Application of culturomics in fungal isolation from mangrove sediments.

BACKGROUND: Fungi play a crucial role in ecosystems, and they have been widely considered a promising source for natural compounds that are crucial for drug discovery. Fungi have a high diversity, but about 95% of them remain unknown to science. The description rate of fungi is very low, mainly due to the inability of most fungi to grow in artificial media, which could not provide a sufficiently similar environment to their natural habitats. Moreover, many species in nature are in a state of low metabolic activity which cannot readily proliferate without proper resuscitation. Previously developed culturomics techniques are mostly designed and applicable for bacteria, with few attempts for fungal isolation because of their significantly larger cell size and hyphal growth properties. RESULTS: This study attempted to isolate previously uncultured and rare fungi from mangrove sediments using newly developed fungal enrichment culture method (FECM) and fungal isolation chips (FiChips). Comparison of fungal community composition at different enrichment stages showed that FECM had great influence on fungal community composition, with rare taxa increased significantly, thus improving the isolation efficiency of previously uncultured fungi. Similarly, in situ cultivation using FiChips has a significant advantage in detecting and culturing rare fungi, as compared to the conventional dilution plate method (DPM). In addition, based on morphological comparisons and phylogenetic analyses, we described and proposed 38 new ascomycetous taxa, including three new families, eight new genera, 25 new species, and two new combinations (presented in additional file 1). CONCLUSIONS: Our study demonstrated that mangrove sediments harbor a high diversity of fungi, and our new isolation approaches (FECM and FiChips) presented a high efficiency in isolating hitherto uncultured fungi, which is potentially usable for fungal isolation in other similar environments. Video Abstract.

Genomic characterization of three marine fungi, including Emericellopsis atlantica sp. nov. with signatures of a generalist lifestyle and marine biomass degradation.

Marine fungi remain poorly covered in global genome sequencing campaigns; the 1000 fungal genomes (1KFG) project attempts to shed light on the diversity, ecology and potential industrial use of overlooked and poorly resolved fungal taxa. This study characterizes the genomes of three marine fungi: Emericellopsis sp. TS7, wood-associated Amylocarpus encephaloides and algae-associated Calycina marina. These species were genome sequenced to study their genomic features, biosynthetic potential and phylogenetic placement using multilocus data. Amylocarpus encephaloides and C. marina were placed in the Helotiaceae and Pezizellaceae (Helotiales), respectively, based on a 15-gene phylogenetic analysis. These two genomes had fewer biosynthetic gene clusters (BGCs) and carbohydrate active enzymes (CAZymes) than Emericellopsis sp. TS7 isolate. Emericellopsis sp. TS7 (Hypocreales, Ascomycota) was isolated from the sponge Stelletta normani. A six-gene phylogenetic analysis placed the isolate in the marine Emericellopsis clade and morphological examination confirmed that the isolate represents a new species, which is described here as E. atlantica. Analysis of its CAZyme repertoire and a culturing experiment on three marine and one terrestrial substrates indicated that E. atlantica is a psychrotrophic generalist fungus that is able to degrade several types of marine biomass. FungiSMASH analysis revealed the presence of 35 BGCs including, eight non-ribosomal peptide synthases (NRPSs), six NRPS-like, six polyketide synthases, nine terpenes and six hybrid, mixed or other clusters. Of these BGCs, only five were homologous with characterized BGCs. The presence of unknown BGCs sets and large CAZyme repertoire set stage for further investigations of E. atlantica. The Pezizellaceae genome and the genome of the monotypic Amylocarpus genus represent the first published genomes of filamentous fungi that are restricted in their occurrence to the marine habitat and form thus a valuable resource for the community that can be used in studying ecological adaptions of fungi using comparative genomics.

Leptosphaerulina species isolated from golf turfgrass in China, with description of L. macrospora, sp. nov.

