Mechanism of echinocandin resistance in Candida glabrata

Niimi K., Woods M.A., Maki K., Hatakenaka K., Nakayama H., Chibana H., Niimi M., Monk B.C., Cannon R.D.

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The echinocandins inhibit the synthesis of β -1,3-g1ucan, an essential component of fungal cell walls. The incidence of candin-resistant pathogenic fungi is rare but has been associated with mutations in the echinocandin resistance region (the hot spot) of β -1,3-g1ucan synthase catalytic subunits encoded by FKS genes. We analysed the FKS genes in a micafungin-resistant Candida glabrata clinical isolate. Single nucleotide changes were detected in both C glabrata genes that are syntenic orthologues of the S. cerevisiae FKS1 and FKS2 genes. One mutation was predicted to cause an amino acid change in the hot spot of CgFKS1 and the other a premature stop codon in CgFKS2. We hypothesized that clinically significant candin resistance in C glabrata may require single nucleotide changes in both CgFKS1 and CgFKS2. Relationships between the observed mutations and candin resistance were assessed experimentally by reproducing the nucleotide changes in the CgFKS genes of a candin-susceptible C glabrata strain (micafungin MTC = 0.031 µg/ml) using site-directed mutagenesis, Introduction of the hot spot mutation into the CgFKS1 gene alone conferred intermediate resistance (MIC = 0.5 µg/ml) whereas the introduction of a premature stop codon in CgFKS2 alone had no effect on susceptibility, However, the insertion of both mutations conferred high level resistance (micafungin MIC = 2 µg/ml) equivalent to that of the clinical isolate, and cross-resistance to caspofungin. The phenotypes and candin susceptibilities of δ fks1 deletant and δ fks2 deletant indicate that the two CgFKS genes are functionally redundant, with each encoding a β -1,3 glucan synthase catalytic subunit. Like S. cerevisiae, the deletion of both CgFKS1 and CgFKS2 was found to be lethal. Clinically significant micafungin resistance in C. glabrata appears to be rare because CgFKS1 and CgFKS2 are differentially expressed and mutations in both CgFKS1 and CgFKS2 are required for the acquisition of high-level candin resistance.

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17th International Society for Human and Animal Mycology
    • ISHAM 17th (2009)