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Arbuscular mycorrhizal (AM) fungi - Glomeromycota and Endogonomycetes - comprise multiple species and higher-level taxa that have remained undescribed. We propose a mixed morphology- and DNA-based classification framework to promote taxonomic communication and shed light into the phylogenetic structure of these ecologically essential fungi. Based on eDNA samples and long reads as type materials, we describe 15 new species and corresponding genera (Pseudoentrophospora kesseensis, Hoforsa rebekkae, Kahvena rebeccae, Kelottijaervia shannonae, Kungsaengena shadiae, Langduoa dianae, Lehetua indrekii, Lokruma stenii, Moostea stephanieae, Nikkaluokta mahdiehiae, Parnigua craigii, Riederberga sylviae, Ruua coralieae, Tammsaarea vivikae and Unemaeea nathalieae), the genus Parvocarpum as well as 19 families (Pseudoentrophosporaceae, Hoforsaceae, Kahvenaceae, Kelottijaerviaceae, Kungsaengenaceae, Langduoaceae, Lehetuaceae, Lokrumaceae, Moosteaceae, Nikkaluoktaceae, Parniguaceae, Riederbergaceae, Ruuaceae, Tammsaareaceae, Unemaeeaceae, Bifigurataceae, Planticonsortiaceae, Jimgerdemanniaceae and Vinositunicaceae) and 17 orders (Hoforsales, Kahvenales, Kelottijaerviales, Kungsaengenales, Langduoales, Lehetuales, Lokrumales, Moosteales, Nikkaluoktales, Parniguales, Riederbergales, Ruuales, Tammsaareales, Unemaeeales, Bifiguratales and Densosporales), and propose six combinations (Diversispora bareae, Diversispora nevadensis, Fuscutata cerradensis, Fuscutata reticulata, Viscospora deserticola and Parvocarpum badium) based on phylogenetic evidence. We highlight further knowledge gaps in the phylogenetic structure of AM fungi and propose an alphanumeric coding system for preliminary communication and reference-based eDNA quality-filtering of the remaining undescribed genus- and family-level groups. Using AM fungi as examples, we hope to offer a sound, mixed framework for classification to boost research in the alpha taxonomy of fungi, especially the “dark matter fungi”.

Abstract In our changing world, understanding plant community responses to global change drivers is critical for predicting future ecosystem composition and function. Plant functional traits promise to be a key predictive tool for many ecosystems, including grasslands; however, their use requires both complete plant community and functional trait data. Yet, representation of these data in global databases is sparse, particularly beyond a handful of most used traits and common species. Here we present the CoRRE Trait Data, spanning 17 traits (9 categorical, 8 continuous) anticipated to predict species’ responses to global change for 4,079 vascular plant species across 173 plant families present in 390 grassland experiments from around the world. The dataset contains complete categorical trait records for all 4,079 plant species obtained from a comprehensive literature search, as well as nearly complete coverage (99.97%) of imputed continuous trait values for a subset of 2,927 plant species. These data will shed light on mechanisms underlying population, community, and ecosystem responses to global change in grasslands worldwide.

Abstract To adapt to the complex belowground environment, plants make trade-offs between root resource acquisition and defence ability. This includes forming partnerships with different types of root associating microorganisms, such as arbuscular mycorrhizal and ectomycorrhizal fungi. These trade-offs, by mediating root chemistry, exert legacy effects on nutrient release during decomposition, which may, in turn, affect the ability of new roots to re-acquire resources, thereby generating a feedback loop. However, the linkages at the basis of this potential feedback loop remain largely unquantified. Here, we propose a trait-based root ‘acquisition-defence-decomposition’ conceptual framework and test the strength of relevant linkages across 90 angiosperm tree species. We show that, at the plant species level, the root-fungal symbiosis gradient within the root economics space, root chemical defence (condensed tannins), and root decomposition rate are closely linked, providing support to this framework. Beyond the dichotomy between arbuscular mycorrhizal-dominated versus ectomycorrhizal-dominated systems, we suggest a continuous shift in feedback loops, from ‘high arbuscular mycorrhizal symbiosis-low defence-fast decomposition-inorganic nutrition’ by evolutionarily ancient taxa to ‘high ectomycorrhizal symbiosis-high defence-slow decomposition-organic nutrition’ by more modern taxa. This ‘acquisition-defence-decomposition’ framework provides a foundation for testable hypotheses on multidimensional linkages between species’ belowground strategies and ecosystem nutrient cycling in an evolutionary context.

Abstract Novel methods for sampling and characterizing biodiversity hold great promise for re-evaluating patterns of life across the planet. The sampling of airborne spores with a cyclone sampler, and the sequencing of their DNA, have been suggested as an efficient and well-calibrated tool for surveying fungal diversity across various environments. Here we present data originating from the Global Spore Sampling Project, comprising 2,768 samples collected during two years at 47 outdoor locations across the world. Each sample represents fungal DNA extracted from 24 m3 of air. We applied a conservative bioinformatics pipeline that filtered out sequences that did not show strong evidence of representing a fungal species. The pipeline yielded 27,954 species-level operational taxonomic units (OTUs). Each OTU is accompanied by a probabilistic taxonomic classification, validated through comparison with expert evaluations. To examine the potential of the data for ecological analyses, we partitioned the variation in species distributions into spatial and seasonal components, showing a strong effect of the annual mean temperature on community composition.

Abstract Forest soils harbor hyper-diverse microbial communities which fundamentally regulate carbon and nutrient cycling across the globe. Directly testing hypotheses on how microbiome diversity is linked to forest carbon storage has been difficult, due to a lack of paired data on microbiome diversity and in situ observations of forest carbon accumulation and storage. Here, we investigated the relationship between soil microbiomes and forest carbon across 238 forest inventory plots spanning 15 European countries. We show that the composition and diversity of fungal, but not bacterial, species is tightly coupled to both forest biotic conditions and a seven-fold variation in tree growth rates and biomass carbon stocks when controlling for the effects of dominant tree type, climate, and other environmental factors. This linkage is particularly strong for symbiotic endophytic and ectomycorrhizal fungi known to directly facilitate tree growth. Since tree growth rates in this system are closely and positively correlated with belowground soil carbon stocks, we conclude that fungal composition is a strong predictor of overall forest carbon storage across the European continent.

