Evaluation of MRSA-Screen, a simple anti-PBP 2a slide latex agglutination kit, for rapid detection of methicillin resistance in Staphylococcus aureus.
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The MRSA-Screen test (Denka Seiken Co., Ltd., Tokyo, Japan), consisting of a slide latex agglutination kit that detects PBP 2a with a monoclonal antibody, was blindly compared to the oxacillin disk diffusion test, the oxacillin-salt agar screen, and PCR of the mecA gene for the detection of methicillin resistance in Staphylococcus aureus. A total of 120 methicillin-susceptible S. aureus (MSSA) and 80 methicillin-resistant S. aureus (MRSA) isolates, defined by the absence or presence of the mecA gene, respectively, were tested. The MRSA-Screen test, the oxacillin disk diffusion test, and the oxacillin-salt agar screening test showed sensitivities of 100, 61.3, and 82.5% and specificities of 99.2, 96.7, and 98.3%, respectively. We conclude that the MRSA-Screen is a very accurate, reliable, and fast test (15 min) for differentiation of MRSA from MSSA colonies on agar plates. (+info)
Misclassification of susceptible strains of Staphylococcus aureus as methicillin-resistant S. aureus By a rapid automated susceptibility testing system.
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Eight Staphylococcus aureus strains initially identified by Vitek GPS-BS or GPS-SA cards as resistant to oxacillin, but susceptible to most non-beta-lactam antibiotics, were found on further testing to be susceptible to oxacillin and ceftizoxime by disk diffusion tests. For all these strains, the MICs of oxacillin were +info)
Oxacillin susceptibility testing of staphylococci directly from Bactec Plus blood cultures by the BBL Crystal MRSA ID system.
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The BBL Crystal MRSA ID test (Becton Dickinson) was applied directly to blood culture vials containing clusters of gram-positive cocci. The sensitivity and specificity of the test were 84 and 100% and 54 and 100% for vials containing Staphylococcus aureus and coagulase-negative staphylococci, respectively. This test is a reliable method for direct detection of methicillin resistance in positive blood culture vials when S. aureus is identified in parallel by rapid identification procedures. (+info)
Disk with high oxacillin content discriminates between methicillin-resistant and borderline methicillin-susceptible Staphylococcus aureus strains in disk diffusion assays using a low salt concentration.
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A separation between mecA+ strains of Staphylococcus aureus and strains lacking mecA was achieved by the disk diffusion assay and the agar dilution method, utilizing disks containing 5 microg of oxacillin and inocula of approximately 5 x 10(5) CFU/spot, respectively, provided that agar with 0 to 0.5% NaCl and incubation at 30 degrees C were employed. The 5-microg oxacillin disks clearly discriminated between borderline methicillin-susceptible and mecA+ strains. The oxacillin MICs were more affected by the inoculum density and salt concentration than were the methicillin MICs, and oxacillin MICs of 4 to 16 microg/ml were obtained for strains lacking mecA. Significantly higher levels of beta-lactamase production and reduced oxacillin susceptibilities were recorded for strains lacking mecA, in particular strains of phage group V, when agar with >/=2% NaCl was used than when agar with 0 to 0.5% NaCl was employed. The results indicate that the borderline methicillin-susceptible phenotype is a salt-dependent in vitro phenomenon of questionable clinical relevance. (+info)
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.
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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. (+info)
Topoisomerase sequences of coagulase-negative staphylococcal isolates resistant to ciprofloxacin or trovafloxacin.
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Coagulase-negative staphylococcal isolates (n = 188) were screened for susceptibility to oxacillin, ciprofloxacin, and trovafloxacin, a new fluoroquinolone. At an oxacillin concentration of >/=4 microg/ml, 43% were methicillin resistant; of these, 70% were ciprofloxacin resistant (MIC, >/=4 microg/ml). Of the methicillin-resistant, ciprofloxacin-resistant isolates, 46% were susceptible to /=8 microg/ml) and increased trovafloxacin MICs (0.25 to 2 microg/ml) could be conferred by the combined presence of single mutations in each gyrA and grlA gene. Trovafloxacin MICs of >/=8 microg/ml also occurred, but these required an additional mutation in grlA. (+info)
Comparative efficacies of liposomal amikacin (MiKasome) plus oxacillin versus conventional amikacin plus oxacillin in experimental endocarditis induced by Staphylococcus aureus: microbiological and echocardiographic analyses.
