Autoimmune amelogenesis imperfecta in patients with APS-1 and coeliac disease.

Ameloblasts are specialized epithelial cells in the jaw that have an indispensable role in tooth enamel formation-amelogenesis1. Amelogenesis depends on multiple ameloblast-derived proteins that function as a scaffold for hydroxyapatite crystals. The loss of function of ameloblast-derived proteins results in a group of rare congenital disorders called amelogenesis imperfecta2. Defects in enamel formation are also found in patients with autoimmune polyglandular syndrome type-1 (APS-1), caused by AIRE deficiency3,4, and in patients diagnosed with coeliac disease5-7. However, the underlying mechanisms remain unclear. Here we show that the vast majority of patients with APS-1 and coeliac disease develop autoantibodies (mostly of the IgA isotype) against ameloblast-specific proteins, the expression of which is induced by AIRE in the thymus. This in turn results in a breakdown of central tolerance, and subsequent generation of corresponding autoantibodies that interfere with enamel formation. However, in coeliac disease, the generation of such autoantibodies seems to be driven by a breakdown of peripheral tolerance to intestinal antigens that are also expressed in enamel tissue. Both conditions are examples of a previously unidentified type of IgA-dependent autoimmune disorder that we collectively name autoimmune amelogenesis imperfecta.

CRISPR/Cas9-Based Mutagenesis of Histone H3.1 in Spinal Dynorphinergic Neurons Attenuates Thermal Sensitivity in Mice.

Burn injury is a trauma resulting in tissue degradation and severe pain, which is processed first by neuronal circuits in the spinal dorsal horn. We have recently shown that in mice, excitatory dynorphinergic (Pdyn) neurons play a pivotal role in the response to burn-injury-associated tissue damage via histone H3.1 phosphorylation-dependent signaling. As Pdyn neurons were mostly associated with mechanical allodynia, their involvement in thermonociception had to be further elucidated. Using a custom-made AAV9_mutH3.1 virus combined with the CRISPR/cas9 system, here we provide evidence that blocking histone H3.1 phosphorylation at position serine 10 (S10) in spinal Pdyn neurons significantly increases the thermal nociceptive threshold in mice. In contrast, neither mechanosensation nor acute chemonociception was affected by the transgenic manipulation of histone H3.1. These results suggest that blocking rapid epigenetic tagging of S10H3 in spinal Pdyn neurons alters acute thermosensation and thus explains the involvement of Pdyn cells in the immediate response to burn-injury-associated tissue damage.

CRISPR-Cas9-Based Mutagenesis of the Mucormycosis-Causing Fungus Lichtheimia corymbifera.

Lichtheimia corymbifera is considered as one of the most frequent agents of mucormycosis. The lack of efficient genetic manipulation tools hampers the characterization of the pathomechanisms and virulence factors of this opportunistic pathogenic fungus. Although such techniques have been described for certain species, the performance of targeted mutagenesis and the construction of stable transformants have remained a great challenge in Mucorales fungi. In the present study, a plasmid-free CRISPR-Cas9 system was applied to carry out a targeted gene disruption in L. corymbifera. The described method is based on the non-homologous end-joining repair of the double-strand break caused by the Cas9 enzyme. Using this method, short, one-to-five nucleotide long-targeted deletions could be induced in the orotidine 5'-phosphate decarboxylase gene (pyrG) and, as a result, uracil auxotrophic strains were constructed. These strains are applicable as recipient strains in future gene manipulation studies. As we know, this is the first genetic modification of this clinically relevant fungus.

CRISPR-Cas9-mediated disruption of the HMG-CoA reductase genes of Mucor circinelloides and subcellular localization of the encoded enzymes.

