<Emphasis Type="Italic">Monotropastrum humile</Emphasis> var. <Emphasis Type="Italic">humile</Emphasis> is associated with diverse ectomycorrhizal Russulaceae fungi in Japanese forests

Monotropastrum humile is nearly lacking in chlorophyll and obtains its nutrients, including carbon sources, from associated mycorrhizal fungi. We analyzed the mycorrhizal fungal affinity and species diversity of M. humile var. humile mycorrhizae to clarify how the plant population survives in Japanese forest ecosystems. We classified 78 samples of adult M. humile var. humile individuals from Hokkaido, Honshu, and Kyusyu Islands into 37 root mycorrhizal morphotypes. Of these, we identified 24 types as Russula or Lactarius fungal taxa in the Russulaceae, Basidiomycetes, but we could not identify the remaining 13 types as to their genus in the Basidiomycetes. The number of fungal species on M. humile var. humile was the highest in the plant subfamily. The diversity of fungal species revealed its increased trends in natural forests at the stand level, fagaceous vegetation, and cool-temperate climate. The most frequently observed fungus colonized mainly samples collected from sub-alpine forests; the second most frequently observed fungus colonized samples collected from sub-alpine to warm-temperate forests. These results suggest that Japanese M. humile populations are associated with specific but diverse fungi that are common ectomycorrhizal symbionts of various forest canopy trees, indicating a tripartite mycorrhizal relationship in the forest ecosystem.     

Mycorrhizal features and fungal partners of four mycoheterotrophic Monotropoideae (Ericaceae) species from Yunnan, China

We provide a preliminary report of the mycobionts found within four Monotropoideae (Ericaceae) species from China: Monotropa uniflora, Hypopitys monotropa, Monotropastrum humile and Monotropastrum sciaphilum (a rare endemic species never previously studied for mycorrhizae). Such achlorophyllous Monotropoideae plants obtain their carbohydrates from mycorrhizal fungi linking them to surrounding trees, on which these fungi form ectomycorrhizae. Since Monotropoideae were rarely studied in continental Asia, the root systems of the four species sampled in Yunnan were examined using morphological and molecular methods. All the roots of these four species exhibit a typical monotropoid mycorrhizal morphology, including a fungal mantle, a Hartig net and hyphal pegs. In M. uniflora and M. humile mycorrhizae, cystidia typical of Russula symbionts covered the fungal mantle. ITS barcoding revealed that Russulales were the most frequent colonizers in all species, but Hypopitys monotropa displayed various additional mycorrhizal taxa. Moreover, a few additional ectomycorrhizal and saprotrophic Basidiomycota taxa were identified in the three other species, challenging that these four Monotropoideae species are as strictly fungal specific as the other Monotropoideae species hitherto studied. Moreover, a comparison with accompanying fungus sporocarps revealed that the fruiting fungal community significantly differed from that associated with the Monotropoideae roots, so that a clear fungal preference was evident. Finally, four fungal species were found on more than one Monotropoideae species: this contrasted with previous reports of sympatrically growing mycoheterotrophic plants, which did not reveal any overlap. This again challenges the idea of strict fungal specificity.     

Monotropa uniflora plants of eastern Massachusetts form mycorrhizae with a diversity of russulacean fungi

Plant species in the subfamily Monotropoideae are mycoheterotrophs; they obtain fixed carbon from photosynthetic plants via a shared mycorrhizal network. Previous findings show mycoheterotrophic plants exhibit a high level of specificity to their mycorrhizal fungi. In this study we explore the association of mycorrhizal fungi and Monotropa uniflora (Monotropoideae: Ericaceae) in eastern North America. We collected M. uniflora roots and nearby basidiomycete sporocarps from four sites within a 100 km2 area in eastern Massachusetts. We analyzed DNA sequences of the internal transcribed spacer region (ITS) from the fungal nuclear ribosomal gene to assess the genetic diversity of fungi associating with M. uniflora roots. In this analysis we included 20 ITS sequences from Russula sporocarps collected nearby, 44 sequences of Russula or Lactarius species from GenBank and 12 GenBank sequences of fungi isolated from M. uniflora roots in previous studies. We found that all 56 sampled M. uniflora mycorrhizal fungi were members of the Russulaceae, confirming previous research. The analysis showed that most of the diversity of mycorrhizal fungi spreads across the genus Russula. ITS sequences of the mycorrhizal fungi consisted of 20 different phylotypes: 18 of the genus Russula and two of Lactarius, based on GenBank searches. Of the sampled plants, 57% associated with only three of the 20 mycorrhizal fungi detected in roots, and of the 25 sporocarp phylotypes collected three, were associated with M. uniflora. Furthermore the results indicate that the number of different fungal phylotypes associating with M. uniflora of eastern North America is higher than that of western North America but patterns of fungal species abundance might be similar between mycorrhizae from the two locations.     

