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Predatory interactions among microbes are major evolutionary driving forces for biodiversity. The fungivorous amoeba Protostelium aurantium has a wide fungal food spectrum including foremost pathogenic members of the genus Candida. Here we show that upon phagocytic ingestion by the amoeba, Candida parapsilosis is confronted with an oxidative burst and undergoes lysis within minutes of processing in acidified phagolysosomes. On the fungal side, a functional genomic approach identified copper and redox homeostasis as primary targets of amoeba predation, with the highly expressed copper exporter gene CRP1 and the peroxiredoxin gene PRX1 contributing to survival when encountered with P. aurantium. The fungicidal activity was largely retained in intracellular vesicles of the amoebae. Following their isolation, the content of these vesicles induced immediate killing and lysis of C. parapsilosis in vitro. Proteomic analysis identified 56 vesicular proteins from P. aurantium. Although completely unknown proteins were dominant, many of them could be categorised as hydrolytic enzymes targeting the fungal cell wall, indicating that fungal cell wall structures are under selection pressure by predatory phagocytes in natural environments. TAKE AWAY: The amoeba Protostelium aurantium feeds on fungi, such as Candida parapsilosis. Ingested yeast cells are exposed to reactive oxygen species. A copper exporter and a peroxiredoxin contribute to fungal defence. Yeast cells undergo intracellular lysis. Lysis occurs via a cocktail of hydrolytic enzymes from intracellular vesicles., (© 2021 John Wiley & Sons Ltd.)

The human pathogenic fungus Candida albicans is a frequent cause of mucosal infections. Although the ability to transition from the yeast to the hypha morphology is essential for virulence, hypha formation and host cell invasion per se are not sufficient for the induction of epithelial damage. Rather, the hypha-associated peptide toxin, candidalysin, a product of the Ece1 polyprotein, is the critical damaging factor. While synthetic, exogenously added candidalysin is sufficient to damage epithelial cells, the level of damage does not reach the same level as invading C. albicans hyphae. Therefore, we hypothesized that a combination of fungal attributes is required to deliver candidalysin to the invasion pocket to enable the full damaging potential of C. albicans during infection. Utilising a panel of C. albicans mutants with known virulence defects, we demonstrate that the full damage potential of C. albicans requires the coordinated delivery of candidalysin to the invasion pocket. This process requires appropriate epithelial adhesion, hyphal extension and invasion, high levels of ECE1 transcription, proper Ece1 processing and secretion of candidalysin. To confirm candidalysin delivery, we generated camelid V H Hs (nanobodies) specific for candidalysin and demonstrate localization and accumulation of the toxin only in C. albicans-induced invasion pockets. In summary, a defined combination of virulence attributes and cellular processes is critical for delivering candidalysin to the invasion pocket to enable the full damage potential of C. albicans during mucosal infection. TAKE AWAYS: Candidalysin is a peptide toxin secreted by C. albicans causing epithelial damage. Candidalysin delivery to host cell membranes requires specific fungal attributes. Candidalysin accumulates in invasion pockets created by invasive hyphae. Camelid nanobodies enabled visualisation of candidalysin in the invasion pocket., (© 2021 The Authors. Cellular Microbiology published by John Wiley & Sons Ltd.)

Candida albicans is a common opportunistic fungal pathogen that causes a wide range of infections from superficial mucosal to hematogenously disseminated candidiasis. The hyphal form plays an important role in the pathogenic process by invading epithelial cells and causing tissue damage. Notably, the secretion of the hyphal toxin candidalysin is essential for both epithelial cell damage and activation of mucosal immune responses. However, the mechanism of candidalysin-induced cell death remains unclear. Here, we examined the induction of cell death by candidalysin in oral epithelial cells. Fluorescent imaging using healthy/apoptotic/necrotic cell markers revealed that candidalysin causes a rapid and marked increase in the population of necrotic rather than apoptotic cells in a concentration dependent manner. Activation of a necrosis-like pathway was confirmed since C. albicans and candidalysin failed to activate caspase-8 and -3, or the cleavage of poly (ADP-ribose) polymerase. Furthermore, oral epithelial cells treated with candidalysin showed rapid production of reactive oxygen species, disruption of mitochondria activity and mitochondrial membrane potential, ATP depletion and cytochrome c release. Collectively, these data demonstrate that oral epithelial cells respond to the secreted fungal toxin candidalysin by triggering numerous cellular stress responses that induce necrotic death. TAKE AWAYS: Candidalysin secreted from Candida albicans causes epithelial cell stress. Candidalysin induces calcium influx and oxidative stress in host cells. Candidalysin induces mitochondrial dysfunction, ATP depletion and epithelial necrosis. The toxicity of candidalysin is mediated from the epithelial cell surface., (© 2021 The Authors. Cellular Microbiology published by John Wiley & Sons Ltd.)

