Project description:The principal opportunistic human fungal pathogen Candida albicans forms biofilms resistant to antifungal therapeutics. Biofilms are a class of soft matter with viscoelastic properties and response to flow, but little is known regarding the genes contributing to these rheological phenotypes in fungal biofilms. Here, we identify C. albicans genes with deletion phenotypes of altered biofilm viscoelasticity. We analyzed mutants deleted for genes contributing to cell wall structure or extracellular matrix (ECM) production, and we identified increased elastic moduli, indicative of higher viscoelasticity, in strains singly deleted for PMR1, KRE5, and ALG11. PMR1 encodes a secretory pathway calcium pump. KRE5 encodes a UDP-glucose:glycoprotein glucosyltransferase, and ALG11 encodes alpha-1,2-mannosyltransferase. These mutants form less biofilm ECM by weight relative to wild type when cultured on agar. For these strains, biofilm morphology is smooth, with reduced hyphal formation. The mutants exhibit decreased resistance to the antifungal agent fluconazole relative to wild type biofilm cultures. To identify intracellular changes underlying these altered rheological properties, we globally profiled transcript levels in the respective mutants. Genes encoding membrane proteins were enriched in the set of transcripts differentially abundant in the alg11 deletion mutant. RNA levels are altered for genes associated with translation in the pmr1 deletion mutant and protein catabolism in the kre5 deletion strain. Genes involved in lipid metabolism and filamentous development are differentially expressed in cells from alg11, kre5, and pmr1 deletion mutant biofilms. Collectively, the data indicate C. albicans biofilm rheology as a phenotype affected by ECM production and cell morphology, while identifying genes for the investigation of mechanisms underlying properties of fungal biofilm viscoelasticity.
Project description:Candida auris is frequently associated with biofilm-related invasive infections. The resistant profile of these biofilms necessitates innovative therapeutic options, where quorum sensing may be a potential target. Farnesol and tyrosol are two fungal quorum-sensing molecules with antifungal effects at supraphysiological concentrations. Here, we performed genome-wide transcript profiling with C. auris biofilms following farnesol or tyrosol exposure using transcriptome sequencing (RNA-Seq). Since transition metals play a central role in fungal virulence and biofilm formation, levels of intracellular calcium, magnesium, and iron were determined following farnesol or tyrosol treatment using inductively coupled plasma optical emission spectrometry. Farnesol caused an 89.9% and 73.8% significant reduction in the calcium and magnesium content, respectively, whereas tyrosol resulted in 82.6%, 76.6%, and 81.2% decrease in the calcium, magnesium, and iron content, respectively, compared to the control. Genes involved in biofilm events, glycolysis, ergosterol biosynthesis, fatty acid oxidation, iron metabolism, and autophagy were primarily affected in treated cells. To prove ergosterol quorum-sensing molecule interactions, microdilution-based susceptibility testing was performed, where the complexation of farnesol, but not tyrosol, with ergosterol was impeded in the presence of exogenous ergosterol, resulting in a minimum inhibitory concentration increase in the quorum-sensing molecules. This study revealed several farnesol- and tyrosol-specific responses, which will contribute to the development of alternative therapies against C. auris biofilms.