Identification of Rosellinia species as producers of cyclodepsipeptide PF1022 A and resurrection of the genus Dematophora as inferred from polythetic taxonomy

Rosellinia (Xylariaceae) is a large, cosmopolitan genus comprising over 130 species that have been defined based mainly on the morphology of their sexual morphs. The genus comprises both lignicolous and saprotrophic species that are frequently isolated as endophytes from healthy host plants, and important plant pathogens. In order to evaluate the utility of molecular phylogeny and secondary metabolite profiling to achieve a better basis for their classification, a set of strains was selected for a multi-locus phylogeny inferred from a combination of the sequences of the internal transcribed spacer region (ITS), the large subunit (LSU) of the nuclear rDNA, beta-tubulin (TUB2) and the second largest subunit of the RNA polymerase II (RPB2). Concurrently, various strains were surveyed for production of secondary metabolites. Metabolite profiling relied on methods with high performance liquid chromatography with diode array and mass spectrometric detection (HPLC-DAD/MS) as well as preparative isolation of the major components after re-fermentation followed by structure elucidation using nuclear magnetic resonance (NMR) spectroscopy and high resolution mass spectrometry (HR-MS). Two new and nine known isopimarane diterpenoids were identified during our mycochemical studies of two selected Dematophora strains and the metabolites were tested for biological activity. In addition, the nematicidal cyclodepsipeptide PF1022 A was purified and identified from a culture of Rosellinia corticium, which is the first time that this endophyte-derived drug precursor has been identified unambiguously from an ascospore-derived isolate of a Rosellinia species. While the results of this first HPLC profiling were largely inconclusive regarding the utility of secondary metabolites as genus-specific chemotaxonomic markers, the phylogeny clearly showed that species featuring a dematophora-like asexual morph were included in a well-defined clade, for which the genus Dematophora is resurrected. Dematophora now comprises all previously known important plant pathogens in the genus such as D. arcuata, D. bunodes, D. necatrix and D. pepo, while Rosellinia s. str. comprises those species that are known to have a geniculosporium-like or nodulisporium-like asexual morph, or where the asexual morph remains unknown. The extensive morphological studies of L.E. Petrini served as a basis to transfer several further species from Rosellinia to Dematophora, based on the morphology of their asexual morphs. However, most species of Rosellinia and allies still need to be recollected in fresh state, cultured, and studied for their morphology and their phylogenetic affinities before the infrageneric relationships can be clarified.Fil: Wittstein, Kathrin. Helmholtz-Zentrum für Infektionsforschung GmbH; Alemania. German Centre for Infection Research; AlemaniaFil: Cordsmeier, Arne. Helmholtz-Zentrum für Infektionsforschung GmbH; Alemania. Universitat Erlangen Nuremberg; AlemaniaFil: Lambert, Christopher. Helmholtz-Zentrum für Infektionsforschung GmbH; Alemania. German Centre for Infection Research; AlemaniaFil: Wendt, L.. Helmholtz-Zentrum für Infektionsforschung GmbH; Alemania. German Centre for Infection Research; AlemaniaFil: Sir, Esteban Benjamin. Universidad Nacional de Tucumán. Instituto de Bioprospección y Fisiología Vegetal. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet Noa Sur. Instituto de Bioprospección y Fisiología Vegetal; ArgentinaFil: Weber, J.. Helmholtz-Zentrum für Infektionsforschung GmbH; Alemania. German Centre for Infection Research; AlemaniaFil: Wurzler, N.. Helmholtz-Zentrum für Infektionsforschung GmbH; Alemania. German Centre for Infection Research; AlemaniaFil: Petrini, Liliane E.. Via al Perato 15c; SuizaFil: Stadler, Marc. Helmholtz-Zentrum für Infektionsforschung GmbH; Alemania. German Centre for Infection Research; Alemani

