Detection of clinical vancomycin-resistant enterococci in Denmark by multiplex PCR and sandwich hybridization.

Since the first cases of human infection with vancomycin-resistant enterococci (VRE) were reported in the late eighties, there has been a dramatic increase in VRE all over the world. So far, there have not been any reports of clinical VRE in Denmark. In this study we have investigated 131 clinically important enterococci sent to Statens Serum Institut from all over Denmark during the period July 1995 to May 1997. The susceptibility to vancomycin, teicoplanin, ampicillin and gentamicin was tested by the agar dilution method. In addition, two methods were developed to detect the different genotypes of glycopeptide resistance described in enterococci: a multiplex PCR assay for detection of vanA, vanB, vanC-1, vanC-2/3 ligase genes including 16S rRNA gene control primers and a sandwich hybridization assay to confirm vanA and vanB PCR-positive strains. The highest frequency of resistance to the tested antibiotics was found in the Enterococcus faecium group. Four strains were found with acquired resistance to glycopeptides: one E. faecium and one E. gallinarum were vanA positive, and two E. faecium isolates were vanB positive. These strains were isolated from different hospitals in different periods of time, and all patients recovered from their infections with VRE. Today, the PCR and sandwich hybridization methods are used for screening of vancomycin-resistant enterococci in humans as part of the Danish surveillance programme.