Pseudohyphal Growth of the Emerging Pathogen Candida auris Is Triggered by Genotoxic Stress through the S Phase Checkpoint

Abstract

The morphogenetic switching between yeast cells and filaments (true hyphae and pseudohyphae) is a key cellular feature required for full virulence in many polymorphic fungal pathogens, such as Candida albicans. In the recently emerged yeast pathogen Candida auris, occasional elongation of cells has been reported. However, environmental conditions and genetic triggers for filament formation have remained elusive. Here, we report that induction of DNA damage and perturbation of replication forks by treatment with genotoxins, such as hydroxyurea, methyl methanesulfonate, and the clinically relevant fungistatic 5-fluorocytosine, cause filamentation in C. auris. The filaments formed were characteristic of pseudohyphae and not parallel-sided true hyphae. Pseudohyphal growth is apparently signaled through the S phase checkpoint and, interestingly, is Tup1 independent in C. auris. Intriguingly, the morphogenetic switching capability is strain specific in C. auris, highlighting the heterogenous nature of the species as a whole. IMPORTANCE Candida auris is a newly emerged fungal pathogen of humans. This species was first reported in 2009 when it was identified in an ear infection of a patient in Japan. However, despite intense interest in this organism as an often multidrug-resistant fungus, there is little knowledge about its cellular biology. During infection of human patients, fungi are able to change cell shape from ellipsoidal yeast cells to elongated filaments to adapt to various conditions within the host organism. There are different types of filaments, which are triggered by reactions to different cues. Candida auris fails to form filaments when exposed to triggers that stimulate yeast filament morphogenesis in other fungi. Here, we show that it does form filaments when its DNA is damaged. These conditions might arise when Candida auris cells interact with host immune cells or during growth in certain host tissues (kidney or bladder) or during treatment with antifungal drugs.

A Zinc Cluster Transcription Factor Contributes to the Intrinsic Fluconazole Resistance of Candida auris

“…The use of electroporation for transformation of C. auris also does not seem to be the problem, because electroporation was also used in the studies by Grahl et al, Kim et al,Rybak et al,and Iyer et al (15,16,25,26). It is possible that homologous recombination is less efficient in strains from clade III and clade IV, used to generate mutants in our present study, as opposed to clade I strains, which were the parents of the mutants generated in the six previous studies (15,16,(23)(24)(25)(26). Alternatively, the frequency of specific marker integration may vary considerably depending on the target locus.…”

“…Rybak et al (16) also used a CRISPR-Cas9 system in combination with a replacement cassette containing the caSAT1 selection marker and short (50-bp) flanking homology regions to delete CDR1 and MDR1 in two clade I isolates but did not report the frequency of specific gene replacement. Integration specificity was also not detailed in two very recent studies in which the NAT1 marker and long flanking regions were used for gene deletions in C. auris (23,26). The reason for the low frequency of specific marker integration into the target locus in our experiments is not evident.…”

“…Since C. auris is a haploid species, we anticipated that the generation of specific gene deletion mutants would be straightforward, as also inferred from the successful construction of C. auris mutants by other researchers (15,16,(23)(24)(25)(26). Unexpectedly, the vast majority of our transformants had unspecifically inserted the five different gene deletion cassettes at ectopic sites in the genome instead of integrating them at the target locus, suggesting that homologous recombination is much less efficient in C. auris than in C. albicans.…”

Fun(gi)omics: Advanced and Diverse Technologies to Explore Emerging Fungal Pathogens and Define Mechanisms of Antifungal Resistance

“…A dominant virulence strategy in Candida spp., specifically, C. albicans, is to morphologically adapt via the production of phenotype-switching metabolites to generate true hyphae. The novel MDR C. auris lacks the ability to form true hyphae but instead forms pseudohyphae (104,105). GC-MS analysis highlighted a different metabolic profile of C. auris compared to that of C. albicans under hypha-forming conditions, most significant was the production of hypha-inhibiting metabolites and biofilmforming molecules such as tyrosol (96).…”

Comparative Evaluations of the Pathogenesis of Candida auris Phenotypes and Candida albicans Using Clinically Relevant Murine Models of Infections

“…Unlike Candida albicans, C. auris does not undergo morphological switching between yeast and hyphal forms (5,17), and the lack of filamentation may explain the distinct niches of colonization between the species; while C. albicans can colonize and infect mucosal surfaces, C. auris primarily colonizes skin (13,16,20,21). However, occasional elongation of cells in C. auris into a filamentous and/or pseudohyphal form has been reported in response to temperature, cell cycle arrest, or depletion of Hsp90 (22,23) and DNA damage from exposure to antimicrobial agents (24). Although C. auris does not form true hyphae, it can adhere and form robust biofilms on surfaces, albeit the biofilms are less complex than those formed by the hypha-producing C. albicans (13,14,25).…”