Title Concentration of CCCP Should Be Optimized to Detect the Efflux System in Quinolone-Susceptible Escherichia coli
Author Hyengun Cho, Yoojung Oh, Seohyung Park, and Yeonhee Lee*
Address Department of Biology, Seoul Women’s University, Seoul 139-774, Korea
Bibliography Journal of Microbiology, 39(1),62-66, 2001,
Key Words antibiotic resistance, CCCP, efflux, E. coli, quinolone
Abstract Unlike eukaryotic efflux pumps energized by ATPase, bacterial efflux pumps are energized by the proton motive force. That is the reason why CCCP, an inhibitor of proton motive force, is widely used to study the bacterial efflux pump. In many cases, efflux systems have been observed only in quinoloneresistant bacteria. Most of the quinolone-susceptible strains have been found to maintain little efflux pump. However, some susceptible bacteria showed the increased intracellular quinolone concentration only at a low concentration (0.01 or 0.1 mM) but not at a high concentration (1 mM) of CCCP. If bacterial cells were killed at high concentrations of CCCP and lost the integrity of their membranes, the intracellular quinolone would leak out from cells with no efflux system. The efflux pump system in the quinolone-susceptible strains could not be detected at the same concentration used for sistant bacteria. To test this hypothesis, the intracellular quinolone concentration in the quinolone-susceptible and -resistant strains of Escherichia coli, Pseudomonas aeruginosa, and Staphylococcus aureus was assayed at various concentrations of CCCP. Since the effect of CCCP is very rapid, the survival of bacteria was observed by assaying the DNA synthesis in 5 min. In the case of E. coli, but not P. aeruginosa or S. aureus, the quinolone susceptible strain was more susceptible to CCCP than the quinolone resistant ones, especially when the incubation with CCCP was extended. Decrease of the intracellular quinolone concentration resulted in a false result-no or weak efflux system in the quinolone susceptible strains. Results suggested that the concentration of CCCP should be optimized in order to detect the efflux system in the quinolone susceptible strains of E. coli.
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