In this study, we evaluated the in vitro activity of fosfomycin and 7 other comparator agents against 307 Escherichia coli isolates including ciprofloxacin-resistant or extended-spectrum beta-lactamase (ESBL)-producing isolates. Bacterial isolates were collected from urine and blood from patients at a Korean tertiary-care hospital. Among 307 E. coli isolates, 30.3% were resistant to ciprofloxacin (MIC(90), >32 mg/L) and 7.8% produced ESBLs. The highest resistance rate was observed in ampicillin (69.7%), followed by trimethoprim-sulfamethoxazole (43.0%), and then amoxicillin-clavulanate (32.2%). All isolates were susceptible to imipenem (MIC(90), 0.125 mg/L). All but 1 isolate was susceptible to fosfomycin (MIC(90), 16 mg/L), regardless of the collected sources, ciprofloxacin resistance, and ESBL production. The data showed excellent activity of fosfomycin against E. coli isolates including fluoroquinolone-resistant strains. The clinical usefulness of fosfomycin, as a 1st-line therapy for urinary tract infection, should be evaluated further, especially in regions where ciprofloxacin resistance rates are high.