Interactions between copy number and expression level of genes involved in fluconazole resistance in Candida glabrata

Abstract

Objectives: This study aimed to elucidate the relative involvement of drug resistance gene copy number and overexpression in fluconazole resistance in clinical C. glabrata isolates using a population-based approach.Methods: Fluconazole resistance levels were quantified using the minimal inhibitory concentration (MIC) via Etest method. Both gene expression levels and gene copy number of CgCDR1, CgPDH1, CgERG11, and CgSNQ2 were assessed via quantitative real-time PCR. The influence of the main effects and first-level interactions of both the expression level and copy number of these genes on fluconazole resistance levels were analyzed using a multivariate statistical model.Results: Forty-three C. glabrata isolates were collected from 30 patients during in a hospital survey. In the multivariate analysis, C. glabrata fluconazole MICs were independently increased by CgSNQ2 overexpression (p < 10−4) and the interaction between CgPDH1 gene copy number and CgPDH1 expression level (p = 0.038). In contrast, both CgPDH1 overexpression (p = 0.049) and the interaction between CgSNQ2 and CgERG11 expression (p = 0.003) led to a significant decrease in fluconazole MICs.Conclusion: Fluconazole resistance in C. glabrata involves complex interactions between drug resistance gene expression and/or copy number. The population-based multivariate analysis highlighted the involvement of the CgSNQ2 gene in fluconazole resistance and the complex effect of the other genes such as PDH1 for which overexpression was associated with reduced fluconazole resistance levels, while the interaction between PDH1 overexpression and copy number was associated with increased resistance levels.

Clonal Spread of Candida glabrata Bloodstream Isolates and Fluconazole Resistance Affected by Prolonged Exposure: a 12-Year Single-Center Study in Belgium

“…However, only CgCDR1 was expressed at a significantly higher level in the FLC-resistant C. glabrata BSI isolates, indicating that it is the major FLC efflux pump in C. glabrata, consistent with previous studies (10,(41)(42)(43)45). Although significant, the higher expression of CgCDR1 in resistant strains was not marked and was generally lower than in previous reports (41,42,44), suggesting that other mechanisms are involved in FLC resistance in our set of isolates. Indeed, gainof-function mutations in CgPDR1 were earlier reported to mediate azole resistance in C. glabrata (11)(12)(13)45), and genes other than those encoding ABC efflux pumps were shown to play a role in FLC resistance in C. glabrata.…”

“…FLC resistance in C. glabrata is generally associated with an elevated expression of genes encoding ABC efflux pumps (10-13, 42, 43). Previous studies investigated the importance of these molecular mechanisms but were generally limited to a set of related isolates sequentially obtained from the same patient (13,29,44). In contrast, we examined in a large set of independent BSI isolates whether the level of FLC susceptibility could be correlated to the expression of efflux pumps and if the latter could be associated with a specific genotype.…”

“…Indeed, gainof-function mutations in CgPDR1 were earlier reported to mediate azole resistance in C. glabrata (11)(12)(13)45), and genes other than those encoding ABC efflux pumps were shown to play a role in FLC resistance in C. glabrata. The latter include the ERG11 gene, genes that belong to the major facilitator superfamily and to the aldo-keto-reductase superfamily, together with various complex interactions between different molecular mechanisms (43,44,(46)(47)(48)(49)(50). Moreover, C. glabrata can be involved in breakthrough infections due to its capacity to exhibit inducible resistance (51).…”

Antifungal Therapy: New Advances in the Understanding and Treatment of Mycosis

“…The exact mechanism underlying FLZ resistance in C. glabrata is not well defined; however, different studies indicate an essential role for ABC transporters (Abbes et al, 2013). FLZ-resistant C. glabrata clinical isolates present CDR1 and CDR2 upregulation (Sanglard et al, 1999; Bennett et al, 2004).…”

Mechanisms of antibiotic resistance

“…Sun et al (2013) reported that sterol C-22 desaturase ERG5, which is highly conserved among various fungal species, is involved in azole resistance and may serve as a novel target for antifungal drugs, in particular against Neurospora crassa and Fusarium verticillioides. Using the population-based multivariate analysis, Abbes et al (2013) observed that fluconazole resistance in Candida glabrata involves complex interactions between drug resistance gene expression and/or copy number.…”