Hericium erinaceus, an amazing medicinal mushroom

International audienceMedicinal mushrooms have become a compelling topic because the bioactive compounds they contain promise a plethora of therapeutic properties. Hericium erinaceus commonly known as “Houtou” or “Shishigashira” in China and "Yamabushitake” in Japan, has commonly been prescribed in traditional Chinese medicine (TCM), because its consumption has been shown to be beneficial to human health. The species isfound throughout the northern hemisphere in Europe, Asia, and North America. Hericium erinaceus has been firmly established as an important medicinal mushroom and its numerous bioactive compounds have been developed into food supplements and alternative medicines. However, the correspondence of the active components that cause the observed effects is often not clear. The mushroom as well as the fermented mycelia have been reported to produce several classes of bioactive molecules, including polysaccharides, proteins, lectins, phenols, and terpenoids. Most interestingly, two classes of terpenoid compounds, hericenones and erinacines, from fruiting bodies and cultured mycelia, respectively, have been found to stimulate nerve growth factor (NGF) synthesis. In this review we examine the scientific literature to explore and highlight the scientific facts concerning medicinal properties of H. erinaceus. We provide up-to-date information on this mushroom, including its taxonomy and a summary of bioactive compounds that appear related to the therapeutic potential of H. erinaceus

Screening for inhibitors of mutacin synthesis in Streptococcus mutans using fluorescent reporter strains

Abstract Background Within the polymicrobial dental plaque biofilm, bacteria kill competitors by excreting mixtures of bacteriocins, resulting in improved fitness and survival. Inhibiting their bacteriocin synthesis might therefore be a useful strategy to eliminate specific pathogens. We used Streptococcus mutans, a highly acidogenic inhabitant of dental plaque, as a model and searched for natural products that reduced mutacin synthesis. To this end we fused the promoter of mutacin VI to the GFP+ gene and integrated the construct into the genome of S. mutans UA159 by single homologous recombination. Results The resulting reporter strain 423p - gfp + was used to screen 297 secondary metabolites from different sources, mainly myxobacteria and fungi, for their ability to reduce the fluorescence of the fully induced reporter strain by > 50% while growth was almost unaffected (> 90% of control). Seven compounds with different chemical structures and different modes of action were identified. Erinacine C was subsequently validated and shown to inhibit transcription of all three mutacins of S. mutans. The areas of the inhibition zones of the sensor strains S. sanguinis and Lactococcus lactis were reduced by 35% to 61% in comparison to controls in the presence of erinacine C, demonstrating that the amount of active mutacins in the culture supernatants of S. mutans was reduced. Erinacines are cyathane diterpenes that were extracted from cultures of the edible mushroom Hericium erinaceus. They have anti-inflammatory, antimicrobial and neuroprotective effects. For erinacine C, a new biological activity was found here. Conclusions We demonstrate the successful development of a whole-cell fluorescent reporter for the screening of natural compounds and report that erinacine C suppresses mutacin synthesis in S. mutans without affecting cell viability

Two New Cyathane Diterpenoids from Mycelial Cultures of the Medicinal Mushroom Hericium erinaceus and the Rare Species, Hericium flagellum

Basidiomycetes of the genus Hericium are among the most praised medicinal and edible mushrooms, which are known to produce secondary metabolites with the potential to treat neurodegenerative diseases. This activity has been attributed to the discovery of various terpenoids that can stimulate the production of nerve growth factor (NGF) or (as established more recently) brain-derived neurotrophic factor (BDNF) in cell-based bioassays. The present study reports on the metabolite profiles of a Lion’s Mane mushroom (Hericium erinaceus) strain and a strain of the rare species, Hericium flagellum (synonym H. alpestre). While we observed highly similar metabolite profiles between the two strains that were examined, we isolated two previously undescribed metabolites, given the trivial names erinacines Z1 and Z2. Their chemical structures were elucidated by means of nuclear magnetic resonance (NMR) spectroscopy and high resolution mass spectrometry. Along with six further, previously identified cyathane diterpenes, the novel erinacines were tested for neurotrophin inducing effects. We found that erinacines act on BDNF, which is a neurotrophic factor that has been reported recently by us to be induced by the corallocins, but as well on NGF expression, which is consistent with the literature

Study of three interesting Amanita species from Thailand: Morphology, multiple-gene phylogeny and toxin analysis.