Antibiotic resistance crisis dictates the need for resistance monitoring and the search for new antibiotics. The development of monitoring protocols is hindered by the great diversity of resistance factors, while the “streetlight effect” denies the possibility of discovering novel drugs based on existing databases. In this study, we address these challenges using high-throughput environmental screening viewed from a trait-based ecological perspective. Through an in-depth analysis of the metagenomes of 658 topsoil samples spanning Europe, we explored the distribution of 241 prokaryotic and fungal genes responsible for producing metabolites with antibiotic properties and 485 antibiotic resistance genes. We analyzed the diversity of these gene collections at different levels and modeled the distribution of each gene across environmental gradients. Our analyses revealed several nonparallel distribution patterns of the genes encoding sequential steps of enzymatic pathways synthesizing large antibiotic groups, pointing to gaps in existing databases and suggesting potential for discovering new analogues of known antibiotics. We show that agricultural activity caused a continental-scale homogenization of microbial antibiotic-related machinery, emphasizing the importance of maintaining indigenous ecosystems within the landscape mosaic. Based on the relationships between the proportion of the genes in the metagenomes with the main predictors (soil pH, land cover type, climate temperature and humidity), we illustrate how the properties of chemical structures dictate the distribution of the genes responsible for their synthesis across environments. With this understanding, we propose general principles to facilitate the discovery of antibiotics, including principally new ones, establish abundance baselines for antibiotic resistance genes, and predict their dissemination.

Societal Impact Statement Humankind is facing both climate and biodiversity crises. This article proposes the foundations of a scheme that offers tradable credits for combined aboveground and soil carbon and biodiversity. Multidiversity—as estimated based on high‐throughput molecular identification of soil meiofauna, fungi, bacteria, protists, plants and other organisms shedding DNA into soil, complemented by acoustic and video analyses of aboveground macrobiota—offers a cost‐effective method that captures much of the terrestrial biodiversity. Such a voluntary crediting system would increase the quality of carbon projects and contribute funding for delivering the Kunming‐Montreal Global Biodiversity Framework. Summary Carbon crediting and land offsets for biodiversity protection have been developed to tackle the challenges of increasing greenhouse gas emissions and the loss of global biodiversity. Unfortunately, these two mechanisms are not optimal when considered separately. Focusing solely on carbon capture—the primary goal of most carbon‐focused crediting and offsetting commitments—often results in the establishment of non‐native, fast‐growing monocultures that negatively affect biodiversity and soil‐related ecosystem services. Soil contributes a vast proportion of global biodiversity and contains traces of aboveground organisms. Here, we outline a carbon and biodiversity co‐crediting scheme based on the multi‐kingdom molecular and carbon analyses of soil samples, along with remote sensing estimation of aboveground carbon as well as video and acoustic analyses‐based monitoring of aboveground macroorganisms. Combined, such a co‐crediting scheme could help halt biodiversity loss by incentivising industry and governments to account for biodiversity in carbon sequestration projects more rigorously, explicitly and equitably than they currently do. In most cases, this would help prioritise protection before restoration and help promote more socially and environmentally sustainable land stewardship towards a ‘nature positive’ future.

Fungi are an integral part of the nitrogen and phosphorus cycling in trophic networks, as they participate in biomass decomposition and facilitate plant nutrition through root symbioses. Nutrient content varies considerably between the main fungal habitats, such as soil, plant litter or decomposing dead wood, but there are also large differences within habitats. While some soils are heavily loaded with N, others are limited by N or P. One way in which nutrient availability can be reflected in fungi is their content in biomass. In this study, we determined the C, N, and P content (in dry mass) of fruiting bodies of 214 fungal species to inspect how phylogeny and membership in ecological guilds (soil saprotrophs, wood saprotrophs, and ectomycorrhizal fungi) affect the nutrient content of fungal biomass. The C content of fruiting bodies (415 ± 25 mg g–1) showed little variation (324–494 mg g–1), while the range of N (46 ± 20 mg g–1) and P (5.5 ± 3.0 mg g–1) contents was within one order of magnitude (8–103 mg g–1 and 1.0–18.9 mg g–1, respectively). Importantly, the N and P contents were significantly higher in the biomass of soil saprotrophic fungi compared to wood saprotrophic and ectomycorrhizal fungi. While the average C/N ratio in fungal biomass was 11.2, values exceeding 40 were recorded for some fungi living on dead wood, typically characterized by low N content. The N and P content of fungal mycelium also showed a significant phylogenetic signal, with differences in nutrient content being relatively low within species and genera of fungi. A strong correlation was found between N and P content in fungal biomass, while the correlation of N content and the N-containing fungal cell wall biopolymer—chitin showed only weak significance. The content of macronutrients in fungal biomass is influenced by the fungal life style and nutrient availability and is also limited by phylogeny.

BackgroundThe cultivation of short-rotation tree species on non-forest land is increasing due to the growing demand for woody biomass for the future bioeconomy and to mitigate climate change impacts. However, forest plantations are often seen as a trade-off between climate benefits and low biodiversity. The diversity and composition of soil fungal biota in plantations of hybrid aspen, one of the most planted tree species for short-rotation forestry in Northern Europe, are poorly studied.MethodsThe goal of this study was to obtain baseline knowledge about the soil fungal biota and the edaphic, floristic and management factors that drive fungal richness and communities in 18-year-old hybrid aspen plantations on former agricultural soils and compare the fungal biota with those of European aspen stands on native forest land in a 130-year chronosequence. Sites were categorized as hybrid aspen (17–18-year-old plantations) and native aspen stands of three age classes (8–29, 30–55, and 65-131-year-old stands). High-throughput sequencing was applied to soil samples to investigate fungal diversity and assemblages.ResultsNative aspen forests showed a higher ectomycorrhizal (EcM) fungal OTU richness than plantations, regardless of forest age. Short-distance type EcM genera dominated in both plantations and forests. The richness of saprotrophic fungi was similar between native forest and plantation sites and was highest in the middle-aged class (30–55-year-old stands) in the native aspen stands. The fungal communities of native forests and plantations were significantly different. Community composition varied more, and the natural forest sites were more diverse than the relatively homogeneous plantations. Soil pH was the best explanatory variable to describe soil fungal communities in hybrid aspen stands. Soil fungal community composition did not show any clear patterns between the age classes of native aspen stands.ConclusionWe conclude that edaphic factors are more important in describing fungal communities in both native aspen forest sites and hybrid aspen plantation sites than forest thinning, age, or former land use for plantations. Although first-generation hybrid aspen plantations and native forests are similar in overall fungal diversity, their taxonomic and functional composition is strikingly different. Therefore, hybrid aspen plantations can be used to reduce felling pressure on native forests; however, our knowledge is still insufficient to conclude that plantations could replace native aspen forests from the soil biodiversity perspective.