Terpenoid compounds, such as sterols, carotenoids or the prenyl groups of various proteins are synthesized via the mevalonate pathway. A rate-limiting step of this pathway is the conversion of 3-methylglutaryl-CoA (HMG-CoA) to mevalonic acid catalyzed by the HMG-CoA reductase. Activity of this enzyme may affect several biological processes, from the synthesis of terpenoid metabolites to the adaptation to various environmental conditions. In this study, the three HMG-CoA reductase genes (i.e. hmgR1, hmgR2 and hmgR3) of the ß-carotene producing filamentous fungus, Mucor circinelloides were disrupted individually and simultaneously by a recently developed in vitro plasmid-free CRISPR-Cas9 method. Examination of the mutants revealed that the function of hmgR2 and hmgR3 are partially overlapping and involved in the general terpenoid biosynthesis. Moreover, hmgR2 seemed to have a special role in the ergosterol biosynthesis. Disruption of all three genes affected the germination ability of the spores and the sensitivity to hydrogen peroxide. Disruption of the hmgR1 gene had no effect on the ergosterol production and the sensitivity to statins but caused a reduced growth at lower temperatures. By confocal fluorescence microscopy using strains expressing GFP-tagged HmgR proteins, all three HMG-CoA reductases were localized in the endoplasmic reticulum.

ECMM CandiReg-A ready to use platform for outbreaks and epidemiological studies.

BACKGROUND: Recent outbreaks of Candida auris further exemplify that invasive Candida infections are a substantial threat to patients and healthcare systems. Even short treatment delays are associated with higher mortality rates. Epidemiological shifts towards more resistant Candida spp. require careful surveillance. OBJECTIVES: Triggered by the emergence of C auris and by increasing antifungal resistance rates the European Confederation of Medical Mycology developed an international Candida Registry (FungiScope™ CandiReg) to allow contemporary multinational surveillance. METHODS: CandiReg serves as platform for international cooperation to enhance research regarding invasive Candida infections. CandiReg uses the General Data Protection Regulation compliant data platform ClinicalSurveys.net that holds the electronic case report forms (eCRF). Data entry is supported via an interactive macro created by the software that can be accessed via any Internet browser. RESULTS: CandiReg provides an eCRF for invasive Candida infections that can be used for a variety of studies from cohort studies on attributable mortality to evaluations of guideline adherence, offering to the investigators of the 28 ECMM member countries the opportunity to document their cases of invasive Candida infection. CandiReg allows the monitoring of epidemiology of invasive Candida infections, including monitoring of multinational outbreaks. Here, we describe the structure and management of the CandiReg platform. CONCLUSION: CandiReg supports the collection of clinical information and isolates to improve the knowledge on epidemiology and eventually to improve management of invasive Candida infections. CandiReg promotes international collaboration, improving the availability and quality of evidence on invasive Candida infection and contributes to improved patient management.

Exophiala dermatitidis Endophthalmitis: Case Report and Literature Review.

We report a case of a 59-year-old male patient with a postoperative fungal infection of the left eye. A dark-pigmented yeast, Exophiala dermatitidis (previously known as Wangiella dermatitidis), was identified from the culture of the biopsy taken from the posterior capsule. The infection was successfully eradicated by a combination of surgical and medical (i.e., voriconazole and fluconazole) treatment. This is the first report of successfully treated E. dermatitidis endophthalmitis, which demonstrates that a prompt and aggressive antifungal therapy combined with surgical intervention is necessary to prevent vision loss in cases of endophthalmitis due to Exophiala species. Beside the case description, we also aim to provide a literature review of previously reported eye infections caused by Exophiala species in order to help the future diagnosis and management of the disease.

An Organic Solvent-Tolerant Lipase with Both Hydrolytic and Synthetic Activities from the Oleaginous Fungus Mortierella echinosphaera.

