Eduardo Nouhra

Global diversity and geography of soil fungi

Introduction The kingdom Fungi is one of the most diverse groups of organisms on Earth, and they are integral ecosystem agents that govern soil carbon cycling, plant nutrition, and pathology. Fungi are widely distributed in all terrestrial ecosystems, but the distribution of species, phyla, and functional groups has been poorly documented. On the basis of 365 global soil samples from natural ecosystems, we determined the main drivers and biogeographic patterns of fungal diversity and community composition. Direct and indirect effects of climatic and edaphic variables on plant and fungal richness. Line thickness corresponds to the relative strength of the relationships between the variables that affect species richness. Dashed lines indicate negative relationships. MAP, mean annual precipitation; Fire, time since last fire; Dist. equator, distance from the equator; Ca, soil calcium concentration; P, soil phosphorus concentration; pH, soil pH. Rationale We identified soil-inhabiting fungi using 454 Life Sciences (Branford, CN) pyrosequencing and through comparison against taxonomically and functionally annotated sequence databases. Multiple regression models were used to disentangle the roles of climatic, spatial, edaphic, and floristic parameters on fungal diversity and community composition. Structural equation models were used to determine the direct and indirect effects of climate on fungal diversity, soil chemistry, and vegetation. We also examined whether fungal biogeographic patterns matched paradigms derived from plants and animals—namely, that species’ latitudinal ranges increase toward the poles (Rapoport’s rule) and diversity increases toward the equator. Last, we sought group-specific global biogeographic links among major biogeographic regions and biomes using a network approach and area-based clustering. Results Metabarcoding analysis of global soils revealed fungal richness estimates approaching the number of species recorded to date. Distance from equator and mean annual precipitation had the strongest effects on richness of fungi, including most fungal taxonomic and functional groups. Diversity of most fungal groups peaked in tropical ecosystems, but ectomycorrhizal fungi and several fungal classes were most diverse in temperate or boreal ecosystems, and many fungal groups exhibited distinct preferences for specific edaphic conditions (such as pH, calcium, or phosphorus). Consistent with Rapoport’s rule, the geographic range of fungal taxa increased toward the poles. Fungal endemicity was particularly strong in tropical regions, but multiple fungal taxa had cosmopolitan distribution. Conclusions Climatic factors, followed by edaphic and spatial patterning, are the best predictors of soil fungal richness and community composition at the global scale. Richness of all fungi and functional groups is causally unrelated to plant diversity, with the exception of ectomycorrhizal root symbionts, suggesting that plant-soil feedbacks do not influence the diversity of soil fungi at the global scale. The plant-to-fungi richness ratio declined exponentially toward the poles, indicating that current predictions—assuming globally constant ratios—overestimate fungal richness by 1.5- to 2.5-fold. Fungi follow similar biogeographic patterns as plants and animals, with the exception of several major taxonomic and functional groups that run counter to overall patterns. Strong biogeographic links among distant continents reflect relatively efficient long-distance dispersal compared with macro-organisms. Fungi play major roles in ecosystem processes, but the determinants of fungal diversity and biogeographic patterns remain poorly understood. Using DNA metabarcoding data from hundreds of globally distributed soil samples, we demonstrate that fungal richness is decoupled from plant diversity. The plant-to-fungus richness ratio declines exponentially toward the poles. Climatic factors, followed by edaphic and spatial variables, constitute the best predictors of fungal richness and community composition at the global scale. Fungi show similar latitudinal diversity gradients to other organisms, with several notable exceptions. These findings advance our understanding of global fungal diversity patterns and permit integration of fungi into a general macroecological framework. Global metagenomics detects hotspots of fungal diversity and macroecological patterns and indicates that plant and fungal diversity are uncoupled. [Also see Perspective by Wardle and Lindahl] Assessing fungal diversity worldwide Fungi are hyperdiverse but poorly known, despite their ecological and economic impacts. Tedersoo et al. collected nearly 15,000 topsoil samples from 365 sites worldwide and sequenced their genomes (see the Perspective by Wardle and Lindahl). Overall, they found a striking decline in fungal species richness with distance from the equator. For some specialist groups though, diversity depended more on the abundance of host plants than host diversity or geography. The findings reveal a huge gap between known and described species and the actual numbers of distinct fungi in the worlds soils. Science, this issue 10.1126/science.1256688; see also p. 1052

