Successful therapy of experimental endocarditis caused by vancomycin-resistant Staphylococcus aureus with a combination of vancomycin and beta-lactam antibiotics

Abstract

VRS1 is the first isolated strain of vancomycin-resistant Staphylococcus aureus (VRSA) found to carry the vanA gene complex previously described in Enterococcus. Under vancomycin pressure, VRS1 makes aberrant cell walls consisting of stem tetrapeptide and depsipeptide that lack the terminal D-Ala-D-Ala residues targeted by vancomycin. Previous data have suggested that this aberrant cell wall is not cross-linked by PBP2a, the enzyme responsible for cell wall transpeptidation in the presence of beta-lactam antibiotics. We examined the efficacy of treating VRS1 with a combination of vancomycin and beta-lactam antibiotics in vitro and in vivo. We found that the MIC of oxacillin for VRS1 decreased from >256 microg/ml to <1 microg/ml in the presence of vancomycin. Using the rabbit model of endocarditis, we treated VRS1-infected rabbits with nafcillin alone, vancomycin alone, or a combination of nafcillin and vancomycin. Treatment with nafcillin in combination with vancomycin cleared bloodstream infections within 24 h and sterilized 12/13 spleens (92%), as well as 8/13 kidneys (62%), following 3 days of treatment. Mean aortic valve vegetation counts were reduced 3.48 log(10) CFU/g with the combination therapy (compared to untreated controls) and were significantly lower than with either vancomycin or nafcillin given alone. VRS1 was extremely virulent in this model, as no untreated rabbits survived the 3-day trial. Treatment of clinical infections due to VRSA with the combination of vancomycin and beta-lactams may be an option, based on these results.

FIG. 1.

Susceptibility testing of VRS1. An overnight culture of VRS1 grown in the presence (D) or absence (A, B, and C) of 32 μg/ml vancomycin was diluted to 0.5 McFarland standard and swabbed onto brain heart infusion agar with (C and D) or without (A and B) 16 μg/ml vancomycin. Oxacillin (A, C, and D) or vancomycin (B) Etest strips were placed on the plates, and the plates were incubated at 37°C for 24 h. The marks within the zone of clearing on plates C and D are not colonies.

FIG. 2.

Time-kill curves for VRS1. VRS1 was grown in the absence of antibiotic for 2 h. Antibiotics were added at 0 h. Control (⧫), nafcillin at 25 μg/ml (▴), vancomycin at 30 μg/ml (▪), and nafcillin at 25 μg/ml in combination with vancomycin at 30 μg/ml (×) are shown.

FIG. 3.

Bacterial counts in valve vegetations after 3- or 7-day treatment. Control (⧫), nafcillin at 25 μg/ml (▴), vancomycin at 30 μg/ml (▪), and nafcillin at 25 μg/ml in combination with vancomycin at 30 μg/ml (×) are shown. The horizontal lines represent the mean log₁₀ CFU/gram. Each symbol represents one rabbit. *, P < 0.05 compared to the control group. **, P < 0.05 compared to the control, nafcillin, or vancomycin group. ***, P < 0.05 compared to the vancomycin group (7-day).

FIG. 4.

Percent positive blood cultures by treatment day for rabbits with endocarditis caused by VRS1. Control (⧫), nafcillin at 25 μg/ml (▴), vancomycin at 30 μg/ml (▪), and nafcillin at 25 μg/ml in combination with vancomycin at 30 μg/ml (×) are shown.

FIG. 5.

Survival curves for rabbits with endocarditis caused by VRS1. Control (⧫), nafcillin at 25 μg/ml (▴), vancomycin at 30 μg/ml (▪), and nafcillin at 25 μg/ml in combination with vancomycin at 30 μg/ml (×) are shown.