Lipase enzymes of the oleaginous fungal group Mortierella are rarely studied. However, considering that most commercial lipases are derived from filamentous fungal sources, their investigation can contribute to the cost-effective development of new biotechnological processes. Here, an extracellular lipase with a molecular mass of 30 kDa was isolated from Mortierella echinosphaera CBS 575.75 and characterized. The purified lipase exhibited an optimal p-nitrophenyl palmitate (pNPP)-hydrolyzing activity at 25 °C and pH 6.6-7.0 and proved to be highly stable at temperatures up to 40 °C and under broad pH conditions. The enzyme was active under low temperatures, retaining 32.5% of its activity at 10 °C, and was significantly stable in polar and non-polar organic solvents. The Km, Vmax, and kcat for pNPP were 0.336 mM, 30.4 µM/min, and 45.7 1/min for pNPP and 0.333 mM, 36.9 µM/min, and 55.6 1/min for pNP-decanoate, respectively. The pNPP hydrolysis was inhibited by Hg2+, N-bromosuccinimide, and sodium dodecyl sulfate, while ethylenediaminetetraacetic acid and metal ions, such as Ca2+, Mg2+, Na⁺, and K⁺ enhanced the activity. The purified lipase had non-regioselective activity and wide substrate specificity, showing a clear preference for medium-chained p-nitrophenyl esters. Besides its good transesterification activity, the enzyme appeared as a suitable biocatalyst to operate selective esterification reactions to long-chained alkyl esters. Adsorption to Accurel MP1000 improved the storage stability of the enzyme at 5 °C. The immobilized lipase displayed tolerance to a non-aqueous environment and was reusable for up to five cycles without significant loss in its synthetic and hydrolytic activities. These findings confirm the applicability of both the free and the immobilized enzyme preparations in future research.

Recurrent Scedosporium apiospermum mycetoma successfully treated by surgical excision and terbinafine treatment: a case report and review of the literature.

BACKGROUND: Scedosporium apiospermum is an emerging opportunistic filamentous fungus, which is notorious for its high levels of antifungal-resistance. It is able to cause localized cutaneous or subcutaneous infections in both immunocompromised and immunocompetent persons, pulmonary infections in patients with predisposing pulmonary diseases and invasive mycoses in immunocompromised patients. Subcutaneous infections caused by this fungus frequently show chronic mycetomatous manifestation. CASE REPORT: We report the case of a 70-year-old immunocompromised man, who developed a fungal mycetomatous infection on his right leg. There was no history of trauma; the aetiological agent was identified by microscopic examination and ITS sequencing. This is the second reported case of S. apiospermum subcutaneous infections in Hungary, which was successfully treated by surgical excision and terbinafine treatment. After 7 months, the patient remained asymptomatic. Considering the antifungal susceptibility and increasing incidence of the fungus, Scedosporium related subcutaneous infections reported in the past quarter of century in European countries were also reviewed. CONCLUSIONS: Corticosteroid treatment represents a serious risk factor of S. apiospermum infections, especially if the patient get in touch with manure-enriched or polluted soil or water. Such infections have emerged several times in European countries in the past decades. The presented data suggest that besides the commonly applied voriconazole, terbinafine may be an alternative for the therapy of mycetomatous Scedosporium infections.

In vitro susceptibility of Scedosporium isolates to N-acetyl-L-cysteine alone and in combination with conventional antifungal agents.

In recent years, Scedosporium species have been more commonly recognized from severe, difficult-to-treat human infections, such as upper respiratory tract and pulmonary infections. To select an appropriate therapeutic approach for these infections is challenging, because of the commonly observed resistance of the causative agents to several antifungal drugs. Therefore, to find a novel strategy for the treatment of pulmonary Scedosporium infections the in vitro antifungal effect of a mucolytic agent, N-acetyl-L-cysteine and its in vitro combinations with conventional antifungals were investigated. Synergistic and indifferent interactions were registered in 23 and 13 cases, respectively. Antagonism was not revealed between the compounds.

Psathyloma, a new genus in Hymenogastraceae described from New Zealand.

