Abstract
Background: Thymol is a phenolic compound used for its wide spectrum antimicrobial activity. There is a limited understanding of the antimicrobial mechanisms underlying thymol activity. To investigate this, E. coli strain JM109 was exposed to thymol at sub-lethal concentrations and after 16 rounds of exposure, isolates with a 2-fold increased minimal inhibitory concentration (MIC) were recovered (JM109-Thyr). The phenotype was stable after multiple sub-cultures without thymol. Results: Cell morphology studies by scanning electron microscopy (SEM) suggest that thymol renders bacterial cell membranes permeable and disrupts cellular integrity. 1H Nuclear magnetic resonance (NMR) data showed an increase in lactate and the lactic acid family amino acids in the wild type and JM109-Thyr in the presence of thymol, indicating a shift from aerobic respiration to fermentation. Sequencing of JM109-Thyr defined multiple mutations including a stop mutation in the acrR gene resulting in a truncation of the repressor of the AcrAB efflux pump. AcrAB is a multiprotein complex traversing the cytoplasmic and outer membrane, and is involved in antibiotic clearance. Conclusions: Our data suggests that thymol tolerance in E. coli induces morphological, metabolic and genetic changes to adapt to thymol antimicrobial activity. Keywords: Escherichia coli, Thymol, Resistance, Acriflavine resistance regulator, Efflux pump