Effect of RecA inactivation and detoxification systems on the evolution of ciprofloxacin resistance in Escherichia coli

Abstract

Suppression of SOS response and overproduction of reactive oxygen species (ROS) through detoxification system suppression enhance the activity of fluoroquinolones. Objectives: To evaluate the role of both systems in the evolution of resistance to ciprofloxacin in an isogenic model of Escherichia coli. Methods: Single-gene deletion mutants of E. coli BW25113 (wild-type) (DrecA, DkatG, DkatE, DsodA, DsodB), double-gene (DrecA-DkatG, DrecA-DkatE, DrecA-DsodA, DrecA-DsodB, DkatG-DkatE, DsodB-DsodA) and triplegene (DrecA-DkatG-DkatE) mutants were included. The response to sudden high ciprofloxacin pressure was evaluated by mutant prevention concentration (MPC). The gradual antimicrobial pressure response was evaluated through experimental evolution and antibiotic resistance assays. Results: For E. coli BW25113 strain, DkatE, DsodB and DsodB/DsodA mutants, MPC values were 0.25 mg/L. The DkatG, DsodA, DkatG/katE and DrecA mutants showed 2-fold reductions (0.125 mg/L). The DkatG/DrecA, DkatE/ DrecA, DsodA/DrecA, DsodB/DrecA and DkatG/DkatE/DrecA strains showed 4-8-fold reductions (0.03-0.06 mg/L) relative to the wild-type. Gradual antimicrobial pressure increased growth capacity for DsodA and DsodB and DsodB/DsodA mutants (no growth in 4mg/L) compared with the wild-type (no growth in the range of 0.5-2mg/L). Accordingly, increased growth was observed with the mutants DrecA/DkatG (no growth in 2mg/L), DrecA/DkatE (no growth in 2mg/L), DrecA/DsodA (no growth in 0.06mg/L), DrecA/DsodB (no growth in 0.25mg/L) and DrecA/ DkatG/DkatE (no growth in 0.5mg/L) comparedwith DrecA (no growth in the range of 0.002-0.015mg/L). Conclusions: After RecA inactivation, gradual exposure to ciprofloxacin reduces the evolution of resistance. After suppression of RecA and detoxification systems, sudden high exposure to ciprofloxacin reduces the evolution of resistance in E. coli.