Background: Xpert MTB/RIF Ultra (Ultra) is a novel molecular assay which is replacing Xpert MTB/RIF (Xpert) as a front-line test for tuberculosis (TB) diagnosis. We conducted a field evaluation of the diagnostic accuracy of Ultra using two cohorts of participants (from passive and active case finding interventions) in a high TB/HIV burden area in Southern Mozambique. Methods: Respiratory specimens obtained from symptomatic adults accessing health care services (passive case finding (PCF) cohort), and from household and community close contacts (active case finding (ACF) cohort), were tested for smear microscopy, culture, Xpert and Ultra (same specimen). Liquid and solid culture results served as a composite reference standard. We also explored trace results’ impact on test specificity by their recategorization to negative (in all cases or just in previously treated individuals) and active follow-up. Findings: A total of 1419 and 252 participants were enrolled in the ACF and PCF cohorts respectively. For the PCF cohort, Ultra showed higher sensitivity compared to Xpert, overall (0·95 (95% CI: 0·90, 0·98) vs 0·88 (0·82, 0·93); p<0·001) and among smear negative patients (0·84 (95% CI: 0·71, 0·93) vs 0·63 (95% CI: 0·48, 0·76) p<0·001). Conversely, specificity of Ultra was lower and similar than Xpert’s (0·98 (0·97, 0·99) vs 0·96 (0·95, 0·97); p= 0·008). In the ACF-group, sensitivity and specificity for both tests were the same (0·67 (95% CI: 0·22,0·96), although Ultra doubled Xpert in the number of microbiologically confirmed samples (4·7% (12/252) vs 2·7% (7/252)). Conditional recategorization of trace results, maintained differences in specificity in the PCF cohort. Interpretation: Our results add evidence on the improved sensitivity of Ultra and suggest that overall differences in specificity among assays might be driven by misclassification of true-positive results. These findings support the use of Ultra as initial front-line test for diagnosis in health care settings, and advocate for a comprehensive clinical and epidemiological evaluation of patients with single trace results in order to inform treatment initiation. Funding Statement: We acknowledge support from the Spanish Ministry of Science, Innovation and Universities through the “Centro de Excelencia Severo Ochoa 2019-2023” Program (CEX2018-000806-S), and support from the Generalitat de Catalunya through the CERCA Program. B.S receives a pre-doctoral fellowship from the Secretariat of Universities and Research, Ministry of Enterprise and Knowledge of the Government of Catalonia and co-funded by European Social Fund (AGAUR). This project has received consumables donations from Cepheid. Cepheid was not involved in the data analysis or decision to submit this manuscript for publication. Declaration of Interests: The authors have no conflicts of interest to disclose. Ethics Approval Statement: The protocol was approved by CISM’s Internal Scientific Committee, CISM’s Internal Bioethics Committee (CIBS – Comite Institucional de Bioetica para a Saude) and the National Bioethics Committee (CNBS - Comite Nacional de Bioetica para a Saude, reference 369/CNBS/17).