A new genus Psathyloma is described based on collections of agarics from New Zealand. We describe two new species in the genus, Ps. leucocarpum and Ps. catervatim, both of which have been known and tentatively named for a long time awaiting a formal description. Morphological traits and phylogenetic analyses reveal that Psathyloma forms a strongly supported sister clade to Hebeloma, Naucoria and Hymenogaster Morphologically Psathyloma resembles Hebeloma from which it differs mainly by producing smooth basidiospores with a germ pore. The geographical range of the genus has been demonstrated to include several regions in the southern hemisphere. A survey of published environmental sequences reveals that Psathyloma spp. were isolated from ectomycorrhizal root tips from Tasmania and Argentina, indicating an ectomycorrhizal association with southern beech.

TLR signalling can modify the mineralization of tooth germ.

OBJECTIVE: The aim of this work is to investigate the possible role of Toll-like receptor 4 (TLR4) during the development of mouse tooth germ. TLR4 is well known to inhibit mineralization and cause inflammation in mature odontoblasts and dental pulp cells. However, unlike these pathological functions of TLR4, little is known about the developmental function(s) of TLR4 during tooth development. MATERIALS AND METHODS: TLR4 expression was studied via Western blot in developing lower mouse incisors from E13.5 to E18.5. To generate functional data about the effects of TLR4, a specific agonist (LPS) was applied to the medium of in vitro tooth germ cultures, followed by Western blot, histochemical staining, ELISA assay, in situ hybridization and RT-qPCR. RESULTS: Increased accumulation of biotin-labelled LPS was detected in the enamel organ and in preodontoblasts. LPS treatment induced degradation of the inhibitor molecule (IκB) of the NF-κB signalling pathway. However, no morphological alterations were detected in cultured tissue after LPS addition at the applied dosage. Activation of TLR4 inhibited the mineralization of enamel and dentin, as demonstrated by alizarin red staining and as decreased levels of collagen type X. mRNA expression of ameloblastin was elevated after LPS administration. CONCLUSION: These results demonstrate that TLR4 may decrease the mineralization of hard tissues of the tooth germ and may trigger the maturation of ameloblasts; it can give valuable information to understand better congenital tooth abnormalities.

In vitro antifungal activity of antipsychotic drugs and their combinations with conventional antifungals against Scedosporium and Pseudallescheria isolates.

In the present study, in vitro antifungal activities of five antipsychotic drugs (i.e., chlorpromazine hydrochloride, CPZ; trifluoperazine hydrochloride, TPZ; amantadine hydrochloride; R-(-)-deprenyl hydrochloride, and valproic acid sodium salt) and five conventional antifungal drugs (i.e., amphotericin B, AMB; caspofungin, CSP; itraconazole; terbinafine, TRB and voriconazole, VRC) were investigated in broth microdilution tests against four clinical and five environmental Scedosporium and Pseudallescheria isolates. When used alone, phenothiazines CPZ and TPZ exerted remarkable antifungal effects. Thus, their in vitro combinations with AMB, CSP, VRC, and TRB were also examined against the clinical isolates. In combination with antifungal agents, CPZ was able to act synergistically with AMB and TRB in cases of one and two isolates, respectively. In all other cases, indifferent interactions were revealed. Antagonism was not observed between the tested agents. These combinations may establish a more effective and less toxic therapy after further in vitro and in vivo studies for Scedosporium and Pseudallescheria infections.

Molecular identification and antifungal susceptibility of Curvularia australiensis, C. hawaiiensis and C. spicifera isolated from human eye infections.

A reliable identification method was developed for three closely related Curvularia species, which are frequently isolated from human keratomycoses. Since the traditionally used morphological method and the increasingly used internal transcribed spacer (ITS)-based molecular method proved to be insufficient to discern C. australiensis, C. hawaiiensis and C. spicifera, other molecular targets, such as ß-tubulin, translation elongation factor 1-α and the nuclear ribosomal intergenic spacer (IGS), were tested. Among them, the use of the highly divergent IGS sequence is suggested and the species-specific discriminating characters were determined in appropriate reference strains. It was also concluded that C. hawaiiensis and C. spicifera can be predominantly isolated from eye infections among the three species. The in vitro antifungal susceptibility of 10 currently used antifungal agents against 32 Curvularia isolates was also investigated. MICs were determined in each case. Isolates of C. spicifera proved to be less susceptible to the tested antifungals than those of C. hawaiiensis, which underline the importance of the correct identification of these species.

