Genome-wide analysis of experimentally evolvedCandida aurisreveals multiple novel mechanisms of multidrug-resistance

Abstract

AbstractCandida aurisis globally recognized as an opportunistic fungal pathogen of high concern, due to its extensive multidrug-resistance (MDR). Still, molecular mechanisms of MDR are largely unexplored. This is the first account of genome wide evolution of MDR inC. aurisobtained through serialin vitroexposure to azoles, polyenes and echinocandins. We show the stepwise accumulation of multiple novel mutations in genes known and unknown in antifungal drug resistance, albeit almost all new forC. auris. Echinocandin resistance evolved through a codon deletion inFKS1accompanied by a substitution inFKS1hot spot 3. Mutations inERG3andCIS2further increased the echinocandin MIC. Decreased azole susceptibility was acquired through a gain of function mutation in transcription factorTAC1byielding overexpression of the drug efflux pump Cdr1; a segmental duplication of chromosome 1 containingERG11; and a whole chromosome 5 duplication, which containsTAC1b. The latter was associated with increased expression ofERG11, TAC1bandCDR2, but notCDR1. The simultaneous emergence of nonsense mutations inERG3andERG11, presumably leading to the abrogation of ergosterol synthesis, was shown to decrease amphotericin B susceptibility, accompanied with fluconazole cross resistance. A mutation inMEC3, a gene mainly known for its role in DNA damage homeostasis, further increased the polyene MIC. Overall, this study shows the alarming potential and diversity for MDR development inC. auris, even in a clade until now not associated with MDR (clade II), hereby stressing its clinical importance and the urge for future research.ImportanceC. aurisis a recently discovered human fungal pathogens and has shown an alarming potential for multi- and pan-resistance towards all classes of antifungals most commonly used in the clinic. Currently,C. aurishas been globally recognized as a nosocomial pathogen of high concern due to this evolutionary potential. So far, this is the first study in which the stepwise progression of MDR inC. aurisis monitoredin vitro. Multiple novel mutations in known ‘resistance genes’ and genes previously not or vaguely associated with drug resistance reveal rapid MDR evolution in aC. aurisclade II isolate. Additionally, this study shows thatin vitroexperimental evolution can be a powerful tool to discover new drug resistance mechanisms, although it has its limitations.