Reduced susceptibility to polyenes associated with a missense mutation in the ERG6 gene in a clinical isolate of Candida glabrata with pseudohyphal growth.
Little information is available about the molecular mechanisms responsible for polyene resistance in pathogenic yeasts. A clinical isolate of Candida glabrata with a poor susceptibility to polyenes, as determined by disk diffusion method and confirmed by determination of MIC, was recovered from a patient treated with amphotericin B. Quantitative analysis of sterols revealed a lack of ergosterol and an accumulation of late sterol intermediates, suggesting a defect in the final steps of the ergosterol pathway. Sequencing of CgERG11, CgERG6, CgERG5, and CgERG4 genes revealed exclusively a unique missense mutation in CgERG6 leading to the substitution of a cysteine by a phenylalanine in the corresponding protein. In addition, real-time reverse transcription-PCR demonstrated an overexpression of genes encoding enzymes involved in late steps of the ergosterol pathway. Moreover, this isolate exhibited a pseudohyphal growth whatever the culture medium used, and ultrastructural changes of the cell wall of blastoconidia were seen consisting in a thinner inner layer. Cell wall alterations were also suggested by the higher susceptibility of growing cells to Calcofluor white. Additionally, complementation of this isolate with a wild-type copy of the CgERG6 gene restored susceptibility to polyenes and a classical morphology. Together, these results demonstrated that mutation in the CgERG6 gene may lead to a reduced susceptibility to polyenes and to a pseudohyphal growth due to the subsequent changes in sterol content of the plasma membrane.
Vandeputte
Patrick
P
Groupe d'Etude des Interactions Hôte-Parasite, UPRES-EA 3142, Laboratoire de Parasitologie-Mycologie, Centre Hospitalier Universitaire, 4 rue Larrey, 49933 Angers Cedex 9, France. patrick.vandeputte@etud.univ-angers.fr
Tronchin
Guy
G
Bergès
Thierry
T
Hennequin
Christophe
C
Chabasse
Dominique
D
Bouchara
Jean-Philippe
JP
D000935:Antifungal Agents / Q000494:pharmacology*; D041221:Candida glabrata / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development; D002177:Candidiasis / Q000382:microbiology*; D017931:DNA Primers; D005800:Genes, Fungal / Q000235:genetics; D005816:Genetic Complementation Test; D008780:Methyltransferases / Q000235:genetics*; D008826:Microbial Sensitivity Tests; D008969:Molecular Sequence Data; D020125:Mutation, Missense / Q000502:physiology*; D010641:Phenotype; D011090:Polyenes / Q000494:pharmacology*; D012331:RNA, Fungal / Q000096:biosynthesis / Q000235:genetics; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013261:Sterols / Q000378:metabolism
D000935:Antifungal Agents; D017931:DNA Primers; D011090:Polyenes; D012331:RNA, Fungal; D012333:RNA, Messenger; D013261:Sterols; D008780:Methyltransferases; C022670:delta 24-sterol methyltransferase