Amanita ballerina and A. brunneitoxicaria spp. nov. are introduced from Thailand. Amanita fuligineoides is also reported for the first time from Thailand, increasing the known distribution of this taxon. Together, those findings support our view that many taxa are yet to be discovered in the region. While both morphological characters and a multiple-gene phylogeny clearly place A. brunneitoxicaria and A. fuligineoides in sect. Phalloideae (Fr.) Quél., the placement of A. ballerina is problematic. On the one hand, the morphology of A. ballerina shows clear affinities with stirps Limbatula of sect. Lepidella. On the other hand, in a multiple-gene phylogeny including taxa of all sections in subg. Lepidella, A. ballerina and two other species, including A. zangii, form a well-supported clade sister to the Phalloideae sensu Bas 1969, which include the lethal "death caps" and "destroying angels". Together, the A. ballerina-A. zangii clade and the Phalloideae sensu Bas 1969 also form a well-supported clade. We therefore screened for two of the most notorious toxins by HPLC-MS analysis of methanolic extracts from the basidiomata. Interestingly, neither α-amanitin nor phalloidin was found in A. ballerina, whereas Amanita fuligineoides was confirmed to contain both α-amanitin and phalloidin, and A. brunneitoxicaria contained only α-amanitin. Together with unique morphological characteristics, the position in the phylogeny indicates that A. ballerina is either an important link in the evolution of the deadly Amanita sect. Phalloideae species, or a member of a new section also including A. zangii

Erinacine C Activates Transcription from a Consensus ETS DNA Binding Site in Astrocytic Cells in Addition to NGF Induction.

Medicinal mushrooms of the genus Hericium are known to produce secondary metabolites with homeostatic properties for the central nervous system. We and others have recently demonstrated that among these metabolites cyathane diterpenoids and in particular erinacine C possess potent neurotrophin inducing properties in astrocytic cells. Yet, the signaling events downstream of erinacine C induced neurotrophin acitivity in neural-like adrenal phaeochromocytoma cells (PC12) cells have remained elusive. Similar, signaling events activated by erinacine C in astrocytic cells are unknown. Using a combination of genetic and pharmacological inhibitors we show that erinacine C induced neurotrophic activity mediates PC12 cell differentiation via the TrkA receptor and likely its associated PLCγ-, PI3K-, and MAPK/ERK pathways. Furthermore, a small library of transcriptional activation reporters revealed that erinacine C induces transcriptional activation mediated by DNA consensus binding sites of selected conserved transcription factor families. Among these, transcription is activated from an ETS consensus in a concentration dependent manner. Interestingly, induced ETS-consensus transcription occurs in parallel and independent of neurotrophin induction. This finding helps to explain the many pleiotropic functions of cyathane diterpenoids. Moreover, our studies provide genetic access to cyathane diterpenoid functions in astrocytic cells and help to mechanistically understand the action of cyathanes in glial cells

KdmB, a Jumonji Histone H3 Demethylase, Regulates Genome-Wide H3K4 Trimethylation and Is Required for Normal Induction of Secondary Metabolism in Aspergillus nidulans.

Histone posttranslational modifications (HPTMs) are involved in chromatin-based regulation of fungal secondary metabolite biosynthesis (SMB) in which the corresponding genes-usually physically linked in co-regulated clusters-are silenced under optimal physiological conditions (nutrient-rich) but are activated when nutrients are limiting. The exact molecular mechanisms by which HPTMs influence silencing and activation, however, are still to be better understood. Here we show by a combined approach of quantitative mass spectrometry (LC-MS/MS), genome-wide chromatin immunoprecipitation (ChIP-seq) and transcriptional network analysis (RNA-seq) that the core regions of silent A. nidulans SM clusters generally carry low levels of all tested chromatin modifications and that heterochromatic marks flank most of these SM clusters. During secondary metabolism, histone marks typically associated with transcriptional activity such as H3 trimethylated at lysine-4 (H3K4me3) are established in some, but not all gene clusters even upon full activation. KdmB, a Jarid1-family histone H3 lysine demethylase predicted to comprise a BRIGHT domain, a zinc-finger and two PHD domains in addition to the catalytic Jumonji domain, targets and demethylates H3K4me3 in vivo and mediates transcriptional downregulation. Deletion of kdmB leads to increased transcription of about ~1750 genes across nutrient-rich (primary metabolism) and nutrient-limiting (secondary metabolism) conditions. Unexpectedly, an equally high number of genes exhibited reduced expression in the kdmB deletion strain and notably, this group was significantly enriched for genes with known or predicted functions in secondary metabolite biosynthesis. Taken together, this study extends our general knowledge about multi-domain KDM5 histone demethylases and provides new details on the chromatin-level regulation of fungal secondary metabolite production