Human and plant pathogenic fungi have a major impact on public health and agriculture. Although these fungi infect very diverse hosts and are often highly adapted to specific host niches, they share surprisingly similar mechanisms that mediate immune evasion, modulation of distinct host targets and exploitation of host nutrients, highlighting that successful strategies have evolved independently among diverse fungal pathogens. These attributes are facilitated by an arsenal of fungal factors. However, not a single molecule, but rather the combined effects of several factors enable these pathogens to establish infection. In this review, we discuss the principles of human and plant fungal pathogenicity mechanisms and discuss recent discoveries made in this field., (© 2020 The Authors. Cellular Microbiology published by John Wiley & Sons Ltd.)

Biotin is an important cofactor for multiple enzymes in central metabolic processes. While many bacteria and most fungi are able to synthesise biotin de novo, Candida spp. are auxotrophic for this vitamin and thus require efficient uptake systems to facilitate biotin acquisition during infection. Here we show that Candida glabrata and Candida albicans use a largely conserved system for biotin uptake and regulation, consisting of the high-affinity biotin transporter Vht1 and the transcription factor Vhr1. Both species induce expression of biotin-metabolic genes upon in vitro biotin depletion and following phagocytosis by macrophages, indicating low biotin levels in the Candida-containing phagosome. In line with this, we observed reduced intracellular proliferation of both Candida cells pre-starved of biotin and deletion mutants lacking VHR1 or VHT1 genes. VHT1 was essential for the full virulence of C. albicans during systemic mouse infections, and the lack of VHT1 led to reduced fungal burden in C. glabrata-infected brains and C. albicans-infected brains and kidneys. Together, our data suggest a critical role of Vht1-mediated biotin acquisition for C. glabrata and C. albicans during intracellular growth in macrophages and systemic infections., (© 2020 The Authors. Cellular Microbiology published by John Wiley & Sons Ltd.)

Membrane bilayers of eukaryotic cells are an amalgam of lipids and proteins that distinguish organelles and compartmentalise cellular functions. The mammalian cell has evolved mechanisms to sense membrane tension or damage and respond as needed. In the case of the plasma membrane and phagosomal membrane, these bilayers act as a barrier to microorganisms and are a conduit by which the host interacts with pathogens, including fungi such as Candida, Cryptococcus, Aspergillus, or Histoplasma species. Due to their size, morphological flexibility, ability to produce long filaments, secrete pathogenicity factors, and their potential to replicate within the phagosome, fungi can assault host membranes in a variety of physical and biochemical ways. In addition, the recent discovery of a fungal pore-forming peptide toxin further highlights the importance of membrane biology in the outcomes between host and fungal cells. In this review, we discuss the apparent "stretching" of membranes as a sophisticated biological response and the role of vesicular transport in combating membrane stress and damage. We also review the known pathogenicity factors and physical properties of fungal pathogens in the context of host membranes and discuss how this may contribute to pathogenic interactions between fungal and host cells., (© 2019 John Wiley & Sons Ltd.)

Candida albicans is an important human opportunistic fungal pathogen which is frequently found as part of the normal human microbiota. It is well accepted that the fungus interacts with other components of the resident microbiota and that this impacts the commensal or pathogenic outcome of C. albicans colonization. Different types of interactions, including synergism or antagonism, contribute to a complex balance between the multitude of different species. Mixed biofilms of C. albicans and streptococci are a well-studied example of a mutualistic interaction often potentiating the virulence of the individual members. In contrast, other bacteria like lactobacilli are known to antagonize C. albicans, and research has just started elucidating the mechanisms behind these interactions. This scenario is even more complicated by a third player, the host. This review focuses on interactions between C. albicans and gram-positive bacteria whose investigation will without doubt ultimately help understanding C. albicans infections., (© 2016 John Wiley & Sons Ltd.)

Candida albicans and C. glabrata are the two most common pathogenic yeasts of humans, yet they are phylogenetically, genetically and phenotypically very different. In this review, we compare and contrast the strategies of C. albicans and C. glabrata to attach to and invade into the host, obtain nutrients and evade the host immune response. Although their strategies share some basic concepts, they differ greatly in their outcome. While C. albicans follows an aggressive strategy to subvert the host response and to obtain nutrients for its survival, C. glabrata seems to have evolved a strategy which is based on stealth, evasion and persistence, without causing severe damage in murine models. However, both fungi are successful as commensals and as pathogens of humans. Understanding these strategies will help in finding novel ways to fight Candida, and fungal infections in general., (© 2012 Blackwell Publishing Ltd.)