Molecular identification of the mycorrhizal fungi of the epiparasitic plant Monotropastrum humile var. glaberrimum (Ericaceae)

Achlorophyllous monotropoid plants (Monotropoideae, Ericaceae) are epiparasites that obtain all of their carbon from their host plants via connections with mycorrhizal fungi. The mycorrhizal fungi of the epiparasitic monotropoid Monotropastrum humile var. glaberrima were identified based on mitochondrial, large ribosomal DNA sequences, and were compared with those of another variety, M. humile var. humile. The fungi that inhabit M. humile var. glaberrimum belong to the Thelephoraceae, whereas that of M. humile var. humile is a member of the Russulaceae. Two explanations are possible for this phenomenon: a misunderstanding of the taxonomic position of M. humile var. glaberrimum, or a change in the fungal partner within the Monotropastrum.     

How are plant and fungal communities linked to each other in belowground ecosystems? A massively parallel pyrosequencing analysis of the association specificity of root-associated fungi and their host plants

In natural forests, hundreds of fungal species colonize plant roots. The preference or specificity for partners in these symbiotic relationships is a key to understanding how the community structures of root‐associated fungi and their host plants influence each other. In an oak‐dominated forest in Japan, we investigated the root‐associated fungal community based on a pyrosequencing analysis of the roots of 33 plant species. Of the 387 fungal taxa observed, 153 (39.5%) were identified on at least two plant species. Although many mycorrhizal and root‐endophytic fungi are shared between the plant species, the five most common plant species in the community had specificity in their association with fungal taxa. Likewise, fungi displayed remarkable variation in their association specificity for plants even within the same phylogenetic or ecological groups. For example, some fungi in the ectomycorrhizal family Russulaceae were detected almost exclusively on specific oak (Quercus) species, whereas other Russulaceae fungi were found even on “non‐ectomycorrhizal” plants (e.g., Lyonia and Ilex). Putatively endophytic ascomycetes in the orders Helotiales and Chaetothyriales also displayed variation in their association specificity and many of them were shared among plant species as major symbionts. These results suggest that the entire structure of belowground plant–fungal associations is described neither by the random sharing of hosts/symbionts nor by complete compartmentalization by mycorrhizal type. Rather, the colonization of multiple types of mycorrhizal fungi on the same plant species and the prevalence of diverse root‐endophytic fungi may be important features of belowground linkage between plant and fungal communities.     

Effect of environmental gradient in coastal vegetation on communities of arbuscular mycorrhizal fungi associated with Ixeris repens (Asteraceae)

The community structure of arbuscular mycorrhizal (AM) fungi associated with Ixeris repens was studied in coastal vegetation near the Tottori sand dunes in Japan. I. repens produces roots from a subterranean stem growing near the soil surface which provides an opportunity to examine the effects of an environmental gradient related to distance from the sea on AM fungal communities at a regular soil depth. Based on partial sequences of the nuclear large subunit ribosomal RNA gene, AM fungi in root samples were divided into 17 phylotypes. Among these, five AM fungal phylotypes in Glomus and Diversispora were dominant near the seaward forefront of the vegetation. Redundancy analysis of the AM fungal community showed significant relationships between the distribution of phylotypes and environmental variables such as distance from the sea, water-soluble sodium in soil, and some coexisting plant species. These results suggest that environmental gradients in the coastal vegetation can be determinants of the AM fungal community.     

