Root-associated Fungi Research Articles

Host plant—ectomycorrhizal fungus combination drives resource allocation in willow: Evidence for complex species interaction from a simple experiment

Root-associated mycorrhizal fungi affect plant growth and resource allocation. Our major aim was to explore the plant-internal mechanisms behind the effects of mycorrhizal colonization on leaf chemistry of willow (Salix spp.). Combinations of 2 willow varieties (Loden, Tora) and 2 ectomycorrhizal fungal species (Hebeloma fastibile, Tricholoma cingulatum) were grown under controlled conditions. Host plant variety and fungal species effects on host resource allocation (biomass and leaf chemistry) varied in a complex way. Shoot biomass growth and allocation was mostly affected by host plant variety, whereas leaf and root biomass allocation were strongly affected by mycorrhizal treatment. Leaf biomass production was affected by willow variety, mycorrhizal treatment, and the interaction between them. The results indicate a strong effect of mycorrhizal colonization on host plant biomass allocation, which can mediate mycorrhizal effects on leaf chemistry. For example, leaf biomass allocation was inversely correlated with foliar concentrations of salicylic acid, suggesting a functional link between the two. However, the apparent complexity of the host plant variety-fungal species interaction makes any prediction of their possible outcome very difficult, especially in an ecological context. Still, fungal species identity seems to be more important for the responses of Salix to mycorrhizal colonization than the amount of fungal biomass.

Diversity of root-associated fungi of mature Habenaria radiata and Epipactis thunbergii colonizing manmade wetlands in Hiroshima Prefecture, Japan

Throughout the industrialized world, wetland species face the greatest risk of extinction from altered environmental conditions and loss of habitat. Manmade wetlands are often the only feasible strategy to provide habitat for these species. Wetland orchids are particularly susceptible to environmental degradation due to potentially limited availability of specialized pollinators and mycorrhizal symbionts. Here, we assess the fungal symbiont diversity of two orchid species, Habenaria radiata and Epipactis thunbergii, occupying three manmade wetlands in Hiroshima Prefecture, Japan to determine if orchids colonizing reconstructed habitats associate with a phylogenetically diverse or narrow suite of fungal symbionts. We collected three individuals each of H. radiata and E. thunbergii, respectively, growing at the first pond, six H. radiata from a second pond, and two E. thunbergii from a third pond. We identified fungal taxa using PCR and DNA sequencing techniques. Habenaria radiata associated with a phylogenetically diverse suite of fungi; in comparison, E. thunbergii associated with a phylogenetically narrow range of fungi dominated by the Tulasnellaceae. These common wetland orchid species readily colonize manmade wetlands, and we propose sampling soils for the presence of appropriate mycorrhizal fungi to determine limitations on orchid population regeneration due to mycorrhizal specificity.

Microfungi associated with withering willow wood in ground contact near Syowa Station, East Antarctica for 40 years

Data are rather lacking on the diversity of microfungi associated with exotic plant substrates transported to continental Antarctica. We examined the diversity and species composition of microfungi associated with withering woody shoots of saplings of Salix spp. (willows) transplanted and in ground contact near Syowa Station, East Antarctica for more than 40 years. The willow saplings originated from Hokkaido, Northern Japan, and were experimentally transplanted in 1967–1968, but died within a few years. Dead willow shoots, unbranched and standing on bare ground for approximately 50 years, were used for the isolation of fungi with the surface disinfection method. A total of 43 isolates were retrieved from 32 (78 %) of the 41 shoots tested. The fungal isolates were classified into 18 molecular operational taxonomic units (MOTUs) based on the similarity of rDNA ITS sequences at the 97 % criterion. Leotiomycetes was the most common class in terms of the number of isolates and MOTUs, followed by Dothideomycetes, Sordariomycetes, and Eurotiomycetes. Molecular phylogenetic affinities suggested that the closest relatives of the MOTUs were saprobic and root-associated fungi. The result of the present study suggested that Cadophora luteo-olivacea is widespread in soils throughout Antarctica and likely indigenous.

