Upregulation of AcrEF in Quinolone Resistance Development in Escherichia coli When AcrAB-TolC Function Is Impaired

We studied mechanisms of drug resistance development in Escherichia coli strains lacking efflux pump components. E. coli K12 deletion mutants were subjected to increasing concentrations of ciprofloxacin (CIP) to determine the frequency of target gene mutations. We generated a series of mutants that were selected based on their minimum inhibitory concentrations (MICs) to CIP, as well as their corresponding point mutations in target genes. The mutants displayed a number of target modifications and, in particular, gyrA mutations altering codons Ser83Leu, Asp87Gly, and Asp87His as well as a change in parC at 78 (substitution of Gly for Asp). All these mutations were related to drug resistance. When exposed to CIP, mutants lacking efflux pump genes acrA and acrB demonstrated a low level of resistance that was because of point mutations in the target genes. High-level resistance was achieved with a 100- to 500-fold increase in expression of efflux pump genes acrE and acrF that compensated for the loss of AcrA and AcrB, and thus resulted in an obvious increase of CIP MIC. We demonstrate that an intact AcrAB-TolC efflux pump is crucial to the development of bacterial resistance. Its activity is complemented by expression of the alternative AcrEF efflux pump.