Multiplex polymerase chain reaction for detection of genes for Staphylococcus aureus thermonuclease and methicillin resistance and correlation with oxacillin resistance.
A multiplex polymerase chain reaction (mPCR) was used for simultaneous amplification of the staphylococcal nuc gene, encoding the thermostable nuclease (TNase), and the mecA gene, encoding the penicillin-binding protein 2a which is associated with staphylococcal methicillin resistance. A total of 219 staphylococcal strains were tested and the mPCR data were compared with coagulase production and in vitro oxacillin susceptibility. The agreement was 100% for coagulase production and nuc amplification, and 97.7%, 96.8 and 97.3% for mecA amplification and oxacillin resistance tested with MIC determination, disk diffusion and agar screen methods, respectively. Discrepant results were due to non-S. aureus isolates with borderline MICs of oxacillin (1-8 micrograms/ml). In a pilot test the mPCR simultaneously amplified both genes of staphylococci in blood cultures. This mPCR is a rapid and reliable method for single-step identification of cultures of MRSA and may prove to be useful for direct application on clinical specimens.
Brakstad
O G
OG
Applied Chemistry Division, SINTEF, Trondheim, Norway.
Maeland
J A
JA
Tveten
Y
Y
D017931:DNA Primers; D004269:DNA, Bacterial / Q000032:analysis / Q000302:isolation & purification; D004352:Drug Resistance, Microbial / Q000235:genetics*; D004587:Electrophoresis, Agar Gel; D005798:Genes, Bacterial; D016106:Methicillin Resistance / Q000235:genetics*; D008826:Microbial Sensitivity Tests; D008836:Micrococcal Nuclease / Q000235:genetics*; D010068:Oxacillin / Q000633:toxicity*; D016133:Polymerase Chain Reaction / Q000379:methods*; D013210:Staphylococcus / Q000235:genetics; D013211:Staphylococcus aureus / Q000187:drug effects / Q000201:enzymology* / Q000235:genetics*
D017931:DNA Primers; D004269:DNA, Bacterial; D008836:Micrococcal Nuclease; D010068:Oxacillin