Towards global patterns in the diversity and community structure of ectomycorrhizal fungi

Global species richness patterns of soil micro-organisms remain poorly understood compared to macro-organisms. We use a global analysis to disentangle the global determinants of diversity and community composition for ectomycorrhizal (EcM) fungi-microbial symbionts that play key roles in plant nutrition in most temperate and many tropical forest ecosystems. Host plant family has the strongest effect on the phylogenetic community composition of fungi, whereas temperature and precipitation mostly affect EcM fungal richness that peaks in the temperate and boreal forest biomes, contrasting with latitudinal patterns of macro-organisms. Tropical ecosystems experience rapid turnover of organic material and have weak soil stratification, suggesting that poor habitat conditions may contribute to the relatively low richness of EcM fungi, and perhaps other soil biota, in most tropical ecosystems. For EcM fungi, greater evolutionary age and larger total area of EcM host vegetation may also contribute to the higher diversity in temperate ecosystems. Our results provide useful biogeographic and ecological hypotheses for explaining the distribution of fungi that remain to be tested by involving next-generation sequencing techniques and relevant soil metadata.

Historical biogeography and diversification of truffles in the Tuberaceae and their newly identified southern hemisphere sister lineage.

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.

Large-scale fungal diversity assessment in the Andean Yungas forests reveals strong community turnover among forest types along an altitudinal gradient.

The Yungas, a system of tropical and subtropical montane forests on the eastern slopes of the Andes, are extremely diverse and severely threatened by anthropogenic pressure and climate change. Previous mycological works focused on macrofungi (e.g. agarics, polypores) and mycorrhizae in Alnus acuminata forests, while fungal diversity in other parts of the Yungas has remained mostly unexplored. We carried out Ion Torrent sequencing of ITS2 rDNA from soil samples taken at 24 sites along the entire latitudinal extent of the Yungas in Argentina. The sampled sites represent the three altitudinal forest types: the piedmont (400–700 m a.s.l.), montane (700–1500 m a.s.l.) and montane cloud (1500–3000 m a.s.l.) forests. The deep sequence data presented here (i.e. 4 108 126 quality‐filtered sequences) indicate that fungal community composition correlates most strongly with elevation, with many fungi showing preference for a certain altitudinal forest type. For example, ectomycorrhizal and root endophytic fungi were most diverse in the montane cloud forests, particularly at sites dominated by Alnus acuminata, while the diversity values of various saprobic groups were highest at lower elevations. Despite the strong altitudinal community turnover, fungal diversity was comparable across the different zonal forest types. Besides elevation, soil pH, N, P, and organic matter contents correlated with fungal community structure as well, although most of these variables were co‐correlated with elevation. Our data provide an unprecedented insight into the high diversity and spatial distribution of fungi in the Yungas forests.

Phylogenetic relationships of the Gomphales based on nuc-25S-rDNA, mit-12S-rDNA, and mit-atp6-DNA combined sequences