Expression of Xanthophyllomyces dendrorhous cytochrome-P450 hydroxylase and reductase in Mucor circinelloides.

Carotenoids are natural pigments that act as powerful antioxidants and have various beneficial effects on human and animal health. Mucor circinelloides (Mucoromycotina) is a carotenoid producing zygomycetes fungus, which accumulates ß-carotene as the main carotenoid but also able to produce the hydroxylated derivatives of ß-carotene (i.e. zeaxanthin and ß-cryptoxanthin) in low amount. These xanthophylls, together with the ketolated derivatives of ß-carotene (such as canthaxanthin, echinenone and astaxanthin) have better antioxidant activity than ß-carotene. In this study our aim was to modify and enhance the xanthophyll production of the M. circinelloides by expression of heterologous genes responsible for the astaxanthin biosynthesis. The crtS and crtR genes, encoding the cytochrome-P450 hydroxylase and reductase, respectively, of wild-type and astaxanthin overproducing mutant Xanthophyllomyces dendrorhous strains were amplified from cDNA and the nucleotide and the deduced amino acid sequences were compared to each other. Introduction of the crtS on autonomously replicating plasmid in the wild-type M. circinelloides resulted enhanced zeaxanthin and ß-cryptoxanthin accumulation and the presence of canthaxanthin, echinenone and astaxanthin in low amount; the ß-carotene hydroxylase and ketolase activity of the X. dendrorhous cytochrome-P450 hydroxylase in M. circinelloides was verified. Increased canthaxanthin and echinenone production was observed by expression of the gene in a canthaxanthin producing mutant M. circinelloides. Co-expression of the crtR and crtS genes led to increase in the total carotenoid and slight change in xanthophyll accumulation in comparison with transformants harbouring the single crtS gene.

Enhanced production of industrial enzymes in Mucoromycotina fungi during solid-state fermentation of agricultural wastes/by-products.

Cellulolytic, lipolytic and proteolytic enzyme production of zygomycetes Mucor corticolus, Rhizomucor miehei, Gilbertella persicaria and Rhizopus niveus were investigated using agro-industrial wastes as substrates. Solid-state cultures were carried out on untreated corn residues (stalk and leaf) as single substrate (SSF1) or corn residues and wheat bran in mixed fermentation (SSF2). Rapid production of endoglucanase (CMCase) was observed with maximal activity reaching after about 48-h fermentation, while cellobiohydrolase (CBH) and ß-glucosidase enzymes generally had their peak after 72-h incubation. Highest filter paper degrading (FPase), CMCase, CBH and ß-glucosidase activities obtained were (U g⁻¹ dss) 17.3, 74.1, 12.2 and 158.3, for R. miehei, G. persicaria, M. corticolus and Rh. niveus, respectively. M. corticolus proved to be the best lipolytic enzyme producer in SSF1 presenting 447.6 U g⁻¹ dss yield, while R. miehei showed 517.7 U g⁻¹ dss activity in SSF2. Rh. niveus exhibited significantly greater protease production than the other strains. Suc-AAPF-pNA hydrolyzing activities of this strain were 1.1 and 1.96 U g⁻¹ dss in SSF1 and SSF2, respectively. We conclude that the used corn stalk and leaf residues could potentially be applicable as strong inducers for cellulase and lipase production by Mucoromycotina fungi.

Transcription of the three HMG-CoA reductase genes of Mucor circinelloides.

