Identification of enterococcal isolates by temperature gradient gel electrophoresis and partial sequence analysis of PCR-amplified 16S rDNA variable V6 regions.

Based on partial sequence analysis of PCR-amplified 16S rDNA variable V6 regions of 14 enterococcal type strains, Enterococcus faecalis, Enterococcus mundtii, Enterococcus gallinarum, Enterococcus avium, Enterococcus raffinosus and Enterococcus saccharolyticus showed characteristic sequence motifs which made it possible to separate them into six individual species lines. Furthermore, two species cluster groups could be identified, including (i) Enterococcus faecium, Enterococcus durans, Enterococcus hirae, Enterococcus malodoratus, and (ii) Enterococcus casseliflavus/Enterococcus flavescens, Enterococcus pseudoavium, Enterococcus dispar and Enterococcus sulfureus. There were identical DNA sequences in the V6 region within each group. Temporal temperature gradient gel electrophoresis (TTGE) of the PCR products from 16 type strains, 12 enterococcal reference strains and 8 clinical isolates revealed that a single nucleotide divergence in DNA sequences was sufficient for separation, identification and division of the studied enterococcal strains into corresponding TTGE profiles. It was concluded that partial DNA sequence analysis and TTGE profiling of PCR-amplified 16S rDNA variable V6 regions provide useful tools for the identification of clinically important Enterococcus spp.