Identification of methicillin-resistant strains of staphylococci by polymerase chain reaction

Abstract

A simple and reliable method using a polymerase chain reaction (PCR) was devised to identify methicillinresistant staphylococci. By using lysates of the strain to be tested as templates and 22-mer oligonucleotides as primers, a 533-bp region of mecA, the structural gene of a low-affinity penicillin-binding protein (PBP 2'), was amplified by PCR and detected by agarose gel electrophoresis. Results obtained by this method were compared with those obtained by broth microdilution MIC determination for 210 and 100 clinical isolates of Staphylococcus aureus and coagulase-negative staphylococci, respectively. Of 99 mecA-negative S. aureus isolates, 100% of the strains were methicillin susceptible and 98% of the strains were oxacillin susceptible. Three strains (3%) of 111 mecA-positive S. aureus isolates exhibited almost the same susceptibility to ,(-lactams as the mecA-negative ones and did not produce detectable amounts of PBP 2' despite the presence of the mecA gene. One of them yielded typically methicillin-resistant variants at a low frequency with concomitant recovery of PBP 2' production. The mecA gene was also found in coagulase-negative Staphylococcus epidermidis, Staphylococcus haemolyticus, Staphylococcus sciuri, Staphylococcus saprophyticus, and Staphylococcus caprae and conferred resistance on most of the bacteria. * Corresponding author. species were identified by Staphyogram (Terumo, Tokyo, Japan), an identification kit, and the coagulase test. S. aureus SR3626, SR3633, SR3636, SR3639, SR3665, SR3681, and SR3716 were used as mecA-positive control organisms (12), and S. aureus ATCC 25923, 209P JC-1, and Smith were used as mecA-negative control organisms.

High prevalence of methicillin-resistant Staphylococcus aureus clone ST80-IV in hospital and community settings in Algiers

“…The mecA gene coding for methicillin resistance was detected by PCR as described previously [18]. The staphylococcal chromosomal cassette mec (SCCmec I-IV) was detected as described by Oliveira et al [19], and the SCCmec type V was detected as described previously [20].…”

A review of the characteristics and treatment of methicillin‐resistant Staphylococcus aureus (MRSA) in the horse and a case series of MRSA infection in four horses

“…Presence of the mecA gene can be readily verified in suspected MRSA isolates by a comparatively simple polymerase chain reaction (PCR) assay (Murakami et al 1991), or its product (PBP2a) can be detected by latex agglutination testing (Cavassini et al 1999). It should be noted that the mecA gene is not limited to S. aureus and its presence in a staphylococcal isolate does not necessarily indicate the isolate is MRSA; hence any mecA confirmed isolate should be identified to species level by PCR or other means.…”

Distribution of the exfoliative toxin D gene in clinical Staphylococcus aureus isolates in France

“…Sequences specific for staphylococcal enterotoxin genes (sea-e, seh, sek, sem, sel and seo), the toxic shock syndrome toxin gene (tst), the exfoliative toxin A and B genes (eta, etb), the Panton-Valentine leukocidin (PVL) genes (lukS-PV-lukF-PV), the LukE-LukD leukotoxin gene (lukE-lukD), the c-haemolysin gene (hlg), the c-haemolysin variant gene (hlgv) and accessory gene regulator alleles (agr1-4) were all detected by PCR as described previously [7,8]. The mecA gene coding for methicillin resistance was detected by PCR as described by Murakami et al [9]. All PCR products were resolved by electrophoresis through agarose 1.5% w ⁄ v gels, followed by staining with ethidium bromide and analysis.…”