Synergy of β-Lactams with Vancomycin against Methicillin-Resistant Staphylococcus aureus: Correlation of Disk Diffusion and Checkerboard Methods

Abstract

Modified disk diffusion (MDD) and checkerboard tests were employed to assess the synergy of combinations of vancomycin and ␤-lactam antibiotics for 59 clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) and Mu50 (ATCC 700699).Bacterial inocula equivalent to 0.5 and 2.0 McFarland standard were inoculated on agar plates containing 0, 0.5, 1, and 2 g/ml of vancomycin. Oxacillin-, cefazolin-, and cefoxitin-impregnated disks were applied to the surface, and the zones of inhibition were measured at 24 h. The CLSI-recommended checkerboard method was used as a reference to detect synergy. The MICs for vancomycin were determined using the Etest method, broth microdilution, and the Vitek 2 automated system. Synergy was observed with the checkerboard method in 51% to 60% of the isolates when vancomycin was combined with any ␤-lactam. The fractional inhibitory concentration indices were significantly lower in MRSA isolates with higher vancomycin MIC combinations (P < 0.05). The overall agreement between the MDD and checkerboard methods to detect synergy in MRSA isolates with bacterial inocula equivalent to McFarland standard 0.5 were 33.0% and 62.5% for oxacillin, 45.1% and 52.4% for cefazolin, and 43.1% and 52.4% for cefoxitin when combined with 0.5 and 2 g/ml of vancomycin, respectively. Based on our study, the simple MDD method is not recommended as a replacement for the checkerboard method to detect synergy. However, it may serve as an initial screening method for the detection of potential synergy when it is not feasible to perform other labor-intensive synergy tests.V ancomycin and other glycopeptide antibiotics are currently the standard treatment for methicillin-resistant Staphylococcus aureus (MRSA) (1). The emergence of MRSA with reduced vancomycin susceptibility (SA-RVS) raises concerns about the reliability of vancomycin for the treatment of MRSA (2). Resistance during treatment with daptomycin has raised similar concerns about its effectiveness for life-threatening MRSA infections (3). Low serum levels and the bacteriostatic activity of tigecycline against MRSA limit its use for the treatment of MRSA bacteremia (4).The combination of vancomycin and ␤-lactam antibiotics offers a potential option for management of MRSA infections (5). This concept is based on several in vitro and animal studies that demonstrated a synergistic effect of vancomycin-␤-lactam combinations against MRSA (6-11).The current study was designed to determine the utility of a modified disk diffusion (MDD) test to assess synergy and antagonism for combinations of vancomycin and ␤-lactam antibiotics among clinical isolates of MRSA (8, 12). The CLSI checkerboard method was used as the reference for the determination of synergy. We determined the vancomycin MIC breakpoint where synergy is most commonly observed in MRSA isolates. MATERIALS AND METHODSBacterial isolates. Fifty-nine unique clinical isolates of MRSA with vancomycin MICs of Ն0.5 g/ml were obtained from patients at the Kaohsiung Veterans General Hospital in Taiwan ...

Comparison of methods for the detection of in vitro synergy in multidrug-resistant gram-negative bacteria

“…In addition, some studies have shown the benefit of combined therapy despite the resistance of the microorganism to some antimicrobials in the scheme [24]. Synergism has also been described using DA and MIC: MIC methods against Gram-positive organisms-especially S. aureus [25][26][27]; these are likely useful methods against Gram-positive organisms. Unfortunately, for logistical reasons, we could not evaluate in vitro synergy against Gram-positive standard organisms.…”

Comparison of methods for the detection of in vitro synergy in multidrug-resistant Gram-negative bacteria

“…In addition, some studies have shown the benefit of combined therapy despite the resistance of the microorganism to some antimicrobials in the scheme [34]. Synergism has also been described using DA and MIC:MIC methods against Gram-positive organisms -especially S. aureus [35][36][37]; these are likely useful methods against Gram-positive organisms.…”

Comparison of methods for detection of in vitro synergy in multidrug-resistant Gram-negative bacteria

“…These authors evaluated a combination of levofloxacin and piperacillin/tazobactam, finding 77% of agreement between the methods. Synergism has been described using DA and MIC:MIC methods against Gram-positive organisms, especially S. aureus [33][34][35]; likely useful methods against Gram-positive organisms. Unfortunately, because of logistic reasons, we could not evaluate in vitro synergy against Gram-positive standard organisms.…”