Rationale: Fluoroquinolones are the most commonly prescribed antibiotic class in the United States. They have potential to become first-line anti-tuberculosis therapy, but the effect of fluoroquinolone use on fluoroquinolone resistance in M. tuberculosis is not well-characterized. Objective: To determine prevalence of and risk factors for fluoroquinolone-resistant tuberculosis in a large U.S. population. Methods: We identified all persons with culture-confirmed tuberculosis enrolled in TennCare (Medicaid) and reported to the Tennessee Department of Health from January 2002–December 2006. Persons with fluoroquinolone-resistant M. tuberculosis isolates (cases) were compared to those with susceptible isolates (controls). Fluoroquinolone resistance was determined by agar proportion using ofloxacin 2µg/ml. Outpatient fluoroquinolone exposure in the 12 months prior to tuberculosis diagnosis was ascertained from TennCare pharmacy data. Measurements and Main Results: Of 640 study patients, 116 (18%) had fluoroquinolone exposure 12 months prior to diagnosis, and 16 (2.5%;95% CI:1.4– 4.0%) M. tuberculosis isolates were fluoroquinolone-resistant. Among the 54 patients with >10 days of fluoroquinolone exposure, 7 (13%) had fluoroquinolone resistance. In multivariable logistic regression analyses using propensity score to control for age, sex, race, HIV serostatus, and site of disease, >10 days of fluoroquinolone exposure prior to tuberculosis diagnosis was associated with fluoroquinolone resistance (OR 7.0;95% CI:2.3–20.6;P=0.001). Fluoroquinolone exposure for >10 days that occurred >60 days before tuberculosis diagnosis was associated with the highest risk of resistance (20.8%;OR 17.0;95% CI 5.1–56.8;P<0.001 compared to no exposure). Conclusions: Overall, fluoroquinolone resistance was relatively low. However, receipt of fluoroquinolones for > 10 days, particularly > 60 days prior to tuberculosis diagnosis, was associated with a high risk of fluoroquinolone-resistant tuberculosis.