We examined Enterococcus faecalis strains clinically isolated from 100 patients with urinary tract infections (UTIs) for their susceptibility to levofloxacin (LVX) by measuring the MIC and investigated amino acid mutations by direct DNA sequencing, which were then correlated with LVX resistance. Next, we studied risk factors for LVX resistance, such as age, gender, and previous fluoroquinolone use, and investigated the statistical correlation of these risk factors with each amino acid mutation and LVX resistance. Of the 100 isolates tested, 14 isolates showed LVX resistance and all of these isolates had amino acid mutations. We demonstrated that 2 out of 4 mutations (Ser83-to-Ile in gyrA and Ser80-to-Ile in parC) had a significant correlation with LVX resistance. There was a significant relationship between isolates with 2 or 3 amino acid mutations and LVX resistance. In addition, we found a significant correlation between the previous use of fluoroquinolones and LVX resistance or the presence of mutations and also demonstrated that previous use of other types of antibiotics was significantly related to the presence of mutations by multivariate analysis. In conclusion, we found significant correlation between amino acid mutations in E. faecalis, LVX resistance, and risk factors such as previous use of fluoroquinolones.Enterococcus faecalis is one of the most common pathogens in urinary tract infections (UTIs) (7). Recently, enterococcal infections have increased, and E. faecalis accounts for the majority of enterococcal infections (7). Fluoroquinolones have been frequently used to treat E. faecalis UTIs, and the emergence of fluoroquinolone-resistant E. faecalis (QREF) strains has recently been reported in several countries (12). In our institution, the proportion of QREF isolates gradually increased from 12/83 (14.4%) in 2004 to 10/49 (20.4%) in 2007, but this was not a statistically significant difference (P ϭ 0.3755). Rudy et al. demonstrated in analyses of 130 E. faecalis strains isolated from urine that all strains were sensitive to glycopeptides (vancomycin [VAN] and teicoplanin), 96% were sensitive to penicillin, 43% to ciprofloxacin, and 28% to tetracycline (22). Muratani et al. reported a 38% frequency of levofloxacin (LVX)-resistant E. faecalis isolates from UTIs (17).Mutations in the quinolone resistance-determining regions (QRDR) of gyrA coding for DNA gyrase and parC coding for DNA topoisomerase IV, the efflux system, antimicrobial-modifying resistance enzymes, and plasmid-mediated mechanisms are considered to contribute to resistance to fluoroquinolones (3,10,11,18,23,25,27). To our knowledge, no previous report has correlated risk factors with fluoroquinolone resistance and amino acid mutation in the QRDR in E. faecalis strains clinically isolated from the urine of UTI patients.Rattanaumpawan et al. investigated the risk factors for fluoroquinolone resistance in enterococcal UTIs and demonstrated that recent exposure to antibiotics such as fluoroquinolones, extended-spectrum cephalosporins, and c...