Phylogenetic relationships among Geastrales, Gomphales, Hysterangiales, and Phallales were estimated via combined sequences: nuclear large subunit ribosomal DNA (nuc-25S-rDNA), mitochondrial small subunit ribosomal DNA (mit-12S-rDNA), and mitochondrial atp6 DNA (mit-atp6-DNA). Eighty-one taxa comprising 19 genera and 58 species were investigated, including members of the Clathraceae, Gautieriaceae, Geastraceae, Gomphaceae, Hysterangiaceae, Phallaceae, Protophallaceae, and Sphaerobolaceae. Although some nodes deep in the tree could not be fully resolved, some well-supported lineages were recovered, and the interrelationships among Gloeocantharellus, Gomphus, Phaeoclavulina, and Turbinellus, and the placement of Ramaria are better understood. Both Gomphus sensu lato and Ramaria sensu lato comprise paraphyletic lineages within the Gomphaceae. Relationships of the subgenera of Ramaria sensu lato to each other and to other members of the Gomphales were clarified. Within Gomphus sensu lato, Gomphus sensu stricto, Turbinellus, Gloeocantharellus and Phaeoclavulina are separated by the presence/absence of clamp connections, spore ornamentation (echinulate, verrucose, subreticulate or reticulate), and basidiomal morphology (fan-shaped, funnel-shaped or ramarioid). Gautieria, a sequestrate genus in the Gautieriaceae, was recovered as monophyletic and nested with members of Ramaria subgenus Ramaria. This agrees with previous observations of traits shared by these two ectomycorrhizal taxa, such as the presence of fungal mats in the soil. Clavariadelphus was recovered as a sister group to Beenakia, Kavinia, and Lentaria. The results reaffirm relationships between the Geastrales, Gomphales, Hysterangiales, and the Phallales, suggesting extensive convergence in basidiomal morphology among members of these groups. A more extensive sampling that focuses on other loci (protein-coding genes have been shown to be phylogenetically informative) may be useful to answer questions about evolutionary relationships among these fungal groups.

Differential hypogeous sporocarp production from Nothofagus dombeyi and N. pumilio forests in southern Argentina

Mycorrhizal fungi that form hypogeous sporocarps are an important component of the temperate forest soil community. In many regions, such as the Nothofagus forest in the Patagonian Andes, this group of fungi has been poorly studied. Here we examined the spring and autumn community composition of “sequestrate fungi”, based on sporocarp production in pure forests of Nothofagus dombeyi (evergreen) and N. pumilio (deciduous). We investigated the possible relationships between these communities and environmental factors over 2 y. The rarefaction curves and the minimal richness estimates converged at nearly the same level for each forest type, and the asymptotes suggested that the sampling effort was sufficient to capture most of the hypogeous sporocarp richness in these forest stands. In total 27 species were recovered. Basidiomycota, Ascomycota and Glomeromycota respectively accounted for nine, two and one genera. Species richness of hypogeous sporocarps varied in relation to forest type but not to season (fall and spring), whereas sporocarp biomass varied according to an interaction between season and forest type. Species richness and sporocarp biomass were positively correlated with rainfall and negatively correlated with altitude. In addition sporocarp species richness was positively related to number of trees per transect. We found that two different forest stands, each dominated by different species of Nothofagus, exhibited different hypogeous sporocarp communities.

How to know the fungi: combining field inventories and DNA‐barcoding to document fungal diversity

The fungi kingdom is among the most diverse eukaryotic lineages on Earth with estimates of several million extant species (O’Brien et al., 2005; Blackwell, 2011; Taylor et al., 2014). Fungi play critical roles in carbon andnutrient cycling of terrestrial and aquatic ecosystems, and they are important pathogens and mutualists (Read & Perez-Moreno, 2003; Taylor et al., 2012; Grossart et al., 2016). More than 80% of plant species form symbioses with fungi and these symbioses have been crucial to the colonization of terrestrial ecosystems (Field et al., 2015a; Selosse et al., 2015). Despite their impacts on primary ecosystem functions, assessments of fungal biodiversity estimate that only c. 10% of fungal species have been described (Bass & Richards, 2011; Hibbett et al., 2011). Traditionally, specimen-based taxonomic studies have been the only way to discover new species. Because most fungi have microscopic life-stages and convergent morphological features (Rivas-Plata & Lumbsch, 2011; Wynns, 2015), many fungal groups remain severely undersampled. DNA-barcoding and highthroughput sequencing methods have provided a new framework for studying fungal biodiversity (Fierer et al., 2012; Schoch et al., 2012; Myrold et al., 2014), and diversity estimates based on environmental sequences have increased exponentially. Although these ‘sequence-based classification and identification’ methods are a powerful means to rapidly detect hidden diversity, careful interpretation of these data is needed to make accurate inferences (K~oljalg et al., 2013; Lindahl et al., 2013; Nguyen et al., 2015; Hibbett et al., 2016). In particular, many environmental sequences cannot be associated with a known fungal species or lineage. This remains a major challenge to decipher fungal community composition and understand ecological roles of fungi in leaf litter, soil, or inside plants (Yahr et al., 2016). In some cases, these fungi are truly undescribed and their ecological roles are unknown but in other cases they represent described taxa for which no sequence is available (Nagy et al., 2011; Nilsson et al., 2016). DNA barcoding of herbarium specimens and culture collections is extremely valuable to link unidentified sequences to known taxa (e.g. Brock et al., 2009; Nagy et al., 2011; Osmundson et al., 2013; Garnica et al., 2016).DNA sequences have been generated from fungal type specimens > 200 years old (Larsson & Jacobsson, 2004), but in many cases obtaining sequences from historical material is challenging (Dentinger et al., 2010). Today’s threats to biodiversity from habitat loss and climate change are occurring at an unprecedented scale, and it is possible that many species may become extinct before they have been discovered (Costello et al., 2013; Monastersky, 2014). In the need to describe and protect as many species as possible we addressed the following questions: what are the best methods to rapidly document fungal biodiversity? Are traditional, specimen-based approaches still useful?