Leptosphaerulina leaf blight occurs on most turfgrasses. Hitherto, Leptosphaerulina species associated with this disease include L. americana, L. argentinensis, L. australis, and L. trifolii. However, following Koch's postulates, L. australis was confirmed as saprobes but not pathogens, and the other three species have not been tested. The pathogenicity of Leptosphaerulina spp. is still questionable. In this study, we isolated 19 Leptosphaerulina strains from diseased golf turfgrasses in China, and they were identified as L. gaeumannii, L. saccharicola, and a new species, L. macrospora, through multilocus (ITS, 28S, rpb2, and tub2) phylogenetic analyses and morphological observations. Pathogenicity test revealed that the three Leptosphaerulina species identified in this study cannot infect live/healthy turfgrass tissues of Poa pratensis and Agrostis stolonifera and only produced pseudothecia on the dead leaves of stressed seedlings. Considering the results of pathogenicity tests in this and previous studies, we speculate that most Leptosphaerulina species isolated from diseased turfgrass are not pathogens but saprobes. Applying proper management practices to prevent severe turfgrass stress is a key measure to reduce or eliminate the effects of Leptosphaerulina on golf turfgrass.

Citizen science project reveals high diversity in Didymellaceae (Pleosporales, Dothideomycetes).

Fungal communities play a crucial role in maintaining the health of managed and natural soil environments, which directly or indirectly affect the properties of plants and other soil inhabitants. As part of a Citizen Science Project initiated by the Westerdijk Fungal Biodiversity Institute and the Utrecht University Museum, which aimed to describe novel fungal species from Dutch garden soil, the diversity of Didymellaceae, which is one of the largest families in the Dothideomycetes was investigated. A preliminary analysis of the ITS and LSU sequences from the obtained isolates allowed the identification of 148 strains belonging to the family. Based on a multi-locus phylogeny of a combined ITS, LSU, rpb2 and tub2 alignment, and morphological characteristics, 20 different species were identified in nine genera, namely Ascochyta, Calophoma, Didymella, Juxtiphoma, Nothophoma, Paraboeremia, Phomatodes, Stagonosporopsis, and Xenodidymella. Several isolates confirmed to be ubiquitous plant pathogens or endophytes were for the first time identified from soil, such as Ascochyta syringae, Calophoma clematidis-rectae, and Paraboeremia litseae. Furthermore, one new genus and 12 novel species were described from soil: Ascochyta benningiorum sp. nov., Didymella degraaffiae sp. nov., D. kooimaniorum sp. nov., Juxtiphoma kolkmaniorum sp. nov., Nothophoma brennandiae sp. nov., Paraboeremia rekkeri sp. nov., P. truiniorum sp. nov., Stagonosporopsis stuijvenbergii sp. nov., S. weymaniae sp. nov., Vandijckomycella joseae gen. nov. et sp. nov., V. snoekiae sp. nov., and Xenodidymella weymaniae sp. nov. From the results of this study, soil was revealed to be a rich substrate for members of Didymellaceae, several of which were previously known only from diseased or apparently healthy plant hosts.

Pestalotiopsis and allied genera from Camellia, with description of 11 new species from China.

A total of 124 Pestalotiopsis-like isolates associated with symptomatic and asymptomatic tissues of Camellia sinensis and other Camellia spp. from eight provinces in China were investigated. Based on single- and multi-locus (ITS, TEF, TUB2) phylogenies, as well as morphological characters, host associations and geographical distributions, they were classified into at least 19 species in three genera, i.e. Neopestalotiopsis, Pestalotiopsis and Pseudopestalotiopsis. Eight novel species in Pestalotiopsis and three novel species in Pseudopestalotiopsis were described. Our data suggested that the currently widely used loci in Pestalotiopsis-like genera do not consistently provide stable and sufficient resolution tree topologies, especially for Neopestalotiopsis. Moreover, the number, branch pattern and length of the conidial basal appendages were revealed to be phylogenetically informative characters in Pestalotiopsis.