title: Nucleotide substitutions in the Candida albicans ERG11 gene of azole-susceptible and azole-resistant clinical isolates.
abstract: One of the mechanisms of Candida albicans resistance to azole drugs used in antifungal therapy relies on increased expression and presence of point mutations in the ERG11 gene that encodes sterol 14α demethylase (14DM), an enzyme which is the primary target for the azole class of antifungals. The aim of the study was to analyze nucleotide substitutions in the Candida albicans ERG11 gene of azole-susceptible and azole-resistant clinical isolates. The Candida albicans isolates represented a collection of 122 strains selected from 658 strains isolated from different biological materials. Samples were obtained from hospitalized patients. Fluconazole susceptibility was tested in vitro using a microdilution assay. Candida albicans strains used in this study consisted of two groups: 61 of the isolates were susceptible to azoles and the 61 were resistant to azoles. Four overlapping regions of the ERG11 gene of the isolates of Candida albicans strains were amplified and sequenced. The MSSCP (multitemperature single strand conformation polymorphism) method was performed to select Candida albicans samples presenting genetic differences in the ERG11 gene fragments for subsequent sequence analysis. Based on the sequencing results we managed to detect 19 substitutions of nucleotides in the ERG11 gene fragments. Sequencing revealed 4 different alterations: T495A, A530C, G622A and A945C leading to changes in the corresponding amino acid sequence: D116E, K128T, V159I and E266D. The single nucleotide changes in the ERG11 gene did not affect the sensitivity of Candida albicans strains, whereas multiple nucleotide substitutions in the ERG11 gene fragments indicated a possible relation with the increase in resistance to azole drugs.
authors: Chair and Department of General Biology, Medical University of Silesia, Zabrze, Poland.
authors: Joanna Katarzyna
authors: JK
authors: Strzelczyk
authors: Laboratory of Microbiology in Nowy Targ Hospital, Nowy Targ, Poland.
authors: Anna
authors: A
authors: Slemp-Migiel
authors: Chair and Department of General Biology, Medical University of Silesia, Zabrze, Poland and Postgraduate School of Molecular Medicine, Warsaw, Poland.
authors: Magdalena
authors: M
authors: Rother
authors: Chair and Department of General Biology, Medical University of Silesia, Zabrze, Poland.
authors: Karolina
authors: K
authors: Gołąbek
authors: Chair and Department of General Biology, Medical University of Silesia, Zabrze, Poland.
authors: Andrzej
authors: A
authors: Wiczkowski
mesh_terms: D000935:Antifungal Agents / Q000627:therapeutic use; D002176:Candida albicans / Q000187:drug effects* / Q000235:genetics; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D025141:Drug Resistance, Fungal / Q000235:genetics; D015725:Fluconazole / Q000627:therapeutic use*; D005656:Fungal Proteins / Q000235:genetics*; D006801:Humans; D017354:Point Mutation
chemical_list: D000935:Antifungal Agents; D005656:Fungal Proteins; C465525:cytochrome P-450 CYP51, Candida albicans; D015725:Fluconazole; D003577:Cytochrome P-450 Enzyme System