Division of the genus Enterococcus into species groups using PCR-based molecular typing methods.
Broad-range 16S rDNA PCR (BR-PCR) applied to DNA from 32 clinical enterococcal isolates and 12 other enterococci from a clinical reference collection followed by species-specific hybridization analysis identified 25 strains of Enterococcus faecalis and 19 Enterococcus species. Randomly amplified polymorphic DNA (RAPD) analysis using UPGMA clustering on the same material revealed four different clusters at a similarity level of 49%. Based on partial 16S rDNA sequence analysis of variable regions V4 and V9, it was possible to divide the 19 type strains specifying the genus Enterococcus into 12 different 16S rDNA species groups. The type strain distribution then served as a template for the analysis of the other 44 strains which were assigned to four different species groups (a-d) based on their 16S rDNA motifs. There was good agreement with the RAPD clusters. Species group a was an individual species line containing 25 strains that were identified as E. faecalis. Group b also represented an individual species line of 12 strains identified as E. faecium. The remaining seven strains that formed species groups c and d could not be fully identified to species by this analysis. It was concluded that BR-PCR of 16S rDNA followed by partial sequence analysis of the PCR products is a reliable technique for the identification and classification of enterococci. Further division of unresolved species groups should be achievable if regions other than V4 and V9 of 16S rDNA are also analysed.
Monstein
Hans-Jürg
HJ
Division of Clinical Microbiology, Faculty of Health Sciences, University Hospital, S-581 85 Linköping, Sweden.
Quednau
Mikael
M
Laboratory of Food Hygiene, Department of Food Technology, University of Lund, S-221 00 Lund, Sweden.
Samuelsson
Annika
A
Division of Clinical Microbiology, Faculty of Health Sciences, University Hospital, S-581 85 Linköping, Sweden.
Ahrné
Siv
S
Laboratory of Food Hygiene, Department of Food Technology, University of Lund, S-221 00 Lund, Sweden.
Isaksson
Barbro
B
Division of Clinical Microbiology, Faculty of Health Sciences, University Hospital, S-581 85 Linköping, Sweden.
Jonasson
Jon
J
Division of Clinical Microbiology, Faculty of Health Sciences, University Hospital, S-581 85 Linköping, Sweden.
D015373:Bacterial Typing Techniques; D001483:Base Sequence; D015139:Blotting, Southern; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics; D004275:DNA, Ribosomal / Q000032:analysis / Q000235:genetics; D004587:Electrophoresis, Agar Gel; D016983:Enterococcus / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D016908:Gram-Positive Bacterial Infections / Q000382:microbiology*; D006801:Humans; D017720:Molecular Epidemiology; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D012336:RNA, Ribosomal, 16S / Q000235:genetics; D019105:Random Amplified Polymorphic DNA Technique; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D013045:Species Specificity
D004269:DNA, Bacterial; D004275:DNA, Ribosomal; D012336:RNA, Ribosomal, 16S