Communities of arbuscular mycorrhizal fungi in Stipa krylovii (Poaceae) in the Mongolian steppe

We examined arbuscular mycorrhizal (AM) fungi colonizing the roots of Stipa krylovii, a grass species dominating the grasslands of the steppe zone in Hustai and Uvurkhangai in Mongolia. The AM fungal communities of the collected S. krylovii roots were examined by molecular analysis based on the partial sequences of a small subunit of ribosomal RNA gene as well as AM fungal colonization rates. Almost all AM fungi detected were in Glomus-group A, and were divided into 10 phylotypes. Among them, one phylotype forming a clade with G. intraradices and G. irregulare was the most dominant. Furthermore, it was also found that most of the phylotypes include AM fungi previously detected in high altitude regions in the Eurasian Continent. Significant correlations were found among soil total N, total plant biomass and AM fungal colonization ratio, which suggested that higher plant biomass may be required for the proliferation of AM fungi in the environment. Meanwhile, redundancy analysis on AM fungal distribution and environmental variables suggested that the effect of plant biomass and most soil chemical properties on the AM fungal communities were not significant.     

Wheat cultivars form distinctive communities of root-associated arbuscular mycorrhiza in a conventional agroecosystem

Background and aims

The effect of plant species on their root-associated arbuscular mycorrhizal (AM) fungi is well studied, but how this effect operates at the cultivar level remains poorly understood. This study investigates how wheat cultivars shape their AM fungal communities.

Methods

Twenty-one new wheat cultivars were traditionally cultivated in a dryland of northwestern China, and their agronomic traits, soil characteristics and the abundance and community composition of AM fungi were measured.

Results

Both spore community in soils and AM fungal phylotypes inside roots were significantly influenced by cultivar even though hyphal abundance, spore density and AM fungal diversity were similar across cultivars. Three out of 16 AM fungal phylotypes interacted with most cultivars, whilst some phylotypes preferred to colonize cultivars with similar agronomic traits. Six wheat cultivars, all which had hosted 6 AM fungal phylotypes, seemed to be generalists. Nestedness analysis and stochastic model fitting revealed that the AM fungal communities colonizing roots were codetermined by deterministic and stochastic processes.

Conclusions

A complex pattern of cultivar-AM fungal interactions was observed in this study, and our results highlight that the host effect on the community assembly of AM fungi could be operating on the level of plant cultivar.     

Local adaptation to soil hypoxia determines the structure of an arbuscular mycorrhizal fungal community in roots from natural CO₂ springs

The processes responsible for producing and maintaining the diversity of natural arbuscular mycorrhizal (AM) fungal communities remain largely unknown. We used natural CO(2) springs (mofettes), which create hypoxic soil environments, to determine whether a long-term, directional, abiotic selection pressure could change AM fungal community structure and drive the selection of particular AM fungal phylotypes. We explored whether those phylotypes that appear exclusively in hypoxic soils are local specialists or widespread generalists able to tolerate a range of soil conditions. AM fungal community composition was characterized by cloning, restriction fragment length polymorphism typing, and the sequencing of small subunit rRNA genes from roots of four plant species growing at high (hypoxic) and low (control) geological CO(2) exposure. We found significant levels of AM fungal community turnover (β diversity) between soil types and the numerical dominance of two AM fungal phylotypes in hypoxic soils. Our results strongly suggest that direct environmental selection acting on AM fungi is a major factor regulating AM fungal communities and their phylogeographic patterns. Consequently, some AM fungi are more strongly associated with local variations in the soil environment than with their host plant's distribution.     

Diversity and host preference of fungi co-inhabiting Cenococcum mycorrhizae

Diverse fungal assemblages colonize the fine feeder roots of woody plants, including mycorrhizal fungi, fungal root endophytes and soil saprotrophs. The fungi co-inhabiting Cenococcum geophilum ectomycorrhizae (ECM) of Abies balsamea, Betula papyrifera and Picea glauca were studied at two boreal forest sites in Eastern Canada by direct PCR of ITS rDNA. 50 non-Cenococcum fungal sequence types were detected, including several potentially mycorrhizal species as well as fungal root endophytes. Non-melanized ascomycetes dominated, in contrast to the dark septate endophytes (DSE) reported in most culture dependent studies. The results demonstrate significant differences in root associated fungal assemblages among the host species studied. Fungal diversity was also host dependent, with P. glauca roots supporting a more diverse community than A. balsamea. Differences in root associated fungal communities may well influence ecological interactions among host plant species.     

