An optimal method for generating stable vancomycin heteroresistance and intermediate susceptibility in methicillin-resistant Staphylococcus aureusblood isolates under in-vitro vancomycin pressure

Objectives: The relevance of vancomycin intermediately-susceptible (VISA) and hetero-resistant (hVISA) methicillin-resistantStaphylococcus aureus (MRSA) remains uncertain because of their low frequency. In vitro attempts to generate reducedsusceptibility have been inconsistent. We describe a simple method for generating VISA/hVISA.Materials and methods: Twenty-four SCCmec type II and IV MRSA blood isolates plus USA100 and USA300 controls werecultured (107 CFU/ml) in BHI broth with 0, 2 and 3 mg/L vancomycin for 10 days followed by 10 days vancomycin-freepassage.We monitored vancomycin minimum inhibitory concentration (MIC; Etest) and population analysis profile-areaunder the curve (PAP-AUC) ratios of tested isolate-Mu3 during and after vancomycin pressure (VP). PAP-AUC ratios <0.9,0.9-1.3 and >1.3 were considered consistent with susceptible, hVISA and VISA, respectively.Results: VP at 2 mg/L (VP-2) increased MIC to 3 mg/L in 20 (83.3%) isolates and raised PAP-AUC ratio to hVISA (n=18; 75.0%)and VISA (six; 25.0%) ranges. VP-3 increased MIC to 3 and 4 mg/L in 19 (79.2%) and three (12.5%) isolates, respectively andraised PAP-AUC ratio to hVISA (n=3; 12.5%) and VISA (n=19; 79.2%) ranges. SCCmec IV isolates had lower pre-exposurePAP-AUC ratios (0.48±0.14 vs. 0.69±0.10; p<0.001) and lower MIC (1.6±0.2 vs. 1.8±0.2 mg/L; p=0.1) but MIC rise and VISA/hVISA emergence was comparable. MIC and PAP-AUC ratio rises were stable in 20/22 isolates during drug-free passages.Conclusions: VISA/hVISA readily emerges among MRSA SCCmec type II and IV isolates under VP. VP at the MIC level generateshVISA while pressure at slightly higher level results in the VISA phenotype. J Microbiol Infect Dis 2012; 2(4): 129-134Key words: Staphylococcus aureus; heteroresistance; glycopeptides; resistance; bacteremia.