Community composition of root-associated fungi in a Quercus-dominated temperate forest: "codominance" of mycorrhizal and root-endophytic fungi.

In terrestrial ecosystems, plant roots are colonized by various clades of mycorrhizal and endophytic fungi. Focused on the root systems of an oak-dominated temperate forest in Japan, we used 454 pyrosequencing to explore how phylogenetically diverse fungi constitute an ecological community of multiple ecotypes. In total, 345 operational taxonomic units (OTUs) of fungi were found from 159 terminal-root samples from 12 plant species occurring in the forest. Due to the dominance of an oak species (Quercus serrata), diverse ectomycorrhizal clades such as Russula, Lactarius, Cortinarius, Tomentella, Amanita, Boletus, and Cenococcum were observed. Unexpectedly, the root-associated fungal community was dominated by root-endophytic ascomycetes in Helotiales, Chaetothyriales, and Rhytismatales. Overall, 55.3% of root samples were colonized by both the commonly observed ascomycetes and ectomycorrhizal fungi; 75.0% of the root samples of the dominant Q. serrata were so cocolonized. Overall, this study revealed that root-associated fungal communities of oak-dominated temperate forests were dominated not only by ectomycorrhizal fungi but also by diverse root endophytes and that potential ecological interactions between the two ecotypes may be important to understand the complex assembly processes of belowground fungal communities.

Roots and Associated Fungi Drive Long-Term Carbon Sequestration in Boreal Forest

Boreal forest soils function as a terrestrial net sink in the global carbon cycle. The prevailing dogma has focused on aboveground plant litter as a principal source of soil organic matter. Using (14)C bomb-carbon modeling, we show that 50 to 70% of stored carbon in a chronosequence of boreal forested islands derives from roots and root-associated microorganisms. Fungal biomarkers indicate impaired degradation and preservation of fungal residues in late successional forests. Furthermore, 454 pyrosequencing of molecular barcodes, in conjunction with stable isotope analyses, highlights root-associated fungi as important regulators of ecosystem carbon dynamics. Our results suggest an alternative mechanism for the accumulation of organic matter in boreal forests during succession in the long-term absence of disturbance.

Fungal communities influence root exudation rates in pine seedlings

Root exudates are hypothesized to play a central role in belowground food webs, nutrient turnover, and soil C dynamics in forests, but little is known about the extent to which root-associated microbial communities influence exudation rates in trees. We used a novel experimental technique to inoculate loblolly pine (Pinus taeda L.) seedlings with indigenous forest fungi to examine how diverse fungal communities influence exudation. Surface-sterilized seeds were sown in intact, unsieved soil cores for 14weeks to promote root colonization by fungi. After 14weeks, we transferred seedlings and root-associated fungi into cuvettes and measured exudate accumulation in trap solutions. Both the abundance and identity of root-associated fungi influenced exudation. Exudation rates were greatest in root systems least colonized by ectomycorrhizal (ECM) fungi and most colonized by putative pathogenic and saprotrophic fungi. However, the ECM community composition was not a strong determinant of exudation rates. These results suggest that environmental conditions that influence the degree to which tree roots are colonized by pathogenic and saprotrophic vs. mutualistic fungi are likely to mediate fluxes of labile C in forest soils, with consequences for soil biogeochemistry and ecosystem processes.

Assessment of root-associated fungal communities colonizing two species of tropical grasses reveals incongruence to fungal communities of North American native grasses

Abstract This study characterized the root-associated fungal (RAF) communities inhabiting Eustachys petraea and Panicum maximum , two tropical and weedy grass species on an island off the coast of Honduras, Central America. Media-based analyses revealed that fungi colonizing these grasses exhibited similar morphotypes (albeit in different proportions). Conversely, molecular-based analyses suggested that the community of fungal OTUs were dissimilar, with several OTUs commonly present on only one plant and no AMF sequences present in either plant species. When compared to various datasets of RAF obtained from the North American mainland (with similar methods), the Honduran grasses harbored a different set of fungal OTUs. Interestingly, some cosmopolitan taxonomic clades commonly encountered on the mainland were also isolated in Honduras, though based on ITS rDNA the fungi were only related distantly to those on the mainland. Additional work is necessary to determine how the RAF communities change over geographic distance and whether invasive grasses are constitutionally set up to accept relationships with a different suite of RAF species.

