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Fungi are among the most diverse and ecologically important kingdoms in life. However, the distributional ranges of fungi remain largely unknown as do the ecological mechanisms that shape their distributions 1,2 . To provide an integrated view of the spatial and seasonal dynamics of fungi, we implemented a globally distributed standardized aerial sampling of fungal spores 3 . The vast majority of operational taxonomic units were detected within only one climatic zone, and the spatiotemporal patterns of species richness and community composition were mostly explained by annual mean air temperature. Tropical regions hosted the highest fungal diversity except for lichenized, ericoid mycorrhizal and ectomycorrhizal fungi, which reached their peak diversity in temperate regions. The sensitivity in climatic responses was associated with phylogenetic relatedness, suggesting that large-scale distributions of some fungal groups are partially constrained by their ancestral niche. There was a strong phylogenetic signal in seasonal sensitivity, suggesting that some groups of fungi have retained their ancestral trait of sporulating for only a short period. Overall, our results show that the hyperdiverse kingdom of fungi follows globally highly predictable spatial and temporal dynamics, with seasonality in both species richness and community composition increasing with latitude. Our study reports patterns resembling those described for other major groups of organisms, thus making a major contribution to the long-standing debate on whether organisms with a microbial lifestyle follow the global biodiversity paradigms known for macroorganisms 4,5 ., (© 2024. The Author(s).)

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 m 3 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., (© 2024. The Author(s).)

Anaerobic fungi (AF), belonging to the phylum Neocallimastigomycota, are a pivotal component of the digestive tract microbiome of various herbivorous animals. In the last decade, the diversity of AF has rapidly expanded due to the exploration of numerous (novel) habitats. Studies aiming at understanding the role of AF require robust and reliable isolation and cultivation techniques, many of which remained unchanged for decades. Using amplicon sequencing, we compared three different media: medium with rumen fluid (RF), depleted rumen fluid (DRF), and no rumen fluid (NRF) to enrich the AF from the feces of yak, as a rumen control; and Przewalski's horse, llama, guanaco, and elephant, as a non-rumen habitats. The results revealed the selective enrichment of Piromyces and Neocallimastix from the feces of elephant and llama, respectively, in the RF medium. Similarly, the enrichment culture in DRF medium explicitly manifested Piromyces -related sequences from elephant feces. Five new clades (MM1-5) were defined from llama, guanaco, yak, and elephant feces that could as well be enriched from llama and elephant samples using non-conventional DRF and NRF media. This study presents evidence for the selective enrichment of certain genera in medium with RF and DRF from rumen as well as from non-rumen samples. NRF medium is suggested for the isolation of AF from non-rumen environments.

Anaerobic fungi from the herbivore digestive tract ( Neocallimastigomycetes ) are primary lignocellulose modifiers and hold promise for biotechnological applications. Their molecular detection is currently difficult due to the non-specificity of published primer pairs, which impairs evolutionary and ecological research with environmental samples. We developed and validated a Neocallimastigomycetes -specific PCR primer pair targeting the D2 region of the ribosomal large subunit suitable for screening, quantifying, and sequencing. We evaluated this primer pair in silico on sequences from all known genera, in vitro with pure cultures covering 16 of the 20 known genera, and on environmental samples with highly diverse microbiomes. The amplified region allowed phylogenetic differentiation of all known genera and most species. The amplicon is about 350 bp long, suitable for short-read high-throughput sequencing as well as qPCR assays. Sequencing of herbivore fecal samples verified the specificity of the primer pair and recovered highly diverse and so far unknown anaerobic gut fungal taxa. As the chosen barcoding region can be easily aligned and is taxonomically informative, the sequences can be used for classification and phylogenetic inferences. Several new Neocallimastigomycetes clades were obtained, some of which represent putative novel lineages such as a clade from feces of the rodent Dolichotis patagonum (mara).

Grass (Poaceae) pollen is the most important outdoor aeroallergen, 1 exacerbating a range of respiratory conditions, including allergic asthma and rhinitis ("hay fever"). 2-5 Understanding the relationships between respiratory diseases and airborne grass pollen with a view to improving forecasting has broad public health and socioeconomic relevance. It is estimated that there are over 400 million people with allergic rhinitis 6 and over 300 million with asthma, globally, 7 often comorbidly. 8 In the UK, allergic asthma has an annual cost of around US$ 2.8 billion (2017). 9 The relative contributions of the >11,000 (worldwide) grass species (C. Osborne et al., 2011, Botany Conference, abstract) to respiratory health have been unresolved, 10 as grass pollen cannot be readily discriminated using standard microscopy. 11 Instead, here we used novel environmental DNA (eDNA) sampling and qPCR 12-15 to measure the relative abundances of airborne pollen from common grass species during two grass pollen seasons (2016 and 2017) across the UK. We quantitatively demonstrate discrete spatiotemporal patterns in airborne grass pollen assemblages. Using a series of generalized additive models (GAMs), we explore the relationship between the incidences of airborne pollen and severe asthma exacerbations (sub-weekly) and prescribing rates of drugs for respiratory allergies (monthly). Our results indicate that a subset of grass species may have disproportionate influence on these population-scale respiratory health responses during peak grass pollen concentrations. The work demonstrates the need for sensitive and detailed biomonitoring of harmful aeroallergens in order to investigate and mitigate their impacts on human health., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Allergic rhinitis is an inflammation in the nose caused by overreaction of the immune system to allergens in the air. Managing allergic rhinitis symptoms is challenging and requires timely intervention. The following are major questions often posed by those with allergic rhinitis: How should I prepare for the forthcoming season? How will the season's severity develop over the years? No country yet provides clear guidance addressing these questions. We propose two previously unexplored approaches for forecasting the severity of the grass pollen season on the basis of statistical and mechanistic models. The results suggest annual severity is largely governed by preseasonal meteorological conditions. The mechanistic model suggests climate change will increase the season severity by up to 60%, in line with experimental chamber studies. These models can be used as forecasting tools for advising individuals with hay fever and health care professionals how to prepare for the grass pollen season., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC).)

