UNLABELLED: The aim of the study was to develop a simple, inexpensive, reproducible ethidium bromide (EB)-agar based method that is independent of any specialized instrumentation, for the demonstration of efflux pump activity, which is responsible for antibiotic resistance of bacteria. MATERIALS AND METHODS: A series of agar plates containing varying concentrations of EB were swabbed with strains of Escherichia coli or Staphylococcus aureus, which differed with respect to efflux pump activity. The plates were incubated at different temperatures and time periods and the measurements of fluorescence were used to evaluate the efflux activity of each culture. RESULTS: This simple assay allowed us to identify the efflux of EB in different bacteria following an overnight incubation. The minimal concentration of EB that produced fluorescence was significantly greater at 37 degrees C than at 4 degrees C, suggesting the presence of an energy-dependent pump. The method was shown to simultaneously identify strains of a mixed culture that differed from each other with respect to the activity of their efflux pumps. CONCLUSION: The method, in conjunction with the use of antibiotic-containing disks, provides an additional advantage for the easy identification and selection of colonies that differ with respect to antibiotic susceptibility and degree of efflux pump activity. Because the method is very reproducible it may form the basis for interlaboratory standardization of efflux pump activity of multi-drug resistant (MDR) clinical isolates.
UNLABELLED: The aim of the study was to develop a simple, inexpensive, reproducible ethidium bromide (EB)-agar based method that is independent of any specialized instrumentation, for the demonstration of efflux pump activity, which is responsible for antibiotic resistance of bacteria. MATERIALS AND METHODS: A series of agar plates containing varying concentrations of EB were swabbed with strains of Escherichia coli or Staphylococcus aureus, which differed with respect to efflux pump activity. The plates were incubated at different temperatures and time periods and the measurements of fluorescence were used to evaluate the efflux activity of each culture. RESULTS: This simple assay allowed us to identify the efflux of EB in different bacteria following an overnight incubation. The minimal concentration of EB that produced fluorescence was significantly greater at 37 degrees C than at 4 degrees C, suggesting the presence of an energy-dependent pump. The method was shown to simultaneously identify strains of a mixed culture that differed from each other with respect to the activity of their efflux pumps. CONCLUSION: The method, in conjunction with the use of antibiotic-containing disks, provides an additional advantage for the easy identification and selection of colonies that differ with respect to antibiotic susceptibility and degree of efflux pump activity. Because the method is very reproducible it may form the basis for interlaboratory standardization of efflux pump activity of multi-drug resistant (MDR) clinical isolates.
Authors: Ana Martins; Gabriella Spengler; Liliana Rodrigues; Miguel Viveiros; Jorge Ramos; Marta Martins; Isabel Couto; Séamus Fanning; Jean-Marie Pagès; Jean Michel Bolla; Joseph Molnar; Leonard Amaral Journal: PLoS One Date: 2009-08-17 Impact factor: 3.240
Authors: Marta Martins; Matthew P McCusker; Miguel Viveiros; Isabel Couto; Séamus Fanning; Jean-Marie Pagès; Leonard Amaral Journal: Open Microbiol J Date: 2013-03-22
Authors: Miguel Viveiros; Myrielle Dupont; Liliana Rodrigues; Isabel Couto; Anne Davin-Regli; Marta Martins; Jean-Marie Pagès; Leonard Amaral Journal: PLoS One Date: 2007-04-11 Impact factor: 3.240
Authors: Laura Paixão; Liliana Rodrigues; Isabel Couto; Marta Martins; Pedro Fernandes; Carla C C R de Carvalho; Gabriel A Monteiro; Filipe Sansonetty; Leonard Amaral; Miguel Viveiros Journal: J Biol Eng Date: 2009-10-16 Impact factor: 4.355