Accuracy of antibiotyping using standard antibiograms compared with 16S–23S ribosomal spacer PCR for diagnosis of MRSA

Abstract

Molecular methods for typing methicillin-resistant Staphylococcus aureus (MRSA) include PCR-based methods, pulsed-field gel electrophoresis (PFGE), spa typing, and multilocus sequence typing (MLST) [1, 2]. These techniques assist in targeted infection control efforts and are, moreover, indispensable for tracking genotypes on a large scale. Nevertheless, in vitro antimicrobial susceptibility patterns are still used in epidemiological investigations since antibiograms are rapidly delivered to infection control specialists without the need for additional investigations. The obvious disadvantage of antibiotyping, however, is the variability of resistance expression, which is also influenced by horizontal transmission and loss of extrachromosomal genetic elements. In addition, selective pressure tends to homogenize the antibiogram pattern over time. Some studies have revealed that by considering the diameters of growth inhibition, it is possible to more accurately type MRSA beyond the categorical classifications of susceptible and resistant [3]. However, the documentation and reporting of inhibition zone diameters is not a standard procedure in most hospitals. The aim of the study presented here was to assess the positive and negative predictive values and the sensitivity and specificity of antibiotyping MRSA as compared with 16S–23S ribosomal spacer PCR. The University Hospital Vienna is a 2,140-bed tertiarycare teaching hospital with an MRSA rate of 12.8% [4]. From January 2000 to December 2001, 268 patient-unique MRSA isolates were collected. The isolates were derived from the following sources: swabs of wounds and screening samples (n=132), respiratory tract (n=50), urine (n=42), intravascular catheter tips (n=17), blood (n=12), and miscellaneous other clinical materials (n=15). Staphylococcus aureus was cultured and identified according to standard procedures. Susceptibility testing for oxacillin was performed with oxacillin disks (1 μg) according to the guidelines of the Clinical and Laboratory Standards Institute (CLSI: formerly NCCLS) [5]. Strains identified as MRSA were confirmed by multiplex PCR amplification of the S. aureus-specific gene-fragment SA442 and fragments of the mecA and femB genes [6, 7]. Antimicrobial susceptibility testing with erythromycin, clindamycin, gentamicin, amikacin, ciprofloxacin, rifampicin, and trimethoprim was performed using the Kirby– Bauer disk diffusion method on Mueller–Hinton agar according to CLSI guidelines [5]. The compounds fosfomycin (120 μg disk) and fusidic acid (10 μg disk), which currently lack CLSI-defined breakpoints for susceptibility, were also included in the study, and the inhibition zones required for susceptibility were ≥10 mm for fosfomycin and ≥14 mm for fusidic acid. For further analysis, intermediate-susceptible and -resistant isolates were uniformly referred to as non-susceptible isolates. Glycopeptide antibiotics were not included in the analysis, because all of the isolates were susceptible to these compounds. F. Daxboeck (*) Clinical Institute for Hygiene and Medical Microbiology, Medical University Vienna, Kinderspitalgasse 15, 1090 Vienna, Austria e-mail: florian.daxboeck@meduniwien.ac.at Tel.: +43-1-404001902 Fax: +43-1-404001907