Abstract Phytopathogenic fungi threaten global food security but the ecological drivers of their global diversity and biogeography remain unknown. Here, we construct and analyse a global atlas of potential phytopathogenic fungi from 20,312 samples across all continents and major oceanic island regions, eleven land cover types, and twelve habitat types. We show a peak in the diversity of phytopathogenic fungi in mid-latitude regions, in contrast to the latitudinal diversity gradients observed in aboveground organisms. Our study identifies climate as an important driver of the global distribution of phytopathogenic fungi, and our models suggest that their diversity and invasion potential will increase globally by 2100. Importantly, phytopathogen diversity will increase largely in forest (37.27-79.12%) and cropland (34.93-82.51%) ecosystems, and this becomes more pronounced under fossil-fuelled industry dependent future scenarios. Thus, we recommend improved biomonitoring in forests and croplands, and optimised sustainable development approaches to reduce potential threats from phytopathogenic fungi.

Abstract Factors driving microbial community composition and diversity are well established but the relationship with microbial functioning is poorly understood, especially at large scales. We analysed microbial biodiversity metrics and distribution of potential functional groups along a gradient of increasing land-use perturbation, detecting over 79,000 bacterial and 25,000 fungal OTUs in 715 sites across 24 European countries. We found the lowest bacterial and fungal diversity in less-disturbed environments (woodlands) compared to grasslands and highly-disturbed environments (croplands). Highly-disturbed environments contain significantly more bacterial chemoheterotrophs, harbour a higher proportion of fungal plant pathogens and saprotrophs, and have less beneficial fungal plant symbionts compared to woodlands and extensively-managed grasslands. Spatial patterns of microbial communities and predicted functions are best explained when interactions among the major determinants (vegetation cover, climate, soil properties) are considered. We propose guidelines for environmental policy actions and argue that taxonomical and functional diversity should be considered simultaneously for monitoring purposes.

Fungal metabarcoding of substrates such as soil, wood, and water is uncovering an unprecedented number of fungal species that do not seem to produce tangible morphological structures and that defy our best attempts at cultivation, thus falling outside the scope of the International Code of Nomenclature for algae, fungi, and plants. The present study uses the new, ninth release of the species hypotheses of the UNITE database to show that species discovery through environmental sequencing vastly outpaces traditional, Sanger sequencing-based efforts in a strongly increasing trend over the last five years. Our findings challenge the present stance of some in the mycological community – that the current situation is satisfactory and that no change is needed to “the code” – and suggest that we should be discussing not whether to allow DNA-based descriptions (typifications) of species and by extension higher ranks of fungi, but what the precise requirements for such DNA-based typifications should be. We submit a tentative list of such criteria for further discussion. The present authors hope for a revitalized and deepened discussion on DNA-based typification, because to us it seems harmful and counter-productive to intentionally deny the overwhelming majority of extant fungi a formal standing under the International Code of Nomenclature for algae, fungi, and plants.

Afforestation is proposed as one of the most effective climate solutions for carbon sequestration. As a majority of threatened species are linked to forests, afforestation can also contribute to mitigate the biodiversity crisis. There is however a caveat: the agricultural legacy (high nutrient availability, altered soil biota structure and function) of new forests constrains the development of forest-adapted species, affects tree growth and stability, and delays environmental benefits from afforestation.We hypothesize that inoculation of former arable land with soil (including microbiome, fauna and seeds/rhizomes of understory vegetation) from old forests along with targeted tree species mixtures will improve productivity and more rapidly restore forest-adapted communities. This will ultimately result in diverse, stable and resilient multifunctional forests.We will test this hypothesis and develop applied inoculation methods by: i) exploring soil biota and benchmarking biodiversity in existing afforestation research Chronosequence platforms (chronosequences and sites with increasing distance to other forests); ii) conducting inoculation experiments in mesocosms to measure seedling performance and, above- and belowground linkages; iii) establishing field-scale inoculation experiments in new and existing afforestations to test short- and long-term inoculation success on forest productivity, biodiversity and soil functioning at the ecosystem scale; iv) incorporating the landscape context into guidelines and tools for spatially explicit prioritization of areas for assisted dispersal.The aims are to resolve barriers for successful restoration and develop landscape-scale afforestation strategies that optimize productivity and biodiversity for the planning and implementation of green infrastructure; and produce basic knowledge on the tree, understory vegetation, soil fauna and microbiome nexus and its effect on forest productivity, biodiversity and soil functions (N-retention, C-sequestration, methane uptake).

To help solve the actual problem of global climate warming, it is important to comprehensively study soil organic carbon (SOC), soil fungi, and other parameters at different depths in the soil. This study was aimed at investigating the chemical and microbiological parameters and their interactions at various soil depths (0–5 to 195–200 cm) in an Arenosol in a Scots pine stand in southwestern Lithuania, with a focus on the main groups of fungi and their influence on SOC. The highest diversity of soil fungi species was found at a depth of 50–55 cm. Saprotrophs were dominant at all investigated soil depths. Ectomycorrhizal fungi were mostly abundant at depths of up to 50–55 cm. The C:N ratio gradually decreased down to 50–55 cm, then increased in deeper soil layers (from 50–55 to 195–200 cm). This means that the most active mineralization processes occur at depths of between 0 and 55 cm. Carbon stabilization processes occur at depths of 100–105 to 195–200 cm, and most of this carbon does not enter the atmosphere nor contribute to the process of climate change.

IntroductionTraditional approaches to collecting large-scale biodiversity data pose huge logistical and technical challenges. We aimed to assess how a comparatively simple method based on sequencing environmental DNA (eDNA) characterises global variation in plant diversity and community composition compared with data derived from traditional plant inventory methods.MethodsWe sequenced a short fragment (P6 loop) of the chloroplast trnL intron from from 325 globally distributed soil samples and compared estimates of diversity and composition with those derived from traditional sources based on empirical (GBIF) or extrapolated plant distribution and diversity data.ResultsLarge-scale plant diversity and community composition patterns revealed by sequencing eDNA were broadly in accordance with those derived from traditional sources. The success of the eDNA taxonomy assignment, and the overlap of taxon lists between eDNA and GBIF, was greatest at moderate to high latitudes of the northern hemisphere. On average, around half (mean: 51.5% SD 17.6) of local GBIF records were represented in eDNA databases at the species level, depending on the geographic region.DiscussioneDNA trnL gene sequencing data accurately represent global patterns in plant diversity and composition and thus can provide a basis for large-scale vegetation studies. Important experimental considerations for plant eDNA studies include using a sampling volume and design to maximise the number of taxa detected and optimising the sequencing depth. However, increasing the coverage of reference sequence databases would yield the most significant improvements in the accuracy of taxonomic assignments made using the P6 loop of the trnL region.

