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Invasive candidiasis is among the most life-threatening infections in patients in intensive care units. Although Candida albicans is the leading cause of candidaemia, the incidence of Candida parapsilosis infections is also rising, particularly among the neonates. Due to differences in their biology, these species employ different antifungal resistance and virulence mechanisms and also induce dissimilar immune responses. Previously, it has been suggested that core virulence effecting transcription regulators could be attractive ligands for future antifungal drugs. Although the virulence regulatory mechanisms of C. albicans are well studied, less is known about similar mechanisms in C. parapsilosis. In order to search for potential targets for future antifungal drugs against this species, we analyzed the fungal transcriptome during host-pathogen interaction using an in vitro infection model. Selected genes with high expression levels were further examined through their respective null mutant strains, under conditions that mimic the host environment or influence pathogenicity. As a result, we identified several mutants with relevant pathogenicity affecting phenotypes. During the study we highlight three potentially tractable signaling regulators that influence C. parapsilosis pathogenicity in distinct mechanisms. During infection, CPAR2_100540 is responsible for nutrient acquisition, CPAR2_200390 for cell wall assembly and morphology switching and CPAR2_303700 for fungal viability. ; The project was subsidized by the European Union and co-financed by the European Social Fund. AG was funded by NKFIH NN 113153, by GINOP-2.3.2-15-2016-00035, by GINOP-2.3.3-15-2016-00006. AG and LN were also founded by CNPq (Program Science without borders - 407380/2013-2). TN was supported by the Postdoctoral Fellowship by the Hungarian Academy of Sciences. Research at TG lab was partially funded by grants from Spanish Ministry of Economy and Competitiveness BFU2015-67107 cofounded by European Regional Development Fund (ERDF); from the European Union Seventh Framework Programme (FP7/2007-2013) under grant agreements FP7-PEOPLE-2013-ITN-606786 "ImResFun" and from the European Union's Horizon 2020 research and innovation programme under the Marie Sklodowska-Curie grant agreement No H2020-MSCA-ITN-2014-642095. JDN is partially supported by US NIH grants R21 AI124797-02 and AI52733-06A2. We would like to thank Prof. Geraldine Butler for the KO strategy and parental strains necessary for mutant strain generation. We would like to thank Chetna Tyagi for the help with the in silico data analyses, Mónika Homolya for contributing to the chitinase and chitin synthase expression experiments, Dr. Sándor Kocsubé for the summary table and Tamás Petkovits for the help with the SEM experiments.

This is the final version of the article. Available from Springer Nature via the DOI in this record. ; Aquatic chytrid fungi threaten amphibian biodiversity worldwide owing to their ability to rapidly expand their geographical distributions and to infect a wide range of hosts. Combating this risk requires an understanding of chytrid host range to identify potential reservoirs of infection and to safeguard uninfected regions through enhanced biosecurity. Here we extend our knowledge on the host range of the chytrid Batrachochytrium dendrobatidis by demonstrating infection of a non-amphibian vertebrate host, the zebrafish. We observe dose-dependent mortality and show that chytrid can infect and proliferate on zebrafish tissue. We also show that infection phenotypes (fin erosion, cell apoptosis and muscle degeneration) are direct symptoms of infection. Successful infection is dependent on disrupting the zebrafish microbiome, highlighting that, as is widely found in amphibians, commensal bacteria confer protection against this pathogen. Collectively, our findings greatly expand the limited tool kit available to study pathogenesis and host response to chytrid infection. ; N.L. is supported by the Lister Institute of Preventive Medicine. J.C. and A.E. were funded by the Welsh Government and Higher Education Funding Council for Wales through the Sêr Cymru National Research Network for Low Carbon, Energy and Environment AquaWales Project. C.R.T. and M.J.D. were funded by the Leverhulme Trust. Work in the Fisher laboratory is supported by the UK Natural Environmental Research Council (NERC NE/K014455/1) and the Leverhulme Trust. Work in the Mostowy laboratory is supported by a Wellcome Trust Research Career Development Fellowship (WT097411MA) and the Lister Institute of Preventive Medicine.

