Transcriptome Reveals Roles of Lignin-Modifying Enzymes and Abscisic Acid in the Symbiosis of Mycena and Gastrodia elata.

Gastrodia elata is a well-known medicinal and heterotrophic orchid. Its germination, limited by the impermeability of seed coat lignin and inhibition by abscisic acid (ABA), is triggered by symbiosis with fungi such as Mycena spp. However, the molecular mechanisms of lignin degradation by Mycena and ABA biosynthesis and signaling in G. elata remain unclear. In order to gain insights into these two processes, this study analyzed the transcriptomes of these organisms during their dynamic symbiosis. Among the 25 lignin-modifying enzyme genes in Mycena, two ligninolytic class II peroxidases and two laccases were significantly upregulated, most likely enabling Mycena hyphae to break through the lignin seed coats of G. elata. Genes related to reduced virulence and loss of pathogenicity in Mycena accounted for more than half of annotated genes, presumably contributing to symbiosis. After coculture, upregulated genes outnumbered downregulated genes in G. elata seeds, suggesting slightly increased biological activity, while Mycena hyphae had fewer upregulated than downregulated genes, indicating decreased biological activity. ABA biosynthesis in G. elata was reduced by the downregulated expression of 9-cis-epoxycarotenoid dioxygenase (NCED-2), and ABA signaling was blocked by the downregulated expression of a receptor protein (PYL12-like). This is the first report to describe the role of NCED-2 and PYL12-like in breaking G. elata seed dormancy by reducing the synthesis and blocking the signaling of the germination inhibitor ABA. This study provides a theoretical basis for screening germination fungi to identify effective symbionts and for reducing ABA inhibition of G. elata seed germination.

Moniliophthora perniciosa, the mushroom causing witches' broom disease of cacao: Insights into its taxonomy, ecology and host range in Brazil.

Witches' broom caused by Moniliophthora perniciosa is the main disease of cacao (Theobroma cacao) in Brazil. The fungus is known to occur on other host families and these populations have been addressed in the literature as biotypes: C (Malvaceae); H (Malpighiaceae); L (Bignoniaceae) and S (Solanaceae). No complete elucidation of the phylogenetic relationships of isolates obtained from this disparate host range appears in the literature. One member of H (ex Heteropterys acutifolia) has been described as a distinct species. But should other biotypes be also recognized as distinct taxa? In the present study, a survey yielding 24 isolates of M. perniciosa from ten hosts and covering a wide range of geographic regions in Brazil was undertaken. These isolates were compared with those from T. cacao using three DNA regions for the phylogenetic analyses: ITS, LSU and RPB1. Morphology was also examined. All isolates in this study were found to belong to M. perniciosa, including the population from H. acutifolia, formerly treated as Moniliophthora brasiliensis but reduced here to a synonym of M. perniciosa. This species ranged from pathogenic to a previously unreported occurrence as a non-pathogenic endophyte in the Atlantic rainforest tree Allophylus edulis (Sapindaceae). M. perniciosa was recorded on a range of solanaceous hosts (16 species) over a wide variety of ecosystems. The ecological and evolutionary significance of these novel findings are discussed.

Unraveling the mechanisms of resistance to Sclerotium rolfsii in peanut (Arachis hypogaea L.) using comparative RNA-Seq analysis of resistant and susceptible genotypes.

Stem rot, a devastating fungal disease of peanut, is caused by Sclerotium rolfsii. RNA-sequencing approaches have been used to unravel the mechanisms of resistance to stem rot in peanut over the course of fungal infection in resistant (NRCG-CS85) and susceptible (TG37A) genotypes under control conditions and during the course of infection. Out of about 290 million reads, nearly 251 million (92.22%) high-quality reads were obtained and aligned to the Arachis duranensis and Arachis ipaensis genomes with the average mapping of 78.91% and 78.61%, respectively. In total, about 48.6% of genes were commonly regulated, while approximately 21.8% and 29.6% of uniquely regulated genes from A. duranensis and A. ipaensis genomes, respectively, were identified. Several annotated transcripts, such as receptor-like kinases, jasmonic acid pathway enzymes, and transcription factors (TFs), including WRKY, Zinc finger protein, and C2-H2 zinc finger, showed higher expression in resistant genotypes upon infection. These transcripts have a known role in channelizing the downstream of pathogen perception. The higher expression of WRKY transcripts might have induced the systemic acquired resistance (SAR) by the activation of the jasmonic acid defense signaling pathway. Furthermore, a set of 30 transcripts involved in the defense mechanisms were validated with quantitative real-time PCR. This study suggested PAMP-triggered immunity as a probable mechanism of resistance, while the jasmonic acid signaling pathway was identified as a possible defense mechanism in peanut. The information generated is of immense importance in developing more effective ways to combat the stem rot disease in peanut.

