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Two genes recently associated with glycopeptide intermediate resistance in Staphylococcus aureus (GISA) are mprF and tcaA, with inactivation causing shifts in vancomycin resistance. This study reveals that expression levels of both genes are similar in groups of clinical GISA, heteroGISA and glycopeptide susceptible strains, suggesting no association with clinical isolates.

The detection of methicillin-resistant S. aureus (SA) (MRSA) refractory to glycopeptides is a serious clinical issue. The prevalence of hetero-resistant GISA (hGISA) strains at H. Maréchal Joffre, France is reported.858 non-repeat SA were isolated during 1999. 367 (43%) of these, from 257 patients, were MRSA (mean incidence 11.9/1000 admissions). All MSRA detected during 1999 were screened for vancomycin (VAN) resistance (BHI+4 mg/l VAN). Isolates recovered were retested using Etest strips (2 McFarland inoculum on BHI) and population analysis profile/area under the curve (PAP-AUC) analysis with hGISA SA Mu3 as a comparator. 58 selected strains were screened for teicoplanin resistance(TEI) using SFM recommended screen (2 McFarland inoculum on MH+5 mg/L TEI) and MIC (0.5 MF inoculum swabbed on MH agar) methods. 188 (51.3%) grew on VAN screen agar (6.1/1000 admissions). 58 strains (7.6%) possessed Etest VAN MIC > 8 mg/l all others being VAN < 8 mg/l. Of these 58 isolates, 10 were stably heterogeneously resistant to both VAN and teicoplanin (MIC > 8 mg/l). PAP-AUC showed 12 strains to have PAP-AUC ratios > 0.95 but < 1.5 (ie. hGISA, not GISA). All 7 isolates defined as hGISA by both Etest and PAP-AUC comprised 1 PFGE clone (< 3 bands difference). Additionally 2 distinct PFGE types were detected among the other 5 hGISA identified PAP-AUC. The 12 hGISAs, were derived from 12 patients with severe underlying disease. None were on glycopeptide therapy prior to hGISA isolation. This is the first report of endemic hGISA, comprising 3 clonal types. The isolation of hVISA seems not to be associated with patient-specific glycopeptide therapies.

Heterogeneous vancomycin-intermediate Staphylococcus aureus (hVISA) may account for treatment failure with vancomycin and act as a precursor of vancomycin-intermediate or -resistant S. aureus. The activity of vancomycin was assessed against vancomycinsusceptible, hVISA and VISA strains in a dilutional pharmacokinetic model. Over a 48 h period, total bacteria and cells with a vancomycin-intermediate phenotype were quantified. Total counts of hVISA were reduced by vancomycin in a similar way to a vancomycin-susceptible control. The vancomycin-intermediate sub-population was eradicated from the model within one dose interval. Exposure to low vancomycin concentrations did not result in an increase in the proportion of cells which were vancomycin intermediate. Short-term exposure of hVISA to vancomycin at gradient concentrations did not increase the proportion of cells with vancomycin-intermediate phenotype.

One hundred methicillin-resistant Staphylococcus aureus (MRSA) strains, isolated between 1983 and 1999, were tested alongside the vancomycin hetero-resistant S. aureus (hVRSA) strain Mu 3, and vancomycin-resistant S. aureus (VRSA) strain Mu 50, for their resistance to vancomycin. This was achieved using the screening method described by Hiramatsu, gradient plates, agar incorporation, standard Etest, macrodilution Etest and a modified population analysis. Using Hiramatsu's screening method, 5% of the 100 MRSA were identified as VRSA and 5% identified as hVRSA, the gradient plates identified 7% hVRSA, and the standard and macrodilution Etests identified no hVRSA. Mu 3 appeared to be vancomycin-susceptible using both the agar incorporation and standard Etest methods, but was classified as hVRSA using the macrodilution Etest. The modified population analysis reliably detected vancomycin hetero-resistance in Mu 3 and identified no hVRSAs within the 100 MRSA sample.

Isolates of Staphylococcus aureus resistant to vancomycin have been reported but appear to be extremely rare. However, isolates displaying hetero-resistance to vancomycin (hVRSA) are reportedly common in parts of Japan (9.3% of MRSA isolated from a group of university hospitals). We have investigated the reliability of the proposed method for detection of hetero-resistant isolates and the ability of clinical S. aureus isolates to express vancomycin resistance. The original method for identification of hVRSA was found to have poor reproducibility and may select for, rather than detect, vancomycin resistance. There appears to be a spectrum of heterogeneity in the expression of resistance to vancomycin among S. aureus. Until there is a clearer understanding of the mechanism and control of vancomycin resistance in S. aureus, and reliable tests are devised, the clinical relevance of different degrees of hetero-resistance cannot be assessed.

Vancomycin-sensitive, -intermediate, and -heterointermediate methicillin-resistant Staphylococcus aureus isolates were tested by using E-tests to explore the interaction of methicillin and vancomycin. For the vancomycin-intermediate and -heterointermediate strains both drugs showed antagonism at concentrations below their MICs but synergy at methicillin concentrations near the MIC. This property could be used to screen for heterointermediate S. aureus strains.