed infected macrophages by a Fas-independent, granule-dependent mechanism that resulted in killing of bacteria. These data indicate that two phenotypically distinct subsets of human cytolytic T lymphocytes use different mechanisms to kill infected cells and contribute in different ways to host defense against intracellular infection.