Epidemiology of European community-associated methicillin-resistant Staphylococcus aureus clonal complex 80 type IV strains isolated in Denmark from 1993 to 2004.

In Europe, community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) infections have been caused predominantly by isolates belonging to the "European CA-MRSA" clone (sequence type 80, staphylococcal cassette chromosome mec type IV). In this study, the epidemiology of European CA-MRSA was investigated on a nationwide scale, covering the period from 1993 to 2004. Denmark has been a low-prevalence country regarding MRSA since the mid-1970s but has experienced an increase in the number of new MRSA cases in recent years. Our results show that European CA-MRSA contributed to this increase. The isolates primarily caused skin and soft tissue infections (SSTIs) in patients outside hospitals, and transmission between household members was the predominant mode of spread. Although some of the isolates were found in hospitalized patients, nosocomial transmission seemed likely in only one instance, pointing to endogenous infections as an important factor. Compared to the CA-MRSA clone most common in the United States (USA300), the European CA-MRSA clone seems less well adapted to persist in hospital environments. Patients with a recent history of travel or family relation to the Mediterranean or Middle East were highly overrepresented. The epidemiological data indicated that the European CA-MRSA isolates were introduced into Denmark on multiple occasions, paralleled by an increasing level of genetic diversity of the isolates found during the study period. European CA-MRSA has previously been described as a rather uniform clone. However, we found pronounced, diverse pulsed-field gel electrophoresis subtypes, staphylococcal protein A gene (spa) types, and susceptibility patterns.

Increasing incidence but decreasing in-hospital mortality of adult Staphylococcus aureus bacteraemia between 1981 and 2000.

Staphylococcus aureus is a leading cause of bacteraemia. This study analysed temporal trends from 18,702 adult cases of S. aureus bacteraemia in Denmark between 1981 and 2000. After stratification for mode of acquisition, 57% of cases were hospital-acquired (HA), 28% were community-acquired (CA) and 15% were of undetermined acquisition (UA). Incidence rates increased from 18.2 to 30.5 cases/100,000 population. Annual rates increased by 6.4% for CA, by 2.2% for HA and by 3.6% for UA cases, respectively. Case-mortality associated with HA bacteraemia decreased from 36.2% to 20.7% (43% rate reduction, p 0.0001), compared with a decrease from 34.5% to 26.5% (23% rate reduction, p 0.0001) for CA bacteraemia. Following multivariate analysis, age, pneumonia, endocarditis and chronic illness were associated with increased mortality, regardless of the mode of acquisition. Overall, mortality associated with S. aureus bacteraemia declined significantly between 1981 and 2000, but incidence rates doubled, so that the total number of deaths increased. These data emphasise the public health importance of S. aureus bacteraemia and the need for further preventive measures and improved care in order to reduce incidence rates and improve outcomes.

Evaluation of a new 3-h hybridization method for detecting the mecA gene in Staphylococcus aureus and comparison with existing genotypic and phenotypic susceptibility testing methods.

A new 3-h hybridization assay for detection of the staphylococcal mecA gene and the Staphylococcus aureus nuclease gene was evaluated by comparing the assay with existing genotypic and phenotypic methods. A total of 275 S. aureus strains were tested, including 257 epidemiologically unrelated strains (135 mecA-positive and 122 mecA-negative; collection I), and 18 strains with known borderline resistance to methicillin (collection II). Complete agreement was obtained for both collections when comparing the new assay with genotypic methods. We further evaluated a range of phenotypic susceptibility methods recommended in Europe and/or USA using the presence of the mecA gene as the defining standard. For collection I a high degree of agreement was found for both Etests (256 strains) and the oxacillin screen plate test (255 strains); the degree of agreement was lower for agar dilution methicillin (250 strains) and oxacillin 1 microg discs (239 strains). For the borderline strains a high degree of agreement was only obtained by the oxacillin screen plate test (17 of 18 strains). The other tests were less accurate, in the following order: agar dilution methicillin, Etest methicillin, Etest oxacillin and oxacillin discs with disagreement for four, five, nine and 13 strains, respectively. In conclusion, the new hybridization assay is a rapid and exact method for detecting the mecA gene and the S. aureus nuclease gene. This study confirms that phenotypic tests for methicillin resistance in S. aureus strains creates both false-susceptible and false-resistant results, especially for borderline resistant strains.

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.

Glycopeptide susceptibility among Danish Enterococcus faecium and Enterococcus faecalis isolates of animal and human origin and PCR identification of genes within the VanA cluster.

The MICs of vancomycin and avoparcin were determined for isolates of Enterococcus faecium and isolates of Enterococcus faecalis recovered from the feces of humans and animals in Denmark. Two hundred twenty-one of 376 (59%) isolates of E. faecium and 2 of 133 (1.5%) isolates of E. faecalis were resistant to vancomycin (MICs, 128 to > or = 256 micrograms/ml), and all vancomycin-resistant isolates were resistant to avoparcin (MICs, 64 to > or = 256 micrograms/ml). All vancomycin-resistant isolates examined carried the vanA, vanX, and vanR genes, suggesting that a gene cluster similar to that of the transposon Tn1546 was responsible for the resistance.