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Optimal treatment strategies for serious infections caused by Staphylococcus aureus have not been fully characterized. The combination of a beta-lactam plus an aminoglycoside can act synergistically against S. aureus in vitro and in vivo. MiKasome, a new liposome-encapsulated formulation of conventional amikacin, significantly prolongs serum half-life (t1/2) and increases the area under the concentration-time curve (AUC) compared to free amikacin. Microbiologic efficacy and left ventricular function, as assessed by echocardiography, were compared in animals administered either oxacillin alone or oxacillin in combination with conventional amikacin or MiKasome in a rabbit model of experimental endocarditis due to S. aureus. In vitro, oxacillin, combined with either free amikacin or MiKasome, prevented the bacterial regrowth observed with aminoglycosides alone at 24 h of incubation. Rabbits with S. aureus endocarditis were treated with either oxacillin alone (50 mg/kg, given intramuscularly three times daily), oxacillin plus daily amikacin (27 mg/kg, given intravenously twice daily), or oxacillin plus intermittent MiKasome (160 mg/kg, given intravenously, a single dose on days 1 and 4). The oxacillin-alone dosage represents a subtherapeutic regimen against the infecting strain in the endocarditis model (L. Hirano and A. S. Bayer, Antimicrob. Agents Chemother. 35:685-690, 1991), thus allowing recognition of any enhanced bactericidal effects between oxacillin and either aminoglycoside formulation. Treatment was administered for either 3 or 6 days, and animals were sacrificed after each of these time points or at 5 days after a 6-day treatment course (to evaluate for posttherapy relapse). Left ventricular function was analyzed by utilizing serial transthoracic echocardiography during treatment and posttherapy by measurement of left ventricular fractional shortening. At all sacrifice times, both combination regimens significantly reduced S. aureus vegetation counts versus control counts (P < 0.05). In contrast, oxacillin alone did not significantly reduce S. aureus vegetation counts after 3 days of therapy. Furthermore, at this time point, the two combinations were significantly more effective than oxacillin alone (P < 0.05). All three regimens were effective in significantly decreasing bacterial counts in the myocardium during and after therapy compared to controls (P < 0.05). In kidney and spleen abscesses, all regimens significantly reduced bacterial counts during therapy (P < 0.0001); however, only the combination regimens prevented bacteriologic relapse in these organs posttherapy. By echocardiographic analysis, both combination regimens yielded a significant physiological benefit by maintaining normal left ventricular function during treatment and posttherapy compared with oxacillin alone (P < 0.001). These results suggest that the use of intermittent MiKasome (similar to daily conventional amikacin) enhances the in vivo bactericidal effects of oxacillin in a severe S. aureus infection model and preserves selected physiological functions in target end organs. (+info)
Methicillin-resistant Staphylococcus aureus clonal types in the Czech Republic.
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Molecular surveillance studies have documented the extensive spread of methicillin-resistant Staphylococcus aureus (MRSA) clones. Studies carried out by Centro de Epidemiologia Molecular-Network for Tracking Gram-Positive Pathogenic Bacteria (CEM/NET) led to the identification of two international multidrug-resistant strains, which were designated as the Iberian and Brazilian MRSA clones and which were defined by multiple genomic typing methods; these included ClaI restriction digests hybridized with mecA- and Tn554-specific DNA probes and pulsed-field gel electrophoresis (PFGE). The genotypic characteristics of these clones are distinct: the Iberian clone is defined as mecA type I, Tn554 type E (or its variants), and PFGE pattern A (I:E:A), whereas the Brazilian clone is defined as mecA type XI (or its variants), Tn554 type B, and PFGE pattern B (XI:B:B). In this study, we characterized 59 single-patient isolates of MRSA collected during 1996 and 1997 at seven hospitals located in Prague and five other cities in the Czech Republic by using the methodologies mentioned above and by using ribotyping of EcoRI and HindIII digests hybridized with a 16S-23S DNA probe. The Brazilian MRSA clone (XI:B:B) was the major clone (80%) spread in two hospitals located in Prague and one located in Brno; the Iberian MRSA clone (I:E:A or its variant I:DD:A), although less representative (12%), was detected in two hospitals, one in Prague and the other in Plzen. Almost all the strains belonging to clone XI:B:B (45 of 47) corresponded to a unique ribotype, E1H1, whereas most strains of the I:E:A and I:DD:A clonal types (6 of 7) corresponded to ribotype E2H2. (+info)