Mast seeding in European beech (Fagus sylvatica L.) is associated with reduced fungal sporocarp production and community diversity.

Mast seeding is a well-documented phenomenon across diverse forest ecosystems. While its effect on aboveground food webs has been thoroughly studied, how it impacts the soil fungi that drive soil carbon and nutrient cycling has not yet been explored. To evaluate the relationship between mast seeding and fungal resource availability, we paired a Swiss 29-year fungal sporocarp census with contemporaneous seed production for European beech (Fagus sylvatica L.). On average, mast seeding was associated with a 55% reduction in sporocarp production and a compositional community shift towards drought-tolerant taxa across both ectomycorrhizal and saprotrophic guilds. Among ectomycorrhizal fungi, traits associated with carbon cost did not explain species' sensitivity to seed production. Together, our results support a novel hypothesis that mast seeding limits annual resource availability and reproductive investment in soil fungi, creating an ecosystem 'rhythm' to forest processes that is synchronized above- and belowground.

Detecting Pathogenic Phytophthora Species Using Volatile Organic Compounds.

There are several highly damaging Phytophthora species pathogenic to forest trees, many of which have been spread beyond their native range by the international trade of live plants and infested materials. Such introductions can be reduced through the development of better tools capable of the early, rapid, and high-throughput detection of contaminated plants. This study utilized a volatilomics approach (solid-phase microextraction coupled to gas chromatography-mass spectrometry) to differentiate between several Phytophthora species in culture and discriminate between healthy and Phytophthora-inoculated European beech and pedunculate oak trees. We tentatively identified 14 compounds that could differentiate eight Phytophthora species from each other in vitro. All of the Phytophthora species examined, except Phytophthora cambivora, uniquely produced at least one compound not observed in the other species; however, most detected compounds were shared between multiple species. Phytophthora polonica had the most unique compounds and was the least similar of all the species examined. The inoculated seedlings had qualitatively different volatile profiles and could be distinguished from the healthy controls by the presence of isokaurene, anisole, and a mix of three unknown compounds. This study supports the notion that volatiles are suitable for screening plant material, detecting tree pathogens, and differentiating between healthy and diseased material.

Fossil history of fungus host-specificity: Association of conidia of fossil Asterosporium asterospermum with macro- and microremains of Fagus.

Fossil staurosporous conidia almost identical to modern conidia of Asterosporium asterospermum were found from three Central European localities ranging from the Late Oligocene (Germany) to Middle/Late Miocene (Poland). Extant A. asterospermum is strictly host-specific and found only on branches or bark of various Fagus species from Europe, Asia and North America. Conspicuous association of conidia of A. asterospermum with numerous macro- and microremains of Fagus were reported from all the localities where fossil conidia of Asterosporium were found confirming the host-specificity of fossil A. asterospermum to ancient beeches. The host-specific relationship of A. asterospermum and beech was presumably established early in the history of the Fagus genus.

Ectomycorrhizal diversity, taxon-specific traits and root N uptake in temperate beech forests.

Roots of forest trees are colonized by a diverse spectrum of ectomycorrhizal (EM) fungal species differing in their nitrogen (N) acquisition abilities. Here, we hypothesized that root N gain is the result of EM fungal diversity or related to taxon-specific traits for N uptake. To test our hypotheses, we traced 15 N enrichment in fine roots, coarse roots and taxon-specific ectomycorrhizas in temperate beech forests in two regions and three seasons, feeding 1 mM NH4 NO3 labelled with either 15 NH4 + or 15 NO3 - . We morphotyped > 45 000 vital root tips and identified 51 of 53 detected EM species by sequencing. EM root tips exhibited strong, fungal taxon-specific variation in 15 N enrichment with higher NH4 + than NO3 - enrichment. The translocation of N into the upper parts of the root system increased with increasing EM fungal diversity. Across the growth season, influential EM species predicting root N gain were not identified, probably due to high temporal dynamics of the species composition of EM assemblages. Our results support that root N acquisition is related to EM fungal community-level traits and highlight the importance of EM diversity for tree N nutrition.

Bacterial, but not fungal, communities show spatial heterogeneity in European beech (Fagus sylvatica L.) deadwood.