Pterulaceae was formally proposed to group six coralloid and dimitic genera: Actiniceps (= Dimorphocystis) , Allantula , Deflexula , Parapterulicium , Pterula, and Pterulicium . Recent molecular studies have shown that some of the characters currently used in Pterulaceae do not distinguish the genera. Actiniceps and Parapterulicium have been removed, and a few other resupinate genera were added to the family. However, none of these studies intended to investigate the relationship between Pterulaceae genera. In this study, we generated 278 sequences from both newly collected and fungarium samples. Phylogenetic analyses supported with morphological data allowed a reclassification of Pterulaceae where we propose the introduction of Myrmecopterula gen. nov. and Radulomycetaceae fam. nov., the reintroduction of Phaeopterula , the synonymisation of Deflexula in Pterulicium, and 53 new combinations. Pterula is rendered polyphyletic requiring a reclassification; thus, it is split into Pterula , Myrmecopterula gen. nov., Pterulicium and Phaeopterula . Deflexula is recovered as paraphyletic alongside several Pterula species and Pterulicium, and is sunk into the latter genus. Phaeopterula is reintroduced to accommodate species with darker basidiomes. The neotropical Myrmecopterula gen. nov. forms a distinct clade adjacent to Pterula , and most members of this clade are associated with active or inactive attine ant nests. The resupinate genera Coronicium and Merulicium are recovered in a strongly supported clade close to Pterulicium . The other resupinate genera previously included in Pterulaceae , and which form basidiomes lacking cystidia and with monomitic hyphal structure ( Radulomyces , Radulotubus and Aphanobasidium ), are reclassified into Radulomycetaceae fam. nov. Allantula is still an enigmatic piece in this puzzle known only from the type specimen that requires molecular investigation. A key for the genera of Pterulaceae and Radulomycetaceae fam. nov. is also provided here., Competing Interests: Competing interestsThe authors declare that they have no competing interests., (© The Author(s) 2020.)

Stellera chamaejasme L. is the most problematic weed in China's grasslands. Its root exudates affect co-occurring plants and thus may also affect soil fungi. Soils (0-20 cm depth) on two adjacent sites, one invaded the other uninvaded, were compared for a range of physiochemical parameters and by DNA sequencing of fungal communities. At the invaded site, relationships between S. chamaejasme abundance, soil physiochemical factors, and fungal communities were further investigated to determine whether these relationships corroborated conclusions on the basis of site differences that could be translated into functional variation. Results showed that the invaded soils had lower N, P, organic matter, fungal alpha diversity, and relative abundance of arbuscular mycorrhizal fungi (AMF), but greater abundance of pathogenic fungi. Organic matter and P were the edaphic factors most strongly linked to site differences in total fungal communities. Within the invaded site, organic matter rather than S. chamaejasme cover was closely linked to total fungal composition. However, on this site, a number of fungal species that had various ecological functions and that differentiated the two sites were related to S. chamaejasme cover. This study indicates that lower fertility soils may be more susceptible to invasion by S. chamaejasme. Although the influence of S. chamaejasme on total fungal community composition was limited, there was evidence of effects on particular fungal species. Further research is needed to determine whether these effects influence S. chamaejasme invasiveness., Competing Interests: The authors declare no conflict of interest. The funders had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript, or in the decision to publish the results.

Survival and growth of the anaerobic gut fungi (AGF; Neocallimastigomycota) in the herbivorous gut necessitate the possession of multiple abilities absent in other fungal lineages. We hypothesized that horizontal gene transfer (HGT) was instrumental in forging the evolution of AGF into a phylogenetically distinct gut-dwelling fungal lineage. The patterns of HGT were evaluated in the transcriptomes of 27 AGF strains, 22 of which were isolated and sequenced in this study, and 4 AGF genomes broadly covering the breadth of AGF diversity. We identified 277 distinct incidents of HGT in AGF transcriptomes, with subsequent gene duplication resulting in an HGT frequency of 2 to 3.5% in AGF genomes. The majority of HGT events were AGF specific (91.7%) and wide (70.8%), indicating their occurrence at early stages of AGF evolution. The acquired genes allowed AGF to expand their substrate utilization range, provided new venues for electron disposal, augmented their biosynthetic capabilities, and facilitated their adaptation to anaerobiosis. The majority of donors were anaerobic fermentative bacteria prevalent in the herbivorous gut. This study strongly indicates that HGT indispensably forged the evolution of AGF as a distinct fungal phylum and provides a unique example of the role of HGT in shaping the evolution of a high-rank taxonomic eukaryotic lineage. IMPORTANCE The anaerobic gut fungi (AGF) represent a distinct basal phylum lineage (Neocallimastigomycota) commonly encountered in the rumen and alimentary tracts of herbivores. Survival and growth of anaerobic gut fungi in these anaerobic, eutrophic, and prokaryote-dominated habitats necessitates the acquisition of several traits absent in other fungal lineages. We assess here the role of horizontal gene transfer as a relatively fast mechanism for trait acquisition by the Neocallimastigomycota postsequestration in the herbivorous gut. Analysis of 27 transcriptomes that represent the broad diversity of Neocallimastigomycota identified 277 distinct HGT events, with subsequent gene duplication resulting in an HGT frequency of 2 to 3.5% in AGF genomes. These HGT events have allowed AGF to survive in the herbivorous gut by expanding their substrate utilization range, augmenting their biosynthetic pathway, providing new routes for electron disposal by expanding fermentative capacities, and facilitating their adaptation to anaerobiosis. HGT in the AGF is also shown to be mainly a cross-kingdom affair, with the majority of donors belonging to the bacteria. This study represents a unique example of the role of HGT in shaping the evolution of a high-rank taxonomic eukaryotic lineage., (Copyright © 2019 American Society for Microbiology.)

