Tumour necrosis factor-alpha (TNF-alpha) plays a critical role in the recruitment and activation of mononuclear cells in mycobacterial infection. The role of membrane TNF, in host resistance against Mycobacterium bovis bacille Calmette-Guérin (BCG), was tested in knock-in mice in which the endogenous TNF was replaced by a non-cleavable and regulated allele (Delta1-12, TNF(tm/tm)). While 100% of mice with complete TNF deficiency (TNF(-/-)) succumbed to infection, 50% of TNF(tm/tm) mice were able to control M. bovis BCG infection and survived the experimental period. Membrane expressed TNF allowed a substantial recruitment of activated T cells and macrophages with granuloma formation and expression of bactericidal inducible nitric oxide synthase (iNOS). Using virulent Mycobacterium tuberculosis infection we confirm that membrane TNF conferred partial protection. Infection in TNF(tm/tm) double transgenic mice with TNF-R1 or TNF-R2 suggest protection is mediated through TNF-R2 signalling. Therefore, the data suggest that membrane-expressed TNF plays a critical role in host defence to mycobacterial infection and may partially substitute for soluble TNF.