The wetland soil microbiome has a major impact on greenhouse gas emissions. Here the authors characterize how a group of archaea contribute to N2O emissions and find that climate and land use changes could promote these organisms.

This study aimed to determine the differences and drivers of oomycete diversity and community composition in alder- and birch-dominated park and natural forest soils of the Fennoscandian and Baltic countries of Estonia, Finland, Lithuania, Norway, and Sweden. For this, we sequenced libraries of PCR products generated from the DNA of 111 soil samples collected across a climate gradient using oomycete-specific primers on a PacBio high-throughput sequencing platform. We found that oomycete communities are most affected by temperature seasonality, annual mean temperature, and mean temperature of the warmest quarter. Differences in composition were partly explained by the higher diversity of Saprolegniales in Sweden and Norway, as both total oomycete and Saprolegniales richness decreased significantly at higher longitudes, potentially indicating the preference of this group of oomycetes for a more temperate maritime climate. None of the evaluated climatic variables significantly affected the richness of Pythiales or Peronosporales. Interestingly, the relative abundance and richness of Pythiales was higher at urban sites compared to forest sites, whereas the opposite was true for Saprolegniales. Additionally, this is the first report of Phytophthora gallica and P. plurivora in Estonia. Our results indicate that the composition of oomycetes in soils is strongly influenced by climatic factors, and, therefore, changes in climate conditions associated with global warming may have the potential to significantly alter the distribution range of these microbes, which comprise many important pathogens of plants.

Our knowledge of microbial biogeography has advanced in recent years, yet we lack knowledge of the global diversity of some important functional groups. Here, we used environmental DNA from 327 globally collected soil samples to investigate the biodiversity patterns of nitrogen-fixing bacteria by focusing on the nifH gene but also amplifying the general prokaryotic 16S SSU region. Globally, N-fixing prokaryotic communities are driven mainly by climatic conditions, with most groups being positively correlated with stable hot or seasonally humid climates. Among soil parameters, pH, but also soil N content were most often shown to correlate with the diversity of N-fixer groups. However, specific groups of N-fixing prokaryotes show contrasting responses to the same variables, notably in Cyanobacteria that were negatively correlated with stable hot climates, and showed a U-shaped correlation with soil pH, contrary to other N-fixers. Also, the non-N-fixing prokaryotic community composition was differentially correlated with the diversity and abundance of N-fixer groups, showing the often-neglected impact of biotic interactions among bacteria.

Abstract Microbes dominate terrestrial ecosystems via their great species diversity and vital ecosystem functions, such as biogeochemical cycling and mycorrhizal symbiosis. Fungi and other organisms form diverse association networks. However, the roles of species belonging to different kingdoms in multi‐kingdom community networks have remained largely elusive. In light of the integrative microbiome initiative, we inferred multiple‐kingdom biotic associations from high elevation timberline soils using the SPIEC‐EASI method. Biotic interactions among plants, nematodes, fungi, bacteria, and archaea were surveyed at the community and network levels. Compared to single‐kingdom networks, multi‐kingdom networks and their associations increased the within‐kingdom and cross‐kingdom edge numbers by 1012 and 10,772, respectively, as well as mean connectivity and negative edge proportion by 15.2 and 0.8%, respectively. Fungal involvement increased network stability (i.e., resistance to node loss) and connectivity, but reduced modularity, when compared with those in the single‐kingdom networks of plants, nematodes, bacteria, and archaea. In the entire multi‐kingdom network, fungal nodes were characterized by significantly higher degree and betweenness than bacteria. Fungi more often played the role of connector, linking different modules. Consistently, structural equation modeling and multiple regression on matrices corroborated the “bridge” role of fungi at the community level, linking plants and other soil biota. Overall, our findings suggest that fungi can stabilize the self‐organization process of multi‐kingdom networks. The findings facilitate the initiation and carrying out of multi‐kingdom community studies in natural ecosystems to reveal the complex above‐ and belowground linkages.

Abstract Data sharing is one of the cornerstones of modern science that enables large-scale analyses and reproducibility. We evaluated data availability in research articles across nine disciplines in Nature and Science magazines and recorded corresponding authors’ concerns, requests and reasons for declining data sharing. Although data sharing has improved in the last decade and particularly in recent years, data availability and willingness to share data still differ greatly among disciplines. We observed that statements of data availability upon (reasonable) request are inefficient and should not be allowed by journals. To improve data sharing at the time of manuscript acceptance, researchers should be better motivated to release their data with real benefits such as recognition, or bonus points in grant and job applications. We recommend that data management costs should be covered by funding agencies; publicly available research data ought to be included in the evaluation of applications; and surveillance of data sharing should be enforced by both academic publishers and funders. These cross-discipline survey data are available from the plutoF repository.

Mycorrhizal symbioses have evolved repeatedly in diverse fungal lineages. A large phylogenomic analysis sheds light on genomic changes associated with transitions from saprotrophy to symbiosis, including divergent genetic innovations underlying the convergent origins of the ectomycorrhizal guild.

Abstract Fungi are a key component of tropical biodiversity. However, due to their inconspicuous and largely subterranean nature, they are usually neglected in biodiversity inventories. The goal of this study was to identify the key determinants of fungal richness, community composition, and turnover in tropical rainforests. We tested specifically for the effect of soil properties, habitat, and locality in Amazonia. For these analyses, we used high‐throughput sequencing data of short and long reads of fungal DNA present in soil and organic litter samples, combining existing and novel genomic data. Habitat type (phytophysiognomy) emerges as the strongest factor explaining fungal community composition. Naturally open areas—campinas—are the richest habitat overall. Soil properties have different effects depending on the soil layer (litter or mineral soil) and the choice of genetic marker. We suggest that campinas could be a neglected hotspot of fungal diversity. An underlying cause for their rich diversity may be the overall low soil fertility, which increases the reliance on biotic interactions essential for nutrient absorption in these environments, notably ectomycorrhizal fungi–plant associations. Our results highlight the advantages of using both short and long DNA reads produced through high‐throughput sequencing to characterize fungal diversity. While short reads can suffice for diversity and community comparison, long reads add taxonomic precision and have the potential to reveal population diversity.

