Characteristics of Extracellular Vesicles Released by the Pathogenic Yeast-Like Fungi Candida glabrata, Candida parapsilosis and Candida tropicalis

Abstract

Candida spp. yeast-like fungi are opportunistic pathogens in humans and have been recently found to release extracellular vesicles (EVs) that are involved in many vital biological processes in fungal cells. These include communication between microorganisms and host–pathogen interactions during infection. The production of EVs and their content have been significantly characterized in the most common candidal species Candida albicans, including the identification of numerous virulence factors and cytoplasmic proteins in the EV cargo. We have here conducted the isolation and proteomic characterization of EVs produced by the clinically important non-albicans Candida species C. glabrata, C. tropicalis and C. parapsilosis. With the use of ultracentrifugation of the cell-free culture supernatant, the candidal EVs were collected and found to be a heterogeneous population of particles for each species with sizes ranging from 60–280 nm. The proteinaceous contents of these vesicles were analyzed using LC-MS/MS, with particular attention paid to surface-expressed proteins that would come into immediate and direct contact with host cells. We thereby identified 42 extracellular and surface-connected proteins from C. glabrata, 33 from C. parapsilosis, and 34 from C. tropicalis, including membrane-associated transporters, glycoproteins and enzymes involved in the organization of the fungal cell wall, as well as several cytoplasmic proteins, including alcohol dehydrogenase, enolase, glyceraldehyde-3-phosphate dehydrogenase, phosphoglycerate kinase and pyruvate kinase, for which the vesicular transport is a possible mechanism underlying their non-classical secretion.

Comparative molecular and immunoregulatory analysis of extracellular vesicles fromCandida albicansandCandida auris

“…As reported previously, EVs from C. albicans are round and bilayered particles (Figure 1A) 9 . Similar results were observed for EVs from both C. auris isolates (Figure 1B and 1C), consistent with the reported morphology of other fungal EVs 7,9,12,16,20,23,25,44 . EVs were also analyzed by dynamic light scattering (DLS) to evaluate their global size.…”

“…The literature reports two distinct populations of EVs from other species of Candida, a smaller one ranging usually from 50-70 nm, and a larger group between 100 and 800 nm 9,25, 44 . We found that C. auris releases EVs encompassed by lipid bilayers with size and shape consistent with those from other species.…”

Structural Insights into the Interactions of Candidal Enolase with Human Vitronectin, Fibronectin and Plasminogen

“…However, the mechanisms underlying the presence and relatively stable attachment of candidal enolase at the cell wall have not yet been identified. Current hypotheses include a sorting role of the 169 amino acid N-terminal domain of this enzyme [25] or the transport of enolase among the cargo of extracellular vesicles [69].…”

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

“…In a downstream analysis by this research group, the extracellular vesicular proteome of these non-albicans Candida spp. was characterized, in which a large portion of identified proteins similarly were classified as moonlighting proteins and understandably featured a large overlap with proteins identified on the cell surface (90). Among the moonlighting proteins identified here, some have immunogenic properties and roles in adhesion to human extracellular matrix proteins (e.g., fibronectin, vitronectin, and laminin).…”