Discovery of a new species of the Hypoxylon rubiginosum complex from Iran and antagonistic activities of Hypoxylon spp. against the Ash Dieback pathogen, Hymenoscyphus fraxineus, in dual culture

Abstract

During a survey of xylarialean fungi in Northern Iran, several specimens that showed affinities to the Hypoxylon rubiginosum complex were collected and cultured. A comparison of their morphological characters, combined with a chemotaxonomic study based on high performance liquid chromatography, coupled with diode array detection and mass spectrometry (HPLC-DAD/MS) and a multi-locus phylogeny based on ITS, LSU, rbp2 and tub2 DNA sequences, revealed a new species here described as Hypoxylon guilanense. In addition, Hypoxylon rubiginosumsensu stricto was also encountered. Concurrently, an endophytic isolate of the latter species showed strong antagonistic activities against the Ash Dieback pathogen, Hymenoscyphus fraxineus, in a dual culture assay in our laboratory. Therefore, we decided to test the new Iranian fungi for antagonistic activities against the pathogen, along with several cultures of other Hypoxylon species that are related to H. rubiginosum. Our results suggest that the antagonistic effects of Hypoxylon spp. against Hym. fraxineus are widespread and that they are due to the production of antifungal phomopsidin derivatives in the presence of the pathogen.

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

“…Furthermore, one non-supported clade containing H. fragiforme and allies (Hx3) was placed as sister group next to a clade containing mostly other genera like Pyrenopolyporus (P, 100% support), Daldinia and allies (99%/100%; E, D, R, T), Hypomontagnella (Hy, 100%) and other nested sequences with non- or low supported positions of Hypoxylon and Entonaema (E, Hx4, Hx5, Hx6) with moderate BS and high likelihood-ratio support (95%/58%). These patterns were already described by Wendt et al [ 2 ], Sir et al [ 20 ], and Pourmoghaddam et al [ 39 ]. The H. invadens -derived sequences were placed next to H. trugodes with maximum support (Hx5).…”

Resolution of the Hypoxylon fuscum Complex (Hypoxylaceae, Xylariales) and Discovery and Biological Characterization of Two of Its Prominent Secondary Metabolites

“…Morphological analyses of microscopic characters were carried out as described by Pourmoghaddam et al [ 21 ]. Pigment colors were determined as described in the latter monograph, with colour codes following Rayner [ 22 ].…”

“…Stromata from Hypoxylon specimens were extracted using acetone as described previously [ 23 ] and subsequently analysed by high-performance liquid chromatography coupled to a diode array and electrospray mass spectrometric detection system (HPLC/DAD-ESIMS) with instrument settings as described recently [ 21 ]. Resulting UV/Vis and mass spectrometric data were compared with internal databases, comprising standards of known Hypoxylaceae and literature data [ 10 , 11 , 12 ].…”

“…Iran (especially the north of Iran) includes subtropical regions and houses numerous species of Xylariales. Only few surveys on their diversity have so far been conducted in the country [ 18 , 19 , 20 , 21 ], and even the Iranian species of Hypoxylon are still poorly known and need further examination.…”

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

“…Endophytes belonging to the genus Hypoxylon and its associated asexual stage Nodulisporium have also been repeatedly reported to produce biologically active volatiles [121,122], even though the taxonomy of most of these strains and the identity of many of the volatile antibiotics produced by these fungi remain to be settled. Recently, an endophytic isolate that was unambiguously identified as H. rubiginosum was found to possess striking activities in dual culture against the Ash dieback pathogen, Hymenoscyphus fraxineus, and the active principles were identified after preparative isolation and classical structure elucidation (NMR spectroscopy and HR mass spectrometry) as phomopsidin derivatives [123,124]. In a follow-up study, the production of phomopsidins (85)(86) in presence of the pathogen was observed in various strains of species that are phylogenetically related to H. rubiginosum including the new species, H. guilanense, but not in some distantly related species like H. fuscum.…”