Peatlands account for 15 to 30% of the world's soil carbon (C) stock and are important controls over global nitrogen (N) cycles. However, C and N concentrations are known to vary among peatlands contributing to the uncertainty of global C inventories, but there are few global studies that relate peatland classification to peat chemistry. We analyzed 436 peat cores sampled in 24 countries across six continents and measured C, N, and organic matter (OM) content at three depths down to 70 cm. Sites were distinguished between northern (387) and tropical (49) peatlands and assigned to one of six distinct broadly recognized peatland categories that vary primarily along a pH gradient. Peat C and N concentrations, OM content, and C:N ratios differed significantly among peatland categories, but few differences in chemistry with depth were found within each category. Across all peatlands C and N concentrations in the 10-20 cm layer, were 440 ± 85.1 g kg-1 and 13.9 ± 7.4 g kg-1, with an average C:N ratio of 30.1 ± 20.8. Among peatland categories, median C concentrations were highest in bogs, poor fens and tropical swamps (446-532 g kg-1) and lowest in intermediate and extremely rich fens (375-414 g kg-1). The C:OM ratio in peat was similar across most peatland categories, except in deeper samples from ombrotrophic tropical peat swamps that were higher than other peatlands categories. Peat N concentrations and C:N ratios varied approximately two-fold among peatland categories and N concentrations tended to be higher (and C:N lower) in intermediate fens compared with other peatland types. This study reports on a unique data set and demonstrates that differences in peat C and OM concentrations among broadly classified peatland categories are predictable, which can aid future studies that use land cover assessments to refine global peatland C and N stocks.

Coarse woody debris (CWD) provides food and shelter to a large proportion of forest biota and is considered vital for biodiversity during periods of harsh weather. However, its importance in long-term stressed ecosystems remains largely unknown. In this work, we explored the contribution of CWD to fungal diversity along the gradient of boreal forest degradation caused by 77 years of heavy industrial emissions. We analyzed the diversity and composition of fungi in 270 samples of well-decayed Picea abies and Abies sibirica logs, as well as forest litter both adjacent to and distant from the logs. Compared with forest litter, the wood had higher water content and possessed substantially lower concentrations of heavy metals, which suggests its potential favorability for biota in polluted areas. The pollution-induced loss of fungal diversity in forest litter reached 34% and was stronger in the microhabitats not influenced by CWD. Meanwhile, wood fungal communities lost less than 10% of their total richness and even increased in alpha diversity. These processes led to the diversity and compositional convergence of fungal communities from different microhabitats and substrates in polluted areas. Despite this, the importance of wood and CWD-influenced microhabitats for fungal diversity maintenance was low. Apart from wood-associated fungi, the taxa whose diversity increased in the wood of polluted areas were ectomycorrhizal fungi and eurytopic soil saprotrophs (Mucoromycota, Mortierellomycota, Eurotiomycetes, and Helotiales) that easily tolerate highly toxic litter. Within the majority of pollution-sensitive soil saprotrophic groups, only terricolous Tricholomataceae benefit from CWD as microrefugia. Upon considering the ecological variability within low-rank taxa, the importance of decayed logs as safe sites can be high for certain soil-inhabiting fungal groups in polluted areas.

With the developments in DNA nanoball sequencing technologies and the emergence of new platforms, there is an increasing interest in their performance in comparison with the widely used sequencing-by-synthesis methods. Here, we test the consistency of metabarcoding results from DNBSEQ-G400RS (DNA nanoball sequencing platform by MGI-Tech) and NovaSeq 6000 (sequencing-by-synthesis platform by Illumina) platforms using technical replicates of DNA libraries that consist of COI gene amplicons from 120 soil DNA samples. By subjecting raw sequencing data from both platforms to a uniform bioinformatics processing, we found that the proportion of high-quality reads passing through the filtering steps was similar in both datasets. Per-sample operational taxonomic unit (OTU) and amplicon sequence variant (ASV) richness patterns were highly correlated, but sequencing data from DNBSEQ-G400RS harbored a higher number of OTUs. This may be related to the lower dominance of most common OTUs in DNBSEQ data set (thus revealing higher richness by detecting rare taxa) and/or to a lower effective read quality leading to generation of spurious OTUs. However, there was no statistical difference in the ASV and post-clustered ASV richness between platforms, suggesting that additional denoising step in the ASV workflow had effectively removed the ‘noisy’ reads. Both OTU-based and ASV-based composition were strongly correlated between the sequencing platforms, with essentially interchangeable results. Therefore, we conclude that DNBSEQ-G400RS and NovaSeq 6000 are both equally efficient high-throughput sequencing platforms to be utilized in studies aiming to apply the metabarcoding approach, but the main benefit of the former is related to lower sequencing cost.

Mycorrhizas—mutualistic relationships formed between fungi and most plant species—are functionally linked to soil carbon stocks. Here the authors map the global distribution of mycorrhizal plants and quantify links between mycorrhizal vegetation patterns and terrestrial carbon stocks.

The number of invasive alien pest and pathogen species affecting ecosystem functioning, human health and economies has increased dramatically over the last decades. Discoveries of invasive pests and pathogens previously unknown to science or with unknown host associations yet damaging on novel hosts highlights the necessity of developing novel tools to predict their appearance in hitherto naïve environments. The use of sentinel plant systems is a promising tool to improve the detection of pests and pathogens before introduction and to provide valuable information for the development of preventative measures to minimize economic or environmental impacts. Though sentinel plantings have been established and studied during the last decade, there still remains a great need for guidance on which tools and protocols to put into practice in order to make assessments accurate and reliable. The sampling and diagnostic protocols chosen should enable as much information as possible about potential damaging agents and species identification. Consistency and comparison of results are based on the adoption of common procedures for sampling design and sample processing. In this paper, we suggest harmonized procedures that should be used in sentinel planting surveys for effective sampling and identification of potential pests and pathogens. We also review the benefits and limitations of various diagnostic methods for early detection in sentinel systems, and the feasibility of the results obtained supporting National Plant Protection Organizations in pest and commodity risk analysis.

Tree species identity is one of the key factors driving ectomycorrhizal (EcM) fungal richness and community composition in boreal and temperate forest ecosystems, but little is known about the influence of tree species combinations and their neighborhood effects on EcM communities. To advance our understanding of host plant effects on EcM fungi, the roots of silver birch, Scots pine, and Norway spruce were analyzed using high-throughput sequencing across mature boreal forest exploratory plots of monocultures and two- and three-species mixtures in Finland. Our analyses revealed that tree species identity was an important determinant of EcM fungal community composition, but tree species richness had no significant influence on EcM fungal richness and community composition. We found that EcM fungal community composition associated with spruce depends on neighboring tree species. Our study suggests that at a regional-scale tree species identity is the primary factor determining community composition of root-associated EcM fungi alongside with tree species composition effects on EcM fungal community of spruce in mixed stands.