Dimethyl Labeling-Based Quantitative Proteomics of Recalcitrant Cocoa Pod Tissue.

Dimethyl labeling is a type of stable-isotope labeling suitable for creating isotopic variants of peptides and thus be utilized for quantitative proteomics experiments. Labeling is achieved through a reductive amination/alkylation reaction using the low-cost reagents formaldehyde and cyanoborohydride, resulting in dimethylation of free amine groups of Lys and N-termini. Availability of isotopomeric forms of these reagents allows for the generation of up to six different isotopic variants. Here we describe the application of dimethylation to create two isotopic variants, light and heavy, differing in 4 Da, to label the total tryptic digest peptides of cocoa pod extracted from healthy pods from cultivars susceptible and resistant to the fungal disease called "frosty pod" caused by Moniliophthora roreri.

Hydrosoluble phylloplane components of Theobroma cacao modulate the metabolism of Moniliophthora perniciosa spores during germination.

The surface of plants forms a defense barrier that directly inhibits the first point of contact of microorganisms with the host. To understand this defense mechanism in Moniliophthora perniciosa interaction with Theobroma cacao cv Catongo, the aim of this study was to compare the changes in protein expression in basidiospores of the fungus M. perniciosa in response the leaf water washes (LWW) of two contrasting cocoa varieties for resistance to witches' broom disease. A total of 8.1 × 108 basidiospores were used for each treatment containing washed leaf material. Germinated basidiospores in the absence of LWW were used as control. The proteomic analysis was performed by the 2D-PAGE technique combined with mass spectrometry (MS). Protein extraction was based on the SDS-dense method followed by sonication for cell disruption and phenol extraction. Sixty-four proteins had accumulation of variation when compared to the control (no LWW). Proteins were identified associated with energy (ATP synthase) and protein (BiP) metabolism, whose accumulation was reduced by basidiospores germinated in leaf wash from Catongo cocoa. The reduction in ATP synthase of the basidiospores germinated the Catongo LWW suggests a shift from aerobic to fermentative metabolism of the fungus in response to components of the LWW. Furthermore, proteins involved in virulence were identified along with fungal resistance to polyketide cyclase, glycoside hydrolase, multidrug transporter protein (SFM) and proteins related to oxidative stress and fermentation, such as catalase A and alcohol dehydrogenase (ADH). The data showed an effect of cocoa phylloplane substances on the germination of fungal basidiospores through differences in protein expression patterns in the presence of LWW of the CCN51 and Catongo genotypes. These results may reveal mechanisms of resistance, host susceptibility and pathogen virulence.

Chocolate Under Threat from Old and New Cacao Diseases.

Theobroma cacao, the source of chocolate, is affected by destructive diseases wherever it is grown. Some diseases are endemic; however, as cacao was disseminated from the Amazon rain forest to new cultivation sites it encountered new pathogens. Two well-established diseases cause the greatest losses: black pod rot, caused by several species of Phytophthora, and witches' broom of cacao, caused by Moniliophthora perniciosa. Phytophthora megakarya causes the severest damage in the main cacao producing countries in West Africa, while P. palmivora causes significant losses globally. M. perniciosa is related to a sister basidiomycete species, M. roreri which causes frosty pod rot. These Moniliophthora species only occur in South and Central America, where they have significantly limited production since the beginnings of cacao cultivation. The basidiomycete Ceratobasidium theobromae causing vascular-streak dieback occurs only in South-East Asia and remains poorly understood. Cacao swollen shoot disease caused by Cacao swollen shoot virus is rapidly spreading in West Africa. This review presents contemporary research on the biology, taxonomy and genomics of what are often new-encounter pathogens, as well as the management of the diseases they cause.