Increasing feed efficiency is a key target in ruminant science which requires a better understanding of rumen microbiota. This study investigated the effect of a shift from a non-grazing to a grazing diet on the rumen bacterial, methanogenic archaea, fungal, and protozoal communities. A systems biology approach based on a description of the community structure, core microbiota, network analysis, and taxon abundance linked to the rumen fermentation was used to explore the benefits of increasing depth of the community analysis. A total of 24 sheep were fed ryegrass hay supplemented with concentrate (CON) and subsequently ryegrass pasture (PAS) following a straight through experimental design. Results showed that concentrate supplementation in CON-fed animals (mainly starch) promoted a simplified rumen microbiota in terms of network density and bacterial, methanogen and fungal species richness which favored the proliferation of amylolytic microbes and VFA production (+48%), but led to a lower (ca. 4-fold) ammonia concentration making the N availability a limiting factor certain microbes. The adaptation process from the CON to the PAS diet consisted on an increase in the microbial concentration (biomass of bacteria, methanogens, and protozoa), diversity (+221, +3, and +21 OTUs for bacteria, methanogens, and fungi, respectively), microbial network complexity (+18 nodes and +86 edges) and in the abundance of key microbes involved in cellulolysis ( Ruminococcus, Butyrivibrio , and Orpinomyces ), proteolysis ( Prevotella and Entodiniinae), lactate production ( Streptococcus and Selenomonas ), as well as methylotrophic archaea (Methanomassiliicoccaceae). This microbial adaptation indicated that pasture degradation is a complex process which requires a diverse consortium of microbes working together. The correlations between the abundance of microbial taxa and rumen fermentation parameters were not consistent across diets suggesting a metabolic plasticity which allowed microbes to adapt to different substrates and to shift their fermentation products. The core microbiota was composed of 34, 9, and 13 genera for bacteria, methanogens, and fungi, respectively, which were shared by all sheep, independent of diet. This systems biology approach adds a new dimension to our understanding of the rumen microbial interactions and may provide new clues to describe the mode of action of future nutritional interventions.

An extended incubation strategy to culture slow growing members of anaerobic fungi resulted in the isolation of a novel anaerobic fungus from the rumen of a goat after 15 days. The novel genus, represented by type strain G1SC, showed filamentous monocentric thallus development and produced uniflagellate zoospores, hence, showing morphological similarity to the genera Piromyces , Buwchfawromyces , Oontomyces and Pecoramyces . However, strain G1SC showed genetic similarity to the genus Anaeromyces , which, though produces uniflagellate zoospore, also exhibits polycentric thallus development. Moreover, unlike Anaeromyces , strain G1SC did not show hyphal constrictions, instead produced a branched, determinate and anucleate rhizoidal system. This fungus also displayed extensive sporangial variations, both exogenous and endogenous type of development, short and long sporangiophores and produced septate sporangia. G1SC utilised various complex and simple substrates, including rice straw and wheat straw and produced H 2 , CO 2 , formate, acetate, lactate, succinate and ethanol. Phylogenetic analysis, using internal transcribed spacer 1 (ITS1) and D1/D2 domain of large-subunit (LSU) rRNA locus, clearly showed a separate lineage for this strain, near Anaeromyces . The ITS1 based geographical distribution studies indicated detection of environmental sequences similar (93-96%) to this strain from cattle faeces. Based on morphological and molecular characterisation results of strain G1SC, we propose a novel anaerobic fungus Liebetanzomycespolymorphus gen. et sp. nov. , in the phylum Neocallimastigomycota .

Behavioural fever is a common response to immune challenge in ectotherms and confers survival benefits. However, costs accrue rapidly as body temperature rises. Thus, the magnitude of adaptive fever responses might reflect the balance of costs and benefits. We investigated behavioural fever in desert locusts, Schistocerca gregaria, infected with the entomopathogenic fungus Metarhizium acridum. We first tracked the time course of behavioural fever in infected locusts, demonstrating that body temperatures rose on the day following inoculation (day 1), and reached peak intensity on the day after that (day 2). Subsequently, the magnitude of fever responses varied during a day, and locusts tended to exhibit high-intensity fever responses in the mornings when basking was first possible. We speculate that this may have resulted from increased fungal load caused by unimpeded growth overnight when locusts could not fever. We next inoculated locusts with different M. acridum doses ranging from 0 to ca. 75,000 conidia. The magnitude of their behavioural fever responses on day 2 post-inoculation was positively related to fungal dose. Thus, we demonstrate dose-dependency in the behavioural fever responses of desert locusts and suggest that this may reflect the adaptive deployment of behavioural fever to minimize costs relative to benefits.