A diverse fungal community associated with Pseudorchis albida (Orchidaceae) roots

Abstract In addition to orchid mycorrhizal fungi (OrMF), the roots of orchids harbour plant fungal endophytes termed root-associated fungi (RAF). In the present study, the endangered photosynthetic orchid Pseudorchis albida was screened for OrMF and RAF using culture-dependent (isolations from root sections and pelotons) and culture-independent (cloning from root sections) techniques. The efficiency of the different approaches for detecting the fungi and the effect of the sampling season (summer or autumn) were evaluated. In total, 66 distinct OTUs of mycorrhizal and non-mycorrhizal fungi were found, which, to our knowledge, is the highest diversity of RAF that has yet been detected in a single orchid species. The OrMF community was dominated by Tulasnella species, which were mainly detected by isolation from pelotons or cloning from root sections. The roots and tubers showed higher mycorrhizal colonization in summer, corroborating the frequent reports of Tulasnella from pelotons in this season. In contrast, two helotialean fungi, Varicosporium elodeae and Leohumicola sp., the latter of which was repeatedly isolated from pelotons, were significantly more abundant in the autumn.

A variety of melanised root-associated fungi from the Sydney basin form endophytic associations with Trifolium subterraneum

Abstract Knowledge of the abundance, diversity, and plant interactions of melanised root-associated fungi remains limited. The objective of this study was to isolate a wide variety of melanised root-associated fungi within the Sydney basin (NSW, Australia) and assess growth response of Trifolium subterraneum to inoculation with individual isolates. Of 902 root-associated fungi isolated from plant roots, 118 were melanised. All but two of these fungi were re-isolated from inoculated T. subterraneum seedlings after 7 weeks in a controlled environment. Approximately 60 % of the melanised root-associated fungi did not reduce plant growth. Twenty-four isolates tended to increase plant growth and were tentatively identified as predominantly ascomycetes, and one zygomycete. Melanised root-associated fungi appeared to form complex interactions with T. subterraneum, the natures of which remain to be further explored. Melanised root-associated fungi could potentially play key ecological roles including positively influencing edaphic conditions.

Proteomic changes and endophytic micromycota during storage of organically and conventionally grown carrots

The physiological state of carrot roots during extended cold-storage is decisive for high postharvest quality. We have investigated differences in the proteome and micromycota of organically and conventionally grown carrots during six months of storage. The levels of only 15 proteins changed in level during storage. Proteins involved in cold stress adaptation and cytoskeleton components changed; these changes in specific protein levels occurred mainly during the first month demonstrating adaptation to storage conditions and that the carrots were subsequently stable, indicating stable carrot quality. The changes observed were similar in the two cropping systems.Using both biological isolation and a fungal PCR targeting the ITS region, we identified several endophytic species belonging to the Ascomycota. The most frequently encountered taxa were Tetracladium, Leptodontidium, Nectriaceae and Phoma which are known to occur as root endophytes or as root-associated fungi. As for the proteomics data, no consistent statistically significant differences in micromycota were observed between the two cropping systems. We conclude that cropping system did not have an influence on the postharvest quality of the carrots during six months of cold storage.

Efficacy of root-associated fungi and PGPR on the growth of Pisum sativum (cv. Arkil) and reproduction of the root-knot nematode Meloidogyne incognita.