Deadwood decomposition and other environmental processes mediated by microbial communities are generally studied with composite sampling strategies, where deadwood is collected from multiple locations in a large volume, that produce an average microbial community. In this study, we used amplicon sequencing to compare fungal and bacterial communities sampled with either traditional, composite samples, or small, 1 cm3 cylinders from a discrete location within decomposing European beech (Fagus sylvatica L.) tree trunks. We found that bacterial richness and evenness is lower in small samples when compared to composite samples. There was no significant difference in fungal alpha diversity between different sampling scales, suggesting that visually defined fungal domains are not restricted to a single species. Additionally, we found that composite sampling may obscure variation in community composition and this affects the understanding of microbial associations that are detected. For future experiments in environmental microbiology, we recommend that scale is explicitly considered as a factor and properly selected to correspond with the questions asked. Studies of microbial functions or associations may require samples to be collected at a finer scale than is currently practised.

Natural durability of timber species exposed to xylophagous fungi in southern Durango, Mexico.

Introduction: Wood is a natural resource used for construction and the manufacture of many products. This material is exposed to damage due to biotic and abiotic factors. An important biotic factor is wood-degrading fungi that generate large economic losses. The objectives of this study were to determine the effect of xylophagous fungi (Coniophora puteana and Trametes versicolor) on the natural durability of six timber species in southern Durango, Mexico, and to establish differences between fungal effects on each tree species. Materials and Methods: Samples of Pinus durangensis, P. cooperi, P. strobiformis, Juniperus deppeana, Quercus sideroxyla, and Alnus acuminata were exposed to fungi for 4 months under laboratory conditions according to European Standard EN350-1. Samples of Fagus sylvatica were used as control. Durability was determined as the percentage of wood mass loss for each species. Welch ANOVA tests were performed to establish differences among tree species. Welch t-tests were used to prove loss mass differences between fungi for each tree species. Results: The most resistant species to C. puteana were P. durangensis, J. deppeana, P. cooperi and P. strobiformis, showing mean mass losses lower than 8.08%. The most resistant species to T. versicolor were J. deppeana, P. strobiformis and P. durangensis (mean mass losses lower than 7.39%). Pinus strobiformis and Q. sideroxyla were more susceptible to C. puteana effect; in contrast, P. durangensis and P. cooperi showed more damage due to T. versicolor degradation. Conclusions: Woods of P. durangensis, P. cooperi, P. strobiformis and Juniperus deppeana are well adapted to infection by these xylophagous fungi and are therefore highly recommended for commercial use in southern Durango, Mexico.

Both abundant and rare fungi colonizing Fagus sylvatica ectomycorrhizal root-tips shape associated bacterial communities.

Ectomycorrhizal fungi live in close association with their host plants and form complex interactions with bacterial/archaeal communities in soil. We investigated whether abundant or rare ectomycorrhizal fungi on root-tips of young beech trees (Fagus sylvatica) shape bacterial/archaeal communities. We sequenced 16S rRNA genes and fungal internal transcribed spacer regions of individual root-tips and used ecological networks to detect the tendency of certain assemblies of fungal and bacterial/archaeal taxa to inhabit the same root-tip (i.e. modularity). Individual ectomycorrhizal root-tips hosted distinct fungal communities associated with unique bacterial/archaeal communities. The structure of the fungal-bacterial/archaeal association was determined by both, dominant and rare fungi. Integrating our data in a conceptual framework suggests that the effect of rare fungi on the bacterial/archaeal communities of ectomycorrhizal root-tips contributes to assemblages of bacteria/archaea on root-tips. This highlights the potential impact of complex fine-scale interactions between root-tip associated fungi and other soil microorganisms for the ectomycorrhizal symbiosis.

Metagenomes, metatranscriptomes and microbiomes of naturally decomposing deadwood.

Deadwood represents significant carbon (C) stock in a temperate forests. Its decomposition and C mobilization is accomplished by decomposer microorganisms - fungi and bacteria - who also supply the foodweb of commensalist microbes. Due to the ecosystem-level importance of deadwood habitat as a C and nutrient stock with significant nitrogen fixation, the deadwood microbiome composition and function are critical to understanding the microbial processes related to its decomposition. We present a comprehensive suite of data packages obtained through environmental DNA and RNA sequencing from natural deadwood. Data provide a complex picture of the composition and function of microbiome on decomposing trunks of European beech (Fagus sylvatica L.) in a natural forest. Packages include deadwood metagenomes, metatranscriptomes, sequences of total RNA, bacterial genomes resolved from metagenomic data and the 16S rRNA gene and ITS2 metabarcoding markers to characterize the bacterial and fungal communities. This project will be of use to microbiologists, environmental biologists and biogeochemists interested in the microbial processes associated with the transformation of recalcitrant plant biomass.

