Genome-guided manipulation of regulators of morphogenesis in a C. albicans strain with low virulence is not sufficient to trigger a high-virulence phenotype

As a member of the microbiome, C. albicans colonizes the oral cavity and other mucosal surfaces of the human body. While commensalism is tightly controlled by the host immune system, the fungal determinants enabling the fungus to colonize the host mucosa without causing tissue damage and inflammation remain less clear. In search of genetic determinants that may underly the commensal lifestyle of the low-virulent C. albicans isolate 101, we identified a small sequence duplication in one of the BRG1 allele resulting in a truncated loss-of-function allele (BRG1TRUNC). Replacing BRG1TRUNC by the full-length allele (BRG1FL) resulted in a modest increase in filamentation, but did not alter the phenotype of the fungus in the oral mucosa of experimentally colonized mice. Spontaneous outgrowth of highly virulent variants of the 101 strain with the BRG1TRUNC alleles replaced by a full-length allele identified the Cyr1-cAMP-PKA signalling pathway as a modulator of fungal virulence. Although the Glu-to-Lys mutation in CYR1 (CYR1E1541K), which converts Cyr1 into a hyperactive adenylate cyclase, greatly increased filamentation, the expression of hyphae-associated genes, and host cell damage when tested in vitro, it was insufficient to increase virulence of C. albicans strain 101 in the oral mucosa in vivo, irrespective of the BRG1 status. Together, this highlights that the low-virulent nature of strain 101 is firmly anchored and cannot be overcome by manipulating BRG1 and CYR1, two genes with known roles in C. albicans virulence. IMPORTANCEDuring homeostasis, the fungus Candida albicans establishes mutualistic interactions with its human host. It can however also adopt a pathogenic state and cause infections with diverse clinical manifestions that pose a significant challenge for diagnosis and therapy. Understanding the fungal determinants that underlie C. albicans colonization under steady state conditions may thus provide new avenues for modulating the fungus-host interaction in candidiasis patients to restore homeostasis. Here, we report gene variants that distinguish high-virulent from low-virulent C. albicans. Gene-exchange mutants provided evidence for the contribution of a BRG1 loss-of-function allele and a CYR1 gain-of-function mutation toward fungal pathogenicity. However, in vivo in an experimental model of C. albicans oral colonization, none of these gene variants individually or in combination were sufficient to change the virulence profile of the fungus. These findings indicate that C. albicans mucosal colonization is regulated by a complex gene network rather than by single genetic determinants.