The effects of root-associated fungi (Aspergillus awamori and Glomus mosseae) and plant growth promoting rhizobacteria (PGPR) (Pseudomonas putida, Pseudomonas alcaligenes and Paenibacillus polymyxa) were studied alone and in combination in glasshouse experiments on the growth of pea, enzyme activity (peroxidase and catalase) and reproduction of root-knot nematode Meloidogyne incognita. Application of A. awamori, G. mosseae and PGPR caused a significant increase in pea growth and enzyme activities of both nematode inoculated and uninoculated plants. A. awamori was more effective in reducing galling and improving the growth of nematode inoculated plants than P. alcaligenes or P. polymyxa. The greatest increase in growth, enzyme activities of nematode-inoculated plants and reduction in galling and nematode multiplication was observed when A. awamori was used with P. putida or G. mosseae as compared to the other combinations tested. Percentage root colonization was higher when AM fungus inoculated plants were treated with P. putida both in presence and absence of nematode.

Arbuscular mycorrhizal colonization of giant sequoia (Sequoiadendron giganteum) in response to restoration practices

Interactions with soil microbiota determine the success of restoring plants to their native habitats. The goal of our study was to understand the effects of restoration practices on interactions of giant sequoia Sequoiadendron giganteum with arbuscular mycorrhizal (AM) fungi (Glomeromycota). Natural regeneration of Sequoiadendron is threatened by the absence of severe fires that create forest canopy gaps. Generating artificial canopy gaps offers an alternative tool for giant sequoia restoration. We investigated the effect of regeneration practices, including (i) sapling location within gaps, (ii) gap size and (iii) soil substrate, on AM fungal colonization of giant sequoia sapling roots in a native giant sequoia grove of the Sierra Nevada, California. We found that the extent of AM fungal root colonization was positively correlated with sapling height and light availability, which were related to the location of the sapling within the gap and the gap size. While colonization frequency by arbuscules in saplings on ash substrate was higher relative to saplings in mineral soil, the total AM fungal root colonization was similar between the substrates. A negative correlation between root colonization by Glomeromycota and non-AM fungal species indicated antagonistic interactions between different classes of root-associated fungi. Using DNA genotyping, we identified six AM fungal taxa representing genera Glomus and Ambispora present in Sequoiadendron roots. Overall, we found that AM fungal colonization of giant sequoia roots was associated with availability of plant-assimilated carbon to the fungus rather than with the AM fungal supply of mineral nutrients to the roots. We conclude that restoration practices affecting light availability and carbon assimilation alter feedbacks between sapling growth and activity of AM fungi in the roots.

The dark side is not fastidious--dark septate endophytic fungi of native and invasive plants of semiarid sandy areas.

Dark septate endophytic (DSE) fungi represent a frequent root-colonizing fungal group common in environments with strong abiotic stress, such as (semi)arid ecosystems. This work aimed to study the DSE fungi colonizing the plants of semiarid sandy grasslands with wood steppe patches on the Great Hungarian Plain. As we may assume that fungi colonizing both invasive and native species are generalists, root associated fungi (RAF) were isolated from eight native and three invasive plant species. The nrDNA sequences of the isolates were used for identification. To confirm that the fungi were endophytes an artificial inoculation system was used to test the isolates: we considered a fungus as DSE if it colonized the roots without causing a negative effect on the plant and formed microsclerotia in the roots. According to the analyses of the ITS sequence of nrDNA the 296 isolates clustered into 41 groups. We found that 14 of these 41 groups were DSE, representing approximately 60% of the isolates. The main DSE groups were generalist and showed no specificity to area or season and colonized both native and invasive species, demonstrating that exotic plants are capable of using the root endophytic fungi of the invaded areas. The DSE community of the region shows high similarity to those found in arid grasslands of North America. Taking into account a previous hypothesis about the common root colonizers of those grasslands and our results reported here, we hypothesize that plants of (semi)arid grasslands share common dominant members of the DSE fungal community on a global scale.