Protein phosphorylation and membrane proteins play an important role in the infection of plants by phytopathogenic fungi, given their involvement in signal transduction cascades. Botrytis cinerea is a well-studied necrotrophic fungus taken as a model organism in fungal plant pathology, given its broad host range and adverse economic impact. To elucidate relevant events during infection, several proteomics analyses have been performed in B. cinerea, but they cover only 10% of the total proteins predicted in the genome database of this fungus. To increase coverage, we analysed by LC-MS/MS the first-reported overlapped proteome in phytopathogenic fungi, the "phosphomembranome" of B. cinerea, combining the two most important signal transduction subproteomes. Of the 1112 membrane-associated phosphoproteins identified, 64 and 243 were classified as exclusively identified or overexpressed under glucose and deproteinized tomato cell wall conditions, respectively. Seven proteins were found under both conditions, but these presented a specific phosphorylation pattern, so they were considered as exclusively identified or overexpressed proteins. From bioinformatics analysis, those differences in the membrane-associated phosphoproteins composition were associated with various processes, including pyruvate metabolism, unfolded protein response, oxidative stress response, autophagy and cell death. Our results suggest these proteins play a significant role in the B. cinerea pathogenic cycle. ; This research has been financed by the Spanish Government under project AGL2015-65684-C2-2-R supported by the Spanish Ministry of Economy and Competitiveness. Eva Liñeiro was supported by an FPI grant from the University of Cádiz (2010-152). The CRG/UPF Proteomics Unit is a member of the ProteoRed PRB3 consortium which is supported by grant PT17/0019 of the PE I+D+i 2013–2016 from the Instituto de Salud Carlos III (ISCIII) and ERDF. We acknowledge support from the Spanish Ministry of Economy and Competitiveness, "Centro de Excelencia Severo Ochoa 2013–2017", SEV-2012-0208, and "Secretaria d'Universitats i Recerca del Departament d'Economia i Coneixement de la Generalitat de Catalunya" (2017SGR595).

44 p.-7 fig. ; Multiple fungal species penetrate substrates and accomplish host invasion through the fast, permanent and unidirectional extension of filamentous cells known as hyphae. Polar growth of hyphae results, however, in a significant increase in the distance between the polarity site, which also receives the earliest information about ambient conditions, and nuclei, where adaptive responses are executed. Recent studies demonstrate that these long distances are overcome by signal transduction pathways which convey sensory information from the polarity site to nuclei, controlling development and pathogenesis. The present review compares the striking connections of the mechanisms for long-distance communication in hyphae with those from neurons, and discusses the importance of their study in order to understand invasion and dissemination processes of filamentous fungi, and design strategies for developmental control in the future. ; Work at the UPV/EHU lab was funded by the University of the Basque Country (grant EHUA15/08) and the Basque Government (grant IT599-13). Work at CIB-CSIC was funded by MINECO (BFU2015-66806-R). ; Peer reviewed

This is the final version of the article. Available from Frontiers Media via the DOI in this record. ; One of the key challenges faced by microbial pathogens is invasion of host tissue. Fungal pathogens adopt a number of distinct strategies to overcome host cell defenses, including the development of specialized infection structures, the secretion of proteins that manipulate host responses or cellular organization, and the ability to facilitate their own uptake by phagocytic mechanisms. Key to many of these adaptations is the considerable morphogenetic plasticity displayed by pathogenic species. Fungal pathogens can, for example, shift their growth habit between non-polarized spores, or yeast-like cells, and highly polarized hyphal filaments. These polarized filaments can then elaborate differentiated cells, specialized to breach host barriers. Septins play fundamental roles in the ability of diverse fungi to undergo shape changes and organize the F-actin cytoskeleton to facilitate invasive growth. As a consequence, septins are increasingly implicated in fungal pathogenesis, with many septin mutants displaying impairment in their ability to cause diseases of both plants and animals. In this mini-review, we show that a common feature of septin mutants is the emergence of extra polar outgrowths during morphological transitions, such as emergence of germ tubes from conidia or branches from hyphae. We propose that because septins detect and stabilize membrane curvature, they prevent extra polar outgrowths and thereby focus fungal invasive force, allowing substrate invasion. ; NT is funded by the European Research Council under the European Union's Seventh Framework Programme (FP7/2007-2013)/ERC grant agreement no. 294702 GENBLAST. MM was funded by NSF Grant IOS1051730 from the Developmental Systems Cluster.