The genus Parapterulicium was first introduced to accommodate two Brazilian species of coralloid fungi with affinities to Pterulaceae (Agaricales). Despite the coralloid habit and the presence of skeletal hyphae, other features, notably the presence of gloeocystidia, dichophyses and papillate hyphal ends, differentiate this genus from Pterulaceae sensu stricto. Fieldwork in Brazil resulted in the rediscovery of two coralloid fungi identifiable as Parapterulicium , the first verified collections of this genus since Corner's original work in the 1950s. Molecular phylogenetic analyses of nrITS and nrLSU sequences from these modern specimens revealed affinities with the /peniophorales clade in the Russulales, rather than Pterulaceae. The presence of distinctive hyphal elements, homologous to the defining features of /peniophorales, is consistent with the phylogenetic evidence and thus clearly distinguished Parapterulicium and its type species P.subarbusculum from Pterulaceae, placing this genus within /peniophorales. Parapterulicium was also found to be polyphyletic so Baltazaria gen. nov. is proposed to accommodate P.octopodites , Scytinostromagalactinum , S.neogalactinum and S.eurasiaticogalactinum also within /peniophorales.

Anaerobic fungi (phylum Neocallimastigomycota) are common inhabitants of the digestive tract of mammalian herbivores, and in the rumen, can account for up to 20% of the microbial biomass. Anaerobic fungi play a primary role in the degradation of lignocellulosic plant material. They also have a syntrophic interaction with methanogenic archaea, which increases their fiber degradation activity. To date, nine anaerobic fungal genera have been described, with further novel taxonomic groupings known to exist based on culture-independent molecular surveys. However, the true extent of their diversity may be even more extensively underestimated as anaerobic fungi continue being discovered in yet unexplored gut and non-gut environments. Additionally many studies are now known to have used primers that provide incomplete coverage of the Neocallimastigomycota. For ecological studies the internal transcribed spacer 1 region (ITS1) has been the taxonomic marker of choice, but due to various limitations the large subunit rRNA (LSU) is now being increasingly used. How the continued expansion of our knowledge regarding anaerobic fungal diversity will impact on our understanding of their biology and ecological role remains unclear; particularly as it is becoming apparent that anaerobic fungi display niche differentiation. As a consequence, there is a need to move beyond the broad generalization of anaerobic fungi as fiber-degraders, and explore the fundamental differences that underpin their ability to exist in distinct ecological niches. Application of genomics, transcriptomics, proteomics and metabolomics to their study in pure/mixed cultures and environmental samples will be invaluable in this process. To date the genomes and transcriptomes of several characterized anaerobic fungal isolates have been successfully generated. In contrast, the application of proteomics and metabolomics to anaerobic fungal analysis is still in its infancy. A central problem for all analyses, however, is the limited functional annotation of anaerobic fungal sequence data. There is therefore an urgent need to expand information held within publicly available reference databases. Once this challenge is overcome, along with improved sample collection and extraction, the application of these techniques will be key in furthering our understanding of the ecological role and impact of anaerobic fungi in the wide range of environments they inhabit.

Same-sex sexual behaviour occurs across diverse animal taxa, but adaptive explanations can be difficult to determine. Here we investigate male-male mounting (MMM) behaviour in female-deprived desert locust males infected with the entomopathogenic fungus Metarhizium acridum. Over a four-week period, infected locusts performed more MMM behaviours than healthy controls. Among infected locusts, the probability of MMM, and the duration of time spent MMM, significantly increased with the mounting locust's proximity to death. In experimental trials, infected locusts were also significantly more likely than controls to attempt to mount healthy males. Therefore, we demonstrate that MMM is more frequent among infected than healthy male locusts, and propose that this may be explained by terminal reproductive effort and a lowered mate acceptance threshold in infected males. However, during experimental trials mounting attempts were more likely to be successful if the mounted locusts were experimentally manipulated to have a reduced capacity to escape. Thus, reduced escape capability resulting from infection may also contribute to the higher frequency of MMM among infected male locusts. Our data demonstrate that pathogen infection can affect same-sex sexual behaviour, and suggest that the impact of such behaviours on host and pathogen fitness will be a novel focus for future research.

Hitherto fungi have rarely been considered in conservation biology, but this is changing as the field moves from addressing single species issues to an integrative ecosystem-based approach. The current emphasis on biodiversity as a provider of ecosystem services throws the spotlight on the vast diversity of fungi, their crucial roles in terrestrial ecosystems, and the benefits of considering fungi in concert with animals and plants. We reviewed the role of fungi in ecosystems and composed an overview of the current state of conservation of fungi. There are 5 areas in which fungi can be readily integrated into conservation: as providers of habitats and processes important for other organisms; as indicators of desired or undesired trends in ecosystem functioning; as indicators of habitats of conservation value; as providers of powerful links between human societies and the natural world because of their value as food, medicine, and biotechnological tools; and as sources of novel tools and approaches for conservation of megadiverse organism groups. We hope conservation professionals will value the potential of fungi, engage mycologists in their work, and appreciate the crucial role of fungi in nature., (© 2014 Society for Conservation Biology.)

Anaerobic fungi (phylum Neocallimastigomycota) inhabit the gastrointestinal tract of mammalian herbivores, where they play an important role in the degradation of plant material. The Neocallimastigomycota represent the earliest diverging lineage of the zoosporic fungi; however, understanding of the relationships of the different taxa (both genera and species) within this phylum is in need of revision. Issues exist with the current approaches used for their identification and classification, and recent evidence suggests the presence of several novel taxa (potential candidate genera) that remain to be characterised. The life cycle and role of anaerobic fungi has been well characterised in the rumen, but not elsewhere in the ruminant alimentary tract. Greater understanding of the 'resistant' phase(s) of their life cycle is needed, as is study of their role and significance in other herbivores. Biotechnological application of anaerobic fungi, and their highly active cellulolytic and hemi-cellulolytic enzymes, has been a rapidly increasing area of research and development in the last decade. The move towards understanding of anaerobic fungi using -omics based (genomic, transcriptomic and proteomic) approaches is starting to yield valuable insights into the unique cellular processes, evolutionary history, metabolic capabilities and adaptations that exist within the Neocallimastigomycota., (© 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.)