BACKGROUND: Precursors of sterols, carotenoids, the prenyl groups of several proteins and other terpenoid compounds are synthesised via the acetate-mevalonate pathway. One of the key enzyme of this pathway is the 3-hydroxy-3-methylglutaryl-CoA (HMG-CoA) reductase, which catalyses the conversion of HMG-CoA to mevalonate. HMG-CoA reductase therefore affects many biological processes, such as morphogenesis, synthesis of different metabolites or adaptation to environmental changes. In this study, transcription of the three HMG-CoA reductase genes (designated as hmgR1, hmgR2 and hmgR3) of the ß-carotene producing Mucor circinelloides has been analysed under various culturing conditions; effect of the elevation of their copy number on the carotenoid and ergosterol content as well as on the sensitivity to statins has also been examined. RESULTS: Transcripts of each gene were detected and their relative levels varied under the tested conditions. Transcripts of hmgR1 were detected only in the mycelium and its relative transcript level seems to be strongly controlled by the temperature and the oxygen level of the environment. Transcripts of hmgR2 and hmgR3 are already present in the germinating spores and the latter is also strongly regulated by oxygen. Overexpression of hmgR2 and hmgR3 by elevating their copy numbers increased the carotenoid content of the fungus and decreased their sensitivity to statins. CONCLUSIONS: The three HMG-CoA reductase genes of M. circinelloides displayed different relative transcript levels under the tested conditions suggesting differences in their regulation. They seem to be especially involved in the adaptation to the changing oxygen tension and osmotic conditions of the environment as well as to statin treatment. Overexpression of hmgR2 and hmgR3 may be used to improve the carotenoid content.

Susceptibility of clinically important dermatophytes against statins and different statin-antifungal combinations.

The investigation of the antifungal activities of drugs whose primary activities are not related to their antimicrobial potential is in the current forefront of research. Statin compounds, which are routinely used as cholesterol-lowering drugs, may also exert direct antimicrobial effects. In this study, the in vitro antifungal activities of various statins (lovastatin, simvastatin, fluvastatin, atorvastatin, rosuvastatin and pravastatin) were examined against one isolate each of four dermatophyte species (Trichophyton mentagrophytes, Trichophyton rubrum, Microsporum canis and Microsporum gypseum). Basically, statins were effective in inhibiting all dermatophyte studied, but were particularly active against M. canis and T. mentagrophytes. Fluvastatin and simvastatin were active against all of the tested fungi causing a complete inhibition of their growth at very low concentrations (6.25-12.5 µg/ml). Lovastatin and rosuvastatin had inhibitory effects at higher concentrations (25-128 µg/ml), while atorvastatin and pravastatin proved the less effective. The in vitro interactions between statins and different antifungals (ketoconazole, itraconazole, fluconazole, amphotericin B, nystatin, griseofulvin, terbinafine and primycin) were also investigated using a standard chequerboard broth microdilution method. Synergetic interactions were observed in several cases, most of them were noticed when statins were combined with terbinafine and the different azoles. Some combinations were particularly active (ketoconazole-simvastatin or terbinafine-simvastatin), as they were found to exert synergistic effect against all of the investigated isolates. The other antifungals showed synergistic interactions with statins in only certain cases. These results suggest that statins exert substantial antifungal effects against dermatophyte fungi and they should be promising components in a combination therapy as they can act synergistically with a number of clinically used antifungal agents.

Repeated diagnostic ultrasound exposure modifies the structural properties of CA1 dendrites and alters the hippocampal transcriptome.

