Minutisphaerales (Dothideomycetes, Ascomycota): a new order of freshwater ascomycetes including a new family, Minutisphaeraceae, and two new species from North Carolina, USA.

Minutisphaera is a recently established genus of freshwater Dothideomycetes characterized by small, globose to subglobose or apothecioid, erumpent to superficial, brown ascomata; fissitunicate, eight-spored, ovoid to obclavate asci; and 1-2-septate, clavate to broadly fusiform, hyaline to pale brown ascospores with or without a gelatinous sheath and filamentous appendages. The genus currently contains two species: M. fimbriatispora, the type species, and M. japonica. The higher-level phylogenetic relationship of Minutisphaera within the Dothideomycetes currently is unresolved. To establish the phylogenetic position of Minutisphaera within the Dothideomycetes and evaluate the phylogenetic affinities of newly collected Minutisphaera-like taxa, we sequenced three rDNA regions-18S, ITS1-5.8SITS2 (ITS) and 28S nuc rDNA, and a protein-coding gene, MCM7, for newly collected strains of Minutisphaera. Based on maximum likelihood and Bayesian analyses of a combined dataset (18S and 28S) composed of 167 taxa, a more refined dataset (28S and MCM7) comprising 52 taxa and a separate ITS dataset, and an examination of morphology, we describe and illustrate two new species of Minutisphaera. The Minutisphaera clade was strongly supported within the Dothideomycetes with likelihood and Bayesian statistics but did not share phylogenetic affinities with any existing taxonomic group within the Dothideomycetes. We therefore establish a new order, Minutisphaerales, and new family, Minutisphaeraceae, for this monophyletic clade of freshwater ascomycetes. Chemical analysis of the organic extract M. aspera (G427) resulted in isolation and characterization of five known secondary metabolites, of which four were dipeptides (1-4) and one an aromatic polyketide (5). Conversely, two aromatic polyketides (5, 6) were isolated and identified from the organic extract of M. parafimbriatispora (G156-4). The isolated compounds were tested for their antimicrobial activity against an array of bacteria and fungi. Compound 6 showed promising activity against Staphylococcus aureus and Mycobacterium smegmatis with minimal inhibitory concentration values of 30 and 60 µg/mL, respectively.

New Insights into Tetraplosphaeriaceae Based on Taxonomic Investigations of Bambusicolous Fungi and Freshwater Fungi.

Species within Tetraplosphaeriaceae have been frequently documented in recent years with the extensive investigations of microfungi along a latitudinal gradient from north to south in the Asian/Australian region. Both bamboo substrates and freshwater habitats serve as extensive reservoirs, hosting a rich diversity of fungi that exhibit broad geographical distributions. The most common fungi in these two environments are generally distributed in distinct families. However, our statistics have revealed an intriguingly distinct preference of Tetraplosphaeriaceae species for inhabiting both bamboo substrates and freshwater habitats. The genera Pseudotetraploa (100%) and Triplosphaeria (100%) exhibit a strong preference, followed by Shrungabeeja (71%) and Quadricrura (67%). Our taxonomic and phylogenetic study of microfungi in southern China have identified four additional novel species, viz., Aquatisphaeria bambusae sp. nov., Pseudotetraploa phyllostachydis sp. nov., Pseudotetraploa yangjiangensis sp. nov., and Tetraploa submersa sp. nov. from bamboo substrates and freshwater habitats. In addition, Aquatisphaeria thailandica has previously been documented from freshwater habitats in Thailand; however, we have once again isolated this species from decaying bamboo substrates in Guangdong, China. The new findings substantiate our hypothesis that the preference of Tetraplosphaeriaceae species for colonizing bamboo substrates and freshwater habitats will be more evident through more extensive investigations conducted in such environments.

Freshwater ascomycetes: Minutisphaera (Dothideomycetes) revisited, including one new species from Japan.

