BACKGROUND: Daptomycin is a novel cyclic lipopeptide antibiotic that exhibits in vitro bactericidal activity against gram-positive pathogens including methicillin-resistant staphylococci and vancomycin-resistant enterococci. The aim of this study is to determine the in vitro activities of daptomycin against recent clinical isolates of methicillin-resistant staphylococci and vancomycin-resistant enterococci in Korea. MATERIALS AND METHODS: A total of 117 clinical strains of methicillin-resistant staphylococci and vancomycin-resistant enterococci were isolated at a tertiary-care hospital in Korea in 2004. Susceptibility to daptomycin was tested by the CLSI broth microdilution method using Mueller-Hinton broth (MHB) which was adjusted to contain a final concentration of 50 microgram/mL of ionized calcium (Ca2+). Susceptibilities to ampicillin, oxacillin, levofloxacin, vancomycin, and linezolid were tested by the CLSI agar dilution method. RESULTS: All isolates of methicillin-resistant S. aureus and coagulase-negative staphylococci were inhibited by 1 microgram/mL of daptomycin, and MIC90s were 1 microgram/mL, which were similar to those of vancomycin and linezolid. MIC90s of daptomycin for vancomycin-resistant E. faecalis and E. faecium were 0.5 microgram/mL and 2 microgram/mL, respectively, and all isolates were susceptible to daptomycin. MIC90s of linezolid and levofloxacin for vancomycin-resistant enterococci were 1-2 microgram/mL and 64 microgram/mL, respectively. Resistance rates of vancomycin-resistant E. faecalis and E. faecium to levofloxacin were 100% and 96%, respectively. Daptomycin MICs in MHB supplemented to 20-25 microgram/ml of Ca2+ were 2-8 fold higher than those in MHB supplemented to 50 microgram/mL of Ca2+. CONCLUSION: Daptomycin is very active in vitro against methicillin-resistant staphylococci and vancomycin-resistant enterococci isolated in Korea, and it is important to test in vitro activity of daptomycin using MHB containing 50 microgram/mL of Ca2+.