title: In vivo evolution of resistance of Pseudomonas aeruginosa strains isolated from patients admitted to an intensive care unit: mechanisms of resistance and antimicrobial exposure.
abstract: OBJECTIVE
  The main objective of this study was to investigate the relationship among the in vivo acquisition of antimicrobial resistance in Pseudomonas aeruginosa clinical isolates, the underlying molecular mechanisms and previous exposure to antipseudomonal agents.
  
  
  METHODS
  PFGE was used to study the molecular relatedness of the strains. The MICs of ceftazidime, cefepime, piperacillin/tazobactam, imipenem, meropenem, ciprofloxacin and amikacin were determined. Outer membrane protein profiles were assessed to study OprD expression. RT-PCR was performed to analyse ampC, mexB, mexD, mexF and mexY expression. The presence of mutations was analysed through DNA sequencing.
  
  
  RESULTS
  We collected 17 clonally related paired isolates [including first positive samples (A) and those with MICs increased ≥4-fold (B)]. Most B isolates with increased MICs of imipenem, meropenem and ceftazidime became resistant to these drugs. The most prevalent resistance mechanisms detected were OprD loss (65%), mexB overexpression (53%), ampC derepression (29%), quinolone target gene mutations (24%) and increased mexY expression (24%). Five (29%) B isolates developed multidrug resistance. Meropenem was the most frequently (71%) received treatment, explaining the high prevalence of oprD mutations and likely mexB overexpression. Previous exposure to ceftazidime showed a higher impact on selection of increased MICs than previous exposure to piperacillin/tazobactam.
  
  
  CONCLUSIONS
  Stepwise acquisition of resistance has a critical impact on the resistance phenotypes of P. aeruginosa, leading to a complex scenario for finding effective antimicrobial regimens. In the clinical setting, meropenem seems to be the most frequent driver of multidrug resistance development, while piperacillin/tazobactam, in contrast to ceftazidime, seems to be the β-lactam least associated with the selection of resistance mechanisms.
authors.affiliation: ISGlobal, Barcelona Ctr. Int. Health Res. (CRESIB), Hospital Clínic - Universitat de Barcelona, Barcelona, Spain.
authors.forename: Mar
authors.initials: M
authors.lastname: Solé
authors.affiliation: ISGlobal, Barcelona Ctr. Int. Health Res. (CRESIB), Hospital Clínic - Universitat de Barcelona, Barcelona, Spain.
authors.forename: Anna
authors.initials: A
authors.lastname: Fàbrega
authors.affiliation: Hospital Clínic, IDIBAPS, University of Barcelona, Barcelona, Spain.
authors.forename: Nazaret
authors.initials: N
authors.lastname: Cobos-Trigueros
authors.affiliation: University Hospital Son Espases, Palma de Mallorca, Spain.
authors.forename: Laura
authors.initials: L
authors.lastname: Zamorano
authors.affiliation: ISGlobal, Barcelona Ctr. Int. Health Res. (CRESIB), Hospital Clínic - Universitat de Barcelona, Barcelona, Spain.
authors.forename: Mario
authors.initials: M
authors.lastname: Ferrer-Navarro
authors.affiliation: ISGlobal, Barcelona Ctr. Int. Health Res. (CRESIB), Hospital Clínic - Universitat de Barcelona, Barcelona, Spain.
authors.forename: Clara
authors.initials: C
authors.lastname: Ballesté-Delpierre
authors.affiliation: ISGlobal, Barcelona Ctr. Int. Health Res. (CRESIB), Hospital Clínic - Universitat de Barcelona, Barcelona, Spain.
authors.forename: Anna
authors.initials: A
authors.lastname: Reustle
authors.affiliation: Hospital Clínic, IDIBAPS, University of Barcelona, Barcelona, Spain.
authors.forename: Pedro
authors.initials: P
authors.lastname: Castro
authors.affiliation: Hospital Clínic, IDIBAPS, University of Barcelona, Barcelona, Spain.
authors.forename: José Maria
authors.initials: JM
authors.lastname: Nicolás
authors.affiliation: University Hospital Son Espases, Palma de Mallorca, Spain.
authors.forename: Antonio
authors.initials: A
authors.lastname: Oliver
authors.affiliation: Hospital Clínic, IDIBAPS, University of Barcelona, Barcelona, Spain.
authors.forename: José Antonio
authors.initials: JA
authors.lastname: Martínez
authors.affiliation: ISGlobal, Barcelona Ctr. Int. Health Res. (CRESIB), Hospital Clínic - Universitat de Barcelona, Barcelona, Spain Hospital Clínic, IDIBAPS, University of Barcelona, Barcelona, Spain jvila@ub.edu.
authors.forename: Jordi
authors.initials: J
authors.lastname: Vila
mesh_terms: D000900:Anti-Bacterial Agents / Q000494:pharmacology / Q000627:therapeutic use*; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics; D024881:Drug Resistance, Bacterial; D016521:Electrophoresis, Gel, Pulsed-Field; D019143:Evolution, Molecular; D020869:Gene Expression Profiling; D006801:Humans; D007362:Intensive Care Units; D008826:Microbial Sensitivity Tests; D058889:Molecular Typing; D011552:Pseudomonas Infections / Q000188:drug therapy* / Q000382:microbiology*; D011550:Pseudomonas aeruginosa / Q000145:classification / Q000187:drug effects* / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D001618:beta-Lactamases / Q000235:genetics
chemical_list: D000900:Anti-Bacterial Agents; D001425:Bacterial Outer Membrane Proteins; D001618:beta-Lactamases