Molecular Mechanisms of Azole Resistance in Four Clinical Candida albicans Isolates

Azole resistance constitutes a serious clinical problem in the management of infections caused by Candida albicans. This study aimed to explore azole-resistant mechanisms in clinical C. albicans isolates collected in Jinan, Shandong, China. In total, 22 samples were collected and analyzed. Among these, four isolates (28A, 28D, 28I, and 28J) exhibited high level of pan-azole-resistance that was Hsp90 dependent. Gene sequencing revealed that the four Hsp90-dependent strains contained different ERG3 mutations that led to four novel amino acid substitutions (S265Y, N322D, N324S, and E355D) in Erg3. The role of these substitutions in azole resistance development was determined by constructing one copy of the mutated ERG3 from the 28A, 28D, and 28I strains into C. albicans CAI4, respectively. The minimum inhibitory concentration value of fluconazole (FLC) against C. albicans CAI4-ERG3^28I increased fourfold compared with the wild-type C. albicans strain, suggesting that the novel combination of substitutions S265Y, N322D, and N324S played an important role in mediating azole resistance in 28I. Besides, we identified several different mechanisms in other three isolates. Strains 28A and 28D displayed increased efflux ability and overexpression of MDR1. Strain 28J showed high level of ERG11 expression, but no mutation in its regulator Upc2 was observed. Our study revealed that multiple factors confer azole resistance in clinical C. albicans isolates and combination therapy should be conducted clinically.