Summary auto-generated
This study evaluated two DNA-based typing methods for identifying enterococcal species, which are important human pathogens with significant antibiotic resistance. The researchers analyzed 66 enterococcal strains representing seven species using arbitrarily primed PCR with primer D11344 and pulsed-field gel electrophoresis (PFGE) of SmaI-digested genomic DNA, comparing results to whole-cell protein analysis by SDS-PAGE. The D11344-primed PCR generated simpler, more easily interpreted band patterns than protein analysis and proved equally discriminative for species identification, allowing visual examination of DNA profiles for clinical isolate identification. PFGE produced no species-specific patterns but was superior for analyzing relationships between individual strains. Importantly, neither PCR nor protein analysis could differentiate between Enterococcus casselifluvus and Enterococcus flavescens, suggesting the latter may represent a biovar rather than a separate species. The study demonstrates that arbitrarily primed PCR with primer D11344 offers a practical, technically simpler alternative to protein electrophoresis for rapid enterococcal species identification in clinical laboratories.
Key findings
- Arbitrarily primed PCR using primer D11344 successfully identified enterococcal species with discrimination equivalent to whole-cell protein analysis
- D11344-primed PCR generated simpler, more easily interpreted band patterns compared to protein electrophoresis, allowing visual identification of unknown isolates
- PFGE was not useful for species-level differentiation but effectively analyzed relationships between individual strains
- Enterococcus casselifluvus and E. flavescens could not be differentiated by either method, suggesting E. flavescens may be a biovar rather than a distinct species
- The PCR method is technically less demanding and more suitable for routine clinical laboratory use than protein electrophoresis analysis
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Abstract
The increasing problems encountered with enterococcal nosocomial infections and the intrinsic and acquired resistance of the enterococci to different antimicrobial compounds highlight the need for a rapid identification technique. Enterococcus faecalis is readily identified by biochemical tests, but species differentiation within the Enterococcus faecium and Enterococcus gallinarum species groups is less well established. In the present study, 66 strains representing the most prevalent human enterococci were used to develop a PCR-based species-specific identification protocol. Whole-cell protein analysis by sodium dodecyl sulfate-polyacrylamide gel electrophoresis was used as a reference method for species identification. In addition, the genomic SmaI macro-restriction fragment distribution of all of the strains was examined by pulsed-field gel electrophoresis (PFGE). Oligonucleotide D11344-primed PCR was as discriminative as whole-cell protein analysis and resulted in more easily interpreted band patterns. This PCR-based technique allowed identification of clinical isolates by visual examination of the DNA profiles obtained. The inability of both methods to discriminate between Enterococcus casseliflavus and Enterococcus flavescens brought into question the species status of E. flavescens. PFGE did not result in species-discriminative DNA bands or band patterns, but proved to be superior for interpretation of interstrain relationships.