Cryoconite holes are known as foci of microbial diversity and activity on polar glacier surfaces, but are virtually unexplored microbial habitats in alpine regions. In addition, whether cryoconite community structure reflects ecosystem functionality is poorly understood. Terminal restriction fragment length polymorphism and Fourier transform infrared metabolite fingerprinting of cryoconite from glaciers in Austria, Greenland and Svalbard demonstrated cryoconite bacterial communities are closely correlated with cognate metabolite fingerprints. The influence of bacterial-associated fatty acids and polysaccharides was inferred, underlining the importance of bacterial community structure in the properties of cryoconite. Thus, combined application of T-RFLP and FT-IR metabolite fingerprinting promises high throughput, and hence, rapid assessment of community structure-function relationships. Pyrosequencing revealed Proteobacteria were particularly abundant, with Cyanobacteria likely acting as ecosystem engineers in both alpine and Arctic cryoconite communities. However, despite these generalities, significant differences in bacterial community structures, compositions and metabolomes are found between alpine and Arctic cryoconite habitats, reflecting the impact of local and regional conditions on the challenges of thriving in glacial ecosystems., (© 2014 Federation of European Microbiological Societies. Published by John Wiley & Sons Ltd. All rights reserved.)

DNA phylogenetic comparisons have shown that morphology-based species recognition often underestimates fungal diversity. Therefore, the need for accurate DNA sequence data, tied to both correct taxonomic names and clearly annotated specimen data, has never been greater. Furthermore, the growing number of molecular ecology and microbiome projects using high-throughput sequencing require fast and effective methods for en masse species assignments. In this article, we focus on selecting and re-annotating a set of marker reference sequences that represent each currently accepted order of Fungi. The particular focus is on sequences from the internal transcribed spacer region in the nuclear ribosomal cistron, derived from type specimens and/or ex-type cultures. Re-annotated and verified sequences were deposited in a curated public database at the National Center for Biotechnology Information (NCBI), namely the RefSeq Targeted Loci (RTL) database, and will be visible during routine sequence similarity searches with NR_prefixed accession numbers. A set of standards and protocols is proposed to improve the data quality of new sequences, and we suggest how type and other reference sequences can be used to improve identification of Fungi. Database URL: http://www.ncbi.nlm.nih.gov/bioproject/PRJNA177353., (Published by Oxford University Press 2013. This work is written by US Government employees and is in the public domain in the US.)

The diversity of highly active bacterial communities in cryoconite holes on three Arctic glaciers in Svalbard was investigated using terminal restriction fragment length polymorphism (T-RFLP) of the 16S rRNA locus. Construction and sequencing of clone libraries allowed several members of these communities to be identified, with Proteobacteria being the dominant one, followed by Cyanobacteria and Bacteroidetes. T-RFLP data revealed significantly different communities in holes on the (cold) valley glacier Austre Brøggerbreen relative to two adjacent (polythermal) valley glaciers, Midtre Lovénbreen and Vestre Brøggerbreen. These population compositions correlate with differences in organic matter content, temperature and the metabolic activity of microbial communities concerned. No within-glacier spatial patterns were observed in the communities identified over the 2-year period and with the 1 km-spaced sampling. We infer that surface hydrology is an important factor in the development of cryoconite bacterial communities.

This contribution is based on the six oral presentations given at the Special Interest Group session on anaerobic fungi held during IMC9. These fungi, recently elevated to the status of a separate phylum (Neocallimastigomycota), distinct from the chytrid fungi, possess several unique traits that make their study both fascinating yet challenging to mycologists. There are several genome sequencing programs underway in the US but these are hampered by the highly AT-rich genomes. Next-generation sequencing has also allowed more detailed investigation of the ecology and diversity of these fungi, and it is apparent that several new taxa beyond the six genera already named exist within the digestive tracts of mammalian herbivores, with others potentially inhabiting other anaerobic niches. By increased collaboration between the various labs studying these fungi, it is hoped to develop a stable taxonomic backbone for these fungi and to facilitate exchange of both cultures and genetic data.

Unlike organic pollutants, heavy metals cannot be degraded and can constitute a persistent environmental hazard. Here, we investigated the success of different remediation strategies in promoting microbial diversity and function with depth in an acidic soil heavily contaminated with Cu, Pb and Zn. Remediation involved the incorporation of either a high- or a low-quality compost or inorganic fertilizer into the topsoil and monitoring of microbial activity and diversity with soil depth over a 4-month period. While changes in topsoil microbial activity were expected, the possible effects on the subsurface microbial community due to the downward movement of metals, nutrients and/or soluble organic matter have not been examined previously. The results showed that both compost additions, especially the low-quality compost, resulted in significantly increased bacterial and fungal diversity (as assessed by terminal restriction fragment length polymorphism) and activity compared with the inorganic and control treatments in the topsoil. Although phospholipid fatty acid profiling indicated that compost addition had promoted enhanced microbial diversity in the subsoil, no concomitant increase in subsoil microbial activity was observed, suggesting that amelioration of the heavy metals remained localized in the topsoil. We conclude that although composts can successfully immobilize heavy metals and promote ecosystem diversity/function, surface incorporation had little remedial effect below the surface layer over the course of our short-term trial.

