title: Concomitant occurrence of itraconazole-resistant and -susceptible strains of Aspergillus fumigatus in routine cultures.
abstract: OBJECTIVE
  Resistance to triazoles in Aspergillus fumigatus has emerged in several countries and is usually mediated by mutations in the cyp51A gene. We determined the presence of both itraconazole-susceptible and -resistant A. fumigatus in routine cultures isolated from environmental and clinical samples.
  
  
  METHODS
  A total of 50 environmental and 16 clinical A. fumigatus isolates obtained from single colonies were tested for itraconazole susceptibility by Etest. Serial dilution and plating for selected A. fumigatus cultures were performed to separate triazole-susceptible and -resistant phenotypes. Antifungal drug susceptibility of subcultures to itraconazole, posaconazole and voriconazole was determined by the broth microdilution method. Itraconazole resistance was determined by the presence of tandem repeats in the promoter region and other resistance-conferring mutations by PCR and/or direct DNA sequencing of cyp51A. Genotyping was performed with a panel of nine microsatellite loci.
  
  
  RESULTS
  PCR amplification of the promoter region identified 1 of 50 environmental and 2 of 16 clinical A. fumigatus isolates as mixed (itraconazole-resistant and -susceptible) cultures, while the remaining isolates yielded patterns that were consistent with their data for susceptibility to itraconazole. Purified subcultures yielded distinct susceptibility profiles, concomitant with genetic determinants of triazole susceptibility/resistance in cyp51A and different microsatellite patterns.
  
  
  CONCLUSIONS
  Concomitant presence of triazole-susceptible and -resistant strains in single colonies of routine A. fumigatus cultures, obtained from environmental and clinical samples, has been conclusively demonstrated.
authors.affiliation: Department of Microbiology, Faculty of Medicine, Kuwait University, Kuwait.
authors.forename: Suhail
authors.initials: S
authors.lastname: Ahmad
authors.affiliation: Department of Microbiology, Faculty of Medicine, Kuwait University, Kuwait.
authors.forename: Leena
authors.initials: L
authors.lastname: Joseph
authors.affiliation: Department of Medical Microbiology and Infectious Diseases, Canisius Wilhelmina Hospital, Nijmegen, The Netherlands.
authors.forename: Ferry
authors.initials: F
authors.lastname: Hagen
authors.affiliation: Department of Medical Microbiology and Infectious Diseases, Canisius Wilhelmina Hospital, Nijmegen, The Netherlands Department of Medical Microbiology, Radboud University Medical Center, Nijmegen, The Netherlands.
authors.forename: Jacques F
authors.initials: JF
authors.lastname: Meis
authors.affiliation: Department of Microbiology, Faculty of Medicine, Kuwait University, Kuwait zkhan@hsc.edu.kw.
authors.forename: Ziauddin
authors.initials: Z
authors.lastname: Khan
mesh_terms: D000935:Antifungal Agents / Q000494:pharmacology*; D001232:Aspergillus fumigatus / Q000145:classification / Q000187:drug effects* / Q000235:genetics / Q000302:isolation & purification; D016000:Cluster Analysis; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics; D025141:Drug Resistance, Fungal; D005656:Fungal Proteins / Q000235:genetics; D005838:Genotype; D006801:Humans; D017964:Itraconazole / Q000494:pharmacology*; D008826:Microbial Sensitivity Tests; D058889:Molecular Typing; D009154:Mutation
chemical_list: D000935:Antifungal Agents; D005656:Fungal Proteins; D017964:Itraconazole; D003577:Cytochrome P-450 Enzyme System; C465526:cytochrome P-450 CYP51A, Aspergillus
keywords: A. fumigatus; cyp51A mutations; invasive aspergillosis; resistance