During investigations of freshwater ascomycetes we found one interesting taxon from Aomori (Japan), as well as three additional taxa from North Carolina (USA), which were morphologically similar to Minutisphaera, a recently described freshwater fungus in the Dothideomycetes. The ascomata of all the collections bore dark hair-like structures around the ostiolar region, obovoid to obclavate bitunicate asci, and one to three septate hyaline to brown ascospores with a sheath (in material from Japan), and with both sheath and appendages (in material from the USA). The apothecial ascomata of these taxa, however, differ from those of the type species of the genus, which are perithecial. Two collections of Minutisphaera-like fungi from the USA were morphologically quite similar but differed in ascospore size. To assess the phylogenetic affinities of Minutisphaera-like taxa with the type species, M. fimbriatispora, we sequenced 18S and 28S nrDNA of five newly collected strains of Minutisphaera. We also sequenced the nrDNA for the entire internal transcribed spacer region of 10 strains to assess interspecific and intraspecific variation with M. fimbriatispora. Additionally we examined the secondary metabolite profiles of two strains from USA. Based on maximum likelihood and Bayesian analyses of combined 18S and 28S, and separate ITS sequences, as well as examination of morphology, we describe and illustrate a new species, M. japonica. One collection from North Carolina is confirmed as M. fimbriatispora, while two other collections are Minutisphaera-like fungi that had a number of similar diagnostic morphological characters but differed only slightly in ascospore sizes. The phylogeny inferred from the internal transcribed spacer region suggested that two out of the three North Carolina collections may be novel and perhaps cryptic species within Minutisphaera. Organic extracts of Minutisphaera from USA, M. fimbriatispora (G155-1) and Minutisphaera-like taxon (G156-1), revealed the presence of palmitic acid and (E)-hexadec-9-en-1-ol as major chemical constituents. We discuss the placement of the Minutisphaera clade within the Dothideomycetes. The description of the genus Minutisphaera is emended to accommodate M. japonica within Minutisphaera.

Multi-Gene Phylogenetic Analyses Revealed Five New Species and Two New Records of Distoseptisporales from China.

Eight hyphomycetes were collected as part of an investigation into the diversity of hyphomycetous fungi in China. Based on morphology and multi-loci (LSU, ITS, tef1α, and rpb2) phylogenetic analyses, five new taxa, including a new Aquapteridospora species A. hyalina and four novel Distoseptispora species, viz D. aquisubtropica, D. septata, D. tropica, and D. wuzhishanensis were introduced in Distoseptisporales (Sordariomycetes). Two new habitat records, viz Distoseptispora pachyconidia and D. xishuangbannaensis were firstly reported. Also provided in this study are detailed descriptions of eight new collections and a revised phylogenetic tree for the Distoseptisporales.

Polyphasic Identification of Distoseptispora with Six New Species from Fresh Water.

Twelve new specimens of sporidesmium-like taxa were collected from freshwater habitats in China and Thailand. Phylogenetic analysis of nuc 28S rDNA (LSU), internal transcribed spacer (ITS), translation elongation factor 1-alpha (TEF1-α) and second-largest subunit of RNA polymerase II (RPB2) sequence data, combined with morphological data, revealed that they are Distoseptispora species. Among them, six new species, including D. aqualignicola, D. aquamyces, D. crassispora, D. curvularia, D. nonrostrata and D. pachyconidia, are introduced. Two new combinations, D. adscendens and D. leonensis, are transferred from Ellisembia. A new habitat and geographical record are reported for D. clematidis, collected from a freshwater habitat in China. New RPB2 sequence data for D. dehongensis are provided.

Five Novel Taxa from Freshwater Habitats and New Taxonomic Insights of Pleurotheciales and Savoryellomycetidae.

Pleurotheciales is the largest order in Savoryellomycetidae with a large proportion of species known from freshwater habitats. In order to investigate the phylogenetic relationships of taxa within Pleurotheciales and contribute to their diversity, submerged wood was collected from freshwater habitats in China (Yunnan Province) and Thailand. Two dematiaceous, sporodochial hyphomycetes and one annulatascales-like ascomycete with unusual morphology as compared to extant ones were discovered. They were subjected to DNA-based phylogenetic analyses and the results revealed three distinct lineages in Savoryellomycetidae. This morpho-phylo taxonomic study supports the establishment of five novel taxa including two novel genera, Obliquifusoideum and Saprodesmium, and three novel species, Coleodictyospora muriformis, Obliquifusoideum guttulatum and Saprodesmium dematiosporum. Coleodictyospora muriformis and S. dematiosporum are placed in Pleurotheciales, while O. guttulatum is referred to Savoryellomycetidae genera incertae sedis. The phylogenetic relationships are also presented for Coleodictyospora and Pseudocoleodictyospora, which raises an intriguing taxonomic issue. These two genera are positioned in two different classes, viz Sordariomycetes and Dothideomycetes, although they are quite similar except for the presence of a conidial sheath. This study expands our knowledge of the fungal diversity of freshwater fungi, and also indicates that Pleurotheciales species are mostly found in freshwater habitats.

Pseudobactrodesmium (Dactylosporaceae, Eurotiomycetes, Fungi) a Novel Lignicolous Genus.

During our ongoing surveys of fungi on submerged wood in the Greater Mekong Subregion, we collected two new species similar to Bactrodesmium longisporum. Pseudobactrodesmium gen. nov. is introduced to accommodate the new species, P. aquaticum, P. chiangmaiensis and B. longisporum is transferred to this genus. Fasciculate conidiophores, enteroblastic conidiogenous cells and subulate to fusiform, phragmoseptate conidia with a tapering apical cell and sheath characterize the genus. Pseudobactrodesmium aquaticum has longer conidia than P. chiangmaiensis. The placement of Pseudobactrodesmium in Dactylosporaceae (Eurotiomycetes) is a novel finding based on analyses of combined LSU, SSU, ITS and RPB2 sequence data. Our study reveals that Pseudobactrodesmium is likely to be a speciose genus with different species in streams around the world.