Soil microbiome has a pivotal role in ecosystem functioning, yet little is known about its build-up from local to regional scales. In a multi-year regional-scale survey involving 1251 plots and long-read third-generation sequencing, we found that soil pH has the strongest effect on the diversity of fungi and its multiple taxonomic and functional groups. The pH effects were typically unimodal, usually both direct and indirect through tree species, soil nutrients or mold abundance. Individual tree species, particularly Pinus sylvestris, Picea abies, and Populus x wettsteinii, and overall ectomycorrhizal plant proportion had relatively stronger effects on the diversity of biotrophic fungi than saprotrophic fungi. We found strong temporal sampling and investigator biases for the abundance of molds, but generally all spatial, temporal and microclimatic effects were weak. Richness of fungi and several functional groups was highest in woodlands and around ruins of buildings but lowest in bogs, with marked group-specific trends. In contrast to our expectations, diversity of soil fungi tended to be higher in forest island habitats potentially due to the edge effect, but fungal richness declined with island distance and in response to forest fragmentation. Virgin forests supported somewhat higher fungal diversity than old non-pristine forests, but there were no differences in richness between natural and anthropogenic habitats such as parks and coppiced gardens. Diversity of most fungal groups suffered from management of seminatural woodlands and parks and thinning of forests, but especially for forests the results depended on fungal group and time since partial harvesting. We conclude that the positive effects of tree diversity on overall fungal richness represent a combined niche effect of soil properties and intimate associations.

Arbuscular mycorrhizal fungi (AMF) strongly affect ecosystem functioning. To understand and quantify the mechanisms of this control, knowledge about the relationship between the actual abundance and community composition of AMF in the soil and in plant roots is needed. We collected soil and root samples in a natural dune grassland to test whether, across a plant community, the abundance of AMF in host roots (measured as the total length of roots colonized) is related to soil AMF abundance (using the neutral lipid fatty acids (NLFA) 16:1ω5 as proxy). Next-generation sequencing was used to explore the role of community composition in abundance patterns. We found a strong positive relationship between the total length of roots colonized by AMF and the amount of NLFA 16:1ω5 in the soil. We provide the first field-based evidence of proportional biomass allocation between intra-and extraradical AMF mycelium, at ecosystem level. We suggest that this phenomenon is made possible by compensatory colonization strategies of individual fungal species. Finally, our findings open the possibility of using AMF total root colonization as a proxy for soil AMF abundances, aiding further exploration of the AMF impacts on ecosystems functioning.

Along with recent developments in high-throughput sequencing (HTS) technologies and thus fast accumulation of HTS data, there has been a growing need and interest for developing tools for HTS data processing and communication. In particular, a number of bioinformatics tools have been designed for analysing metabarcoding data, each with specific features, assumptions and outputs. To evaluate the potential effect of the application of different bioinformatics workflow on the results, we compared the performance of different analysis platforms on two contrasting high-throughput sequencing data sets. Our analysis revealed that the computation time, quality of error filtering and hence output of specific bioinformatics process largely depends on the platform used. Our results show that none of the bioinformatics workflows appears to perfectly filter out the accumulated errors and generate Operational Taxonomic Units, although PipeCraft, LotuS and PIPITS perform better than QIIME2 and Galaxy for the tested fungal amplicon dataset. We conclude that the output of each platform requires manual validation of the OTUs by examining the taxonomy assignment values.

Diplodia sapinea is a cosmopolitan endophyte and opportunistic pathogen having occurred on several conifer species in Europe for at least 200 years. In Europe, disease outbreaks have increased on several Pinus spp. in the last few decades. In this study, the genetic structure of the European and western Asian D. sapinea population were investigated using 13 microsatellite markers. In total, 425 isolates from 15 countries were analysed. A high clonal fraction and low genetic distance between most subpopulations was found. One single haplotype dominates the European population, being represented by 45.3% of all isolates and found in nearly all investigated countries. Three genetically distinct subpopulations were found: Central/North European, Italian and Georgian. The recently detected subpopulations of D. sapinea in northern Europe (Estonia) share several haplotypes with the German subpopulation. The northern European subpopulations (Latvia, Estonia and Finland) show relatively high genetic diversity compared to those in central Europe suggesting either that the fungus has existed in the North in an asymptomatic/endophytic mode for a long time or that it has spread recently by multiple introductions. Considerable genetic diversity was found even among isolates of a single tree as 16 isolates from a single tree resulted in lower clonal fraction index than most subpopulations in Europe, which might reflect cryptic sexual proliferation. According to currently published allelic patterns, D. sapinea most likely originates from North America or from some unsampled population in Asia or central America. In order to enable the detection of endophytic or latent infections of planting stock by D. sapinea, new species-specific PCR primers (DiSapi-F and Diplo-R) were designed. During the search for Diplodia isolates across the world for species specific primer development, we identified D. africana in California, USA, and in the Canary Islands, which are the first records of this species in North America and in Spain.

The relationship between the ecological success of needle pathogens of forest trees and species richness of co-inhabiting endophytic fungi is poorly understood. One of the most dangerous foliar pathogens of pine is Dothistroma septosporum, which is a widely spread threat to northern European forests. We sampled two Pinus sylvestris sites in Estonia and two in Norway in order to analyse the relations between the abundance of D. septosporum and overall fungal richness, specific fungal species composition, time of season, needle age and position in the canopy. In both countries, the overall species richness of fungi was highest in autumn, showing a trend of increase with needle age. The overall species richness in the second-year needles in Estonia and third-year needles in Norway was similar, suggesting that a critical colonization threshold for needle shed in P. sylvestris is breached earlier in Estonia than in Norway. The fungal species richness in P. sylvestris needles was largely affected by Lophodermium conigenum. Especially in older needles, the relative abundance of L. conigenum was significantly higher in spring compared to summer or autumn. The timing of recruitment and colonization mechanisms of different foliage endophytes are shortly discussed.

Culture techniques are vital in both traditional and modern fungal taxonomy. Establishing sexual–asexual links and synanamorphs, extracting DNA and secondary metabolites are mainly based on cultures. However, it is widely accepted that a large number of species are not sporulating in nature while others cannot be cultured. Recent ecological studies based on culture-independent methods revealed these unculturable taxa, i.e., dark taxa. Recent fungal diversity estimation studies suggested that environmental sequencing plays a vital role in discovering missing species. However, Sanger sequencing is still the main approach in determining DNA sequences in culturable species. In this paper, we summarize culture-based and culture-independent methods in the study of ascomycetous taxa. High-throughput sequencing of leaf endophytes, leaf litter fungi and fungi in aquatic environments is important to determine dark taxa. Nevertheless, currently, naming dark taxa is not recognized by the ICN, thus provisional naming of them is essential as suggested by several studies.