Urbanization pressures alter tree rhizosphere microbiomes.

The soil microbial community (SMC) provides critical ecosystem services including organic matter decomposition, soil structural formation, and nutrient cycling. Studies suggest plants, specifically trees, act as soil keystone species controlling SMC structure via multiple mechanisms (e.g., litter chemistry, root exudates, and canopy alteration of precipitation). Tree influence on SMC is shaped by local/regional climate effects on forested environments and the connection of forests to surrounding landscapes (e.g., urbanization). Urban soils offer an ideal analog to assess the influence of environmental conditions versus plant species-specific controls on SMC. We used next generation high throughput sequencing to characterize the SMC of specific tree species (Fagus grandifolia [beech] vs Liriodendron tulipifera [yellow poplar]) across an urban-rural gradient. Results indicate SMC dissimilarity within rural forests suggests the SMC is unique to individual tree species. However, greater urbanization pressure increased SMC similarity between tree species. Relative abundance, species richness, and evenness suggest that increases in similarity within urban forests is not the result of biodiversity loss, but rather due to greater overlap of shared taxa. Evaluation of soil chemistry across the rural-urban gradient indicate pH, Ca+, and organic matter are largely responsible for driving relative abundance of specific SMC members.

Characterization of mating type genes in heterothallic Neonectria species, with emphasis on N. coccinea, N. ditissima, and N. faginata.

Neonectria ditissima and N. faginata are canker pathogens involved in an insect-fungus disease complex of American beech (Fagus grandifolia) in North America commonly known as beech bark disease (BBD). In Europe, both N. ditissima and N. coccinea are involved in BBD on European beech (Fagus sylvatica). Field observations across the range of BBD indicate ascospores to be the dominant spore type in the environment. Several studies report a heterothallic (self-sterile) mating strategy for Neonectria fungi, but one study reported homothallism (self-fertility) for N. ditissima. As such, investigations into mating strategy are important for understanding both the disease cycle and population genetics of Neonectria. This is particularly important in the United States given that over time N. faginata dominates the BBD pathosystem despite high densities of nonbeech hosts for N. ditissima. This study utilized whole-genome sequences of BBD-associated Neonectria spp. along with other publicly available Neonectria and Corinectria genomes and in vitro mating assays to characterize mating type (MAT) locus and confirm thallism for select members of Neonectria and Corinectria. MAT gene-specific primer pairs were developed to efficiently characterize the mating types of additional single-ascospore strains of N. ditissima, N. faginata, and N. coccinea and several other related species lacking genomic data. These assays also confirmed the sexual compatibility among N. ditissima strains from different plant hosts. Maximum likelihood phylogenetic analyses of both MAT1-1-1 and MAT1-2-1 sequences recovered trees with similar topology to previously published phylogenies of Neonectria and Corinectria. The results of this study indicate that all Neonectria and Corinectria tested are heterothallic based on our limited sampling and, as such, thallism cannot help explain the inevitable dominance of N. faginata in the BBD pathosystem.

Fungal diversity in canopy soil of silver beech, Nothofagus menziesii (Nothofagaceae).

Adventitious roots in canopy soils associated with silver beech (Nothofagus menziesii Hook.f. (Nothofagaceae)) form ectomycorrhizal associations. We investigated the extent to which canopy ectomycorrhizal communities contribute to overall diversity of ectomycorrhizal fungi associated with silver beech. Hyphal ingrowth bags were buried for 12 months in canopy and terrestrial soils of five trees at one site. We used amplicon sequencing of the nuclear ribosomal internal transcribed spacer 2 region (ITS2) to assess diversity of both ectomycorrhizal and non-ectomycorrhizal OTUs in hyphal ingrowth bags. There was a significant difference in ectomycorrhizal fungal community diversity between the terrestrial and canopy hyphal ingrowth bag communities. Ectomycorrhizal community composition of the terrestrial and canopy environments was also significantly different. Some ectomycorrhizal taxa were significantly differentially represented in either the terrestrial or canopy environment. The hyphal ingrowth bags also accumulated non-ectomycorrhizal species. The non-ectomycorrhizal fungi also had significantly different diversity and community composition between the canopy and terrestrial environments. Like the ectomycorrhizal community, some non-ectomycorrhizal taxa were significantly differentially represented in either the terrestrial or canopy environment. The canopy soil microhabitat provides a novel environment for growth of ectomycorrhizal adventitious roots and enables the spatial partitioning of ectomycorrhizal and non-ectomycorrhizal fungal diversity in the forest.

