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Molecular analysis of cyp51 from fluconazole-resistant Candida albicans strains
Abstract:
The target enzyme for fluconazole is sterol 14 alpha-demethylase, a cytochrome P450 encoded by cyp51. One mechanism of fluconazole resistance likely to occur in Candida albicans is through an altered target site. To test this hypothesis DNA sequencing of the cyp51 coding sequence from 19 fluconazole-resistant and 19 fluconazole-sensitive C. albicans was undertaken. A number of point mutations were identified in the resistant isolates which were not present in the sensitive ones: F105L (five), E266D (five), K287R (one), G448G (one), G450E (one), G464S (three) and V488I (one). These alterations are discussed in the light of a molecular model of the enzyme regarding potential roles in resistance. It was also demonstrated that sequence-specific primers can be employed to identify polymorphisms which may be associated with resistance; diagnostic tests for resistant strains will prove of value in combating this serious clinical problem.
Insights
Fluconazole resistance in Candida albicans may stem from mutations in the target enzyme, sterol 14 alpha-demethylase (cyp51). DNA sequencing identified several new mutations in resistant strains, aiding in the development of diagnostic tests for this fungal infection.
Area of Science:
- Medical Mycology
- Molecular Biology
- Antimicrobial Resistance
Background:
- Fluconazole is a key antifungal medication targeting sterol 14 alpha-demethylase (CYP51).
- Candida albicans can develop resistance to fluconazole, posing a significant clinical challenge.
- Altered target enzyme sites are a suspected mechanism for fluconazole resistance.
Purpose of the Study:
- To investigate the role of target site alteration in fluconazole resistance in Candida albicans.
- To identify specific mutations in the cyp51 gene associated with fluconazole resistance.
Main Methods:
- DNA sequencing of the cyp51 gene in fluconazole-resistant and sensitive Candida albicans isolates.
- Analysis of point mutations and their correlation with resistance phenotypes.
- Utilizing molecular modeling to assess the impact of mutations on enzyme function.
Main Results:
- Multiple point mutations were identified in the cyp51 gene of fluconazole-resistant isolates, absent in sensitive strains.
- Specific mutations included F105L, E266D, K287R, G448G, G450E, G464S, and V488I.
- Sequence-specific primers were developed to detect resistance-associated polymorphisms.
Conclusions:
- Target site modification via cyp51 mutations is a likely mechanism of fluconazole resistance in Candida albicans.
- Identified mutations provide insights into enzyme-drug interactions and resistance mechanisms.
- Diagnostic tests based on these polymorphisms can aid in managing fluconazole-resistant infections.