Detection and genotyping of vancomycin-resistant Enterococcus spp. by multiplex polymerase chain reaction in Korean aquatic environmental samples.

The distribution characteristics of Enterococcus spp., which are indicators of fecal pollution, were investigated at 33 sites within the 3 major water systems of Korea. Enterococci were detected at concentrations ranging from 1 to 37 CFU/100mL in 41 of 132 samples (31.1%) from the 3 major water systems. The overall average detected concentration was 1.2 CFU/100mL, while the average concentration for all detection sites was 5.3 CFU/100mL. After optimized multiplex polymerase chain reaction (PCR) was performed with newly developed VanA, VanB, VanC-1, and VanC-2/3 primers, concentrations of vancomycin-resistant Enterococcus spp. (VRE) ranging from 1 to 23 CFU/100mL were detected in 17 of 132 samples (12.9%). Of 216 individual enterococcal colonies, 64 (29.6%) displayed the VanC genotype. The results of a susceptibility test to vancomycin showed that the range of the minimal inhibitory concentration (MIC), an indicator of bacterial resistance, was 4 to 24μg/mL, with the average MIC at 9.2±4.5μg/mL. Of the bacterial isolates, 1 colony with the VanC-1 genotype was identified as E. gallinarum by 16S rDNA sequencing, whereas the other 63 colonies had the VanC-2/3 genotype and were identified as E. casseliflavus. Although these results imply that the major head bays of Korea are not contaminated with the highly vancomycin-resistant VanA- or VanB-type VREs, the misuse of antibiotics should be prohibited to minimize the presence of VREs and to maintain a safe water supply for protecting the health of the general population. Based on the study results, we also recommend the implementation of a continuous, broad-spectrum inspection program for Enterococcus spp. and VRE contamination in the major head bays. Furthermore, the multiplex PCR method described in this study can be used effectively for this purpose.