Mrakia fibulata sp. nov., a psychrotolerant yeast from temperate and cold habitats.

Tree fluxes are sugar-rich, sometimes ephemeral, substrates occurring on sites where tree sap (xylem or phloem) is leaking through damages of tree bark. Tree sap infested with microorganisms has been the source of isolation of many species, including the biotechnologically relevant carotenoid yeast Phaffia rhodozyma. Tree fluxes recently sampled in Germany yielded 19 species, including several psychrophilic yeasts of the genus Mrakia. Four strains from tree fluxes represented a potential novel Mrakia species previously known from two isolates from superficial glacial melting water of Calderone Glacier (Italy). The Italian isolates, originally identified as Mrakia aquatica, and two strains from Germany did not show any sexual structures. But another culture collected in Germany produced clamped hyphae with teliospores. A detailed examination of the five isolates (three from Germany and two from Italy) proved them to be a novel yeast species, which is described in this manuscript as Mrakia fibulata sp. nov. (MB 830398), holotype DSM 103931 and isotype DBVPG 8059. In contrast to other sexually reproducing Mrakia species, M. fibulata produces true hyphae with clamp connections. Also, this is the first psychrotolerant Mrakia species which grows above 20 °C. Spring tree fluxes are widespread and can be recognized and sampled by amateurs in a Citizen Science project. This substrate is a prominent source of yeasts, and may harbor unknown species, as demonstrated in the present work. The description of Mrakia fibulata is dedicated to our volunteer helpers and amateurs, like Anna Yurkova (9-years-old daughter of Andrey Yurkov), who collected the sample which yielded the type strain of this species.

The genetic relationships within Apiognomonia errabunda and related species.

Species of Apiognomonia are some of the most ubiquitous leaf-associated fungi of broad-leaved trees of the northern temperate zone. Especially widespread and diverse is Apiognomonia errabunda, found mostly on beech, oak, and linden. This species and its closest relatives are known for their complicated phylogenetic relationships that have caused considerable confusion in the past. In the present paper, we describe the results of a three-gene-based reconstruction of phylogenetic relationships between A. errabunda, A. veneta, and A. hystrix for 373 isolates. Using combined information from the nuc rDNA internal transcribed spacer region ITS1-5.8S-ITS2 (ITS barcode) and intron regions of actin (ACT) and calmodulin (CAL) genes for 263 isolates, we confirmed the occurrence of host-associated lineages within A. errabunda. However, the clustering of isolates by host species was incomplete: some isolates occurred on the "wrong" hosts and a number of isolates carried mixed genetic profile, indicating substantial level of inter-host group recombination. In addition, a number of isolates were identified as putative, pending further verification, interspecies hybrids between the much more divergent A. errabunda and A. hystrix.

Effects of root and leaf litter identity and diversity on oribatid mite abundance, species richness and community composition.

Habitat heterogeneity is an important driver of aboveground species diversity but few studies have investigated effects on soil communities. Trees shape their surrounding by both leaf litter and roots generating small scale heterogeneity and potentially governing community patterns of soil organisms. To assess the role of vegetation for the soil fauna, we studied whether tree species (Fagus sylvatica L., Acer pseudoplatanus L., Fraxinus excelsior L., Tilia cordata Mill.), markedly differing in leaf litter quality and root associated mycorrhizal symbionts, affect oribatid mite communities by shaping below- and aboveground resources and habitat complexity and availability. Oribatid mite abundance, species richness, community structure and the proportion of litter living and parthenogenetic individuals were analyzed and related to microbial biomass and the amount of remaining litter mass. Although leaf litter species with higher nutritional values decomposed considerably faster, microbial biomass only slightly differed between leaf litter species. Neither root species nor leaf litter species affected abundance, species richness or community structure of oribatid mites. However, root species had an effect on the proportion of parthenogenetic individuals with increased proportions in the presence of beech roots. Overall, the results suggest that identity and diversity of vegetation via leaf litter or roots are of minor importance for structuring oribatid mite communities of a temperate forest ecosystem.

Home-Field Advantage in Wood Decomposition Is Mainly Mediated by Fungal Community Shifts at "Home" Versus "Away".