Vaginal candidiasis is an extremely common disease predominantly caused by four phylogenetically diverse species: Candida albicans; Candida glabrata; Candida parapsilosis; and Candida tropicalis. Using a time course infection model of vaginal epithelial cells and dual RNA sequencing, we show that these species exhibit distinct pathogenicity patterns, which are defined by highly species-specific transcriptional profiles during infection of vaginal epithelial cells. In contrast, host cells exhibit a homogeneous response to all species at the early stages of infection, which is characterized by sublethal mitochondrial signalling inducing a protective type I interferon response. At the later stages, the transcriptional response of the host diverges in a species-dependent manner. This divergence is primarily driven by the extent of epithelial damage elicited by species-specific mechanisms, such as secretion of the toxin candidalysin by C. albicans. Our results uncover a dynamic, biphasic response of vaginal epithelial cells to Candida species, which is characterized by protective mitochondria-associated type I interferon signalling and a species-specific damage-driven response. ; M.P., H.H., E.I., J.O.P., T.G., G.B. and B.H. received funding from the European Union Horizon 2020 research and innovation programme under the Marie Skłodowska-Curie grant no. 642095 (OPATHY). B.H. also received support from the German Research Foundation within the Collaborative Research Centre/Transregio 124 FungiNet (project C1). M.S.G. was supported by the German Research Foundation Emmy Noether Programme (project no. 434385622/GR 5617/1-1). We acknowledge the support of the Spanish Ministry of Science, Innovation and Universities (grant no. PGC2018-099921-B-I00) to the European Molecular Biology Laboratory partnership, the Centro de Excelencia Severo Ochoa and the CERCA Programme/Generalitat de Catalunya. We thank C. Kämnitz from the Electron Microscopy Center in Jena for the sample preparation for TEM. The schematic models in Figs. 4–6 were created with images adapted from Servier Medical Art (Servier). ; Peer Reviewed ; Postprint (author's final draft)

Fungal pathogens pose an increasingly worrying threat to human health, food security and ecosystem diversity. To tackle fungal infections and improve current diagnostic and therapeutic tools it is necessary to understand virulence and antifungal drug resistance mechanisms in diverse species. Recent advances in genomics approaches have provided a suitable framework to understand these phenotypes, which ultimately depend on genetically encoded determinants. In this work, we review how the study of genome sequences has been key to ascertain the bases of virulence and drug resistance traits. We focus on the contribution of comparative genomics, population genomics and directed evolution studies. In addition, we discuss how different types of genomic mutations (small or structural variants) contribute to intraspecific differences in virulence or drug resistance. Finally, we review current challenges in the field and anticipate future directions to solve them. In summary, this work provides a short overview of how genomics can be used to understand virulence and drug resistance in fungal pathogens. ; TG group acknowledges support from the Spanish Ministry of Science and Innovation for grant PGC2018-099921-B-I00, cofounded by European Regional Development Fund (ERDF); from the Catalan Research Agency (AGAUR) SGR423; from the European Union's Horizon 2020 research and innovation programme (ERC-2016-724173); from the Gordon and Betty Moore Foundation (Grant GBMF9742); from the 'La Caixa' foundation (Grant LCF/PR/HR21/00737), and from the Instituto de Salud Carlos III (INB Grant PT17/0009/0023 and CIBERINFEC CB21/13/00061- ISCIII-SGEFI/ERDF). MAST received a predoctoral fellowship from the 'Caixa' Foundation (LCF/BQ/DR19/11740023). ; Peer Reviewed ; Postprint (published version)

Abstract
Allergic airways disease (AAD) is a collective term for respiratory disorders that can be exacerbated upon exposure to airborne allergens. The contribution of fungal allergens to the pathogenesis of AAD has become well established over recent years and can be present in the air we breathe as bioaerosols. Understanding the mechanisms allergy to fungal bioaerosols can help to better manage symptoms and conditions, reducing the burden of disease. We conducted a comprehensive review to collate the current understanding of mechanisms involved in the allergic response to fungal bioaerosols in airway epithelia. To be included in the final analysis, studies were compared to defined inclusion/exclusion criteria, relevant data was extracted, and each paper was assessed using a quality scoring tool. The search string provided 440 initial results with 60 studies selected for final analysis. 120 potential target mechanisms were identified within the selected studies and 17 were identified in more than 5 studies. The major pathways identified in allergic response include 1) a role for proteases and the activation of the PAR2 receptor, 2) release of IL33, 3) EGFR pathway and mucin expression, and 4) a skewing towards a Th2 profile promoting eosinophil recruitment. However, there was conflicting evidence when comparing results from individual studies. Thus, further research is required to better understand which fungi/fungal components drive the allergic response.
Funding: This work has been funded under the NIHR Health Protection Research Unit for Environmental Exposures and Health with Imperial College London