Soils contaminated with heavy metals can pose a major risk to freshwaters and food chains. In this study, the success of organic and inorganic intervention strategies to alleviate toxicity in a highly acidic soil heavily contaminated with As, Cu, Pb, and Zn was evaluated over 112 d in a mesocosm trial. Amelioration of metal toxicity was assessed by measuring changes in soil solution chemistry, metal leaching, plant growth, and foliar metal accumulation. Either green waste- or MSW-derived composts increased plant yield and rooting depth, reduced plant metal uptake, and raised the pH and nutrient status of the soil. We conclude that composts are well suited for promoting the re-vegetation of contaminated sites; however, care must be taken to ensure that very short-term leaching pulses of heavy metals induced by compost amendment are not of sufficient magnitude to cause contamination of the wider environment.

We present a 6-gene, 420-species maximum-likelihood phylogeny of Ascomycota, the largest phylum of Fungi. This analysis is the most taxonomically complete to date with species sampled from all 15 currently circumscribed classes. A number of superclass-level nodes that have previously evaded resolution and were unnamed in classifications of the Fungi are resolved for the first time. Based on the 6-gene phylogeny we conducted a phylogenetic informativeness analysis of all 6 genes and a series of ancestral character state reconstructions that focused on morphology of sporocarps, ascus dehiscence, and evolution of nutritional modes and ecologies. A gene-by-gene assessment of phylogenetic informativeness yielded higher levels of informativeness for protein genes (RPB1, RPB2, and TEF1) as compared with the ribosomal genes, which have been the standard bearer in fungal systematics. Our reconstruction of sporocarp characters is consistent with 2 origins for multicellular sexual reproductive structures in Ascomycota, once in the common ancestor of Pezizomycotina and once in the common ancestor of Neolectomycetes. This first report of dual origins of ascomycete sporocarps highlights the complicated nature of assessing homology of morphological traits across Fungi. Furthermore, ancestral reconstruction supports an open sporocarp with an exposed hymenium (apothecium) as the primitive morphology for Pezizomycotina with multiple derivations of the partially (perithecia) or completely enclosed (cleistothecia) sporocarps. Ascus dehiscence is most informative at the class level within Pezizomycotina with most superclass nodes reconstructed equivocally. Character-state reconstructions support a terrestrial, saprobic ecology as ancestral. In contrast to previous studies, these analyses support multiple origins of lichenization events with the loss of lichenization as less frequent and limited to terminal, closely related species.

Anaerobic fungi (Neocallimastigales) are active degraders of fibrous plant material in the rumen. However, only limited information is available relating to how quickly they colonize ingested feed particles. The aim of this study was to determine the dynamics of initial colonization of forage by anaerobic fungi in the rumen and the impact of different postsampling wash procedures used to remove loosely associated microorganisms. Neocallimastigales-specific molecular techniques were optimized to ensure maximal coverage before application to assess the population size (quantitative PCR) and composition (automated ribosomal intergenic spacer analysis) of the colonizing anaerobic fungi. Colonization of perennial ryegrass (PRG) was evident within 5 min, with no consistent effect of time or wash procedure on fungal population composition. Wash procedure had no effect on population size unlike time, which had a significant effect. Colonizing fungal population size continued to increase over the incubation period after an initial lag of c. 4 min. This dynamic differs from that reported previously for rumen bacteria, where substantial colonization of PRG occurred within 5 min. The observed delay in colonization of plant material by anaerobic fungi is suggested to be primarily mediated by the time taken for fungal zoospores to locate, attach and encyst on plant material.

Moniliophthora perniciosa (=Crinipellis perniciosa) causes one of the three main fungal diseases of Theobroma cacao (cacao), the source of chocolate. This pathogen causes Witches' broom disease (WBD) and has brought about severe economic losses in all of the cacao-growing regions to which it has spread with yield reductions that range from 50 to 90%. Cacao production in South America reflects the severity of this pathogen, as the yields in most of the infected regions have not returned to pre-outbreak levels, even with the introduction of resistant varieties. In this review we give a brief historical account and summarize the current state of knowledge focusing on developments in the areas of systematics, fungal physiology, biochemistry, genomics and gene expression in an attempt to highlight this disease. Moniliophthora perniciosa is a hemibiotrophic fungus with two distinct growth phases. The ability to culture a biotrophic-like phase in vitro along with new findings derived from the nearly complete genome and expression studies clearly show that these different fungal growth phases function under distinct metabolic parameters. These new findings have greatly improved our understanding of this fungal/host interaction and we may be at the crossroads of understanding how hemibiotrophic fungal plant pathogens cause disease in other crops., Historical Summary of Wbd: The first WDB symptoms appear to have been described in the diaries of Alexandre Rodrigues Ferreira (described as lagartão; meaning big lizard) from his observations of cacao trees in 1785 and 1787 in Amazonia, which is consistent with the generally accepted idea that M. perniciosa, like its main host T. cacao, evolved in this region. The disease subsequently arrived in Surinam in 1895. WBD moved rapidly, spreading to Guyana in 1906, Ecuador in 1918, Trinidad in 1928, Colombia in 1929 and Grenada in 1948. In each case, cacao production was catastrophically affected with yield reductions of 50-90%. After the arrival of M. perniciosa in Bahia in 1989, Brazil went from being the world's 3rd largest producer of cacao (347 000 tonnes in 1988-1990; c. 15% of the total world production at that time) to a net importer (141 000 tonnes in 1998-2000). Fortunately for chocolate lovers, other regions of the world such as West Africa and South East Asia have not yet been affected by this disease and have expanded production to meet growing world demand (predicted to reach 3 700 000 tonnes by 2010)., Classification: Moniliophthora perniciosa (Stahel) Aime & Phillips-Mora: super-kingdom Eukaryota; kingdom Fungi; phylum Basidiomycota; subphylum Agaricomycotina; class Agaricomycetes; subclass Agaricomycetidae; order Agaricales; family Marasmiaceae; genus Moniliophthora., Useful Websites: http://www.lge.ibi.unicamp.br/vassoura/, http://nt.ars-grin.gov/taxadescriptions/keys/TrichodermaIndex.cfm, http://www.worldcocoafoundation.org/info-center/research-updates.asp, http://www.ars.usda.gov/ba/psi/spcl.

