Evaluation of current methods for detection of staphylococci with reduced susceptibility to glycopeptides

Abstract

The sensitivity and specificity of seven methods (agar dilution, broth microdilution, Etest at 0.5 and 2.0 McFarland (McF) inocula, two agar screening methods, and population studies [PS]) were evaluated in a double-blind study involving 284 methicillin-resistant Staphylococcus aureus (MRSA) strains and 45 Staphylococcus strains with reduced susceptibilities to vancomycin (SRSV). The results were compared to the population analysis profile-area under the curve ratio method (PAP-AUC ratio compared to that of Mu3) as described by Wootton et al. The agar screening method using brain heart infusion agar (6 microg of vancomycin per ml) gave a sensitivity of 22% and a specificity of 97%. A similar method using Mueller-Hinton agar (5 microg of vancomycin per ml) gave a sensitivity of 20% and a specificity of 99%. The PS method detected 34 false positives (12%) and gave a sensitivity of 71% and a specificity of 88%. Etest using 0.5 and 2.0 McF inocula gave sensitivities and specificities of 82 and 93% and of 96 and 97%, respectively. The best Etest interpretative criteria for the 2.0 McF inoculum was > or =8 mg of vancomycin per liter and > or =8 microg teicoplanin per ml or > or =12 microg of teicoplanin per ml. The direct colony suspension inoculum for this method was found to be equally accurate in detecting (hetero-)glycopeptide-intermediate S. aureus compared to the overnight broth inoculum preparation method. Agar dilution and broth microdilution using the NCCLS breakpoint criteria for vancomycin gave sensitivities and specificities of 20 and 100% and of 11 and 100%, respectively. Using the Etest with a 2.0 McF inoculum, six different media were assessed against a selection of SRSV (n = 48) and MRSA (n = 12). Brain heart infusion agar yielded the highest sensitivity and specificity values: 88 and 88%, respectively.

FIG. 1

Typical Etest result when using the macromethod of Mu50 possessing a PAP-AUC of 1.51. The microcolonies appear only after the plates have been incubated for 48 h. In this particular instance and under these conditions, cells can grow at up to a vancomycin concentration of 48 μg/ml.

FIG. 2

Correlation of agar dilution vancomycin MIC values compared to PAP-AUCs. The demarcation line shown delineates hGIS and GIS based on repeated PAP-AUC ratios from strain Mu50. Strains possessing a PAP-AUC ratio of >1.3 have populations as resistant as Mu50 and, by this method, have been characterized as GIS. In some instances, strains with a significant PAP-AUC ratio had a vancomycin MIC of only 4 μg/ml.