Background: It is now generally accepted that Vitamin D Deficiency (VDD) is a worldwide health problem that affects not only musculoskeletal health but also a wide range of acute and chronic diseases. The role of VDD in predisposition of TB has been suspected since a long time. Aim: The aim of the study was to assess the levels of Vitamin D, to find out the prevalence of VDD and the determinants associated with it among newly diagnosed tuberculosis patients attending Directly Observed Treatment Short-course (DOTS) center centre RHTC (Rural Health Training Centre) Najafgarh. Methodology: A cross sectional DOTS center based study was conducted among 400 Tuberculosis patients. The study tool comprised of a pre-designed, pre-tested, semi-structured, interviewer administered questionnaire in Hindi. General physical examination and systemic examination was done for all the study participants.3 ml of blood was drawn from the ante-cubital vein using a 24G hypodermic needle and a 5cc plastic syringe. The Calbiotech, Inc. 25-hydroxy (25-OH) Vitamin D ELISA was used for quantitative determination of total 25-OH Vitamin D in human serum and plasma. The data was entered into Microsoft Excel after preparing a Master-chart. Data analysis was done using SPSS software licensed version 21. Chi-square test and Fischer exact test was used to determine the association. Results: Out of 400 study participants, median age was 31 years ranged from a minimum of 7 years to a maximum of 90 years. Males were 66% and females were 44%. VDD was found in 90.5% patients and VDD was significantly associated with vegetarianism, inadequate consumption of dairy products, inadequate sun exposure and low BMI. Conclusion: VDD was highly prevalent in Tuberculosis patients attending DOTS centre Najafgarh Delhi. Main contributing factors were inadequate sun exposure, vegetarianism, inadequate consumption of dairy products & low BMI.