From formulation to in vivo model: A comprehensive study of a synergistic relationship between vancomycin, carvacrol and cuminaldehyde against Enterococcus faecium

Vancomycin Resistant Enterococcus faecium (VRE) has become endemic in healthcare settings, reducing treatment options for enterococcal infections. New antimicrobials for VRE infections are a high priority, but the development of novel antibiotics is time-consuming and expensive. Essential Oils (EOs) synergistically enhance the activity of some existing antibiotics, suggesting that EO-antibiotic combinations could re-sensitise resistant bacteria and maintain the antibiotic repertoire. The mechanism of re-sensitisation of bacteria to antibiotics by EOs is relatively understudied. Here, the synergistic interactions between carvacrol (1.98 mM) and cuminaldehyde (4.20 mM) were shown to re-establish susceptibility to vancomycin (0.031 mg/L) in VRE, resulting in bactericidal activity (4.73 log10 CFU/mL reduction). Gene expression profiling, coupled with β-galactosidase leakage and salt tolerance assays suggested that cell envelope damage contributes to the synergistic bactericidal effect against VRE. The EO-vancomycin combination was also shown to kill clinical isolates of VRE (2.33-5.25 log10 CFU/mL reduction) and stable resistance did not appear to develop even after multiple passages. The in vivo efficacy of the EO-vancomycin combination was tested in a Galleria mellonella larvae assay; however no antimicrobial action was observed, indicating that further drug development is required for the EO-vancomycin combination to be clinically useful for treatment of VRE infections.