During a mycological survey of the Democratic Republic of the Congo, a fungal specimen that morphologically resembled the American species Hypoxylon papillatum was encountered. A polyphasic approach including morphological and chemotaxonomic together with a multigene phylogenetic study (ITS, LSU, tub2, and rpb2) of Hypoxylon spp. and representatives of related genera revealed that this strain represents a new species of the Hypoxylaceae. However, the multi-locus phylogenetic inference indicated that the new fungus clustered with H. papillatum in a separate clade from the other species of Hypoxylon. Studies by ultrahigh performance liquid chromatography coupled to diode array detection and ion mobility tandem mass spectrometry (UHPLC-DAD-IM-MS/MS) were carried out on the stromatal extracts. In particular, the MS/MS spectra of the major stromatal metabolites of these species indicated the production of hitherto unreported azaphilone pigments with a similar core scaffold to the cohaerin-type metabolites, which are exclusively found in the Hypoxylaceae. Based on these results, the new genus Parahypoxylon is introduced herein. Aside from P. papillatum, the genus also includes P. ruwenzoriensesp. nov., which clustered together with the type species within a basal clade of the Hypoxylaceae together with its sister genus Durotheca.

In this investigation, we explored the diversity of melleolide‐type meroterpenoids produced by Armillaria ostoyae , one of the largest and oldest organisms on Earth, using extracts from liquid and solid fermentation media. The study unveiled three unprecedented dimeric bismelleolides and three novel fatty‐acid‐substituted congeners, along with 11 new and 21 known derivatives. The structures were elucidated by 1D and 2D NMR spectroscopy and HRESI‐MS, and ROESY spectral analysis for relative configurations. Absolute configurations were determined from crystal structures and through ECD spectra comparison. A compound library of melleolide‐type meroterpenoids facilitated metabolomics‐wide associations, revealing production patterns under different culture conditions. The library enabled assessments of antimicrobial and cytotoxic activities, revealing that the Δ ^2,4 double bond is not crucial for antifungal activity. Cytotoxicity was linked to the presence of an aldehyde at C1, but lost with hydroxylation at C13. Chemoinformatic analyses demonstrated the intricate interplay of chemical modifications on biological properties. This study marks the first systematic exploration of Armillaria spp. meroterpenoid diversity by MS‐based untargeted metabolomics, offering insight into structure–activity relationships through innovative chemoinformatics.

Plants of the genus Otoba have been the basis for the treatment of tropical diseases in indigenous communities of countries like Colombia. Despite the lack of knowledge about their bioactive principles, endophytic fungi derived from medicinal plants are a prolific source of innovative chemistry. We systematically investigated the secondary metabolite production of a previously undescribed species of Diaporthe , herein introduced as Diaporthe caliensis sp. nov., using different metabolomics approaches together with classical chemical screening. To get an outline of the chemical space produced by this fungus, an exploratory molecular networking (MN) analysis was undertaken. A major molecular family was found to contain the known 10-membered lactone phomol ( 1 ), together with other putative congeners as compound 3 . After isolation by preparative high-performance liquid chromatography, we confirmed phomol ( 1 ) as the main reason for the antimicrobial activity of the crude extract. The unknown absolute configuration of 1 was determined by the synthesis of α-methoxy-α-trifluoromethylphenylacetyl (MTPA)-esters and chemical degradation experiments. Moreover, caliensolides A ( 2 ) and B ( 3 ) were isolated, and their structures were elucidated as novel butenolides structurally unrelated to 1 . Overall, the initial MN analysis incorrectly clustered compounds 1 and 3 within a single molecular family, despite evident differences in chemical structures and biosynthetic origin. Contrariwise, the unsupervised substructure discovery algorithm MS2LDA provided a deeper understanding of the fragmentation patterns and correctly clustered the polyketide-lactones produced by D. caliensis sp. nov. Our findings encourage the exploration of Colombian fungal diversity, which as demonstrated here could result in the discovery of new natural products. IMPORTANCE: The integration of metabolomics-based approaches into the discovery pipeline has enabled improved mining and prioritization of prolific secondary metabolite producers such as endophytic fungi. However, relying on automated untargeted analysis tools might lead to misestimation of the chemical complexity harbored in these organisms. Our study emphasizes the importance of isolation and structure elucidation of the respective metabolites in addition to deep metabolome analysis for the correct interpretation of untargeted metabolomics approaches such as molecular networking. Additionally, it encourages the further exploration of endophytic fungi from traditional medicinal plants for the discovery of natural products.