The incidence of arbuscular mycorrhiza in two submerged Isoëtes species

We investigated the occurrence of arbuscular mycorrhizal fungi in the roots of Isoëtes lacustris and I. echinospora. These submerged lycopsids are the only macrophyte species inhabiting the bottom of two acidified glacial lakes in the Czech Republic. Arbuscular mycorrhizal (AM) fungi were detected in the roots of both species but the percentage of root colonization was both low and variable. Nevertheless, planting Littorella uniflora in the sediments from Isoëtes rhizosphere revealed high levels of viable AM propagules in both lakes. While AM colonization of Isoëtes roots did not exceed 25%, the average colonization of Littorella roots amounted to more than 80%. Although colonization of quillwort roots by AM fungi is evident, the taxonomic identity and role of these AM fungi in plant growth remain unclear. In addition to AM fungi, root-colonizing dark septate endophytic fungi were observed in both Isoëtes species.     

Effect of plant species on communities of arbuscular mycorrhizal fungi in the Mongolian steppe

Communities of arbuscular mycorrhizal (AM) fungi were investigated in Stipa krylovii, Leymus chinensis (Poaceae), Allium bidentatum (Liliaceae), and Astragalus brevifolius (Fabaceae) in the Mongolian steppe to examine the effect of plant species on the communities in this study. The AM fungal communities were examined by molecular analysis based on the partial sequences of a small subunit of the ribosomal RNA gene. The sequences obtained were divided into 23 phylotypes by the sequence similarity >98%. Many of the AM fungal phylotypes included AM fungi previously detected in high-altitude regions in the Tibet and Loes plateaus, which suggested that these AM fungi may have wide distribution with stressful conditions of aridity and coldness. Among the 23 phylotypes, 12 phylotypes were found in all four plants, and 87.4% of the all obtained sequences were affiliated into these 12 types. For the distribution of the AM fungal phylotypes, overlapping of the phylotypes among the four plant species were significantly higher than that simulated by random chance. These results suggested that AM fungal communities were less diversified among the examined plant species.     

Specificity of fungal associations of Pyroleae and Monotropa hypopitys during germination and seedling development

Mycoheterotrophic plants obtain organic carbon from associated mycorrhizal fungi, fully or partially. Angiosperms with this form of nutrition possess exceptionally small ‘dust seeds’ which after germination develop ‘seedlings’ that remain subterranean for several years, fully dependent on fungi for supply of carbon. Mycoheterotrophs which as adults have photosynthesis thus develop from full to partial mycoheterotrophy, or autotrophy, during ontogeny. Mycoheterotrophic plants may represent a gradient of variation in a parasitism–mutualism continuum, both among and within species. Previous studies on plant–fungal associations in mycoheterotrophs have focused on either germination or the adult life stages of the plant. Much less is known about the fungal associations during development of the subterranean seedlings. We investigated germination and seedling development and the diversity of fungi associated with germinating seeds and subterranean seedlings (juveniles) in five Monotropoideae (Ericaceae) species, the full mycoheterotroph Monotropa hypopitys and the putatively partial mycoheterotrophs Pyrola chlorantha, P. rotundifolia, Moneses uniflora and Chimaphila umbellata. Seedlings retrieved from seed sowing experiments in the field were used to examine diversity of fungal associates, using pyrosequencing analysis of ITS2 region for fungal identification. The investigated species varied with regard to germination, seedling development and diversity of associated fungi during juvenile ontogeny. Results suggest that fungal host specificity increases during juvenile ontogeny, most pronounced in the fully mycoheterotrophic species, but a narrowing of fungal associates was found also in two partially mycoheterotrophic species. We suggest that variation in specificity of associated fungi during seedling ontogeny in mycoheterotrophs represents ongoing evolution along a parasitism–mutualism continuum.     

