ABSTRACT Daptomycin exhibits in vitro bactericidal activity against clinically significant gram-positive bacteria. We employed pharmacodynamic modeling to determine a once-daily dosing regimen of daptomycin that correlates to pharmacodynamic endpoints for different resistant gram-positive clinical strains. An in vitro pharmacodynamic model with an initial inoculum of 6 log 10 CFU/ml was used to simulate daptomycin regimens ranging in dose from 0 to 9 mg/kg of body weight/day, with corresponding exposures reflecting free-daptomycin concentrations in serum. Bacterial density was profiled over 48 h for two methicillin-resistant Staphylococcus aureus (MRSA-67 and -R515), two glycopeptide intermediate-resistant S. aureus (GISA-992 and -147398), and two vancomycin-resistant Enterococcus faecium (VREF-12366 and -SF12047) strains. A sigmoid dose-response model was used to estimate the effective dose required to achieve 50% (ED 50 ) and 80% (ED 80 ) bacterial density reduction at 48 h. Daptomycin MICs for study isolates ranged from 0.125 to 4 μg/ml. Model fitting resulted in an r 2 of >0.80 for all tested isolates. Control growths at 48 h ranged from 7.3 to 8.5 log 10 CFU/ml. Sigmoid relationships were not superimposable between categorical resistant species: ED 50 and ED 80 values were 1.9 and 3.1, 4.2 and 5.6, and 5.4 and 6.8 mg/kg for MRSA, GISA, and VREF isolates, respectively. Doses required to achieve ED 50 and ED 80 values correlated with MIC differences between tested organisms. Corresponding area under the concentration-time curve from 0 to 24 h/MIC exposure ratios demonstrated a wide range of ED 80 values among the tested isolates. Doses ranging between 3 and 7 mg/kg produced significant bactericidal activity (ED 80 ) against these multidrug-resistant S. aureus and E. faecium isolates.