Heterologous expression of an alternative oxidase from Moniliophthora perniciosa in Saccharomyces cerevisiae: Antioxidant function and in vivo platform for the study of new drugs against witches' broom disease.

The fungus Moniliophthora perniciosa is the causal agent of witches' broom disease (WBD), one of the most devastating diseases of cacao, the chocolate tree. Many strategies to control WBD have been tested so far, including the use of agrochemicals such as the strobilurins. Strobilurins are fungicides of the QoI family, and they are used in the control of a wide array of fungal diseases in many different crops, including cereals, field crops, fruits, tree nuts, and vegetables. These drugs act by specifically inhibiting fungal respiration at the Qo site of complex III, which is a component of the main mitochondrial respiratory chain. However, M. perniciosa is resistant to this family of chemicals. It has been postulated that this resistant phenotype is, at least in part, a result of the strong ability of this fungus to counteract the oxidative stress generated by the impairment of the main mitochondrial respiratory chain, through the activation of an alternative oxidase (Mp-AOX). To test this hypothesis, we expressed functional mitochondria-localized Mp-AOX in the model yeast Saccharomyces cerevisiae. We demonstrated that heterologous expression of Mp-AOX strongly inhibits hydrogen peroxide production by mitochondria. It also diminishes the total cell amount of oxidized glutathione (GSSG), resulting in a fifty-fold higher GSH/GSSG ratio in cells expressing Mp-AOX than in wild type cells. In addition, Mp-AOX activity decreases yeast growth rate and leads to low biomass production. Therefore, we propose the use of this heterologous expression system to direct the development of new inhibitors of fungal AOX by comparing the differences in optical density of Mp-AOX-expressing cells in the presence and absence of potential AOX inhibitors. Together, our results confirm the antioxidant role of Mp-AOX and provide an in vivo platform to be used in the screening of new fungicides based on Mp-AOX inhibition.

Macrocyclic Trichothecene Mycotoxins from a Deadly Poisonous Mushroom, Podostroma cornu-damae.

Three new macrocyclic trichothecenes (1-3) and five known related compounds (4-8) were isolated from the MeOH extract of a plate culture of the fungus Podostroma cornu-damae, a deadly poisonous mushroom. Miophytocen D (1) is a rearranged macrocyclic type D trichothecene, featuring a bicyclo-[6.5]dodecahydrocyclopenta[ b]chromene scaffold, and the structures of new compounds (1-3) were delineated by the combination of 1D and 2D NMR spectroscopic experiments and HRESIMS, modified Mosher's esterification, and quantum chemical ECD calculations. The isolated compounds (1-8) were evaluated for cytotoxicity against four human breast cancer cell lines (Bt549, HCC70, MDA-MB-231, and MDA-MB-468). Compounds 4, 6, and 8 exhibited significant cytotoxic effects against the breast cancer cell lines, with IC50 values in the range of 0.02-80 nM, which is stronger than doxorubicin, the positive control, and a structure-activity relationship was suggested.

Acute and developmental toxicity assessment of erincine A-enriched Hericium erinaceus mycelia in Sprague-Dawley rats.

This study aimed to establish an in vitro model to confirm the efficacy of erinacine A-enriched Hericium erinaceus (EAHE) mycelia and investigate its potential adverse effects in a preclinical experimental setting, including an assessment on the oral administration of EAHE mycelia in acute and prenatal developmental toxicity tests. At a single dose of 5000 mg/kg body weight, EAHE mycelia elicited no death or treatment-related signs of toxicity in ten Sprague-Dawley rats of both sexes during the 14 days of the experimental period. After considering the recommended dose range of EAHE mycelia from the acute toxicity test as well as the therapeutic doses, EAHE mycelia was administered to 66 pregnant rats in the low, medium, and high-dose groups by gavage at 875, 1750, and 2625 mg/kg body weight, respectively. All dams were subjected to a Caesarean section on the 20th day of pregnancy, and the fetuses were examined for any morphological abnormalities. Results indicated that weight of uterus, fetal body weight, number of corpora lutea, implantation sites, pre-implantation loss, and post-implantation loss of the treatment groups and the control group exhibited no statistical difference. In addition, no significant differences were observed in the fetal external, organ, and skeletal examinations. Taken together, it can be concluded that EAHE mycelia is considered safe and practically nontoxic for consumption within the appropriate doses and investigation period in this study.

