Aim: The aim of the present study was to evaluate the efficacy and role of Xpert Mycobacterium tuberculosis/rifampicin assay in urinary tuberculosis. Methods: This was a prospective study conducted in the Department of department of urology, IGIMS Patna, Bihar, India, for 12 months. Three urine samples were collected from each patient and were subjected to Xpert MTB/RIF assay, acid-fast bacillus (AFB) smear microscopy, and liquid media (BACTEC Mycobacteria Growth Indicator Tube [MGIT] 960) culture. Imaging and tissue biopsies were performed as clinically indicated. Sensitivity, specificity, positive predictive value, and negative predictive value were calculated using the bootstrap method for 95% confidence intervals for the Xpert assay. Results: A total of 120 patients were included for analysis. Of these, 70 fulfilled the study's diagnostic criteria (composite reference) and were labelled as positive for urinary tuberculosis while 50 patients had a negative evaluation and were considered as negative for urinary tuberculosis. When compared to the composite reference standard, a single Xpert assay had a sensitivity of 70.11% (54.21%-81.02% and a specificity of 100% (95% CI 91.11–100%). The PPV was 100% (CI 95% 87.96–100%), and NPV was 73.98% (95% CI 60.37–83.02%). In contrast, the sensitivity of a single smear examination was only 22.5%. Three serial AFB smear examinations improved the sensitivity to 33.23% (20.88–47.21). When MGIT 960 liquid AFB culture was compared to the composite standard using the other variables, the sensitivity was 57.87% (95% CI 45.12–65.77%). McNemar's test showed that Xpert MTB/RIF assay was significantly superior to three auramine-O-fluorescent smear microscopy examinations (70.11% vs. 33.23%, P =−0.0001) while its better performance over the MGIT 960 AFB culture was not statistically significant (70.11% vs. 57.87%, P = 0.06). Xpert MTB/RIF assay identified 60% of smear-negative and 35% of culture-negative cases. However, it missed 3 of the 40 (7.5%) culture-positive patients. None of the patients had rifampicin resistance either on Xpert MTB/RIF assay or on AFB cultures. Conclusion: Xpert MTB/RIF assay on an early morning first void urine specimen can replace smear microscopy as the initial diagnostic test for urinary tuberculosis.