The species of Scleroderma from Argentina, including a new species from the Nothofagus forest

Five ectomycorrhizal species of Scleroderma were identified from herbarium and field-collected specimens from Argentina. A new hypogeous species, Scleroderma patagonicum, was recorded in association with native Nothofagus spp. in Patagonia. The epigeous species S. albidum, S. areolatum, S. bovista and S. citrinum were associated with various exotic tree species. A phylogenetic analysis based on the ITS region of Scleroderma species, including S. patagonicum, illustrates its distinct status within Scleroderma, including its placement among species with reticulate spores. Descriptions with SEM images of the spores and a key to the species are provided.

Ocurrence of ectomycorrhizal, hypogeous fungi in plantations of exotic tree species in central Argentina

Eleven hypogeous, ectomycorrhizal species of Basidiomycota, including two new species, and one of the Zygomycota were collected in exotic tree plantations in Córdoba Province, Argentina. Descomyces fusisporus sp. nov., D. varians sp. nov., Hydnangium archeri (Berk.) Rodway, H. carneum Wallr., Hysterangium gardneri E. Fisch. and Setchelliogaster tenuipes (Setch.) Pouzar were associated with Eucalyptus spp. Endogone lactiflua Berk., Hymenogaster lycoperdineus Vittad., H. griseus Vittad., H. rehsteineri Bucholtz, Rhizopogon couchii A.H. Sm. and R. roseolus (Corda) Th. Fr., were associated with various northern hemisphere tree species. Descriptions are provided to aid identification of the hypogeous fungi in exotic plantations of Argentina.

Phylogenetic analysis of the genus Modicella reveals an independent evolutionary origin of sporocarp-forming fungi in the Mortierellales.

Most studies of tissue differentiation and development have focused on animals and plants but many fungi form multi-cellular aggregations of spore-bearing tissue known as fruiting bodies or sporocarps. The ability to form sporocarps has arisen independently in several different evolutionary lineages of fungi. Evolutionary relationships of most sporocarp-forming fungi are well known, but the enigmatic zygomycete genus Modicella contains two species of sporocarp-forming fungi for which the phylogenetic affinities have not been explored based on molecular data. Species of Modicella have an uncertain trophic mode and have alternatively been considered members of the order Endogonales (which contains documented species of sporocarp-forming fungi) or the order Mortierellales (which contains no previously documented species of sporocarp-forming fungi). In this study we perform phylogenetic analyses based on ribosomal DNA of Modicella malleola from the Northern Hemisphere and Modicella reniformis from the Southern Hemisphere to determine the evolutionary affinities of the genus Modicella. Our analyses indicate that Modicella is a monophyletic genus of sporocarp-forming fungi nested within the Mortierellales, a group of microfungi with no previously documented sporocarp-forming species. Because Modicella is distantly related to all other known sporocarp-forming fungi, we infer that this lineage has independently evolved the ability form sporocarps. We conclude that the genus Modicella should be a high priority for comparative genomics studies to further elucidate the process of sporocarp formation in fungi.