title: CTX-M-1 β-lactamase expression in Escherichia coli is dependent on cefotaxime concentration, growth phase and gene location.
abstract: OBJECTIVE
  Knowledge about the regulatory mechanisms of CTX-M β-lactamase-encoding genes in Escherichia coli is limited. The objectives of this study were to determine the growth response of CTX-M-1-producing E. coli exposed to cefotaxime and to investigate how blaCTX-M-1 expression at mRNA and protein levels is influenced by cefotaxime concentration, growth phase and gene location (chromosome versus plasmid).
  
  
  METHODS
  Two isogenic E. coli strains, MG1655/CTX-M-1 and MG1655/IncI1/CTX-M-1, containing blaCTX-M-1 on the chromosome and on a wild-type IncI1 plasmid, respectively, were constructed and the MIC of cefotaxime was determined. Growth of the two strains was studied in the presence of increasing concentrations of cefotaxime ranging from 0 to 512 mg/L. The levels of mRNA and protein in different growth phases and at different cefotaxime concentrations were studied by qPCR and selected-reaction-monitoring MS, respectively.
  
  
  RESULTS
  The MICs of cefotaxime were 168 and 252 mg/L for MG1655/CTX-M-1 and MG1655/IncI1/CTX-M-1, respectively. Both strains displayed a prolonged lag phase when exposed to cefotaxime. The mRNA of blaCTX-M-1 and CTX-M-1 protein levels increased in the presence of high cefotaxime concentrations and varied with growth phase. Higher mRNA expression levels were detected for MG1655/CTX-M-1 compared with MG1655/IncI1/CTX-M-1, but a higher protein level was found for MG1655/IncI1/CTX-M-1 compared with MG1655/CTX-M-1, the latter corresponding well with the higher MIC for this strain.
  
  
  CONCLUSIONS
  blaCTX-M-1 mRNA expression and CTX-M-1 protein levels were dependent on cefotaxime concentration, growth phase and gene location. These results provide insight into the expression of cephalosporin resistance in CTX-M-1-producing E. coli, improving our understanding of the relationship between antimicrobial therapy and the expression of resistance mechanisms.
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, 1870 Frederiksberg C, Denmark.
authors.forename: Thea S B
authors.initials: TS
authors.lastname: Kjeldsen
authors.affiliation: Department of Clinical Biochemistry and Pharmacology, Odense University Hospital, 5000 Odense, Denmark Institute of Clinical Research, University of Southern Denmark, 5000 Odense C, Denmark.
authors.forename: Martin
authors.initials: M
authors.lastname: Overgaard
authors.affiliation: Department of Large Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, 1870 Frederiksberg C, Denmark.
authors.forename: Søren S
authors.initials: SS
authors.lastname: Nielsen
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, 1870 Frederiksberg C, Denmark.
authors.forename: Valeria
authors.initials: V
authors.lastname: Bortolaia
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, 1870 Frederiksberg C, Denmark.
authors.forename: Lotte
authors.initials: L
authors.lastname: Jelsbak
authors.affiliation: Department of Systems Biology, Technical University of Denmark, 2800 Lyngby, Denmark Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, 2970 Hørsholm, Denmark.
authors.forename: Morten
authors.initials: M
authors.lastname: Sommer
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, 1870 Frederiksberg C, Denmark.
authors.forename: Luca
authors.initials: L
authors.lastname: Guardabassi
authors.affiliation: Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, 1870 Frederiksberg C, Denmark jeo@sund.ku.dk.
authors.forename: John E
authors.initials: JE
authors.lastname: Olsen
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D002439:Cefotaxime / Q000494:pharmacology*; D002876:Chromosomes, Bacterial; D004926:Escherichia coli / Q000187:drug effects* / Q000201:enzymology* / Q000235:genetics; D020869:Gene Expression Profiling; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects*; D015971:Gene Expression Regulation, Enzymologic / Q000187:drug effects*; D005798:Genes, Bacterial; D013058:Mass Spectrometry; D008826:Microbial Sensitivity Tests; D010957:Plasmids; D060888:Real-Time Polymerase Chain Reaction; D001618:beta-Lactamases / Q000096:biosynthesis*
chemical_list: D000900:Anti-Bacterial Agents; C055184:beta-lactamase TEM-3; D001618:beta-Lactamases; D002439:Cefotaxime
keywords: ESBLs; antimicrobial resistance; regulation