The development of neurons is regulated by several spatiotemporally changing factors, which are crucial to give the ability of neurons to form functional networks. While external physical stimuli may impact the early developmental stages of neurons, the medium and long-term consequences of these influences have yet to be thoroughly examined. Using an animal model, this study focuses on the morphological and transcriptome changes of the hippocampus that may occur as a consequence of fetal ultrasound examination. We selectively labeled CA1 neurons of the hippocampus with in-utero electroporation to analyze their morphological features. Furthermore, certain samples also went through RNA sequencing after repetitive ultrasound exposure. US exposure significantly changed several morphological properties of the basal dendritic tree. A notable increase was also observed in the density of spines on the basal dendrites, accompanied by various alterations in individual spine morphology. Transcriptome analysis revealed several up or downregulated genes, which may explain the molecular background of these alterations. Our results suggest that US-derived changes in the dendritic trees of CA1 pyramidal cells might be connected to modification of the transcriptome of the hippocampus and may lead to an increased dendritic input.

In Vivo Imaging of Acute Hindlimb Ischaemia in Rat Model: A Pre-Clinical PET Study.

BACKGROUND: To better understand ischaemia-related molecular alterations, temporal changes in angiogenic Aminopeptidase N (APN/CD13) expression and glucose metabolism were assessed with PET using a rat model of peripheral arterial disease (PAD). METHODS: The mechanical occlusion of the base of the left hindlimb triggered using a tourniquet was applied to establish the ischaemia/reperfusion injury model in Fischer-344 rats. 2-[18F]FDG and [68Ga]Ga-NOTA-c(NGR) PET imaging performed 1, 3, 5, 7, and 10 days post-ischaemia induction was followed by Western blotting and immunohistochemical staining for APN/CD13 in ischaemic and control muscle tissue extracts. RESULTS: Due to a cellular adaptation to hypoxia, a gradual increase in [68Ga]Ga-NOTA-c(NGR) and 2-[18F]FDG uptake was observed from post-intervention day 1 to 7 in the ischaemic hindlimbs, which was followed by a drop on day 10. Conforming pronounced angiogenic recovery, the NGR accretion of the ischaemic extremities differed significantly from the controls 5, 7, and 10 days after ischaemia induction (p ≤ 0.05), which correlated with the Western blot and immunohistochemical results. No remarkable radioactivity was depicted between the normally perfused hindlimbs of either the ischaemic or the control groups. CONCLUSIONS: The PET-based longitudinal assessment of angiogenesis-associated APN/CD13 expression and glucose metabolism during ischaemia may continue to broaden our knowledge on the pathophysiology of PAD.

The evolution of defense mechanisms correlate with the explosive diversification of autodigesting Coprinellus mushrooms (Agaricales, Fungi).

Bursts of diversification are known to have contributed significantly to the extant morphological and species diversity, but evidence for many of the theoretical predictions about adaptive radiations have remained contentious. Despite their tremendous diversity, patterns of evolutionary diversification and the contribution of explosive episodes in fungi are largely unknown. Here, using the genus Coprinellus (Psathyrellaceae, Agaricales) as a model, we report the first explosive fungal radiation and infer that the onset of the radiation correlates with a change from a multilayered to a much simpler defense structure on the fruiting bodies. We hypothesize that this change constitutes a key innovation, probably relaxing constraints on diversification imposed by nutritional investment into the development of protective tissues of fruiting bodies. Fossil calibration suggests that Coprinellus mushrooms radiated during the Miocene coinciding with global radiation of large grazing mammals following expansion of dry open grasslands. In addition to diversification rate-based methods, we test the hard polytomy hypothesis, by analyzing the resolvability of internal nodes of the backbone of the putative radiation using Reversible-Jump MCMC. We discuss potential applications and pitfalls of this approach as well as how biologically meaningful polytomies can be distinguished from alignment shortcomings. Our data provide insights into the nature of adaptive radiations in general by revealing a deceleration of morphological diversification through time. The dynamics of morphological diversification was approximated by obtaining the temporal distribution of state changes in discrete traits along the trees and comparing it with the tempo of lineage accumulation. We found that the number of state changes correlate with the number of lineages, even in parts of the tree with short internal branches, and peaks around the onset of the explosive radiation followed by a slowdown, most likely because of the decrease in available niches.