Acetophenone derivatives from a freshwater fungal isolate of recently described Lindgomyces madisonensis (G416).

The exploration of freshwater ascomycetes, which have undergone only limited investigation, may provide opportunities both to characterize new genera/species of fungi and to uncover new chemical diversity. In this study, seven acetophenone derivatives, madisone, 4'-methoxymadisone, dehydromadisone, 2″-methoxymadisone, dihydroallovisnaginone, dimadisone, and 4'-methoxydimadisone were characterized from an organic extract of a recently described Lindgomyces madisonensis (G416) culture, which was isolated from submerged wood collected in a stream in North Carolina. Madisone, dehydromadisone, 2″-methoxymadisone, dimadisone and 4'-demethoxydimadisone have not been reported previously, while 4'-methoxymadisone and dihydroallovisnaginone were previously unknown as natural products. Their structures were assigned on the basis of NMR and HRESIMS data, with the structure of madisone supported by X-ray crystallography. The antimicrobial activities of madisone, 4'-methoxymadisone and dihydroallovisnaginone were evaluated against a panel of bacteria and fungi. A heat map analysis of the surface of a G416 culture showed that most of the isolated compounds concentrated in the guttate compared with the vegetative mycelium of the fungus.

Microfungal oasis in an oligotrophic desert: diversity patterns and community structure in three freshwater systems of Cuatro Ciénegas, Mexico.

The Cuatro Ciénegas Basin (CCB) comprises several oligotrophic aquatic ecosystems limited by phosphorus. These aquatic systems are dominated by a high prokaryotic diversity, shaped by the stress of low nutrient supplies and interspecific competition. Although fungi constitute a diverse and important component of microbial diversity, the microfungal diversity in the CCB remains to be unveiled. With the aim to explore microfungal diversity and ecological patterns in this area, we present the first investigation analyzing cultivable taxa from sediment and water, as well as lignocellulolytic taxa obtained from incubated submerged plant debris, and wood panels in three contrasting freshwater systems in the CCB: Churince, Becerra and Pozas Rojas. We chose a culture-based approach to analyze sediment and water samples in order to obtain fungal cultures, providing opportunities for a posteriori studies, and the possibility of ex situ preservation of the diversity. We evaluated sequence data from the nuclear ribosomal internal transcribed spacer including the 5.8 rDNA region for 126 isolates, revealing 37 OTUs. These OTUs were phylogenetically affiliated to several genera in the fungal phyla: Zygomycota, Basidiomycota, and Ascomycota. We recorded two OTUs with saline affinity, agreeing with previous findings on the prokaryotic communities with ancestral marine resemblances. All the studied systems showed moderate diversity levels, however discrepancies among the diversity indexes were observed, due to the occurrence of abundant taxa in the samples. Our results indicated that lignocellulolytic microfungal communities are dominated by transient fungal taxa, as resident species were not recorded perhaps as a result of the long-term strong competition with the highly adapted prokaryotic community. Moreover, the obtained microfungal taxa occurred mostly on the resident plant debris, rather than submerged wood panels, perhaps as a result of the high adaptation to specific environmental conditions. In conclusion, the CCB possess a moderate taxonomical diversity compared to other arid environments, probably as a result of high selective pressures. Nonetheless, due to high spatial and temporal heterogeneity, the functional fungal diversity was considerable as predicted by the intermediate disturbance hypothesis. Decisively, the assessment of microfungal diversity freshwater systems is relevant, since this ecological group of microorganisms represents an important indicator of trophic complexity and biotic interactions among microbial communities, having important implications for understanding eukaryotic survival at the oligotrophic limit for life.

The Acoustic Properties of Water Submerged Lodgepole Pine (Pinus contorta) and Spruce (Picea spp.) Wood and Their Suitability for Use as Musical Instruments.

Wood is a common material used for the manufacture of many products, and submerged wood, in particular, has been used in niche markets and musical instruments. In order to examine if submerged wood in British Columbia, Canada, would be appropriate for use as musical instruments, a study was performed in 2007 on submerged wood from Ootsa Lake, British Columbia, Canada. The results of that study showed the wood was not suitable for musical instruments. In this paper, the wood samples were allowed to age untouched in a laboratory setting and were then retested under the hypothesis that physical acoustic characteristics would improve. It was shown, however, that acoustic properties became less adequate after being left to dry over time. This article describes the density, speed of sound, acoustic constant and characteristic impedance properties for submerged wood and a comparison is made for different applications for musical instruments.