Investigating the Effect of Temperature on Pump-Driven Antibiotic Resistance towards Erythromycin in Escherichia coli

Abstract

Antibiotic resistance is an inevitable by-product from treatment of bacterial (and fungal) infections, however the rate and intensity at which it is emerging is alarming. More and more drugs are being rendered ineffective, where poor treatment behaviours, such as overuse, are being held accountable. Coupled with a dry antibiotic pipeline, we are increasingly seeing ourselves approach a post-antibiotic era. As research is being conducted into discovering new antibiotics, we also need to find ways to preserve the ones we still have. Therefore it is equally important to identify and study potential selection pressures, both environmental and clinical, that contribute to the rise in resistance. To provide a preliminary insight on temperature as a possible selection pressure, this research project aimed to investigate the effect of temperature on the susceptibility of Escherichia coli to the macrolide-class antibiotic erythromycin, and ultimately its effect on the expression of multidrug efflux pump AcrAB-TolC. This was done by exposing E. coli cells to a range of erythromycin concentrations during 24 hours of growth, establishing a minimum inhibitory concentration (MIC) at 30°C and seeing any shifts in the MIC at 37°C. It was found that as temperature increased from 30°C to 37°C, so did the MIC. Thus the cells were more resistant at the higher temperature. Next, to see whether and how AcrAB-TolC was selected for as a result of temperature, the expression of protein AcrB was measured via fluorescence emitted from sfGFP (due to the sfGFP gene being physically fused to the acrB gene). It was found that the range of concentrations that select for pump expression, referred to as the “AcrB expression-selection window”, shifted positively with increasing temperature. This suggests a temperature-dependent nature of resistance selection, therefore this knowledge may help in choosing an effective dose for treatment based on thermal conditions. The outcomes of this research project will help provide a foundation for looking further into temperature, and other selection pressures, and their effect on the rise in antibiotic resistance.