Moniliophthora roreri, causal agent of cacao frosty pod rot.

Taxonomy: Moniliophthora roreri (Cif.) H.C. Evans et al. ; Phylum Basidiomycota; Class Agaricomycetes; Order Agaricales; Family Marasmiaceae; Genus Moniliophthora. Biology: Moniliophthora roreri attacks Theobroma and Herrania species causing frosty pod rot. Theobroma cacao (cacao) is the host of major economic concern. Moniliophthora roreri is a hemibiotroph with a long biotrophic phase (45-90 days). Spore masses, of apparent asexual origin, are produced on the pod surface after initiation of the necrotrophic phase. Spores are spread by wind, rain and human activity. Symptoms of the biotrophic phase can include necrotic flecks and, in some cases, pod malformation, but pods otherwise remain asymptomatic. Relationship to Moniliophthora perniciosa: Moniliophthora roreri and Moniliophthora perniciosa, causal agent of witches' broom disease of cacao, are closely related. Their genomes are similar, including many of the genes they carry which are considered to be important in the disease process. Moniliophthora perniciosa, also a hemibiotroph, has a typical basidiomycete lifestyle and morphology, forming clamp connections and producing mushrooms. Basidiospores infect meristematic tissues including flower cushions, stem tips and pods. Moniliophthora roreri does not form clamp connections or mushrooms and infects pods only. Both pathogens are limited to the Western Hemisphere and are a threat to cacao production around the world. Agronomic importance: Disease losses caused by frosty pod rot can reach 90% and result in field abandonment. Moniliophthora roreri remains in the invasive phase in the Western Hemisphere, not having reached Brazil, some islands within the Caribbean and a few specific regions within otherwise invaded countries. DISEASE MANAGEMENT: The disease can be managed by a combination of cultural (for example, maintenance of tree height and removal of infected pods) and chemical methods. These methods benefit from regional application, but can be cost prohibitive. Breeding for disease resistance offers the greatest potential for frosty pod rot management and new tolerant materials are becoming available.

Mushroom poisoning: a retrospective study concerning 11-years of admissions in a Swiss Emergency Department.

Wild mushroom intoxication is an unusual cause of toxic ingestion in Europe. A great diversity of clinical symptoms may arise depending on the variety of wild mushrooms ingested. These initial symptoms are often non-specific, with frequent gastrointestinal symptoms, and have no direct correlation with the outcome. Therefore, management of mushroom poisoning and risk evaluation are a challenge for emergency clinicians. We retrospectively reviewed all cases of mushroom poisoning identified in the ED database spanning 11 years. Demographic and clinical data, time from consumption to symptoms, type of mushrooms, the number of patients presenting at the same time, treatment(s) provided, length of stay, discharge diagnosis, in-hospital mortality, and serious complications were evaluated. We identify 87 cases of mushroom poisoning. The most common symptoms are nausea and vomiting (71 cases, 82%), followed by diarrhea (68%), syncope (10%), abdominal pain (8%), and hallucinations (7%). Sixty-four patients (74%) exhibited early symptoms (appearance <6 h after ingestion) and 23 (26%) late symptoms (appearance >6 h after ingestion). Eleven patients (13%) required hospitalization over 24 h. Patients with late symptoms tended to have longer in-hospital lengths of stay. Only one patient had Amanita phalloides intoxication, with a favorable outcome. Thirty-eight patients (44%) were involved in cluster presentations. Mushroom poisoning is an unusual but potentially severe form of intoxication. Patients presenting with late-appearing symptoms (>6 h) are associated with a higher risk of A. phalloides intoxication, and therefore require specific investigation and management.