Background: A thoracic artificial lung, the MC3 Biolung, is being developed as a bridge to lung transplantation or as a treatment for acute respiratory insufficiency., Methods: The thoracic artificial lung was tested in 10 sheep with the goal of 7 days of respiratory support. The sheep were recovered from surgery and monitored awake for 7 days. Hemodynamics, blood gases, blood cell counts, and organ function were recorded, and after 7 days, all sheep were euthanized and necropsied., Results: Seven sheep survived the full duration. Cardiac output and mean arterial blood pressure were unchanged, averaging 4.7 +/- 0.8 L/min and 98 +/- 10 mm Hg, respectively. Arterial oxygen tension and device oxygen transfer rate were also unchanged, averaging 110 +/- 26 mm Hg and 97.7 +/- 35 mL/min, respectively. Arterial carbon dioxide tension was within normal ranges during the entire experiment, averaging 37.4 +/- 3.8 mm Hg. Artificial lung blood flow decreased from 51% +/- 14% of cardiac output on day 1 to 30% +/- 16% by day 7 because of changes in natural and artificial lung resistance. White blood cell counts were significantly elevated on days 5 and 7, and lastly, kidney and liver function remained normal, although signs of kidney infarction or hemorrhage were noted., Conclusions: The thoracic artificial lung is suitable for 7-day attachment, but improvements in blood biocompatibility are warranted.

Chytridiomycota (chytrids) is the only phylum of true Fungi that reproduces with motile spores (zoospores). Chytrids currently are classified into five orders based on habitat, zoospore characters and life cycles. In this paper we estimate the phylogeny of the chytrids with DNA sequences from the ribosomal RNA operon (18S+5.8S+28S subunits). To our surprise the morphologically reduced parasites Olpidium and Rozella comprise two entirely new, and separate, lineages on the fungal tree. Olpidium brassicae groups among the Zygomycota, and Rozella spp. are the earliest branch to diverge in the fungal kingdom. The phylogeny also suggests that Chytridiomycota is not monophyletic and there are four major lineages of chytrids: Rozella spp., Olpidium brassicae, the Blastocladiales and a "core chytrid clade" containing the remaining orders and families and the majority of flagellated fungi. Within the core chytrid group 11 subclades can be identified, each of which correlates well with zoospore ultrastructure or morphology. We provide a synopsis of each clade and its morphological circumscription. The Blastocladiales appears to be the sister taxon of most nonflagellated fungi. Based on molecular phylogenetic and ultrastructural characters this order is elevated to a phylum, the Blastocladiomycota.

The ancestors of fungi are believed to be simple aquatic forms with flagellated spores, similar to members of the extant phylum Chytridiomycota (chytrids). Current classifications assume that chytrids form an early-diverging clade within the kingdom Fungi and imply a single loss of the spore flagellum, leading to the diversification of terrestrial fungi. Here we develop phylogenetic hypotheses for Fungi using data from six gene regions and nearly 200 species. Our results indicate that there may have been at least four independent losses of the flagellum in the kingdom Fungi. These losses of swimming spores coincided with the evolution of new mechanisms of spore dispersal, such as aerial dispersal in mycelial groups and polar tube eversion in the microsporidia (unicellular forms that lack mitochondria). The enigmatic microsporidia seem to be derived from an endoparasitic chytrid ancestor similar to Rozella allomycis, on the earliest diverging branch of the fungal phylogenetic tree.

Silage quality is typically assessed by the measurement of several individual parameters, including pH, lactic acid, acetic acid, bacterial numbers, and protein content. The objective of this study was to use a holistic metabolic fingerprinting approach, combining a high-throughput microtiter plate-based fermentation system with Fourier transform infrared (FT-IR) spectroscopy, to obtain a snapshot of the sample metabolome (typically low-molecular-weight compounds) at a given time. The aim was to study the dynamics of red clover or grass silage fermentations in response to various inoculants incorporating lactic acid bacteria (LAB). The hyperspectral multivariate datasets generated by FT-IR spectroscopy are difficult to interpret visually, so chemometrics methods were used to deconvolute the data. Two-phase principal component-discriminant function analysis allowed discrimination between herbage types and different LAB inoculants and modeling of fermentation dynamics over time. Further analysis of FT-IR spectra by the use of genetic algorithms to identify the underlying biochemical differences between treatments revealed that the amide I and amide II regions (wavenumbers of 1,550 to 1,750 cm(-1)) of the spectra were most frequently selected (reflecting changes in proteins and free amino acids) in comparisons between control and inoculant-treated fermentations. This corresponds to the known importance of rapid fermentation for the efficient conservation of forage proteins.

The ability of two different flow cytometers, the Microcyte (Optoflow) and the PAS-III (Partec), to differentiate sporangia of the late-blight pathogen Phytophthora infestans from other potential airborne particles was compared. With the PAS-III, light scatter and intrinsic fluorescence parameters could be used to differentiate sporangia from conidia of Alternaria or Botrytis spp., rust urediniospores, and pollen of grasses and plantain. Differentiation between P. infestans sporangia and powdery mildew conidia was not possible by these two methods but, when combined with analytical rules evolved by genetic programming methods, could be achieved after staining with the fluorescent brightener Calcofluor white M2R. The potential application of these techniques to the prediction of late-blight epiphytotics in the field is discussed.

