PDR1 Gain-of-function Mutations Lead To Azole-resistance And Increased Adhesion In Clinical Candida Glabrata Strains

Objective: To investigate the relationship between clinical usage of antifungal agents and drug resistance emergence of Candida glabra,to analyze the effect of gain-of-function(GOF)mutations on azole susceptiblity and adhesion ability to epithelial cells,as well as to clarify the regulation of transcription factor Pdr1 on adhesin EPA1.Methods: Four pairs of isolates were selected.Pairs of each strain were collected before and after antifungal treatment from the same patients.Antifungal susceptibilities were tested by broth dilution method.PDR1 genes were amplified by PCR and further sequenced.The transcript levels of related genes were detected by real-time PCR.The target gene was disrupted by homologous recombination strategy.In addition,molecular clones and epithelial cells adhesion assays were also performed.Results: The azole-susceptible strains obtained resistance to fluconazole after antifungal treatment,and the GOF mutations of PDR1 in azole-resistant strains were G1099 D,G346D,L344 S and P927 S respectively.The increased transcript levels of efflux pump genes CDR1 and CDR2 were the most important cause of azole resistance.Different GOF mutations in PDR1 led to distinct expression profiles of drug-related genes.The G1099 D and P927 S mutations in PDR1 caused enhanced adherence to epithelial cells,which was related to strain background.Epa1 was an important adhesin which was regulated by Pdr1 directly.Conclusions: Irrational use of antifungal agents tends to induce the emergence of azole resistance in Candida glabra.GOF mutations in PDR1 confer drug resistance and increased adherence in the clinical strains,which is an important evolutionary process of viabilities and pathogenicity.