Synergistic essential oil-antibiotic combinations as a strategy to maintain the efficacy of current antibiotics against Enterococcus sp.

Objectives This study aimed to investigate synergistic combinations of vancomycin and Essential Oils (EOs) as potential antimicrobial formulations against Vancomycin-Sensitive Enterococcus faecium (VSE) and Vancomycin-Resistant E. faecium (VRE). This was to be achieved by (1) screening vancomycin and EOs for synergistic interactions, (2) determining the efficacy of lead combinations against VRE clinical isolates, and (3) establishing the in vivo efficacy of the combination.

Methods Antimicrobial interactions between vancomycin and the EOs oregano, cumin and rosewood and EO compounds carvacrol, cuminaldehyde and linalool were determined using the checkerboard method. Combinations with synergistic activity were then investigated by a time-kill assay. Efficacy of a lead combination against 10 VRE clinical isolates was determined by measuring bacterial survival after 24 hours’ contact the combination. The in vivo efficacy of the lead combination was determined using a G. mellonella larvae assay. The combination was injected into the haemolymph of larvae previously inoculated with VRE. Larvae were incubated at 37°C and mortality was recorded over 5 days.

Results Synergistic activity against VSE and VRE was observed for a combination of 4.20 mM cuminaldehyde, 1.98 mM carvacrol and 0.031 mg/L vancomycin. The minimum inhibitory concentration of vancomycin in this combination was below the EUCAST (2018) breakpoint for vancomycin susceptibility in Enterococcus sp. The combination reduced VSE by 5.83 log10 and VRE by 5.07 log10 Colony Forming Units (CFU)/mL over 24 hours. The combination was also antimicrobial against 10 VRE clinical isolates, with 2.33-5.25 log10 CFU/mL reductions observed. However, the combination did not significantly (p<0.05) improve survival of larvae compared to controls.

Conclusions A combination of carvacrol, cuminaldehyde and vancomycin was synergistic against VSE and VRE, reducing the effective dose of vancomycin. As such, combining EOs and antibiotics could be a useful strategy to maintain the current antibiotic repertoire against antibiotic resistant bacteria. However, the lead combination was not significantly antimicrobial in vivo, suggesting further drug development is required for this combination to be useful.

References EUCAST (2018). Breakpoint tables for interpretation of MICs and zone diameters, Version 8.1. Vaxjo, Sweden: EUCAST.