Phylogenetic Assignment of the Fungicolous Hypoxylon invadens (Ascomycota, Xylariales) and Investigation of its Secondary Metabolites

Abstract

The ascomycete Hypoxylon invadens was described in 2014 as a fungicolous species growing on a member of its own genus, H.fragiforme, which is considered a rare lifestyle in the Hypoxylaceae. This renders H.invadens an interesting target in our efforts to find new bioactive secondary metabolites from members of the Xylariales. So far, only volatile organic compounds have been reported from H.invadens, but no investigation of non-volatile compounds had been conducted. Furthermore, a phylogenetic assignment following recent trends in fungal taxonomy via a multiple sequence alignment seemed practical. A culture of H.invadens was thus subjected to submerged cultivation to investigate the produced secondary metabolites, followed by isolation via preparative chromatography and subsequent structure elucidation by means of nuclear magnetic resonance (NMR) spectroscopy and high-resolution mass spectrometry (HR-MS). This approach led to the identification of the known flaviolin (1) and 3,3-biflaviolin (2) as the main components, which had never been reported from the order Xylariales before. Assessment of their antimicrobial and cytotoxic effects via a panel of commonly used microorganisms and cell lines in our laboratory did not yield any effects of relevance. Concurrently, genomic DNA from the fungus was used to construct a multigene phylogeny using ribosomal sequence information from the internal transcribed spacer region (ITS), the 28S large subunit of ribosomal DNA (LSU), and proteinogenic nucleotide sequences from the second largest subunit of the DNA-directed RNA polymerase II (RPB2) and β-tubulin (TUB2) genes. A placement in a newly formed clade with H.trugodes was strongly supported in a maximum-likelihood (ML) phylogeny using sequences derived from well characterized strains, but the exact position of said clade remains unclear. Both, the chemical and the phylogenetic results suggest further inquiries into the lifestyle of this unique fungus to get a better understanding of both, its ecological role and function of its produced secondary metabolites hitherto unique to the Xylariales.

Screening and evaluation of antidiabetic activities of endophytic fungi associated with Etlingera elatior

“…The Daldinia sequences are monophyletic. This cladogram supports a study by Becker et al (2020), who reported that Xylariaceae is separated into numerous main clades of Hypoxylon and Daldinia taxa, forming another distinct clade accommodated in the Hypoxylaceae. Daldinia and Hypoxylon are included in a distinct family of filamentous Ascomycota, Xylariales, containing taxa of Daldinia and Hypoxylon as a sizable monophyletic group (Hsieh et al 2005;Becker et al 2020).…”

Segregation of the genus Parahypoxylon (Hypoxylaceae, Xylariales) from Hypoxylon by a polyphasic taxonomic approach

“…The combined rooted phylogenetic tree showed a clade consisting of the core members of the Hypoxylaceae, such as Hypoxylon, Daldinia, Pyrenopolyporus, Hypomontagnella, Jackrogersella, Rostrohypoxylon, Thamnomyces and Ruwenzoria with medium BS and high PP support (1/90), which was placed in a sister position to a clade consisting of members of Parahypoxylon gen. nov., and Durotheca (Hypoxylaceae) at the base of the tree with strong support (1/100). The genus Hypoxylon could be confirmed as paraphyletic, as has been described already by Wendt et al (2018), Lambert et al (2019), andBecker et al (2020). The sequences assigned to Parahypoxylon ruwenzoriense formed a highly supported (1/100) cluster with the sequences derived from Parahypoxylon papillatum.…”

Recent progress in biodiversity research on the Xylariales and their secondary metabolism

“…The fungicolous H. invadens , which was only described in 2014, exhibits a unique lifestyle by growing on a member of its own genus, H. fragiforme [ 102 ]. So far, only volatile organic compounds have been described from H. invadens [ 103 ], but a recent investigation into its secondary metabolites revealed the known naphthoquinones flaviolin ( 73 ) and 3,3′-biflaviolin ( 74 ) to be produced in submerged cultures [ 104 ]. Only known as products of the melanin biosynthesis when the underlying pathway was actively influenced (e.g.…”