The human pathogenic fungus Candida albicans can cause systemic infections by invading epithelial barriers to gain access to the bloodstream. One of the main reservoirs of C. albicans is the gastrointestinal tract and systemic infections predominantly originate from this niche. In this study, we used scanning electron and fluorescence microscopy, adhesion, invasion and damage assays, fungal mutants and a set of fungal and host cell inhibitors to investigate the interactions of C. albicans with oral epithelial cells and enterocytes. Our data demonstrate that adhesion, invasion and damage by C. albicans depend not only on fungal morphology and activity, but also on the epithelial cell type and the differentiation stage of the epithelial cells, indicating that epithelial cells differ in their susceptibility to the fungus. C. albicans can invade epithelial cells by induced endocytosis and/or active penetration. However, depending on the host cell faced by the fungus, these routes are exploited to a different extent. While invasion into oral cells occurs via both routes, invasion into intestinal cells occurs only via active penetration.

Candida albicans is among the most important fungal pathogens in humans. Morphological plasticity has been linked to its pathogenic potential as filamentous forms are associated with tissue invasion and infection. Here we show that human neutrophils discriminate between yeasts and filaments of C. albicans. Whereas filaments induced targeted motility, resulting in the establishment of close contact between neutrophils and fungal cells, yeast forms were largely ignored during coincubation. In transwell assays, C. albicans filaments induced significantly higher migratory activity in neutrophils than yeasts. Neutrophil motility based on actin rearrangement was essential for killing of C. albicans filaments but not involved in killing of yeast forms. Using inhibitors for MAP-kinase cascades, it was shown that recognition of C. albicans filaments by neutrophils is mediated via the MEK/ERK cascade and independent of JNK or p38 activation. Inhibition of the ERK signalling pathway abolished neutrophil migration induced by C. albicans filaments and selectively impaired the ability to kill this morphotype. These data show that invasive filamentous forms of C. albicans trigger a morphotype-specific activation of neutrophils, which is strongly dependent on neutrophil motility. Therefore, human neutrophils are capable of sensing C. albicans invasion and initiating an appropriate early immune response.

Candida albicans is the most common oral fungal pathogen of humans, but the mechanisms by which C. albicans invades and persists within mucosal epithelium are not clear. To understand oral pathogenesis, we characterized the cellular and molecular mechanisms of epithelial-fungus interactions using reconstituted human oral epithelium (RHE). We observed that hyphal formation facilitates epithelial invasion via both active (physical penetration) and passive (induced endocytosis) processes. Genome wide transcript profiling of C. albicans experimental RHE infection was compared with that from 11 patient samples with pseudomembranous candidiasis to identify genes associated with disease development in vivo. Expression profiles reflected the morphological switch and an adaptive response to neutral pH, non-glucose carbon sources and nitrosative stress. We identified several novel infection-associated genes with unknown function. One gene, upregulated in both RHE infection and patients, named EED1, was essential for maintenance of hyphal elongation. Mutants lacking EED1 showed transient cell elongation on epithelial tissue, which enabled only superficial invasion of epithelial cells. Once inside an epithelial cell, Deltaeed1 cells could proliferate as yeasts or pseudohyphae but remained trapped intracellularly. Our results suggest that the adaptive response and morphology of C. albicans play specific roles for host-fungal interactions during mucosal infections.

Candida infections are common, debilitating and often recurring fungal diseases and a problem of significant clinical importance. Candida albicans, the most virulent of the Candida spp., can cause severe mucosal and life-threatening systemic infections in immunocompromised hosts. Attributes that contribute to C. albicans virulence include adhesion, hyphal formation, phenotypic switching and extracellular hydrolytic enzyme production. The extracellular hydrolytic enzymes, especially the secreted aspartyl proteinases (Saps), are one of few gene products that have been shown to directly contribute to C. albicans pathogenicity. Because C. albicans is able to colonize and infect almost every tissue in the human host, it may be crucial for the fungus to possess a number of similar but independently regulated and functionally distinct secreted proteinases to provide sufficient flexibility in order to survive and promote infection at different niche sites. The aim of this review is to explore the functional roles of the C. albicans proteinases and how they may contribute to the host/pathogen interaction in vivo.