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Objectives In Mali early detection and treatment of multidrug-resistant tuberculosis (MDR-TB) are still challenging due to the cost, time and/or complexity associated with regular tests. Microscopic Observation Drug Susceptibility (MODS) is a low-cost assay validated by WHO in 2010. It is a liquid-culture-based assay to detect the ‘cording’ characteristic of Mycobacterium tuberculosis complex and to assess susceptibility to both isoniazid and rifampicin defining multidrug-resistant tuberculosis (MDR-TB). In this study we aimed to evaluate the performance of MODS as diagnostic tool compared with a validated method—Mycobacteria Growth Indicator Tube/Antimicrobial Susceptibility Testing/Streptomycin, Isoniazid, Rifampicin and Ethambutol (MGIT/AST/SIRE). Methods and Results Between January 2010 and October 2015 we included 98 patients with suspected TB in an observational cohort study. The sensitivity and specificity of MODS assay for detecting TB were respectively 94.12% and 85.71% compared with the reference MGIT/7H11 culture, with a Cohen κ coefficient of 0.78 (95% CI 0.517–1.043). The median time to culture positivity for MODS assay and MGIT (plus interquartile range, IQR) was respectively 8 days (IQR 5–11) and 6 days (IQR 5–6). In detecting patients with MDR-TB, the sensitivity and specificity of MODS assay were respectively 100% and 95.92%. The positive predictive value and negative predictive value were, respectively, 66.7% and 100%. The median turnaround times for obtaining MDR-TB results using MODS assay and MGIT/AST/SIRE was respectively 9 days and 35 days. Hence, the MODS assay rapidly identifies MDR-TB in Mali compared with the MGIT/AST/SIRE. Conclusion As an easy, simple, fast and affordable method, the MODS assay could significantly improve the management of TB. [ABSTRACT FROM AUTHOR]