© 2019 Marín-Menguiano et al. ; Fungal pathogenesis depends on accurate secretion and location of virulence factors which drive host colonization. Protein glycosylation is a common posttranslational modification of cell wall components and other secreted factors, typically required for correct protein localization, secretion and function. Thus, the absence of glycosylation is associated with animal and plant pathogen avirulence. While the relevance of protein glycosylation for pathogenesis has been well established, the main glycoproteins responsible for the loss of virulence observed in glycosylation-defective fungi have not been identified. Here, we devise a proteomics approach to identify such proteins and use it to demonstrate a role for the highly conserved protein disulfide isomerase Pdi1 in virulence. We show that efficient Pdi1 N-glycosylation, which promotes folding into the correct protein conformation, is required for full pathogenic development of the corn smut fungus Ustilago maydis. Remarkably, the observed virulence defects are reminiscent of those seen in glycosylation-defective cells suggesting that the N-glycosylation of Pdi1 is necessary for the full secretion of virulence factors. All these observations, together with the fact that Pdi1 protein and RNA expression levels rise upon virulence program induction, suggest that Pdi1 glycosylation is important for normal pathogenic development in U. maydis. Our results provide new insights into the role of glycosylation in fungal pathogenesis. ; MMM was supported by P09- AGR-5241 Junta de Andalucía. IMS was awarded by BES-2014-069149 MINECO AEI/FEDER, UE, Spain. This work was supported by Junta de Andalucía P09- AGR-5241 grant (https://www.juntadeandalucia.es/organismos/economiaconocimientoempresasyuniversidad.html), Spanish Government BIO2013–48858-P and BIO2016-80180-P grants from MINECO AEI/FEDER, UE to JII and Ramon y Cajal program, RyC-2016-19659 to AFA (http://www.mineco.gob.es/portal/site/mineco/).

Cacao (Theobroma cacao L.) is an important commodity worldwide, but its production is limited by destructive diseases such witches' broom, due to the fungus Moniliophthora perniciosa. The high recombination rate and genetic variability of this fungus promoted resistance breakdown of cacao and annihilated the efforts made by the Brazilian government to reduce the disease impact on plantations. According to the literature, pathogenesis related-proteins (PRs) plays an important role in defense/resistance mechanisms of the plant submitted to various biotic and abiotic stresses. A cDNA clone encoding a PR10 (named TcPR10) was obtained from a cacao-M. perniciosa EST library. The corresponding recombinant protein (expressed in Escherichia coli BL21(DE3)) present a strong antifungal activity against M. perniciosa, as previously demonstrated. In this work, we developed a proteomic analysis of the fungus cultivated in the presence of TcPR10, using 2DE-MS/MS. M. perniciosa was grown in CPD 2% agar medium; after 15 days, the fungal hyphae were broken and were grown in presence of 3 ?g/mL of TcPR10 for 1h.Then, the total proteins were extracted using the ADP method, followed by a simple cleaning using the method of SDS-dense and phenol. The quantification was made using a 2-D quantification kit. The proteins were extracted in triplicate and separated using a 12% bi-dimensional SDS-PAGE gel. The 2D map analysis showed approximately 300 "spots" per gel (control and one hour treatment) with differential protein expression pattern. The analysis using a mass spectrometry (naniESI-Q-TOF) was made for the identification of the spots. We identified several proteins involved in fungal metabolism, carbohydrates/proteins metabolism, related proteins to growth and phytotoxics proteins. More spots have been identified to better understand the mechanism of fungi response to protein PR10. (Texte intégral)

We conducted a systematic review of the 2012-2013 multistate fungal meningitis epidemic in the United States from the perspectives of clinical response, outbreak investigation, and epidemiology. Articles focused on clinical response, outbreak investigation, and epidemiology were included, whereas articles focused on compounding pharmacies, legislation and litigation, diagnostics, microbiology, and pathogenesis were excluded. We reviewed 19 articles by use of the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) framework. The source of the fungal meningitis outbreak was traced to the New England Compounding Center in Massachusetts, where injectable methylprednisolone acetate products were contaminated with the predominant pathogen, Exserohilum rostratum. As of October 23, 2013, the final case count stood at 751 patients and 64 deaths, and no additional cases are anticipated. The multisectoral public health response to the fungal meningitis epidemic from the hospitals, clinics, pharmacies, and the public health system at the local, state, and federal levels led to an efficient epidemiological investigation to trace the outbreak source and rapid implementation of multiple response plans. This systematic review reaffirms the effective execution of a multisectoral public health response and efficient delivery of the core functions of public health assessment, policy development, and service assurances to improve population health.