Distribution of arbuscular mycorrhizal fungi in four semi-mangrove plant communities

To better understand the diversity and species composition of arbuscular mycorrhizal fungi (AMF) in mangrove ecosystems, the AMF colonization and distribution in four semi-mangrove plant communities were investigated. Typical AMF hyphal, vesicle and arbuscular structures were commonly observed in all the root samples, indicating that AMF are important components on the landward fringe of mangrove habitats. AMF spores were extracted from the rhizospheric soils, and an SSU rDNA fragment from each spore morph-type was amplified and sequenced for species identification. AMF species composition and diversity in the roots of each semi-mangrove species were also analyzed based on an SSU-ITS-LSU fragment, which was amplified, cloned and sequenced from root samples. In total, 11 unique AMF sequences were obtained from spores and 172 from roots. Phylogenetic analyses indicated that the sequences from the soil and roots were grouped into 5 and 14 phylotypes, respectively. AMF from six genera including Acaulospora, Claroideoglomus, Diversispora, Funneliformis, Paraglomus, and Rhizophagus were identified, with a further six phylotypes from the Glomeraceae family that could not be identified to the genus level. The AMF genus composition in the investigated semi-mangrove communities was very similar to that in the intertidal zone of this mangrove ecosystem and other investigated mangrove ecosystems, implying possible fungal adaptation to mangrove conditions.     

Interactive influence of light intensity and soil fertility on root-associated arbuscular mycorrhizal fungi

Background and aims

Soil nutrients and light have major effects on the economics of arbuscular mycorrhizal (AM) symbioses. This study tests the main and interactive effects of soil fertility and light on AM fungal community.

Methods

We conducted a 3 year mesocosm experiment with a full two factorial design: light (full light or shade) and soil fertility (unfertilized or fertilized), on the Qinghai-Tibetan Plateau. Plant traits, soil characteristics and the AM fungal communities inside roots and in soils were measured.

Results

Shade reduced AM colonization of roots, fertilization reduced the hyphal abundance in the soil, and both factors reduced species richness of AM fungi inside plant roots. Fertilization exacerbated the negative impacts of shade on AM fungal abundance and diversity. We observed 15 phylotypes of AM fungi inside roots and ten morphotypes of AM fungal spores in the soil. Taxa responded differently to shade and fertilization and there was little congruence between the responses of fungi inside the roots and in the spore community.

Conclusions

Our findings indicate that both shade and fertilization reduce the abundance of AM fungi, but the two factors have different effects on the quality of plant roots as habitat for AM fungi.     

Arbuscular mycorrhizal and dark septate endophytic fungi at 5,500 m on a glacier forefront in the Qinghai-Tibet Plateau, China

Arbuscular mycorrhizal (AM) and dark septate endophytic (DSE) fungi are rarely studied at extremely high elevations. Here, AM and DSE colonization in two dominant plant species (Melandrium apetalum and Poa litwinowiana) were microscopically observed on the forefront of Zhadang Glacier (5,500 m above sea level) in the Qinghai-Tibet Plateau, China. The AM fungal taxa were also identified by molecular methods. Both AM and DSE fungi synchronously colonized these two plant species, but AM dominated in M. apetalum and DSE dominated in P. litwinowiana. A total of five AM fungal spore morphotypes (Acaulospora capsicula, Diversispora sp., Glomus constrictum, G. eburneum and Glomus sp.) were found in the rhizosphere soils. Molecular identification revealed two AM fungal phylotypes: one Claroideoglomus phylotype from roots and one Diversispora phylotype from spores. These results extend the elevation at which both AM and DSE are known to occur.     