Over the past two decades, hypoxylaceous specimens were collected from several sites in Thailand. In this study, we examined their affinity to the genus Pyrenopolyporus using macroscopic and microscopic morphological characters, dereplication of their stromatal secondary metabolites using ultrahigh performance liquid chromatography coupled to diode array detection and ion mobility tandem mass spectrometry (UHPLC-DAD-IM-MS/MS), and molecular phylogenetic analyses. We describe and illustrate five novel species and a new record for the country, present multi-locus phylogenetic analyses that show the distinction between the proposed species, and provide proteomic profiles of the fungi using matrix associated laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF/MS) for the first time. Based on our findings, this strategy is useful as a complementary tool to distinguish species between Daldinia and Pyrenopolyporus in a consistent way with the phylogenetic analysis.

During a study of the diversity of soilborne fungi from Spain, a strain belonging to the family Chaetomiaceae (Sordariales) was isolated. The multigene phylogenetic inference using five DNA loci showed that this strain represents an undescribed species of the genus Amesia, herein introduced as A. hispanica sp. nov. Investigation of its secondary metabolome led to the isolation of two new derivatives (2 and 3) of the known antifungal antibiotic dactylfungin A (1), together with the known compound cochliodinol (4). The planar structures of 1–4 were determined by ultrahigh performance liquid chromatography coupled with diode array detection and ion mobility tandem mass spectrometry (UHPLC-DAD-IM-MS/MS) and extensive 1D and 2D nuclear magnetic resonance (NMR) spectroscopy after isolation by HPLC. All isolated secondary metabolites were tested for their antimicrobial and cytotoxic activities. Dactylfungin A (1) showed selective and strong antifungal activity against some of the tested human pathogens (Aspergillus fumigatus and Cryptococcus neoformans). The additional hydroxyl group in 2 resulted in the loss of activity against C. neoformans but still retained the inhibition of As. fumigatus in a lower concentration than that of the respective control, without showing any cytotoxic effects. In contrast, 25″-dehydroxy-dactylfungin A (3) exhibited improved activity against yeasts (Schizosaccharomyces pombe and Rhodotorula glutinis) than 1 and 2, but resulted in the appearance of slight cytotoxicity. The present study exemplifies how even in a well-studied taxonomic group such as the Chaetomiaceae, the investigation of novel taxa still brings chemistry novelty, as demonstrated in this first report of this antibiotic class for chaetomiaceous and sordarialean taxa.

Fungi are known as prolific producers of bioactive secondary metabolites with applications across various fields, including infectious diseases, as well as in biological control. However, some of the well-known species are still underexplored. Our current study evaluated the production of secondary metabolites by the entomopathogenic fungus Beauveria neobassiana from Thailand. The fermentation of this fungus in a liquid medium, followed by preparative high-performance liquid chromatography (HPLC) purification, resulted in the isolation of a new tenellin congener, namely pretenellin C (1), together with five known derivatives (2–6). Their chemical structures were elucidated by 1D and 2D nuclear magnetic resonance (NMR) spectroscopy in combination with high-resolution electrospray ionization mass spectrometry (HR-ESI-MS). We evaluated the antimicrobial and cytotoxic activities from all isolated compounds, as well as their inhibitory properties on biofilm formation by Staphylococcus aureus. Generally, tenellins displayed varying antibiofilm and cytotoxic effects, allowing us to propose preliminary structure-activity relationships (SARs). Among the tested compounds, prototenellin D (2) exhibited the most prominent antibiofilm activity, while its 2-pyridone congener, tenellin (4), demonstrated potent cytotoxic activity against all tested cell lines. Given the fact that the biological activities of the tenellins have so far been neglected in the past, our study could provide a good starting point to establish more concise structure-activity relationships in the near future.