New Real-Time PCR Assay for Rapid Detection of Methicillin- ResistantStaphylococcus aureusDirectly from Specimens Containing a Mixture of Staphylococci
Open Access
- 1 May 2004
- journal article
- research article
- Published by American Society for Microbiology in Journal of Clinical Microbiology
- Vol. 42 (5), 1875-1884
- https://doi.org/10.1128/jcm.42.5.1875-1884.2004
Abstract
Molecular methods for the rapid identification of methicillin-resistant Staphylococcus aureus (MRSA) are generally based on the detection of an S. aureus-specific gene target and the mecA gene. However, such methods cannot be applied for the direct detection of MRSA from nonsterile specimens such as nasal samples without the previous isolation, capture, or enrichment of MRSA because these samples often contain both coagulase-negative staphylococci (CoNS) and S. aureus, either of which can carry mecA. In this study, we describe a real-time multiplex PCR assay which allows the detection of MRSA directly from clinical specimens containing a mixture of staphylococci in mec (SCCmec) right extremity sequences, including three new sequences, were used in combination with a primer and three molecular beacon probes specific to the S. aureus chromosomal orfX gene sequences located to the right of the SCCmec integration site. Of the 1,657 MRSA isolates tested, 1,636 (98.7%) were detected with the PCR assay, whereas 26 of 569 (4.6%) methicillin-susceptible S. aureus (MSSA) strains were misidentified as MRSA. None of the 62 nonstaphylococcal bacterial species or the 212 methicillin-resistant or 74 methicillin-susceptible CoNS strains (MRCoNS and MSCoNS, respectively) were detected by the assay. The amplification of MRSA was not inhibited in the presence of high copy numbers of MSSA, MRCoNS, or MSCoNS. The analytical sensitivity of the PCR assay, as evaluated with MRSA-negative nasal specimens containing a mixture of MSSA, MRCoNS, and MSCoNS spiked with MRSA, was ∼25 CFU per nasal sample. This real-time PCR assay represents a rapid and powerful method which can be used for the detection of MRSA directly from specimens containing a mixture of staphylococci.Keywords
This publication has 53 references indexed in Scilit:
- SHEA Guideline for Preventing Nosocomial Transmission of Multidrug-Resistant Strains ofStaphylococcus aureusandEnterococcusInfection Control & Hospital Epidemiology, 2003
- Rapid Detection of Clostridium difficile in Feces by Real-Time PCRJournal of Clinical Microbiology, 2003
- Comparison of Mortality Associated with Methicillin‐Resistant and Methicillin‐SusceptibleStaphylococcus aureusBacteremia: A Meta‐analysisClinical Infectious Diseases, 2003
- Toward rapid real-time molecular diagnostic to guide smart use of antimicrobialsCurrent Opinion in Microbiology, 2002
- Rapid Identification of Staphylococcus aureus and the mecA Gene from BacT/ALERT Blood Culture Bottles by Using the LightCycler SystemJournal of Clinical Microbiology, 2002
- Rapid Identification of Methicillin-Resistant Staphylococcus aureus from Positive Blood Cultures by Real-Time Fluorescence PCRJournal of Clinical Microbiology, 2001
- Epidemiological Study of Staphylococcal Colonization and Cross-Infection in Two West African HospitalsMicrobial Drug Resistance, 2000
- Molecular Typing of Methicillin-ResistantStaphylococcus aureusby Pulsed-Field Gel Electrophoresis: Comparison of Results Obtained in a Multilaboratory Effort Using Identical Protocols and MRSA StrainsMicrobial Drug Resistance, 2000
- Resistance Rather Than Virulence Selects for the Clonal Spread of Methicillin-ResistantStaphylococcus aureus:Implications for MRSA TransmissionMicrobial Drug Resistance, 2000
- Induced deletions within a cluster of resistance genes in the mec region of the chromosome of Staphylococcus aureusJournal of General Microbiology, 1990