Enterococci are among prominent causes of nosocomial wound infections. Since the rapid detection of causative agents could make earlier administration of choice antibiotics and quick recovery of patients, so the application of rapid diagnostic methods is important. Therefore, this study was designed to evaluate fluorescent in situ hybridization (FISH) for the detection of Enterococcus in wound swab samples. The time needed for FISH procedure is about 3 h. Specimens taken from 33 hospitalized patients were examined by both FISH and culturing procedures. By using conventional culture, 10 of 33 wound samples were culture-positive. Out of these 10 specimens, eight were FISH-positive, but two specimens were FISH-negative for Enterococcus. The remaining 23 wound specimens were Enterococcus negative according to the both methods. Therefore, the specificity of FISH was 100%; however, this method showed 80% sensitivity. Because of high specificity of FISH, the combined application of FISH and cultivation methods would be suggested for detection of enterococci from wound specimens in situations in which rapid diagnosis has an advantage in the treatment of patients.