The home-field advantage (HFA) hypothesis has been used intensively to study leaf litter decomposition in various ecosystems. However, the HFA in woody substrates is still unexplored. Here, we reanalyzed and integrated existing datasets on various groups of microorganisms collected from natural deadwood of two temperate trees, Fagus sylvatica and Picea abies, from forests in which one or other of these species dominates but where both are present. Our aims were (i) to test the HFA hypothesis on wood decomposition rates of these two temperate tree species, and (ii) to investigate if HFA hypothesis can be explained by diversity and community composition of bacteria and in detail N-fixing bacteria (as determined by molecular 16S rRNA and nifH gene amplification) and fungi (as determined by molecular ITS rRNA amplification and sporocarp surveys). Our results showed that wood decomposition rates were accelerated at "home" versus "away" by 38.19% ± 20.04% (mean ± SE). We detected strong changes in fungal richness (increase 36-50%) and community composition (RANOSIM = 0.52-0.60, P < 0.05) according to HFA hypothesis. The changes of fungi were much stronger than for total bacteria and nitrogen fixing for both at richness and community composition levels. In conclusion, our results support the HFA hypothesis in deadwood: decomposition rate is accelerated at home due to specialization of fungal communities produced by the plant community above them. Furthermore, the higher richness of fungal sporocarps and nitrogen-fixing bacteria (nifH) may stimulate or at least stabilize wood decomposition rates at "home" versus "away."

Capsulimonas corticalis gen. nov., sp. nov., an aerobic capsulated bacterium, of a novel bacterial order, Capsulimonadales ord. nov., of the class Armatimonadia of the phylum Armatimonadetes.

An aerobic bacterial strain designated AX-7T was isolated from the trunk surface of a Japanese beech (Fagus crenata). Cells of strain AX-7T were Gram-stain-negative, non-spore-forming, non-motile rods (1.0-1.2 µm in width and 1.2-3.0 µm in length) with peritrichous fimbriae. Cells were capsulated, and a number of them were surrounded by a thick slime layer. During growth, large aggregates formed, and the culture medium became viscous probably owing to exopolysaccharide release from the slime layer. The temperature range for growth was 10-37 °C, with an optimum at 30 °C. The pH range for growth was 5.0-7.0, with an optimum at pH 6.0. Strain AX-7T used various sugars, including polysaccharides, and yeast extract as growth substrates. Strain AX-7T contained menaquinones MK-9 and MK-10 as the respiratory quinones, and C16 : 1ω5c, C16 : 1ω11c, C16 : 0 and C14 : 0 as the major cellular fatty acids. Four unidentified phospholipids and 11 unidentified polar lipids constituted the polar lipids. The DNA G+C content was 61.0 mol%. The cell-wall peptidoglycan contained ll-diaminopimelic acid. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain AX-7T belonged to the class Armatimonadia, its closest relative being Armatimonas rosea YO-36T, with sequence similarity of 88.1%. Based on data from this polyphasic study, we propose that strain AX-7T represents a new genus of a novel species within the novel order Capsulimonadales ord. nov. of the class Armatimonadia, for which the name Capsulimonas corticalis gen. nov., sp. nov. is proposed. The type strain of C. corticalis is AX-7T (=DSM 105890T=NBRC 113044T).

Spruce and beech as local determinants of forest fungal community structure in litter, humus and mineral soil.

Beech forests reaches its native distribution limit in SE Norway, but is expected to expand substantially northwards due to climate warming. This may potentially result in a fundamental transformation of contemporary Northern European forests, with tentative effects on the associated belowground fungi. Fungal communities mediate vital ecosystem processes such as ecosystem productivity and carbon sequestration in boreal forests. To investigate how soil fungi is affected by the vegetation transition from spruce to beech forest, we sampled litter, humus and mineral soil in a forest landscape dominated by beech, spruce or a mixture of these. The fungal communities in the soil samples were analyzed by DNA metabarcoding of the rDNA ITS2 region. Although soil layers were the most important structuring gradient, we found clear differences in fungal species composition between spruce and beech plots. The differences in fungal community composition were most evident in the litter and least in the mineral soil. Decomposers, most notably Mycena, dominated the litter layer while various mycorrhizal fungi dominated the humus and mineral layers. Some ectomycorrhizal taxa, such as Cenoccocum and Russula, were more abundant in spruce forests. Differences in fungal community composition between forest types can potentially have large impacts on carbon sequestration rates.

Highly competitive fungi manipulate bacterial communities in decomposing beech wood (Fagus sylvatica).