Surprising spectra of root-associated fungi in submerged aquatic plants

Similarly to plants from terrestrial ecosystems, aquatic species harbour wide spectra of root-associated fungi (RAF). However, comparably less is known about fungal diversity in submerged roots. We assessed the incidence and diversity of RAF in submerged aquatic plants using microscopy, culture-dependent and culture-independent techniques. We studied RAF of five submerged isoetid species collected in four oligotrophic freshwater lakes in Norway. Levels of dark septate endophytes (DSE) colonization differed among the lakes and were positively related to the organic matter content and negatively related to pH. In total, we identified 41 fungal OTUs using culture-dependent and culture-independent techniques, belonging to Mucoromycotina, Chytridiomycota, Glomeromycota, Ascomycota as well as Basidiomycota. Sequences corresponding to aquatic hyphomycetes (e.g. Nectria lugdunensis, Tetracladium furcatum and Varicosporium elodeae) were obtained. Eight arbuscular mycorrhizal taxa belonging to the orders Archaeosporales, Diversisporales and Glomerales were also detected. However, the vast majority of the fungal species detected (e.g. Ceratobasidium sp., Cryptosporiopsis rhizophila, Leptodontidium orchidicola, and Tuber sp.) have previously been known only from roots of terrestrial plants. The abundance and phylogenetic distribution of mycorrhizal as well as nonmycorrhizal fungi in the roots of submerged plants have reshaped our views on the fungal diversity in aquatic environment.

Morphology, anatomy, and molecular studies of the ectomycorrhiza formed axenically by the fungus Sistotrema sp. (Basidiomycota)

Several species of the corticioid (resupinate) genus Sistotrema of the cantharelloid clade (Basidiomycota) were recently found to be ectomycorrhizal. This changed the traditional assertion that all Sistotrema species are strictly wood rotting and suggests that the genus may be polyphyletic. In the present investigation, a still unknown root tip-associated fungal specimen (EW63) was isolated and found to be associated with an above-ground fruiting body. Sequencing of the ITS and the nucLSU DNA regions and phylogenetic analyses verified that the root-associated fungus and the fruiting body represented the same species, which was found to belong to the genus Sistotrema. To prove the ectomycorrhizal status of this strain, axenic Pinus sylvestris resyntheses in flask cultures were conducted. Growth parameters of the seedlings were determined and the morphology and anatomy of the synthesized mycorrhizas were described. Length and dry mass of the Pinus shoot as well as those of the total root tips were found to be enhanced as a result of the mycorrhizal association. Mycorrhizal frequency was high (51.5%) in these cultures. Mycorrhizal root tips were cottony light ochre with a thin plectenchymatic hyphal mantle. The clamps of the fruiting body hyphae as well as the mycorrhiza were ampullately inflated. This is the first report proving in axenic culture that a fungus belonging to the genus Sistotrema forms true ectomycorrhiza.

Community structure of ericoid mycorrhizas and root-associated fungi of Vaccinium membranaceum across an elevation gradient in the Canadian Rocky Mountains

Abstract The ecological plasticity of Vaccinium membranaceum (Ericaceae) to grow from valley bottoms to alpine habitats allowed us to test if fungal community structure varies along an elevation gradient in east-central British Columbia. Using the shrub as an independent variable, and plant and soil features along the gradient to model a changing climate, communities of root-associated fungi were investigated. The colonized roots from 40 individuals were examined using culture-dependent sequencing and ARISA (automated ribosomal intergenic spacer analysis). Our results demonstrated that high elevation fungal communities, characterised by Rhizoscyphus ericae , differ from lower elevation communities, where Phialocephala fortinii was the most frequently isolated fungus. Co-occurrence analysis indicated that, overall, fungi tended to occur together more often than would be expected by chance. At the scale of the individual host plant, facilitation may play a more important role than competition in shaping fungal communities in these ecosystems.

Short-term impacts of soil freeze-thaw cycles on roots and root-associated fungi of Holcus lanatus and Calluna vulgaris

Background and aims Soil freeze-thaw cycle (FTC) regimes are altered by climate change and known to influence nutrient cycling and plant growth. Here, we explore mechanistic explanations for the changing plant performance of the grass Holcus lanatus and the dwarf shrub Calluna vulgaris.

