AcrA and TolC are important Efflux Components in the development of low level Adaptive Aminoglycoside Resistance in Escherichia coli K-12 Following Sub-inhibitory Kanamycin Pre-Treatment

Abstract

Escherichia coli K-12 contains a number of inducible drug efflux complexes such as the well characterized AcrAB- TolC complex. Drug efflux complexes are important when considering the mechanisms of antibiotic resistance development. The cytoplasmic membrane drug transporter AcrD is known to be involved in the development of aminoglycoside resistance. Since AcrD is a homolog of AcrB, it has been suggested that AcrD also forms a tripartite efflux complex with the periplasmic fusion protein AcrA and outer membrane channel protein TolC. Our study aims to determine the importance of AcrD, AcrA and TolC in the development of adaptive resistance against aminoglycosides and to evaluate their functional interdependence. Minimal inhibitory concentration assays of 3 and 24 h were performed on sub-inhibitory kanamycin pre-treated acrA, acrD and tolC deletion strains, to assess the development of resistance to inhibitory concentrations of kanamycin, streptomycin, ampicillin and tetracycline. A 24 h, long term protection assay indicated that acrA and tolC mutants were more susceptible and impaired in their ability to develop adaptive resistance against aminoglycosides. Protection against aminoglycosides during the 3 h protection assays was not detected, however transient cross resistance to ampicillin was observed. Results suggested that AcrA and TolC are involved in the gradual development of low level resistance to aminoglycosides following sub-inhibitory kanamycin pre-treatment.