Evaluación del efecto tóxico de hongos Basidiomycota en la eclosión de quistes de Artemia franciscana / Evaluation of the toxicity of Basidiomycota fungi on the hatching of Artemia franciscana cysts

Antecedentes. El consumo de hongos silvestres se ha incrementado en los últimos años; sin embargo, no todos son comestibles y algunos son causantes de varios tipos de envenenamiento. Por esto es necesario realizar estudios que aporten información de su toxicidad. Artemia franciscana es un crustáceo que se emplea en ensayos de toxicidad con una gran aplicación en la detección de toxinas fúngicas. Objetivos. Determinar el porcentaje de inhibición de la eclosión y mortalidad de quistes de A. franciscana producidos por extractos de hongos de la división Basidiomycota. Métodos. Se prepararon extractos acuosos de basidiomas de 15 especies de basidiomicetos recolectados en Jalisco (México) y se probó el efecto de diferentes concentraciones sobre quistes de A. franciscana. Se utilizaron dicromato de potasio y agua de mar como controles positivo y negativo, respectivamente. Se determinaron los porcentajes de inhibición de la eclosión y de la mortalidad de los quistes de A. franciscana. Resultados. Trece de las 15 especies estudiadas afectaron en más del 80% la eclosión de los quistes de A. franciscana en todas las concentraciones probadas; en contraste, el dicromato de potasio inhibió la eclosión en menos del 50%. El mayor porcentaje de mortalidad en los quistes fue causado por los extractos acuosos de Amanita virosa, Leucopaxillus amarus y Tylopilus violatinctus, y el menor lo produjo el extracto de Macrolepiota mastoidea. Conclusiones. El ensayo con A. franciscana demostró ser eficaz en la evaluación de la toxicidad de los hongos, con la excepción de Scleroderma texense, que se considera venenoso, y que no resultó tóxico para este crustáceo (AU)

Background. The consumption of wild mushrooms has increased in recent years. However, not all mushrooms are edible and some of them may cause poisoning. Therefore, their toxicity needs to be studied. Artemia franciscana is a crustacean used in toxicity tests including toxins of fungi. Aims. To determine the percentage of inhibition and mortality produced by extracts of several basidiomycetes on the hatching of A. franciscana cysts. Methods. Aqueous extracts were prepared from 15 species of mushrooms collected from Jalisco state, Mexico. Different concentrations of the extracts were assayed in order to test their toxicity. Potassium dichromate and artificial seawater were the positive and negative controls, respectively. The percentages of hatching and mortality of the cysts were evaluated. Results. Inhibition of hatching greater than 80% in all the concentrations tested was found in 13 of the 15 species studied, in contrast to the positive control, which inhibited cyst hatching less than 50% in all cases. The highest percentage of mortality in the cysts was caused by the aqueous extracts of Amanita virosa, Leucopaxillus amarus, and Tylopilus violatinctus, and the lowest by Macrolepiota mastoidea. Conclusions. The brine shrimp bioassay appeared to be useful in the evaluation of the toxicity of several basidiomycetes, with the exception of Scleroderma texense, a mushroom considered poisonous, which showed no toxicity over A. franciscana (AU)

Effects of microclimatic variables on the symptoms and signs onset of Moniliophthora roreri, causal agent of Moniliophthora pod rot in cacao.

Moniliophthora Pod Rot (MPR) caused by the fungus Moniliophthora roreri (Cif.) Evans et al., is one of the main limiting factors of cocoa production in Latin America. Currently insufficient information on the biology and epidemiology of the pathogen limits the development of efficient management options to control MPR. This research aims to elucidate MPR development through the following daily microclimatic variables: minimum and maximum temperatures, wetness frequency, average temperature and relative humidity in the highly susceptible cacao clone Pound-7 (incidence = 86% 2008-2013 average). A total of 55 cohorts totaling 2,268 pods of 3-10 cm length, one to two months of age, were tagged weekly. Pods were assessed throughout their lifetime, every one or two weeks, and classified in 3 different categories: healthy, diseased with no sporulation, diseased with sporulating lesions. As a first step, we used Generalized Linear Mixed Models (GLMM) to determine with no a priori the period (when and for how long) each climatic variable was better related with the appearance of symptoms and sporulation. Then the significance of the candidate variables was tested in a complete GLMM. Daily average wetness frequency from day 14 to day 1, before tagging, and daily average maximum temperature from day 4 to day 21, after tagging, were the most explanatory variables of the symptoms appearance. The former was positively linked with the symptoms appearance when the latter exhibited a maximum at 30°C. The most important variables influencing sporulation were daily average minimum temperature from day 35 to day 58 and daily average maximum temperature from day 37 to day 48, both after tagging. Minimum temperature was negatively linked with the sporulation while maximum temperature was positively linked. Results indicated that the fungal microclimatic requirements vary from the early to the late cycle stages, possibly due to the pathogen's long latent period. This information is valuable for development of new conceptual models for MPR and improvement of control methods.