Abstract Background Fungi are a diverse eukaryotic group of degraders, pathogens, and symbionts, with many lineages known only from DNA sequences in soil, sediments, air, and water. Results We provide rough phylogenetic placement and principal niche analysis for >40 previously unrecognized fungal groups at the order and class level from global soil samples based on combined 18S (nSSU) and 28S (nLSU) rRNA gene sequences. Especially, Rozellomycota (Cryptomycota), Zygomycota s.lat, Ascomycota, and Basidiomycota are rich in novel fungal lineages, most of which exhibit distinct preferences for climate and soil pH. Conclusions This study uncovers the great phylogenetic richness of previously unrecognized order- to phylum-level fungal lineages. Most of these rare groups are distributed in different ecosystems of the world but exhibit distinct ecological preferences for climate or soil pH. Across the fungal kingdom, tropical and non-tropical habitats are equally likely to harbor novel groups. We advocate that a combination of traditional and high-throughput sequencing methods enable efficient recovery and phylogenetic placement of such unknown taxonomic groups.

Fungi are ecologically important in several ecosystem processes, yet their community composition, ecophysiological roles, and responses to changing environmental factors in historical sediments are rarely studied. Here we explored ancient fungal DNA from lake Lielais Svētiņu sediment throughout the Holocene (10.5 kyr) using the ITS metabarcoding approach. Our data revealed diverse fungal taxa and smooth community changes during most of the Holocene with rapid changes occurring in the last few millennia. More precisely, plankton parasitic fungi became more diverse from the Late Holocene (2–4 kyr) which could be related to a shift towards a cooler climate. The Latest Holocene (~2 kyr) showed a distinct increase in the richness of plankton parasites, mycorrhizal, and plant pathogenic fungi which can be associated with an increased transfer rate of plant material into the lake and blooms of planktonic organisms influenced by increased, yet moderate, human impact. Thus, major community shifts in plankton parasites and mycorrhizal fungi could be utilized as potential paleo-variables that accompany host-substrate dynamics. Our work demonstrates that fungal aDNA with predicted ecophysiology and host specificity can be employed to reconstruct both aquatic and surrounding terrestrial ecosystems and to estimate the influence of environmental change.

Here, we describe the taxon hypothesis (TH) paradigm, which covers the construction, identification, and communication of taxa as datasets. Defining taxa as datasets of individuals and their traits will make taxon identification and most importantly communication of taxa precise and reproducible. This will allow datasets with standardized and atomized traits to be used digitally in identification pipelines and communicated through persistent identifiers. Such datasets are particularly useful in the context of formally undescribed or even physically undiscovered species if data such as sequences from samples of environmental DNA (eDNA) are available. Implementing the TH paradigm will to some extent remove the impediment to hastily discover and formally describe all extant species in that the TH paradigm allows discovery and communication of new species and other taxa also in the absence of formal descriptions. The TH datasets can be connected to a taxonomic backbone providing access to the vast information associated with the tree of life. In parallel to the description of the TH paradigm, we demonstrate how it is implemented in the UNITE digital taxon communication system. UNITE TH datasets include rich data on individuals and their rDNA ITS sequences. These datasets are equipped with digital object identifiers (DOI) that serve to fix their identity in our communication. All datasets are also connected to a GBIF taxonomic backbone. Researchers processing their eDNA samples using UNITE datasets will, thus, be able to publish their findings as taxon occurrences in the GBIF data portal. UNITE species hypothesis (species level THs) datasets are increasingly utilized in taxon identification pipelines and even formally undescribed species can be identified and communicated by using UNITE. The TH paradigm seeks to achieve unambiguous, unique, and traceable communication of taxa and their properties at any level of the tree of life. It offers a rapid way to discover and communicate undescribed species in identification pipelines and data portals before they are lost to the sixth mass extinction.

The soil fungal community plays an important role in determining plant growth and health. In this study, we investigated the fungal diversity and community composition in the roots and soil of 21 potato (Solanum tuberosum L.) cultivars using high-throughput sequencing at three different time points across the growing season. In soil and roots, the fungal richness and relative abundance of pathogens and saprotrophs were mainly affected by sampling time. While sampling time affected fungal composition in soil, root fungal communities were also significantly affected by cultivar. The cultivar had the strongest effect on diversity of pathogens and abundance of particular pathogen species. Our results demonstrate changes in soil and root fungal communities of potato over the growing season, as well as highlighting the importance of potato cultivar on root fungal communities and abundance of pathogens.

European ash (Fraxinus excelsior) is threatened by the invasive ascomycete Hymenoscyphus fraxineus originating from Asia. Ash leaf tissues serve as a route for shoot infection but also as a sporulation substrate for this pathogen. Knowledge of the leaf niche partitioning by indigenous fungi and H. fraxineus is needed to understand the fungal community receptiveness to the invasion. We subjected DNA extracted from unwashed and washed leaflets of healthy and diseased European ash to PacBio sequencing of the fungal ITS1-5.8S-ITS2 rDNA region. Leaflets from co-inhabiting rowan trees (Sorbus aucuparia) served as a reference. The overlap in leaflet mycobiomes between ash and rowan was remarkably high, but unlike in rowan, in ash leaflets the sequence read proportion, and the qPCR-based DNA amount estimates of H. fraxineus increased vigorously towards autumn, concomitant with a significant decline in overall fungal richness. The niche of ash and rowan leaves was dominated by epiphytic propagules (Vishniacozyma yeasts, the dimorphic fungus Aureobasidion pullulans and the dematiaceous hyphomycete Cladosporium ramotenellum and H. fraxineus), and endophytic thalli of biotrophs (Phyllactinia and Taphrina species), the indigenous necrotroph Venturia fraxini and H. fraxineus. Mycobiome comparison between healthy and symptomatic European ash leaflets revealed no significant differences in relative abundance of H. fraxineus, but A. pullulans was more prevalent in symptomatic trees. The impacts of host specificity, spatiotemporal niche partitioning, species carbon utilization profiles and life cycle traits are discussed to understand the ecological success of H. fraxineus in Europe. Further, the inherent limitations of different experimental approaches in the profiling of foliicolous fungi are addressed.

