title: Multidrug efflux pumps: expression patterns and contribution to antibiotic resistance in Pseudomonas aeruginosa biofilms.
abstract: Pseudomonas aeruginosa biofilms are intrinsically resistant to antimicrobial chemotherapies. At present, very little is known about the physiological changes that occur during the transition from the planktonic to biofilm mode of growth. The resistance of P. aeruginosa biofilms to numerous antimicrobial agents that are substrates subject to active efflux from planktonic cells suggests that efflux pumps may substantially contribute to the innate resistance of biofilms. In this study, we investigated the expression of genes associated with two multidrug resistance (MDR) efflux pumps, MexAB-OprM and MexCD-OprJ, throughout the course of biofilm development. Using fusions to gfp, we were able to analyze spatial and temporal expression of mexA and mexC in the developing biofilm. Remarkably, expression of mexAB-oprM and mexCD-oprJ was not upregulated but rather decreased over time in the developing biofilm. Northern blot analysis confirmed that these pumps were not hyperexpressed in the biofilm. Furthermore, spatial differences in mexAB-oprM and mexCD-oprJ expression were observed, with maximal activity occurring at the biofilm substratum. Using a series of MDR mutants, we assessed the contribution of the MexAB-OprM, MexCD-OprJ, MexEF-OprN, and MexXY efflux pumps to P. aeruginosa biofilm resistance. These analyses led to the surprising discovery that the four characterized efflux pumps do not play a role in the antibiotic-resistant phenotype of P. aeruginosa biofilms.
authors.affiliation: Department of Microbiology and Immunology, University of Rochester Medical Center, Rochester, New York 14642, USA.
authors.forename: T R
authors.initials: TR
authors.lastname: De Kievit
authors.forename: M D
authors.initials: MD
authors.lastname: Parkins
authors.forename: R J
authors.initials: RJ
authors.lastname: Gillis
authors.forename: R
authors.initials: R
authors.lastname: Srikumar
authors.forename: H
authors.initials: H
authors.lastname: Ceri
authors.forename: K
authors.initials: K
authors.lastname: Poole
authors.forename: B H
authors.initials: BH
authors.lastname: Iglewski
authors.forename: D G
authors.initials: DG
authors.lastname: Storey
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001425:Bacterial Outer Membrane Proteins; D018441:Biofilms / Q000187:drug effects*; D002352:Carrier Proteins; D004352:Drug Resistance, Microbial; D026901:Membrane Transport Proteins; D009876:Operon; D010641:Phenotype; D010957:Plasmids / Q000187:drug effects; D011550:Pseudomonas aeruginosa / Q000187:drug effects* / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D001425:Bacterial Outer Membrane Proteins; D002352:Carrier Proteins; D026901:Membrane Transport Proteins; C118598:MexA protein, Pseudomonas aeruginosa