The bacterial communities in decomposing wood are receiving increased attention, but their interactions with wood-decay fungi are poorly understood. This is the first field study to test the hypothesis that fungi are responsible for driving bacterial communities in beech wood (Fagus sylvatica). A meta-genetic approach was used to characterise bacterial and fungal communities in wood that had been laboratory-colonised with known wood-decay fungi, and left for a year at six woodland sites. Alpha-, Beta- and Gammaproteobacteria and Acidobacteria were the proportionally dominant bacterial taxa, as in previous studies. Pre-colonising wood with decay fungi had a clear effect on the bacterial community, apparently via direct fungal influence; the bacterial and fungal communities present at the time of collection explained nearly 60% of their mutual covariance. Site was less important than fungal influence in determining bacterial communities, but the effects of pre-colonisation were more pronounced at some sites than at others. Wood pH was also a strong bacterial predictor, but was itself under considerable fungal influence. Burkholderiaceae and Acidobacteriaceae showed directional responses against the trend of the bacterial community as a whole.

Leaf litter species identity influences biochemical composition of ectomycorrhizal fungi.

In forest ecosystems, ectomycorrhizal (ECM) fungi are important for plant growth and soil biogeochemical processes. The biochemical composition of ECM mycelium is an important fungal effect trait with consequences for its decomposition rate, and consequently on soil carbon pools and plant nutrition. Although the link between ECM fungi and leaf litter-released nutrients is well known, the response of ECM fungal biochemical composition to different leaf litter species remains poorly understood. To determine how leaf litter quality influences ECM fungi's biochemical profiles, we planted young beech trees in an oak forest and replaced the natural leaf litter with that of European beech (Fagus sylvatica), ash (Fraxinus excelsior), maple (Acer pseudoplatanus), or lime (Tilia cordata). We assessed the biochemical profiles of ECM root tips colonized by common fungal taxa in temperate forests (i.e., Cenococcum geophilum, Inocybe sp., and Lactarius subdulcis), using attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR). ECM fungal biochemical composition changed with leaf litter species. Changes were apparent in the infrared absorption bands assigned to functional groups of lipids, amides, and carbohydrates. C. geophilum and L. subdulcis exhibited large spectral differences corresponding to the initial pattern of leaf litter chemical composition between samples collected in the beech and ash leaf litter treatments. In contrast, Inocybe sp. was influenced by lime, but with no differences between samples from ash or beech leaf litter treatments. Although the spectral bands affected by leaf litter type differed among ECM fungi, they were mainly related to amides, indicating a dynamic response of the fungal proteome to soil nutritional changes. Overall, the results indicate that the biochemical response of ECM fungi to leaf litter species varies among ECM fungal species and suggests that the biochemical composition of ECM mycelium is a fungal response trait, sensitive to environmental changes such as shifts in leaf litter species.

Three new Leptographium spp. (Ophiostomatales) infecting hardwood trees in Norway and Poland.

Species of Leptographium are characterized by mononematous or synnematous conidiophores and are commonly associated with different arthropods. Some of them also produce a sexual state characterised by globose ascomata with elongated necks. Compared to investigations on coniferous trees, the occurrence of Leptographium species on hardwood trees has been poorly studied in Europe. During a survey of ophiostomatoid fungi on various hardwood tree species in Norway and Poland, three unusual species, which fit in the broader morphological description of Leptographium spp., were found in association with Trypodendron domesticum, Trypodendron signatum and Dryocoetes alni, and from wounds on a variety of hardwoods. Phylogenetic analyses of sequence data for six different loci (ITS1-5.8 S-ITS2, ITS2-LSU, ACT, ß-tubulin, CAL, and TEF-1α) showed that these Leptographium species are phylogenetically closely related to the species of the Grosmannia olivacea complex. The first species forms a well-supported lineage that includes Ophiostoma brevicolle, while the two other new taxa resided in a separate lineage; possibly affiliated with Grosmannia francke-grosmanniae. All the new species produce perithecia with necks terminating in ostiolar hyphae and orange-section shaped ascospores with cucullate, gelatinous sheaths. These species also produce dark olivaceous mononematous asexual states in culture. In addition, two of the newly described species have a second type of conidiophore with a short and non-pigmented stipe. The new Leptographium species can be easily distinguished from each other by their appearance and growth in culture. Based on novel morphological characters and distinct DNA sequences, these fungi were recognised as new taxa for which the names Leptographium tardum sp. nov., Leptographium vulnerum sp. nov., and Leptographium flavum sp. nov. are provided.