Evaluation of commercial susceptibility testing methods for in vitro determining daptomycin susceptibility in vancomycin-resistant Enterococcus faecium

Vancomycin-resistant Enterococcus faecium (VR Efm ) is a major cause of invasive healthcare-associated infections. Daptomycin is an important treatment option, but its efficacy may be compromised for isolates with minimum inhibitory concentrations (MICs) of 4–8 mg/L, even at high doses. EUCAST has therefore assigned an “insufficient evidence” category for Enterococcus spp., making accurate MIC determination essential. We compared the performance of UMIC Daptomycin, several automated broth microdilution (BMD) systems, and E-test against the reference BMD method. Eighty-nine VR Efm isolates with borderline daptomycin susceptibility were tested by standard BMD in calcium-adjusted Mueller–Hinton broth and by Phoenix, EUVENC, VITEK-2, UMIC Daptomycin, and E-test. CLSI breakpoints (susceptible-dose dependent ≤ 4 mg/L, R > 4 mg/L) were applied. Essential agreement (EA), categorical agreement (CA), bias, very major errors (vMEs), and major errors were assessed. Phoenix (EA = 92%, CA = 69%) and EUVENC (EA = 87.6%, CA = 74.2%) were the most accurate. UMIC achieved EA = 74.2% and CA = 70.8%. E-test (EA = 68.5%, CA = 66.3%) and VITEK-2 (EA = 67.4%, CA = 64%) showed lower accuracy. All methods displayed negative bias, underestimating MICs and frequently misclassifying isolates with MIC = 8 mg/L, resulting in high vME rates. None of the methods met ISO-20776-2:2021 criteria. Automated BMD systems performed best, but all showed systematic MIC underestimation near the breakpoint. For reliable detection and confirmation of daptomycin resistance in VR Efm , BMD-based MIC methods—manual or automated—are recommended. IMPORTANCE Vancomycin-resistant Enterococcus faecium (VR Efm ) is a major cause of hospital-acquired infections worldwide, posing a serious threat to patient safety and infection control. Daptomycin remains one of the few therapeutic options for severe VR Efm infections, yet its efficacy depends on accurately determining bacterial susceptibility. Even small variations in measured minimum inhibitory concentrations can influence treatment success or failure. In this study, we systematically evaluated several commercial antimicrobial susceptibility testing methods for daptomycin against VR Efm and compared them with the reference broth microdilution (BMD) method. Our results show that many commonly used systems underestimate resistance, particularly for isolates near the clinical breakpoint. This underestimation may lead to inappropriate treatment decisions. The study provides evidence-based recommendations for clinical laboratories and emphasizes the importance of confirmatory BMD testing to ensure reliable results, optimize patient management, and prevent the further spread of antimicrobial resistance.