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Human fungal pathogens are a deadly and underappreciated risk to global health that most severely affect immunocompromised individuals. A virulence attribute shared by some of the most clinically relevant fungal species is their ability to survive inside macrophages and escape from these immune cells. In this review, we discuss the mechanisms behind intracellular survival and elaborate how escape is mediated by lytic and non‐lytic pathways as well as strategies to induce programmed host cell death. We also discuss persistence as an alternative to rapid host cell exit. In the end, we address the consequences of fungal escape for the host immune response and provide future perspectives for research and development of targeted therapies. [ABSTRACT FROM AUTHOR]

Cytolytic proteins and peptide toxins are classical virulence factors of several bacterial pathogens which disrupt epithelial barrier function, damage cells and activate or modulate host immune responses. Such toxins have not been identified previously in human pathogenic fungi. Here we identify the first, to our knowledge, fungal cytolytic peptide toxin in the opportunistic pathogen Candida albicans. This secreted toxin directly damages epithelial membranes, triggers a danger response signalling pathway and activates epithelial immunity. Membrane permeabilization is enhanced by a positive charge at the carboxy terminus of the peptide, which triggers an inward current concomitant with calcium influx. C. albicans strains lacking this toxin do not activate or damage epithelial cells and are avirulent in animal models of mucosal infection. We propose the name 'Candidalysin' for this cytolytic peptide toxin; a newly identified, critical molecular determinant of epithelial damage and host recognition of the clinically important fungus, C. albicans., The ability of mucosal surfaces to discriminate between commensal and pathogenic microbes is essential to human health. The fungus Candida albicans is normally a benign member of the human microbiota, [...]

Although less prevalent than its relative Candida albicans , the yeast Candida glabrata is a successful pathogen of humans, which causes life-threatening candidiasis. It is thus vital to understand the pathogenicity mechanisms and contributing genes in C. glabrata. However, gene complementation as a tool for restoring the function of a previously deleted gene is not standardized in C. glabrata , and it is less frequently used than in C. albicans. In this study, we established a gene complementation strategy using genomic integration at the TRP1 locus. We prove that our approach can not only be used for integration of complementation cassettes, but also for overexpression of markers like fluorescent proteins and the antigen ovalbumin, or of potential pathogenicity-related factors like the biotin transporter gene VHT1. With urea amidolyase Dur1,2 as an example, we demonstrate the application of the gene complementation approach for the expression of sequence-modified genes. With this approach, we found that a lysine-to-arginine mutation in the biotinylation motif of Dur1,2 impairs urea-dependent growth of C. glabrata and C. albicans. Taken together, the TRP1 -based gene complementation approach is a valuable tool for investigating novel gene functions and for elucidating their role in the pathobiology of C. glabrata. [ABSTRACT FROM AUTHOR]

Metals are essential for life, and they play a central role in the struggle between infecting microbes and their hosts. In fact, an important aspect of microbial pathogenesis is the 'nutritional immunity', in which metals are actively restricted (or, in an extended definition of the term, locally enriched) by the host to hinder microbial growth and virulence. Consequently, fungi have evolved often complex regulatory networks, uptake and detoxification systems for essential metals such as iron, zinc, copper, nickel and manganese. These systems often differ fundamentally from their bacterial counterparts, but even within the fungal pathogens we can find common and unique solutions to maintain metal homeostasis. Thus, we here compare the common and species-specific mechanisms used for different metals among different fungal species-focusing on important human pathogens such as Candida albicans, Aspergillus fumigatus or Cryptococcus neoformans, but also looking at model fungi such as Saccharomyces cerevisiae or A. nidulans as well-studied examples for the underlying principles. These direct comparisons of our current knowledge reveal that we have a good understanding how model fungal pathogens take up iron or zinc, but that much is still to learn about other metals and specific adaptations of individual species-not the least to exploit this knowledge for new antifungal strategies. [ABSTRACT FROM AUTHOR]

