Gamma-interferon (IFN-γ) plays a determinant role in activating macrophages that are critical to control Mycobacterium tuberculosis infection. However, M. tuberculosis can escape killing by attenuating the response of macrophages to IFN-γ by blocking the transcription of a subset of IFN-γ inducible genes. This inhibition occurs after signalling through Toll-like receptor 2 (TLR2). While most studies have investigated the inhibition of IFN-γ responsive genes after TLR2 signalling, the present study focuses on the functional implications of inhibition of IFN-γ signalling in macrophages with regard to mycobacteria killing. Here, we provide evidence that exposure of the murine macrophage cell line J774 to the TLR2 ligands; 19-kDa or zymosan, but not the TLR4 ligand LPS, inhibits IFN-γ-induced killing of Mycobacterium bovis Bacillus Calmette-Guérin (BCG). Moreover, exposure of bone marrow-derived macrophages (BMM) from TLR4-deficient and wild-type (WT), but not from TLR2-deficient mice to 19-kDa lipoprotein (19-kDa) or zymosan, results in an impairment of IFN-γ-mediated killing. We demonstrate that 19-kDa and zymosan inhibit the ability of IFN-γ to activate murine macrophages to kill BCG without inhibiting nitric oxide (NO) or tumour necrosis factor (TNF) production. Finally, we demonstrate that the inhibitory effect of 19-kDa on IFN-γ signalling is overcome with increasing amounts of IFN-γ indicating that the refractoriness could be reversed at optimal IFN-γ concentrations. The critical role of TLR2 but not TLR4 signalling in the inhibition of IFN-γ promoted killing of mycobacteria is discussed. © 2007 The Authors.