How efflux pumps affect the sensitivity of Escherichia coli biofilms to antibiotics

Abstract

Bacterial biofilms are an important and clinically significant growth state of bacteria. The relationship between bacterial antibiotic resistance, including the ability of a bacterial strain to resist multiple drug types, and biofilms is not well established. This research project aimed to examine efflux pumps in Escherichia coli biofilms and how they influence the antibiotic sensitivity of a biofilm. This was done with the use of two efflux pump assays, the Nile red real time assay and the Hoechst 33342 assay with comparisons to biofilm viability. The efflux pump assays implemented were modified for use on E. coli biofilms. Efflux pump inhibitors (EPI) were used in conjunction with antibiotic treatments to determine their effect on efflux pumps activity in the biofilm and cell viability. Well-established EPIs of the AcrABTolC pump, CCCP and PAI²N, were used in this research along with the novel inhibitor, capsaicin. The effects of efflux pump inhibitors and antibiotics on efflux pump transporter proteins were tested using single gene deletion mutants. The data collected suggested that the efflux pump activity of E. coli biofilm cells behaves differently to planktonic cells; biofilms appear not as active as planktonic cells. In efflux pump assays, capsaicin behaved similarly to PAI²N, a competitive substrate inhibitor and was less toxic to the E. coli biofilm, PAI²N in combination with antibiotics was the most effective at E. coli biofilm eradication, however it has been reported to have toxic side effects. Capsaicin is currently used pharmacologically and it therefore is a potentially clinically significant EPI. Further, the research suggests that the effects of the sum of the many efflux pumps of E. coli results in the antibiotic resistance of the biofilm, minimising the apparent significance of any single efflux transporter.

Date of Award	2017
Original language	English