We performed a case-control study of risk factors for the acquisition of ciprofloxacin-resistant gram-negative isolates in a Veterans Affairs medical center. Sixty-five patients with resistant isolates and 50 control patients were identified. Prior fluoroquinolone use was significantly more frequent among patients with resistant isolates than it was among controls (58 versus 20%; P = 0.0001). The association with prior quinolone use was stronger in the long-term-care division (81 versus 32%; P = 0.0005) than it was in the acute-care division (29 versus 0%; P = 0.015). On multivariate analysis, prior receipt of a fluoroquinolone was the single most significant risk factor for isolation of a ciprofloxacin-resistant gram-negative organism (P = 0.0001).