We conducted a systematic review of the 2012-2013 multistate fungal meningitis epidemic in the United States from the perspectives of clinical response, outbreak investigation, and epidemiology. Articles focused on clinical response, outbreak investigation, and epidemiology were included, whereas articles focused on compounding pharmacies, legislation and litigation, diagnostics, microbiology, and pathogenesis were excluded. We reviewed 19 articles by use of the PRISMA (Preferred Reporting Items for Systematic Reviews and Meta-Analyses) framework. The source of the fungal meningitis outbreak was traced to the New England Compounding Center in Massachusetts, where injectable methylprednisolone acetate products were contaminated with the predominant pathogen, Exserohilum rostratum. As of October 23, 2013, the final case count stood at 751 patients and 64 deaths, and no additional cases are anticipated. The multisectoral public health response to the fungal meningitis epidemic from the hospitals, clinics, pharmacies, and the public health system at the local, state, and federal levels led to an efficient epidemiological investigation to trace the outbreak source and rapid implementation of multiple response plans. This systematic review reaffirms the effective execution of a multisectoral public health response and efficient delivery of the core functions of public health assessment, policy development, and service assurances to improve population health.

Not Available ; Rhizoctonia solani AG1-1A is a necrotrophic fungus that causes sheath blight disease in rice. The reliable resistant source against this phytopathogenic fungus is not available in the gene pool of rice. Better understanding of pathogen genomics and gene regulatory networks are critical to devise alternate strategies for developing resistance against this noxious pathogen. In this study, miRNA-like RNAs (milRNAs) of an Indian strain of R. solani were identified by deep sequencing of small RNAs. We identified 128 known and 22 novel milRNAs from 20,963,123 sequence reads. These milRNAs showed 1725 target genes in the fungal genome which include genes associated with growth, development, pathogenesis and virulence of R. solani. Notably, these fungal milRNAs showed their target genes in host (rice) genome also which were later verified by qRT-PCR. The host target genes are associated with auxin metabolism, hypersensitive response, defense genes, and genes related to growth and development of rice. Osa-vacuolar-sorting receptor precursor: Rhi-milR-13, Osa-KANADI1:Rhi-milR-124, Osa-isoflavone reductase: Rhi-milR-135, Osa-nuclear transcription factor Y:Rhi-milR-131, Osa-NB-ARC domain containing protein: Rhi-milR-18, and Osa-OsFBX438: Rhi-milR-142 are notable potential regulons of host target genes: fungal milRNAs that need to be investigated for better understanding of the crosstalk of RNAi pathways between R. solani and rice. The detailed expression analysis of 17 milRNAs by qRT-PCR was analysed during infection at different time points of inoculation, at different growth stages of the host, in four different genotypes of the host, and also in four different strains of fungi which revealed differential regulation of milRNAs associated with pathogenesis and virulence. This study highlights several important findings on fungal milRNAs which need to be further studied and characterized to decipher the gene expression and regulation of this economically important phytopathogen. ; Department of Biotechnology (DBT), Government of India (GrantNo. BT/PR6466/COE/34/16/2012).

The hybrid peptide BP178 (KKLFKKILKYLAGPAGIGKFLHSAKKDEL-OH), derived from BP100 (KKLFKKILKYL) and magainin (1-10), and engineered for plant expression, had a strong bactericidal activity but not fungicidal. Moreover, the preventive spray of tomato plants with BP178 controlled infections by the plant pathogenic bacteria Pseudomonas syringae pv. tomato and Xanthomonas campestris pv. vesicatoria, as well as the fungus Botrytis cinerea. The treatment of tomato plants with BP178 induced the expression of several genes according to microarray and RT-qPCR analysis. Upregulated genes coded for several pathogenesis-related proteins, including PR1, PR2, PR3, PR4, PR5, PR6, PR7, PR9, PR10, and PR14, as well as transcription factors like ethylene transcription factors, WRKY, NAC and MYB, involved in the salicylic acid, jasmonic acid, and ethylene-signaling pathways. BP178 induced a similar gene expression pattern to flg15 according to RT-qPCR analysis, whereas the parent peptide BP100 did not trigger such as a strong plant defense response. It was concluded that BP178 was a bifunctional peptide protecting the plant against pathogen infection through a dual mechanism of action consisting of antimicrobial activity against bacterial pathogens and plant defense elicitation on plant host ; We thank the Government of the Generalitat de Catalunya (Xarxa de Referencia en Biotecnología, and Xarxa de Referència en Tecnologia dels Aliments, SGR09626, 2008SGR812, and 2014SGR697) for support. The research group is accredited by SGR 2014-697 and TECNIO net from Catalonia. We also thank the University of Girona for a grant for research improvement 2015–2018