Secretory aspartyl proteinases (Saps) of Candida albicans are key virulence traits which cause inflammasome-dependent, aseptic inflammation in a mouse model of vaginitis. In this paper, neutrophil migration in response to Sap2, Sap6 and chemo-attractive products released from Saptreated vaginal epithelium was measured in vitro, ex vivo and in vivo. Our results show that Sap2 and Sap6 induce neutrophil migration and production of potent chemoattractive chemokines such as IL-8 and MIP-2 by vaginal epithelial cells. Our data suggest that at least part of MIP-2 production depends upon IL-1β activity. The vaginal fluid of Candida-infected mice contained a heat-labile inhibitor of neutrophil candidacidal activity that was absent from the vaginal fluid of Sap-treated mice. Overall, our data provide additional information on the capacity of C. albicans Saps to cause aseptic vaginal inflammation and highlight the potential role of some chemokines released from vaginal epithelial cells in this phenomenon. [ABSTRACT FROM AUTHOR]

Candida glabrata is a successful human opportunistic pathogen which causes superficial but also life-threatening systemic infections. During infection, C. glabrata has to cope with cells of the innate immune system such as macrophages, which belong to the first line of defense against invading pathogens. Candida glabrata is able to survive and even replicate inside macrophages while causing surprisingly low damage and cytokine release. Here, we present an overview of recent studies dealing with the interaction of C. glabrata with macrophages, from phagocytosis to intracellular growth and escape. We review the strategies of C. glabrata that permit intracellular survival and replication, including poor host cell activation, modification of phagosome maturation and phagosome pH, adaptation to antimicrobial activities, and mechanisms to overcome the nutrient limitations within the phagosome. In summary, these studies suggest that survival within macrophages may be an immune evasion and persistence strategy of C. glabrata during infection. [ABSTRACT FROM AUTHOR]

Candida glabrata currently ranks as the second most frequent cause of invasive candidiasis. Our previous work has shown that C. glabrata is adapted to intracellular survival in macrophages and replicates within non-acidified late endosomal-stage phagosomes. In contrast, heat killed yeasts are found in acidified matured phagosomes. In the present study, we aimed at elucidating the processes leading to inhibition of phagosome acidification and maturation. We show that phagosomes containing viable C. glabrata cells do not fuse with pre-labeled lysosomes and possess low phagosomal hydrolase activity. Inhibition of acidification occurs independent of macrophage type (human/murine), differentiation (M1-/M2-type) or activation status (vitamin D3 stimulation). We observed no differential activation of macrophage MAPK or NFκB signaling cascades downstream of pattern recognition receptors after internalization of viable compared to heat killed yeasts, but Syk activation decayed faster in macrophages containing viable yeasts. Thus, delivery of viable yeasts to non-matured phagosomes is likely not triggered by initial recognition events via MAPK or NFκB signaling, but Syk activation may be involved. Although V-ATPase is abundant in C. glabrata phagosomes, the influence of this proton pump on intracellular survival is low since blocking V-ATPase activity with bafilomycin A1 has no influence on fungal viability. Active pH modulation is one possible fungal strategy to change phagosome pH. In fact, C. glabrata is able to alkalinize its extracellular environment, when growing on amino acids as the sole carbon source in vitro. By screening a C. glabrata mutant library we identified genes important for environmental alkalinization that were further tested for their impact on phagosome pH. We found that the lack of fungal mannosyltransferases resulted in severely reduced alkalinization in vitro and in the delivery of C. glabrata to acidified phagosomes. Therefore, protein mannosylation may play a key role in alterations of phagosomal properties caused by C. glabrata. [ABSTRACT FROM AUTHOR]

Although less prevalent than its relative Candida albicans, the yeast Candida glabrata is a successful pathogen of humans, which causes life-threatening candidiasis. It is thus vital to understand the pathogenicity mechanisms and contributing genes in C. glabrata. However, gene complementation as a tool for restoring the function of a previously deleted gene is not standardized in C. glabrata, and it is less frequently used than in C. albicans. In this study, we established a gene complementation strategy using genomic integration at the TRP1 locus. We prove that our approach can not only be used for integration of complementation cassettes, but also for overexpression of markers like fluorescent proteins and the antigen ovalbumin, or of potential pathogenicity-related factors like the biotin transporter gene VHT1. With urea amidolyase Dur1,2 as an example, we demonstrate the application of the gene complementation approach for the expression of sequence-modified genes. With this approach, we found that a lysine-to-arginine mutation in the biotinylation motif of Dur1,2 impairs urea-dependent growth of C. glabrata and C. albicans. Taken together, the TRP1-based gene complementation approach is a valuable tool for investigating novel gene functions and for elucidating their role in the pathobiology of C. glabrata., (© The Author(s) 2020. Published by Oxford University Press on behalf of FEMS.)