Susceptibility to disease after infection by Mycobacterium tuberculosis is influenced by environmental and host genetic factors. Vitamin D metabolism leads to activation of macrophages and restricts the intracellular growth of M. tuberculosis. This effect may be influenced by polymorphisms at three sites in the vitamin D receptor (VDR) gene. We investigated the interaction between serum vitamin D (25-hydroxycholecalciferol) concentrations and VDR genotype on susceptibility to tuberculosis. This study was a hospital-based case-control analysis of Asians of Gujarati origin, a mainly vegetarian immigrant population with a high rate of tuberculosis. We typed three VDR polymorphisms (defined by the presence of restriction endonuclease sites for Taq1, Bsm1, and Fok1) in 91 of 126 untreated patients with tuberculosis and 116 healthy contacts who had been sensitised to tuberculosis. Serum 25-hydroxycholecalciferol was recorded in 42 contacts and 103 patients. 25-hydroxycholecalciferol deficiency was associated with active tuberculosis (odds ratio 2.9 [95% CI 1.3-6.5], p=0.008), and undetectable serum 25-hydroxycholecalciferol (<7 nmol/L) carried a higher risk of tuberculosis (9.9 [1.3-76.2], p=0.009). Although there was no significant independent association between VDR genotype and tuberculosis, the combination of genotype TT/Tt and 25-hydroxycholecalciferol deficiency was associated with disease (2.8 [1.2-6.5]) and the presence of genotype ff or undetectable serum 25-hydroxycholecalciferol was strongly associated with disease (5.1 [1.4-18.4]). 25-hydroxycholecalciferol deficiency may contribute to the high occurrence of tuberculosis in this population. Polymorphisms in the VDR gene also contribute to susceptibility when considered in combination with 25-hydroxycholecalciferol deficiency.