The objectives of this study were to investigate antibiotic resistance in urinary pathogens from Korean patients with uncomplicated acute pyelonephritis (UAPN), and to determine the effect of fluoroquinolone (FQ) resistance on clinical outcome in those patients with UAPN initially treated with FQ. Clinical and microbiologic data for all the APN patients attending 14 hospitals in South Korea in 2008 were collected retrospectively. Urinary pathogens were identified in 719 cases, and Escherichia coli was the most common pathogen (661/719, 91.9%). Antibiotic susceptibilities to several E. coli antibiotics were as follows: ciprofloxacin, 84.1%; trimethoprim-sulfamethoxazola (TMP-SMX), 67.2%; and extended-spectrum beta-lactamase-negative, 92.4%. FQ was the most frequent antibiotic prescribed for UAPN (45.3% intravenously and 53.9% by mouth). We compared clinical outcomes and hospital days in patients with FQ-resistant (32) and FQ-sensitive E. coli (173) who received FQ as initial empirical therapy. Clinical cure was higher in the FQ-sensitive group (78% vs. 91%, p=0.027), and hospital days were longer in the FQ-resistant group (9.6±5.5 days vs. 7±3.5 days, p=0.001). In conclusion, FQ-sensitivity of E. coli from UAPN was 84.1%. FQ treatment of UAPN caused by FQ-resistant E. coli has a lower cure rate and involves longer hospital stay than FQ treatment of cases caused by FQ-sensitive E. coli.