Plant pathogenesis-related proteins of the cacao fungal pathogen Moniliophthora perniciosa differ in their lipid-binding specificities.

Moniliophthora perniciosa is the causative agent of witches' broom disease, which devastates cacao cultures in South America. This pathogenic fungus infects meristematic tissues and derives nutrients from the plant apoplast during an unusually long-lasting biotrophic stage. To survive, the fungus produces proteins to suppress the plant immune response. Proteins of the PR-1 (pathogenesis-related 1)/CAP superfamily have been implicated in fungal virulence and immune suppression. The genome of M. perniciosa encodes 11 homologues of plant PR-1 proteins, designated MpPR-1 proteins, but their precise mode of action is poorly understood. In this study, we expressed MpPR-1 proteins in a yeast model lacking endogenous CAP proteins. We show that some members of the MpPR-1 family bind and promote secretion of sterols, whereas others bind and promote secretion of fatty acids. Lipid binding by purified MpPR-1 occurs with micromolar affinity and is saturable in vitro Sterol binding by MpPR-1 requires the presence of a flexible loop region containing aromatic amino acids, the caveolin-binding motif. Remarkably, MpPR-1 family members that do not bind sterols can be converted to sterol binders by a single point mutation in the caveolin-binding motif. We discuss the possible implications of the lipid-binding activity of MpPR-1 family members with regard to the mode of action of these proteins during M. perniciosa infections.

Saprotrophic proteomes of biotypes of the witches' broom pathogen Moniliophthora perniciosa.

Nine geographically diverse Moniliophthora perniciosa (witches' broom disease pathogen) isolates were cultured in vitro. They included six C-biotypes differing in virulence on cacao (Theobroma cacao), two S-biotypes (solanaceous hosts), and an L-biotype (liana hosts). Mycelial growth rates and morphologies differed considerably, but no characters were observed to correlate with virulence or biotype. In plant inoculations using basidiospores, one C-biotype caused symptoms on tomato (an S-biotype host), adding to evidence of limited host adaptation in these biotypes. Mycelial proteomes were analysed by two-dimensional gel electrophoresis (2-DE), and 619 gel spots were indexed on all replicate gels of at least one strain. Multivariate analysis of gel spots discriminated the L-biotype, but not the S-biotypes, from the remaining strains. The proteomic similarity of the S- and C-biotypes is consistent with their reported lack of phylogenetic distinction. Sequences from tandem mass spectrometry of tryptic peptides from major 2-DE spots were matched with Moniliophthora genome and transcript sequences on NCBI and WBD Transcriptome Atlas databases. Protein-spot identifications indicated that M. perniciosa saprotrophic mycelial proteomes expressed functions potentially connected with a 'virulence life-style', including peroxiredoxin, heat-shock proteins, nitrilase, formate dehydrogenase, a prominent complement of aldo-keto reductases, mannitol-1-phosphate dehydrogenase, and central metabolism enzymes with proposed pathogenesis functions.

Proteomic analysis during of spore germination of Moniliophthora perniciosa, the causal agent of witches' broom disease in cacao.

BACKGROUND: Moniliophthora perniciosa is a phytopathogenic fungus responsible for witches' broom disease of cacao trees (Theobroma cacao L.). Understanding the molecular events during germination of the pathogen may enable the development of strategies for disease control in these economically important plants. In this study, we determined a comparative proteomic profile of M. perniciosa basidiospores during germination by two-dimensional SDS-PAGE and mass spectrometry. RESULTS: A total of 316 proteins were identified. Molecular changes during the development of the germinative tube were identified by a hierarchical clustering analysis based on the differential accumulation of proteins. Proteins associated with fungal filamentation, such as septin and kinesin, were detected only 4 h after germination (hag). A transcription factor related to biosynthesis of the secondary metabolite fumagillin, which can form hybrids with polyketides, was induced 2 hag, and polyketide synthase was observed 4 hag. The accumulation of ATP synthase, binding immunoglobulin protein (BiP), and catalase was validated by western blotting. CONCLUSIONS: In this study, we showed variations in protein expression during the early germination stages of fungus M. perniciosa. Proteins associated with fungal filamentation, and consequently with virulence, were detected in basidiospores 4 hag., for example, septin and kinesin. We discuss these results and propose a model of the germination of fungus M. perniciosa. This research can help elucidate the mechanisms underlying basic processes of host invasion and to develop strategies for control of the disease.

