CORT0C04210 is required for Candida orthopsilosis adhesion to human buccal cells

Candida orthopsilosis is a human fungal pathogen belonging to the Candida parapsilosis sensu lato species complex. C. orthopsilosis annotated genome harbors 3 putative agglutinin-like sequence (ALS) genes named CORT0B00800, CORT0C04210 and CORT0C04220. The aim of this study was to investigate the role played by CORT0C04210 (CoALS4210) in the virulence and pathogenicity of this opportunistic yeast. Heterozygous and null mutant strains lacking one or both copies of CoALS4210 were obtained using the SAT1-flipper cassette strategy and were characterized in in vitro, ex vivo and in vivo models. While no differences between the mutant and the wild-type strains were observed in in vitro growth or in the ability to undergo morphogenesis, the CoALS4210 null mutant showed an impaired adhesion to human buccal epithelial cells compared to heterozygous and wild type strains. When the pathogenicity of CoALS4210 mutant and wild type strains was evaluated in a murine model of systemic candidiasis, no statistically significant differences were observed in fungal burden of target organs. Since gene disruption could alter chromatin structure and influence transcriptional regulation of other genes, two independent CRISPR/Cas9 edited mutant strains were generated in the same genetic background used to create the deleted strains. CoALS4210-edited strains were tested for their in vitro growing ability, and compared with the deleted strain for adhesion ability to human buccal epithelial cells. The results obtained confirmed a reduction in the adhesion ability of C. orthopsilosis edited strains to buccal cells. These findings provide the first evidence that CRISPR/Cas9 can be successfully used in C. orthopsilosis and demonstrate that CoALS4210 plays a direct role in the adhesion of C. orthopsilosis to human buccal cells but is not primarily involved in the onset of disseminated candidiasis.