title: Molecular insights into fosfomycin resistance in Escherichia coli.
abstract: Fosfomycin activity in Escherichia coli depends on several genes of unknown importance for fosfomycin resistance. The objective was to characterize the role of uhpT , glpT , cyaA and ptsI genes in fosfomycin resistance in E. coli.
  
  
  
  WT E. coli BW25113 and null mutants, Δ uhpT , Δ glpT , Δ cyaA , Δ ptsI , Δ glpT-uhpT , Δ glpT-cyaA , Δ glpT-ptsI , Δ uhpT-cyaA , Δ uhpT-ptsI and Δ ptsI-cyaA , were studied. Susceptibility to fosfomycin was tested using CLSI guidelines. Fosfomycin mutant frequencies were determined at concentrations of 64 and 256 mg/L. Fosfomycin in vitro activity was tested using time-kill assays at concentrations of 64 and 307 mg/L (human C max ).
  
  
  
  Fosfomycin MICs were: WT E. coli BW25113 (2 mg/L), Δ glpT (2 mg/L), Δ uhpT (64 mg/L), Δ cyaA (8 mg/L), Δ ptsI (2 mg/L), Δ glpT-uhpT (256 mg/L), Δ glpT-cyaA (8 mg/L), Δ glpT-ptsI (2 mg/L), Δ uhpT-cyaA (512 mg/L), Δ uhpT-ptsI (64 mg/L) and Δ ptsI-cyaA (32 mg/L). In the mutant frequency assays, no mutants were recovered from BW25113. Mutants appeared in Δ glpT , Δ uhpT , Δ cyaA and Δ ptsI at 64 mg/L and in Δ uhpT and Δ cyaA at 256 mg/L. Δ glpT-ptsI , but not Δ glpT-cyaA , Δ uhpT-cyaA or Δ uhpT-ptsI , increased the mutant frequency compared with the highest frequency found in each single mutant. In time-kill assays, all mutants regrew at 64 mg/L. Initial bacterial reductions of 2-4 log 10 cfu/mL were observed for all strains, except for Δ uhpT-ptsI , Δ glpT-uhpT and Δ uhpT-cyaA . Only Δ glpT and Δ ptsI mutants were cleared using 307 mg/L.
  
  
  
  Fosfomycin MIC may not be a good efficacy predictor, as highly resistant mutants may appear, depending on other pre-existing mutations with no impact on MIC.
authors.forename: M
authors.initials: M
authors.lastname: Ballestero-Téllez
authors.forename: F
authors.initials: F
authors.lastname: Docobo-Pérez
authors.affiliation: Unidad intercentros de Enfermedades Infecciosas, Microbiología y Medicina Preventiva, Hospital Universitario Virgen Macarena, Sevilla, Spain.|Instituto de Biomedicina de Sevilla, IBiS/Hospital Universitario Virgen del Rocío/CSIC/Universidad de Sevilla, Sevilla, Spain.
authors.forename: I
authors.initials: I
authors.lastname: Portillo-Calderón
authors.forename: J M
authors.initials: JM
authors.lastname: Rodríguez-Martínez
authors.affiliation: Unidad intercentros de Enfermedades Infecciosas, Microbiología y Medicina Preventiva, Hospital Universitario Virgen Macarena, Sevilla, Spain.
authors.forename: L
authors.initials: L
authors.lastname: Racero
authors.affiliation: Unidad intercentros de Enfermedades Infecciosas, Microbiología y Medicina Preventiva, Hospital Universitario Virgen Macarena, Sevilla, Spain.
authors.forename: M S
authors.initials: MS
authors.lastname: Ramos-Guelfo
authors.affiliation: Instituto de Biomedicina de Sevilla, IBiS/Hospital Universitario Virgen del Rocío/CSIC/Universidad de Sevilla, Sevilla, Spain.|Centro Nacional de Biotecnología, Madrid, Spain.
authors.forename: J
authors.initials: J
authors.lastname: Blázquez
authors.affiliation: Departamento de Medicina, Universidad de Sevilla, Sevilla, Spain.
authors.forename: J
authors.initials: J
authors.lastname: Rodríguez-Baño
authors.forename: A
authors.initials: A
authors.lastname: Pascual
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D004269:DNA, Bacterial / Q000235:genetics; D024881:Drug Resistance, Bacterial / Q000235:genetics*; D004926:Escherichia coli / Q000187:drug effects* / Q000235:genetics*; D029968:Escherichia coli Proteins / Q000235:genetics*; D005578:Fosfomycin / Q000494:pharmacology*; D019450:Genes, MDR; D006801:Humans; D008826:Microbial Sensitivity Tests; D009154:Mutation
chemical_list: D000900:Anti-Bacterial Agents; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D005578:Fosfomycin