title: Interplay between three RND efflux pumps in doxycycline-selected strains of Burkholderia thailandensis.
abstract: BACKGROUND
  Efflux systems are involved in multidrug resistance in most Gram-negative non-fermentative bacteria. We have chosen Burkholderia thailandensis to dissect the development of multidrug resistance phenotypes under antibiotic pressure.
  
  
  RESULTS
  We used doxycycline selection to obtain several resistant B. thailandensis variants. The minimal inhibitory concentrations of a large panel of structurally unrelated antibiotics were determined ± the efflux pump inhibitor phenylalanine-arginine ß-naphthylamide (PAßN). Membrane proteins were identified by proteomic method and the expressions of major efflux pumps in the doxycycline selected variants were compared to those of the parental strains by a quantitative RT-PCR analysis. Doxycycline selected variants showed a multidrug resistance in two major levels corresponding to the overproduction of two efflux pumps depending on its concentration: AmrAB-OprA and BpeEF-OprC. The study of two mutants, each lacking one of these pumps, indicated that a third pump, BpeAB-OprB, could substitute for the defective pump. Surprisingly, we observed antagonistic effects between PAßN and aminoglycosides or some ß-lactams. PAßN induced the overexpression of AmrAB-OprA and BpeAB-OprB pump genes, generating this unexpected effect.
  
  
  CONCLUSIONS
  These results may account for the weak activity of PAßN in some Gram-negative species. We clearly demonstrated two antagonistic effects of this molecule on bacterial cells: the blocking of antibiotic efflux and an increase in efflux pump gene expression. Thus, doxycycline is a very efficient RND efflux pump inducer and PAßN may promote the production of some efflux pumps. These results should be taken into account when considering antibiotic treatments and in future studies on efflux pump inhibitors.
authors.affiliation: Unité de Bactériologie/UMR_MD 1, Institut de Recherche Biomédicale des Armées, Brétigny-sur-Orge, France ; UMR_MD 1, Aix Marseille Université, IRBA, Facultés de Médecine et de Pharmacie, Marseille, France.
authors.forename: Fabrice Vincent
authors.initials: FV
authors.lastname: Biot
authors.affiliation: Unité de Bactériologie/UMR_MD 1, Institut de Recherche Biomédicale des Armées, Brétigny-sur-Orge, France.
authors.forename: Mélanie Monique
authors.initials: MM
authors.lastname: Lopez
authors.affiliation: Pôle de génomique, Institut de Recherche Biomédicale des Armées, Brétigny-sur-Orge, France.
authors.forename: Thomas
authors.initials: T
authors.lastname: Poyot
authors.affiliation: Unité de Bactériologie/UMR_MD 1, Institut de Recherche Biomédicale des Armées, Brétigny-sur-Orge, France ; UMR_MD 1, Aix Marseille Université, IRBA, Facultés de Médecine et de Pharmacie, Marseille, France.
authors.forename: Fabienne
authors.initials: F
authors.lastname: Neulat-Ripoll
authors.affiliation: FR 3479 Plate-forme de Protéomique, CNRS, Aix-Marseille Université, Marseille, France.
authors.forename: Sabrina
authors.initials: S
authors.lastname: Lignon
authors.affiliation: Unité de Bactériologie/UMR_MD 1, Institut de Recherche Biomédicale des Armées, Brétigny-sur-Orge, France ; UMR_MD 1, Aix Marseille Université, IRBA, Facultés de Médecine et de Pharmacie, Marseille, France.
authors.forename: Arnaud
authors.initials: A
authors.lastname: Caclard
authors.affiliation: Unité de Bactériologie/UMR_MD 1, Institut de Recherche Biomédicale des Armées, Brétigny-sur-Orge, France ; UMR_MD 1, Aix Marseille Université, IRBA, Facultés de Médecine et de Pharmacie, Marseille, France.
authors.forename: François Michel
authors.initials: FM
authors.lastname: Thibault
authors.affiliation: Pôle de génomique, Institut de Recherche Biomédicale des Armées, Brétigny-sur-Orge, France.
authors.forename: Andre
authors.initials: A
authors.lastname: Peinnequin
authors.affiliation: UMR_MD 1, Aix Marseille Université, IRBA, Facultés de Médecine et de Pharmacie, Marseille, France.
authors.forename: Jean-Marie
authors.initials: JM
authors.lastname: Pagès
authors.affiliation: Unité de Bactériologie/UMR_MD 1, Institut de Recherche Biomédicale des Armées, Brétigny-sur-Orge, France ; UMR_MD 1, Aix Marseille Université, IRBA, Facultés de Médecine et de Pharmacie, Marseille, France ; Ecole du Val-de-Grâce, Paris, France.
authors.forename: Eric
authors.initials: E
authors.lastname: Valade
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001425:Bacterial Outer Membrane Proteins / Q000096:biosynthesis; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D019117:Burkholderia / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism*; D004151:Dipeptides / Q000494:pharmacology; D004318:Doxycycline / Q000494:pharmacology*; D024901:Drug Resistance, Multiple, Bacterial / Q000187:drug effects / Q000235:genetics; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D008826:Microbial Sensitivity Tests; D009154:Mutation; D010641:Phenotype; D040901:Proteomics
chemical_list: D000900:Anti-Bacterial Agents; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D004151:Dipeptides; C419365:phenylalanine arginine beta-naphthylamide; D004318:Doxycycline