title: Comparative effects of overproducing the AraC-type transcriptional regulators MarA, SoxS, RarA and RamA on antimicrobial drug susceptibility in Klebsiella pneumoniae.
abstract: In Klebsiella pneumoniae, overproduction of RamA and RarA leads to increased MICs of various antibiotics; MarA and SoxS are predicted to perform a similar function. We have compared the relative effects of overproducing these four AraC-type regulators on envelope permeability (a combination of outer membrane permeability and efflux), efflux pump and porin production, and antibiotic susceptibility in K. pneumoniae.
  
  
  
  Regulators were overproduced using a pBAD expression vector. Antibiotic susceptibility was measured using disc testing. Envelope permeability was estimated using a fluorescent dye accumulation assay. Porin and efflux pump production was quantified using proteomics and validated using real-time quantitative RT-PCR.
  
  
  
  Envelope permeability and antibiotic disc inhibition zone diameters both reduced during overproduction of RamA and to a lesser extent RarA or SoxS, but did not change following overproduction of MarA. These effects were associated with overproduction of the efflux pumps AcrAB (for RamA and SoxS) and OqxAB (for RamA and RarA) and the outer membrane protein TolC (for all regulators). Effects on porin production were strain specific.
  
  
  
  RamA is the most potent regulator of antibiotic permeability in K. pneumoniae, followed by RarA then SoxS, with MarA having very little effect. This observed relative potency correlates well with the frequency at which these regulators are reportedly overproduced in clinical isolates.
authors.affiliation: School of Cellular and Molecular Medicine, University of Bristol, Bristol, UK.
authors.forename: Juan-Carlos
authors.initials: JC
authors.lastname: Jiménez-Castellanos
authors.affiliation: School of Cellular and Molecular Medicine, University of Bristol, Bristol, UK Faculty of Biotechnology & Biomolecular Sciences, Universiti Putra Malaysia, Selangor Darul Ehsan, Malaysia.
authors.forename: Wan Nur Ismah
authors.initials: WN
authors.lastname: Wan Ahmad Kamil
authors.affiliation: School of Cellular and Molecular Medicine, University of Bristol, Bristol, UK.
authors.forename: Ching Hei Phoebe
authors.initials: CH
authors.lastname: Cheung
authors.affiliation: School of Cellular and Molecular Medicine, University of Bristol, Bristol, UK.
authors.forename: Maryann S
authors.initials: MS
authors.lastname: Tobin
authors.affiliation: School of Cellular and Molecular Medicine, University of Bristol, Bristol, UK.
authors.forename: James
authors.initials: J
authors.lastname: Brown
authors.affiliation: School of Cellular and Molecular Medicine, University of Bristol, Bristol, UK.
authors.forename: Sophie G
authors.initials: SG
authors.lastname: Isaac
authors.affiliation: Bristol University Proteomics Facility, University of Bristol, Bristol, UK.
authors.forename: Kate J
authors.initials: KJ
authors.lastname: Heesom
authors.affiliation: Division of Infection and Pathway Medicine, University of Edinburgh, Edinburgh, UK.
authors.forename: Thamarai
authors.initials: T
authors.lastname: Schneiders
authors.affiliation: School of Cellular and Molecular Medicine, University of Bristol, Bristol, UK matthewb.avison@bristol.ac.uk.
authors.forename: Matthew B
authors.initials: MB
authors.lastname: Avison
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000096:biosynthesis*; D001693:Biological Transport, Active; D002462:Cell Membrane / Q000502:physiology; D024881:Drug Resistance, Bacterial; D015870:Gene Expression; D020869:Gene Expression Profiling; D005809:Genes, Regulator; D007711:Klebsiella pneumoniae / Q000187:drug effects* / Q000235:genetics*; D008826:Microbial Sensitivity Tests; D010539:Permeability; D018272:Porins / Q000378:metabolism; D020543:Proteome / Q000032:analysis; D040901:Proteomics; D060888:Real-Time Polymerase Chain Reaction
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D018272:Porins; D020543:Proteome