title: Phenotypic and genome-wide analysis of an antibiotic-resistant small colony variant (SCV) of Pseudomonas aeruginosa.
abstract: BACKGROUND
  Small colony variants (SCVs) are slow-growing bacteria, which often show increased resistance to antibiotics and cause latent or recurrent infections. It is therefore important to understand the mechanisms at the basis of this phenotypic switch.
  
  
  RESULTS
  One SCV (termed PAO-SCV) was isolated, showing high resistance to gentamicin and to the cephalosporine cefotaxime. PAO-SCV was prone to reversion as evidenced by emergence of large colonies with a frequency of 10(-5) on media without antibiotics while it was stably maintained in presence of gentamicin. PAO-SCV showed a delayed growth, defective motility, and strongly reduced levels of the quorum sensing Pseudomonas quinolone signal (PQS). Whole genome expression analysis further suggested a multi-layered antibiotic resistance mechanism, including simultaneous over-expression of two drug efflux pumps (MexAB-OprM, MexXY-OprM), the LPS modification operon arnBCADTEF, and the PhoP-PhoQ two-component system. Conversely, the genes for the synthesis of PQS were strongly down-regulated in PAO-SCV. Finally, genomic analysis revealed the presence of mutations in phoP and phoQ genes as well as in the mexZ gene encoding a repressor of the mexXY and mexAB-oprM genes. Only one mutation occurred only in REV, at nucleotide 1020 of the tufA gene, a paralog of tufB, both encoding the elongation factor Tu, causing a change of the rarely used aspartic acid codon GAU to the more common GAC, possibly causing an increase of tufA mRNA translation. High expression of phoP and phoQ was confirmed for the SCV variant while the revertant showed expression levels reduced to wild-type levels.
  
  
  CONCLUSIONS
  By combining data coming from phenotypic, gene expression and proteome analysis, we could demonstrate that resistance to aminoglycosides in one SCV mutant is multifactorial including overexpression of efflux mechanisms, LPS modification and is accompanied by a drastic down-regulation of the Pseudomonas quinolone signal quorum sensing system.
authors.affiliation: Research Group Microbiology, VIB Department of Structural Biology, Vrije Universiteit Brussel, Brussels, Belgium.
authors.forename: Qing
authors.initials: Q
authors.lastname: Wei
authors.forename: Saeed
authors.initials: S
authors.lastname: Tarighi
authors.forename: Andreas
authors.initials: A
authors.lastname: Dötsch
authors.forename: Susanne
authors.initials: S
authors.lastname: Häussler
authors.forename: Mathias
authors.initials: M
authors.lastname: Müsken
authors.forename: Victoria J
authors.initials: VJ
authors.lastname: Wright
authors.forename: Miguel
authors.initials: M
authors.lastname: Cámara
authors.forename: Paul
authors.initials: P
authors.lastname: Williams
authors.forename: Steven
authors.initials: S
authors.lastname: Haenen
authors.forename: Bart
authors.initials: B
authors.lastname: Boerjan
authors.forename: Annelies
authors.initials: A
authors.lastname: Bogaerts
authors.forename: Evy
authors.initials: E
authors.lastname: Vierstraete
authors.forename: Peter
authors.initials: P
authors.lastname: Verleyen
authors.forename: Liliane
authors.initials: L
authors.lastname: Schoofs
authors.forename: Ronnie
authors.initials: R
authors.lastname: Willaert
authors.forename: Valérie N
authors.initials: VN
authors.lastname: De Groote
authors.forename: Jan
authors.initials: J
authors.lastname: Michiels
authors.forename: Ken
authors.initials: K
authors.lastname: Vercammen
authors.forename: Aurélie
authors.initials: A
authors.lastname: Crabbé
authors.forename: Pierre
authors.initials: P
authors.lastname: Cornelis
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D015169:Colony Count, Microbial; D015536:Down-Regulation / Q000187:drug effects / Q000235:genetics; D024881:Drug Resistance, Bacterial / Q000187:drug effects / Q000235:genetics*; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D005798:Genes, Bacterial / Q000235:genetics; D014644:Genetic Variation / Q000187:drug effects; D016680:Genome, Bacterial / Q000235:genetics*; D005839:Gentamicins / Q000494:pharmacology; D008826:Microbial Sensitivity Tests; D020411:Oligonucleotide Array Sequence Analysis; D010641:Phenotype; D020543:Proteome / Q000378:metabolism; D011550:Pseudomonas aeruginosa / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development*; D015363:Quinolones / Q000378:metabolism; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014158:Transcription, Genetic / Q000187:drug effects
chemical_list: C407944:2-heptyl-3-hydroxy-4-quinolone; D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D005839:Gentamicins; D020543:Proteome; D015363:Quinolones