Rapid development of high-throughput (HTS) molecular identification methods has revolutionized our knowledge about taxonomic diversity and ecology of fungi. However, PCR-based methods exhibit multiple technical shortcomings that may bias our understanding of the fungal kingdom. This study was initiated to quantify potential biases in fungal community ecology by comparing the relative performance of amplicon-free shotgun metagenomics and amplicons of nine primer pairs over seven nuclear ribosomal DNA (rDNA) regions often used in metabarcoding analyses. The internal transcribed spacer (ITS) barcodes ITS1 and ITS2 provided greater taxonomic and functional resolution and richness of operational taxonomic units (OTUs) at the 97% similarity threshold compared to barcodes located within the ribosomal small subunit (SSU) and large subunit (LSU) genes. All barcode-primer pair combinations provided consistent results in ranking taxonomic richness and recovering the importance of floristic variables in driving fungal community composition in soils of Papua New Guinea. The choice of forward primer explained up to 2.0% of the variation in OTU-level analysis of the ITS1 and ITS2 barcode data sets. Across the whole data set, barcode-primer pair combination explained 37.6–38.1% of the variation, which surpassed any environmental signal. Overall, the metagenomics data set recovered a similar taxonomic overview, but resulted in much lower fungal rDNA sequencing depth, inability to infer OTUs, and high uncertainty in identification. We recommend the use of ITS2 or the whole ITS region for metabarcoding and we advocate careful choice of primer pairs in consideration of the relative proportion of fungal DNA and expected dominant groups.

The oomycete-specific ITS primers published by Riit et al. (2016) have been put to use in the scientific community working with oomycetes. Recently, however, it has been brought to our attention that the sequences of the primers ITS1oo and ITS3oo shown in the first Figure of the published manuscript are incomplete, when compared to the sequences of the same primers as listed on the UNITE website. This discrepancy is derived from re-checking primer sequences from tube labels that are restricted to the first 18 bases.Closer examination revealed that the sequence of primer ITS1oo in Figure 1 is missing one nucleotide from the 3’ end and the primer ITS3oo is missing two nucleotides from the 3’ end. These errors are expected to reduce relative primer specificity to Oomycetes, which probably results in lower proportion of this group in metabarcoding studies. We hereby provide the updated figure (Figure 1) with correct information. We apologize to all users of these erroneous primers for their suboptimal performance. We are grateful to Dr. Diana Marčiulynienė and Dr. Sannakajsa Velmala for pointing out these problems.

Molecular data form an important research tool in most branches of mycology. A non-trivial proportion of the public fungal DNA sequences are, however, compromised in terms of quality and reliability, contributing noise and bias to sequence-borne inferences such as phylogenetic analysis, diversity assessment, and barcoding. In this paper we discuss various aspects and pitfalls of sequence quality assessment. Based on our observations, we provide a set of guidelines to assist in manual quality management of newly generated, near-full-length (Sanger-derived) fungal ITS sequences and to some extent also sequences of shorter read lengths, other genes or markers, and groups of organisms. The guidelines are intentionally non-technical and do not require substantial bioinformatics skills or significant computational power. Despite their simple nature, we feel they would have caught the vast majority of the severely compromised ITS sequences in the public corpus. Our guidelines are nevertheless not infallible, and common sense and intuition remain important elements in the pursuit of compromised sequence data. The guidelines focus on basic sequence authenticity and reliability of the newly generated sequences, and the user may want to consider additional resources and steps to accomplish the best possible quality control. A discussion on the technical resources for further sequence quality management is therefore provided in the supplementary material.

Truffles have evolved from epigeous (aboveground) ancestors in nearly every major lineage of fleshy fungi. Because accelerated rates of morphological evolution accompany the transition to the truffle form, closely related epigeous ancestors remain unknown for most truffle lineages. This is the case for the quintessential truffle genus Tuber, which includes species with socio-economic importance and esteemed culinary attributes. Ecologically, Tuber spp. form obligate mycorrhizal symbioses with diverse species of plant hosts including pines, oaks, poplars, orchids, and commercially important trees such as hazelnut and pecan. Unfortunately, limited geographic sampling and inconclusive phylogenetic relationships have obscured our understanding of their origin, biogeography, and diversification. To address this problem, we present a global sampling of Tuberaceae based on DNA sequence data from four loci for phylogenetic inference and molecular dating. Our well-resolved Tuberaceae phylogeny shows high levels of regional and continental endemism. We also identify a previously unknown epigeous member of the Tuberaceae--the South American cup-fungus Nothojafnea thaxteri (E.K. Cash) Gamundí. Phylogenetic resolution was further improved through the inclusion of a previously unrecognized Southern hemisphere sister group of the Tuberaceae. This morphologically diverse assemblage of species includes truffle (e.g. Gymnohydnotrya spp.) and non-truffle forms that are endemic to Australia and South America. Southern hemisphere taxa appear to have diverged more recently than the Northern hemisphere lineages. Our analysis of the Tuberaceae suggests that Tuber evolved from an epigeous ancestor. Molecular dating estimates Tuberaceae divergence in the late Jurassic (~156 million years ago), with subsequent radiations in the Cretaceous and Paleogene. Intra-continental diversification, limited long-distance dispersal, and ecological adaptations help to explain patterns of truffle evolution and biodiversity.

Sequence analysis of the ribosomal RNA operon, particularly the internal transcribed spacer (ITS) region, provides a powerful tool for identification of mycorrhizal fungi. The sequence data deposited in the International Nucleotide Sequence Databases (INSD) are, however, unfiltered for quality and are often poorly annotated with metadata. To detect chimeric and low-quality sequences and assign the ectomycorrhizal fungi to phylogenetic lineages, fungal ITS sequences were downloaded from INSD, aligned within family-level groups, and examined through phylogenetic analyses and BLAST searches. By combining the fungal sequence database UNITE and the annotation and search tool PlutoF, we also added metadata from the literature to these accessions. Altogether 35,632 sequences belonged to mycorrhizal fungi or originated from ericoid and orchid mycorrhizal roots. Of these sequences, 677 were considered chimeric and 2,174 of low read quality. Information detailing country of collection, geographical coordinates, interacting taxon and isolation source were supplemented to cover 78.0%, 33.0%, 41.7% and 96.4% of the sequences, respectively. These annotated sequences are publicly available via UNITE (http://unite.ut.ee/) for downstream biogeographic, ecological and taxonomic analyses. In European Nucleotide Archive (ENA; http://www.ebi.ac.uk/ena/), the annotated sequences have a special link-out to UNITE. We intend to expand the data annotation to additional genes and all taxonomic groups and functional guilds of fungi.