title: Automated ribotyping of vancomycin-resistant Enterococcus faecium isolates.
abstract: Vancomycin-resistant Enterococcus faecium (VREF) strains represent an important threat in hospital infections in the United States and are found at high frequencies in both the community and farm animals in Europe. We evaluated automated ribotyping for interlaboratory reproducibility by using the restriction enzymes EcoRI and BamHI and compared ribotyping to both amplification of fragment length polymorphism (AFLP) analysis and multilocus sequence typing (MLST) to assess its discriminatory power and capacity for the identification of epidemiologically important strains. Of 19 (EcoRI) and 16 (BamHI) isolates tested in duplicate in two laboratories, 18 (95%) and 16 (100%), respectively, showed reproducible ribotypes. These high reproducibility rates were obtained only after manual refinement of the automated fingerprint analysis. A group of 49 VREF strains initially selected to represent 32 distinct AFLP types were separated into 28 EcoRI ribotypes, 25 BamHI ribotypes, and 28 sequence types. Ribotyping with EcoRI and BamHI was able to discern the host-specific genogroups recently disclosed by AFLP typing and MLST and to distinguish most strains containing the esp gene, a marker specific for strains causing hospital outbreaks. An expandable ribotype identification library was created. We recommend EcoRI as the enzyme of choice for automated ribotyping of VREF strains. Given the high level of discrimination of VREF strains, the high rate of interlaboratory reproducibility, and the potential for the identification of epidemiologically important genotypes, automated ribotyping appears to be a very valuable approach for characterizing VREF strains.
authors.affiliation: Eijkman-Winkler Institute, University Medical Centre Utrecht, 3584 CX Utrecht, The Netherlands. sbrisse@lab.azu.nl
authors.forename: Sylvain
authors.initials: S
authors.lastname: Brisse
authors.forename: Vivian
authors.initials: V
authors.lastname: Fussing
authors.forename: Ben
authors.initials: B
authors.lastname: Ridwan
authors.forename: Jan
authors.initials: J
authors.lastname: Verhoef
authors.forename: Rob J L
authors.initials: RJ
authors.lastname: Willems
mesh_terms: D000483:Alleles; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D015373:Bacterial Typing Techniques / Q000379:methods; D002415:Cats; D002417:Cattle; D004275:DNA, Ribosomal; D015245:Deoxyribonuclease BamHI / Q000378:metabolism; D015246:Deoxyribonuclease EcoRI / Q000378:metabolism; D016984:Enterococcus faecium / Q000145:classification* / Q000187:drug effects* / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D015203:Reproducibility of Results; D021521:Ribotyping / Q000379:methods*; D012371:Robotics; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA; D020713:Vancomycin Resistance
chemical_list: D001426:Bacterial Proteins; D004275:DNA, Ribosomal; D015245:Deoxyribonuclease BamHI; D015246:Deoxyribonuclease EcoRI