Many candidate regimens for universal treatment of tuberculosis combine novel and existing, widely-used drugs. Appropriate implementation requires evidence-based, context-specific drug-susceptibility testing (DST) strategies.We created a Markov state-transition model of 100,000 adults with TB receiving a novel, fluoroquinolone (FQ)-containing regimen. We estimated clinical outcomes and resource utilization with no FQ-DST, universal use of FQ-DST, or FQ-DST only in patients with rifampin-resistant TB ('targeted FQ-DST'). We considered scenarios of stronger (South Africa) and weaker (Southeast Asia) correlation of fluoroquinolone resistance with rifampin resistance.Relative to no FQ-DST, targeted FQ-DST increased cure of FQ-resistant TB by 7.5% (interquartile range 6.7-9.2%) in South Africa and 1.7% (0.7 -2.5%) in Southeast Asia. However, rare FQ resistance among the more-prevalent rifampin-susceptible TB accounted for 50% of FQ-resistant TB in South Africa and 83% in Southeast Asia. As a result, universal FQ-DST further increased cure of FQ-resistant TB by 3.4% (2.3-5.4%) in South Africa and 5.8% (5.1-6.3%) in Southeast Asia. With targeted FQ-DST, one additional patient was cured per 50 (42-70) tests in South Africa and 44 (37-51) in Southeast Asia. When expanding from targeted to universal FQ-DST, one additional cure required 3500 (2300-5500) tests in South Africa and 410 (370-450) in Southeast Asia. The impact of FQ-DST was sensitive to overall treatment effectiveness, regimen robustness to loss of fluoroquinolone activity, and prevalence of both moxifloxacin and pyrazinamide resistance.FQ-DST improved patient outcomes and was particularly important for high-risk patient groups and less-robust regimens. A universal strategy was favored in generalized epidemics of fluoroquinolone resistance.