Dispersal of arbuscular mycorrhizal fungi and dark septate endophytes by Ctenomys cf. knighti (Rodentia) in the northern Monte Desert of Argentina

In arid and semi-arid environments, root-associated fungi may play a key role in plant communities (e.g., seedling establishment, nutrient acquisition, plant survival and heat tolerance). Several studies have shown the importance of small mammals as consumers and dispersal agents of mycorrhizal fungi spores in tropical and temperate ecosystems. However, little is known about the dispersal of infective propagules of endophytic fungi in arid and semi-arid ecosystems. This study analyzed the potential role of the subterranean rodent Ctenomys cf . knighti as a dispersal agent of root-associated fungi. In order to demonstrate this role, we analyzed: (1) the incidence of colonization of arbuscular mycorrhizae and dark septate endophytes on representative plant species of the Monte Desert; (2) the presence of fungal structures of arbuscular mycorrhizae and dark septate endophytes in C. cf. knighti fecal samples; and (3) the infectivity of the fungal propagules contained in the scat and their growth effects on nine native plant species. Data strongly suggest that this South American subterranean rodent may play a key role as a dispersal agent of arbuscular mycorrhizae and dark septate endophytes fungi in the arid environment of the northern Monte Desert of Argentina.

Precipitation increases the abundance of some groups of root-associated fungal endophytes in a semiarid grassland

Diverse communities of Root-Associated Fungal (RAF) species, including Dark Septate Endophytes (DSE), inhabit grasses in semiarid soils. These aridlands are often distinguished by large but variable precipitation events (monsoons). We sought to compare the composition and fungal load of the RAF community inhabiting the roots of Sporobolus cryptandrus, a dominant forage grass, exposed to variable precipitation events. We used a rainfall manipulation experiment at the Sevilleta National Wildlife Refuge (SNWR) to examine the abundance and composition of the RAF communities using molecular and microscopic techniques. Molecular data reveals that the RAF communities are dominated by Ascomycetes and few sequences were related to Arbuscular Mycorrhizal Fungi (AMF). The most dominant group (Paraphaeosphaeria spp.) clearly increased in abundance in response to water amendments and the addition of similar volumes of water appeared to increase similarity among the RAF communities, irrespective of size and distribution of precipitation events. Microscopic examinations of the roots also revealed an increased fungal load when exposed to elevated moisture, irrespective of rainfall frequency. We suggest that the increase in these cosmopolitan and abundant groups of Ascomycete fungi, in part, represents a common but complicated strategy of water translocation in a variety of soil types, similar to that proposed and shown in the arbuscular mycorrhizal fungi (AMF). Additional work should examine whether many (or most) members of the Ascomycete RAF also can facultatively broker water in the larger biological network in the soil.

Differences in fungal communities associated to Festuca paniculata roots in subalpine grasslands

Mycorrhizal fungi or endphytes colonize plant roots and their occurrence and composition depend on biotic and abiotic characteristics of the ecosystem. We investigated the composition of these microbial communities associated with Festuca paniculata, a slow growing species, which dramatically impacts functional plant diversity and the recycling of organic matter in subalpine grasslands. F. paniculata individuals from both mown and unmown grasslands were randomly collected and the microscopic observation of the plant roots revealed a difference in fungal colonization according to management. The ITS regions of root-associated fungi were amplified, cloned and sequenced. Bioinformatic analysis revealed a total of 43 and 35 phylotypes in mown and unmown grasslands respectively, highlighting a remarkable difference in the composition between both fungal communities. The phylotypes were assigned to 9 classes in which two classes Eurotiomycetes and Lecanoromycetes were specific to mown grasslands, while Tremellomycetes were specific to unmown grasslands and only five phylotypes were common to both locations. The comparative analysis of fungal lifestyles indicated the dominance of saprobes and a large proportion of endophytes compared to the mycorrhizal fungi (7/1 and 11/2 phylotypes in mown and unmown grasslands, respectively). Endophyte richness was greater in the unmown gassland than in the mown grassland and their relative proportion was twice higher. Our results suggest that endophytes may offer potential resources to F. paniculata and play an important role in the regulation of plant diversity.