Disseminated Hormographiella aspergillata infection with involvement of the lung, brain, and small intestine following allogeneic hematopoietic stem cell transplantation: case report and literature review.

Disseminated infection by Hormographiella aspergillata is extremely rare and small intestine involvement has not been reported previously. A 51-year-old man with myelodysplastic syndrome developed pneumonia after cord blood cell transplantation. Fungal growth from the biopsied lung was identified as H. aspergillata by morphology and the gene analysis. Although antifungal agents including voriconazole and liposomal amphotericin B were administered, he died of disseminated H. aspergillata infection. We review the literature and discuss the treatment and prognosis.

Imprint Desorption Electrospray Ionization Mass Spectrometry Imaging for Monitoring Secondary Metabolites Production during Antagonistic Interaction of Fungi.

Direct analysis of microbial cocultures grown on agar media by desorption electrospray ionization mass spectrometry (DESI-MS) is quite challenging. Due to the high gas pressure upon impact with the surface, the desorption mechanism does not allow direct imaging of soft or irregular surfaces. The divots in the agar, created by the high-pressure gas and spray, dramatically change the geometry of the system decreasing the intensity of the signal. In order to overcome this limitation, an imprinting step, in which the chemicals are initially transferred to flat hard surfaces, was coupled to DESI-MS and applied for the first time to fungal cocultures. Note that fungal cocultures are often disadvantageous in direct imaging mass spectrometry. Agar plates of fungi present a complex topography due to the simultaneous presence of dynamic mycelia and spores. One of the most devastating diseases of cocoa trees is caused by fungal phytopathogen Moniliophthora roreri. Strategies for pest management include the application of endophytic fungi, such as Trichoderma harzianum, that act as biocontrol agents by antagonizing M. roreri. However, the complex chemical communication underlying the basis for this phytopathogen-dependent biocontrol is still unknown. In this study, we investigated the metabolic exchange that takes place during the antagonistic interaction between M. roreri and T. harzianum. Using imprint-DESI-MS imaging we annotated the secondary metabolites released when T. harzianum and M. roreri were cultured in isolation and compared these to those produced after 3 weeks of coculture. We identified and localized four phytopathogen-dependent secondary metabolites, including T39 butenolide, harzianolide, and sorbicillinol. In order to verify the reliability of the imprint-DESI-MS imaging data and evaluate the capability of tape imprints to extract fungal metabolites while maintaining their localization, six representative plugs along the entire M. roreri/T. harzianum coculture plate were removed, weighed, extracted, and analyzed by liquid chromatography-high-resolution mass spectrometry (LC-HRMS). Our results not only provide a better understanding of M. roreri-dependent metabolic induction in T. harzianum, but may seed novel directions for the advancement of phytopathogen-dependent biocontrol, including the generation of optimized Trichoderma strains against M. roreri, new biopesticides, and biofertilizers.

Genomic analyses and expression evaluation of thaumatin-like gene family in the cacao fungal pathogen Moniliophthora perniciosa.

Thaumatin-like proteins (TLPs) are found in diverse eukaryotes. Plant TLPs, known as Pathogenicity Related Protein (PR-5), are considered fungal inhibitors. However, genes encoding TLPs are frequently found in fungal genomes. In this work, we have identified that Moniliophthora perniciosa, a basidiomycete pathogen that causes the Witches' Broom Disease (WBD) of cacao, presents thirteen putative TLPs from which four are expressed during WBD progression. One of them is similar to small TLPs, which are present in phytopathogenic basidiomycete, such as wheat stem rust fungus Puccinia graminis. Fungi genomes annotation and phylogenetic data revealed a larger number of TLPs in basidiomycetes when comparing with ascomycetes, suggesting that these proteins could be involved in specific traits of mushroom-forming species. Based on the present data, we discuss the contribution of TLPs in the combat against fungal competitors and hypothesize a role of these proteins in M. perniciosa pathogenicity.