Thea S B Kjeldsen1, Martin Overgaard2, Søren S Nielsen3, Valeria Bortolaia1, Lotte Jelsbak1, Morten Sommer4, Luca Guardabassi1, John E Olsen5. 1. Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, 1870 Frederiksberg C, Denmark. 2. Department of Clinical Biochemistry and Pharmacology, Odense University Hospital, 5000 Odense, Denmark Institute of Clinical Research, University of Southern Denmark, 5000 Odense C, Denmark. 3. Department of Large Animal Sciences, Faculty of Health and Medical Sciences, University of Copenhagen, 1870 Frederiksberg C, Denmark. 4. Department of Systems Biology, Technical University of Denmark, 2800 Lyngby, Denmark Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, 2970 Hørsholm, Denmark. 5. Department of Veterinary Disease Biology, Faculty of Health and Medical Sciences, University of Copenhagen, 1870 Frederiksberg C, Denmark jeo@sund.ku.dk.
Abstract
OBJECTIVES: 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.
OBJECTIVES: 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: Thea S B Møller; Martin Overgaard; Søren S Nielsen; Valeria Bortolaia; Morten O A Sommer; Luca Guardabassi; John E Olsen Journal: BMC Microbiol Date: 2016-03-12 Impact factor: 3.605
Authors: Thea S B Møller; Gang Liu; Anders Boysen; Line E Thomsen; Freja L Lüthje; Sisse Mortensen; Jakob Møller-Jensen; John E Olsen Journal: Front Microbiol Date: 2017-11-29 Impact factor: 5.640