Objective: An association between reduced susceptibility to the echinocandin drug caspofungin (MIC > 8 ug/ml) and amino acid substitutions in the Fks1p subunit of the beta (1,3)-D-glucan synthase (GS) was recently reported (Park et al. 2005 Antimicrob. Agents Chemother. 49: 3264-3273). Specific mutations in FKS1 genes were identified from both laboratory and clinical isolates of C. albicans, and two hot spot regions affecting susceptibility were defined as Phe641-Pro649 and the residues around Arg1361. To better understand the universality of FKS1 mutations for reduced echinocandin susceptibility, we examined cross-reactivity of mutants to micafungin and characterized the hot-spot regions in isolates of non-C. albicans species showing reduced drug susceptibility. Methods: Antifungal susceptibility testing on yeasts was performed using the broth micro dilution method of the NCCLS document M27-A2. Purification of glucan synthase and IC50 values were determined as described by Park et al. 2005. A 2.4 Kb region of FKS1 from C. glabrata, C. guilliermondii, C. krusei, C. lipolytica, C. parapsilosis, C. rugosa, C. tropicalis and C. dubliniensis was PCR amplified using consensuses FKS1 primers. All PCR products were subjected to automated DNA sequencing in both 5' and 3' directions. Results: The fks1 mutants showed comparable reduced susceptibility to both caspofungin and micafungin in growth assays, and specific mutations were associated with a > 1000-fold reduction in the inhibition of GS activity by both echinocandin drugs. Characteristic mutations associated with reduced susceptibility in C. albicans were also found in C. tropicalis, C. dubliensis and C. krusei, which correlated with reduced IC50 values in assays of glucan synthase activity. Conclusion: The data supports the notion that reduced susceptibility to echinocandin drugs in clinical isolates can arise from amino acid substitutions in defined regions of the highly conserved Fks1p subunit of glucan synthase.
Full conference title:
16th European Congress of Clinical Microbiology and Infectious Diseases
- ECCMID 16th (2006)