Candida parapsilosis and Candida albicans are human fungal pathogens that belong to the CTG clade in the Saccharomycotina. In contrast to C. albicans, relatively little is known about the virulence properties of C. parapsilosis, a pathogen particularly associated with infections of premature neonates. We describe here the construction of C. parapsilosis strains carrying double allele deletions of 100 transcription factors, protein kinases and species-specific genes. Two independent deletions were constructed for each target gene. Growth in >40 conditions was tested, including carbon source, temperature, and the presence of antifungal drugs. The phenotypes were compared to C. albicans strains with deletions of orthologous transcription factors. We found that many phenotypes are shared between the two species, such as the role of Upc2 as a regulator of azole resistance, and of CAP1 in the oxidative stress response. Others are unique to one species. For example, Cph2 plays a role in the hypoxic response in C. parapsilosis but not in C. albicans. We found extensive divergence between the biofilm regulators of the two species. We identified seven transcription factors and one protein kinase that are required for biofilm development in C. parapsilosis. Only three (Efg1, Bcr1 and Ace2) have similar effects on C. albicans biofilms, whereas Cph2, Czf1, Gzf3 and Ume6 have major roles in C. parapsilosis only. Two transcription factors (Brg1 and Tec1) with well-characterized roles in biofilm formation in C. albicans do not have the same function in C. parapsilosis. We also compared the transcription profile of C. parapsilosis and C. albicans biofilms. Our analysis suggests the processes shared between the two species are predominantly metabolic, and that Cph2 and Bcr1 are major biofilm regulators in C. parapsilosis.

The C. parapsilosis sensu lato group involves three closely related species, C. parapsilosis sensu stricto, C . orthopsilosis and C . metapsilosis . Although their overall clinical importance is dramatically increasing, there are few studies regarding the virulence properties of the species of the psilosis complex. In this study, we tested 63 C. parapsilosis sensu stricto, 12 C . metapsilosis and 18 C . orthopsilosis isolates for the ability to produce extracellular proteases, secrete lipases and form pseudohyphae. Significant differences were noted between species, with the C . metapsilosis strains failing to secrete lipase or to produce pseudohyphae. Nine different clinical isolates each of C. parapsilosis sensu stricto, C . orthopsilosis and C . metapsilosis were co-cultured with immortalized murine or primary human macrophages. C. parapsilosis sensu stricto isolates showed a significantly higher resistance to killing by primary human macrophages compared to C . orthopsilosis and C . metapsilosis isolates. In contrast, the killing of isolates by J774.2 mouse macrophages did not differ significantly between species. However, C. parapsilosis sensu stricto isolates induced the most damage to murine and human macrophages, and C . metapsilosis strains were the least toxic. Furthermore, strains that produced lipase or pseudohyphae were most resistant to macrophage-mediated killing and produced the most cellular damage. Finally, we used 9 isolates of each of the C. parapsilosis sensus lato species to examine their impact on the survival of Galleria mellonella larvae. The mortality rate of G . mellonella larvae infected with C . metapsilosis isolates was significantly lower than those infected with C. parapsilosis sensu stricto or C . orthopsilosis strains. Taken together, our findings demonstrate that C . metapsilosis is indeed the least virulent member of the psilosis group, and also highlight the importance of pseudohyphae and secreted lipases during fungal-host interactions.

Prostaglandins are C20 fatty acid metabolites with diverse biological functions. In mammalian cells, prostaglandins are produced from arachidonic acid (AA) via cyclooxygenases (COX1 and COX2). Although fungi do not possess cyclooxygenase homologues, several pathogenic species are able to produce prostaglandins from host-derived arachidonic acid. In this study, we characterized the prostaglandin profile of the emerging human pathogen Candida parapsilosis with HPLC-MS and compared it to that of C. albicans. We found that both species synthesized prostaglandins (mainly PGD 2 and PGE 2 ) from exogenous AA. Furthermore, as OLE2 has been associated with prostaglandin synthesis in C. albicans, we generated homozygous OLE2 deletion mutants in C. parapsilosis and examined their PGE 2 production. However, the PGE 2 production of the OLE2 KO strain was similar to that of wild type (WT), indicating that OLE2 is not required for prostaglandin synthesis in C. parapsilosis. Interestingly, analyses of the fatty acid composition of WT and OLE2 KO cells by gas chromatography (GC) highlighted the accumulation of palmitoleic and oleic acid in the OLE2 deletion mutant. The OLE2 KO cells were killed more efficiently by human monocytes-derived macrophages (MDMs) as well as induced higher interleukin-10 (IL-10) secretion, indicating that OLE2 affects the virulence of C. parapsilosis. Taken together, these results contribute to the better understanding of fatty acid biosynthesis pathways in C. parapsilosis.

Candida parapsilosis sensu stricto, Candida orthopsilosis and Candida metapsilosis are human fungal pathogens with clinical importance. The recently reclassified three closely related species have significant variation in virulence, clinical prevalence and susceptibility characteristics to different antifungal compounds. The aim of this study was to investigate the in vitro activity of atorvastatin and fluvastatin against C. metapsilosis, C. orthopsilosis and C. parapsilosis. Susceptibility tests showed that C. parapsilosis was the most sensitive while C. orthopsilosis was the least susceptible species to both drugs. On the basis of the differential sensitivity, we developed a simple, reliable and highly cost-effective plate assay to distinguish these closely related species. Applying this method, 54 isolates belonging to the C. parapsilosis sensu lato complex deposited in Szeged Microbial Collection could be sorted into the three species with 100 % probability.

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