The enormous variety of substances which may be added to forage in order to manipulate and improve the ensilage process presents an empirical, combinatorial optimization problem of great complexity. To investigate the utility of genetic algorithms for designing effective silage additive combinations, a series of small-scale proof of principle silage experiments were performed with fresh ryegrass. Having established that significant biochemical changes occur over an ensilage period as short as 2 days, we performed a series of experiments in which we used 50 silage additive combinations (prepared by using eight bacterial and other additives, each of which was added at six different levels, including zero [i.e. , no additive]). The decrease in pH, the increase in lactate concentration, and the free amino acid concentration were measured after 2 days and used to calculate a "fitness" value that indicated the quality of the silage (compared to a control silage made without additives). This analysis also included a "cost" element to account for different total additive levels. In the initial experiment additive levels were selected randomly, but subsequently a genetic algorithm program was used to suggest new additive combinations based on the fitness values determined in the preceding experiments. The result was very efficient selection for silages in which large decreases in pH and high levels of lactate occurred along with low levels of free amino acids. During the series of five experiments, each of which comprised 50 treatments, there was a steady increase in the amount of lactate that accumulated; the best treatment combination was that used in the last experiment, which produced 4.6 times more lactate than the untreated silage. The additive combinations that were found to yield the highest fitness values in the final (fifth) experiment were assessed to determine a range of biochemical and microbiological quality parameters during full-term silage fermentation. We found that these combinations compared favorably both with uninoculated silage and with a commercial silage additive. The evolutionary computing methods described here are a convenient and efficient approach for designing silage additives.

Four pairs of primers were designed for PCR amplification of known polymorphic regions of the mitochondrial genome of Phytophthora infestans. Digestion of the amplified products with restriction enzymes allows identification of previously identified haplotypes. Product P2 cut with MspI uniquely identifies haplotypes Ib and IIa, while types Ia and IIb are differentiated by digestion of product P4 with EcoRI. Digestion of products P1 and P3 gave results similar to that with digestion of P4, but amplification of these products was less robust. Thus, all four common haplotypes are identified by amplifying and digesting products P2 and P4. Identification of haplotypes was also possible from DNA extracted directly from small, late-blight lesions on both tomato and potato leaves, making isolation of the fungus unnecessary. A rapid and efficient method of monitoring changes in the pathogen population is facilitated. These PCR primers were also useful for differentiating other Phytophthora species.

To use single-pass cDNA sequencing to characterize low-frequency cDNA clones from a region of the brain that includes the primary site of neurodegeneration in human Parkinson disease, we have developed a prescreening procedure using single brain region first-strand cDNA probes. Selection of cDNA clones giving low hybridization signals allowed the elimination of clones resulting from abundant messages and enrichment for clones corresponding to low-copy messages. Comparative sequencing of standard and prescreened cDNA libraries (191 and 124 clones, respectively) showed that this procedure raised the frequency of novel sequences encountered from 54 to 81%. The increased proportion of novel ESTs justifies the labor of prescreening. Automation of this procedure will accelerate the molecular description of genes expressed in any brain region, or any tissue, and represents a way to maximize access to cDNA sequences for human and mouse genome characterization. In total, the comparative sequencing experiments generated 207 new mouse and 11 new rat brain ESTs.

A detailed investigation of the Liana biotype (L-biotype) of Crinipellis perniciosa (Stahel) Sing, was conducted in Ecuador. Basidiocarps of the L-biotype were consistently found in association with stems or debris of the liana Arrabidaea verrucosa (Standl) A. Gentry, although no symptoms associated with witches'broom disease, were observed. Debris colonized by the L-biotype were often connected via pseudosclerotial pads to living sterns of A. verrucosa. Pairings between 87 isolates obtained from basidiocarp stipe tissues or liana bark cores from three sites permitted 37 somatic compatibility groupings (SCGs) to be defined. In all cases these were found to be very restricted in geographical distribution, a situation which contrasts markedly with the distribution of SCGs in the pathogenic and non-outercrossing eocoa biotype. Mating pairings between single basidiospore isolates confirmed chat the outercrossing mechanism of the L-biotype is muitialletic and that no significant biological barriers to gene flow exist between coastal and Amazonian populations. Several lines of evidence suggested that mycelia belonging to the same SCG (= genet) were isogenic. Correlations between field observations and the distributions of both SCGs and mating type factors art- discussed in relation to patterns of dispersal, establishment and spread of this fungus.

Mortality in Anglesey from cancer (all sites), cancer of trachea, bronchus, and lung, cancer of stomach, and cancer of all other sites has been compared with mortality in England and Wales at two periods; 1949-53 (before fluoridation) and 1979-83 (after 20 years and more of fluoridation). No evidence has been found to support the hypothesis that water fluoridation affects cancer mortality.

Early neonatal mortality in England and Wales in the second quarter of 1970 after a major influenza epidemic was slightly but significantly higher than in the corresponding quarter of the previous year. An increase was also noted in the first quarter of 1970. Analysis of infant mortality and an index of influenza prevalence over the past quarter-century indicates that similar increases occurred in relation to four of the other five major influenza epidemics during the period, the exception being the "Asian 'flu" epidemic of the autumn of 1957. It is suggested that the increased mortality in 1970 was the consequence of an increase in the prematurity rate, but we have no evidence to indicate whether the effect is specifically due to the virus or is nonspecific in nature.