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Hoffmann, R. A wiki for the life sciences where authorship matters. Nature Genetics (2008)
 
 
 
 
 

Comparison of susceptibility testing methods with mecA gene analysis for determining oxacillin (methicillin) resistance in clinical isolates of Staphylococcus aureus and coagulase-negative Staphylococcus spp.

Ninety-nine clinical staphylococcal isolates (58 coagulase-negative Staphylococcus spp. [CoNS] and 41 Staphylococcus aureus isolates) were evaluated for susceptibility to oxacillin. The following susceptibility testing methods, media, and incubation conditions were studied: agar dilution by using Mueller-Hinton (MH) medium (Difco) supplemented with either 0, 2, or 4% NaCl and incubation at 30 or 35 degrees C in ambient air for 24 or 48 h; disk diffusion by using commercially prepared MH medium (Difco) and MH II agar (BBL) and incubation at 35 degrees C in ambient air for 24 or 48 h; and agar screen (spot or swab inoculation) by using commercially prepared agar (Remel) or MH agar (Difco) prepared in-house, each containing 4% NaCl and 6 microg of oxacillin/ml (0.6-microg/ml oxacillin was also studied with MH agar prepared in-house for the agar swab method and CoNS isolates) and incubation at 35 degrees C in ambient air for 24 or 48 h for swab inoculation and at 30 or 35 degrees C in ambient air for 24 or 48 h for spot inoculation. The results for these methods were compared to the results for mecA gene detection by a PCR method. Given the ability to support growth and the results for susceptibility testing (the breakpoint for susceptible isolates was </=2 microg/ml), the best methods for CoNS isolates were (i) agar dilution by using MH medium supplemented with 4% NaCl and incubation at 35 degrees C for 48 h (no growth failures were noted, and sensitivity was 97.6%) and (ii) agar screen (swab inoculation) by using MH medium prepared in-house supplemented with 4% NaCl and containing 0.6 microg oxacillin/ml and incubation at 35 degrees C for 48 h (one isolate that did not carry the mecA gene did not grow, and the sensitivity was 100%). All but one (agar dilution without added NaCl and incubation at 30 degrees C for 48 h) of the methods tested revealed all oxacillin-resistant S. aureus isolates, and no growth failures occurred with any method. If the breakpoint for susceptibility was lowered to </=1 microg/ml for agar dilution methods, more CoNS isolates with oxacillin resistance related to the mecA gene were detected when 0 or 2% NaCl agar supplementation was used. Only one CoNS isolate with mecA gene-associated resistance was not detected by using agar dilution and MH medium supplemented with 4% NaCl with incubation for 48 h. When the breakpoint for susceptibility was decreased 10-fold (from 6.0 to 0.6 microg of oxacillin per ml) for the agar swab screen method, fully 100% of the CoNS isolates that carried the mecA gene were identified.[1]

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