Sharing of Diverse Mycorrhizal and Root-Endophytic Fungi among Plant Species in an Oak-Dominated Cool–Temperate Forest

Most terrestrial plants interact with diverse clades of mycorrhizal and root-endophytic fungi in their roots. Through belowground plant–fungal interactions, dominant plants can benefit by interacting with host-specific mutualistic fungi and proliferate in a community based on positive plant–mutualistic fungal feedback. On the other hand, subordinate plant species may persist in the community by sharing other sets (functional groups) of fungal symbionts with each other. Therefore, revealing how diverse clades of root-associated fungi are differentially hosted by dominant and subordinate plant species is essential for understanding plant community structure and dynamics. Based on 454-pyrosequencing, we determined the community composition of root-associated fungi on 36 co-occurring plant species in an oak-dominated forest in northern Japan and statistically evaluated the host preference phenotypes of diverse mycorrhizal and root-endophytic fungi. An analysis of 278 fungal taxa indicated that an ectomycorrhizal basidiomycete fungus in the genus Lactarius and a possibly endophytic ascomycete fungus in the order Helotiales significantly favored the dominant oak (Quercus) species. In contrast, arbuscular mycorrhizal fungi were generally shared among subordinate plant species. Although fungi with host preferences contributed to the compartmentalization of belowground plant–fungal associations, diverse clades of ectomycorrhizal fungi and possible root endophytes were associated not only with the dominant Quercus but also with the remaining plant species. Our findings suggest that dominant-ectomycorrhizal and subordinate plant species can host different subsets of root-associated fungi, and diverse clades of generalist fungi can counterbalance the compartmentalization of plant–fungal associations. Such insights into the overall structure of belowground plant–fungal associations will help us understand the mechanisms that facilitate the coexistence of plant species in natural communities.     

Mycorrhizal associations in woody plant species at the Mt. Usu volcano,Japan

We investigated the association between ectomycorrhizal (ECM) and arbuscular mycorrhizal (AM) fungi and pioneer woody plant species in areas devastated by the eruption of Mt. Usu, Japan, in 2000. We observed eight woody plant species at the research site, most of which were associated with ECM and/or AM fungi. In particular, dominant woody plant species Populus maximowiczii, Salix hultenii var. angustifolia and Salix sachalinensis were consistently associated with ECM fungi and erratically associated with AM fungi. We found one to six morphotypes in the roots of each ECM host and, on average, two in the roots of each seedling, indicating low ECM fungal diversity. ECM colonization ranged from 17 to 42% of root tips. Using morphotyping and molecular analyses, 15 ECM fungi were identified. ECM fungi differed greatly between hosts. However, Laccaria amethystea, Hebeloma mesophaeum, Thelephora terrestris and other Thelephoraceae had high relative colonization, constituting the majority of the ECM colonization in the roots of each plant species. These ECM fungi may be important for the establishment of pioneer woody plant species and further revegetation at Mt. Usu volcano.     

Fungi associated with aeroponic roots in caves and mines of New Brunswick

We provide the first analysis of the fungi associated with a very special habitat, the aeroponic roots found in caves and mines in New Brunswick, Canada. Fungal diversity was assessed by Illumina sequencing using three complementary primer sets targeting ribosomal RNA genes, and roots were identified using the non-coding trnH-psbA spacer. Early colonizing ectomycorrhizal fungi such as Agaricales, Helotiales, Pezizales, and Thelephorales were predominant. Saprotrophs, endophytes and plant pathogens were also present, but Glomeromycota (arbuscular mycorrhizal fungi) were not detected. Fungal root communities were generally most similar within sites. Fungal diversity was inversely correlated with winter dark zone temperatures and distance from the entrance. By using a combination of three primer sets, we detected more fungal taxa than with any one primer set. This study adds to the understanding of these subterranean ecosystems and suggests that future studies investigate factors limiting the presence of late-stage ectomycorrhizal fungi and Glomeromycota.     

Ericoid mycorrhizal colonization and associated fungal communities along a wetland gradient in the Acadian forest of Eastern Canada

Wetlands provide numerous ecosystem services, and ericaceous plants are important components of these habitats. However, the ecology of fungi associated with ericaceous roots in these habitats is poorly known. To investigate fungi associated with ericaceous roots in wetlands, ericoid mycorrhizal colonization was quantified, and fungal communities were characterized on the roots of Gaultheria hispidula and Kalmia angustifolia along two upland – forested wetland transects in spring and fall. Ericoid mycorrhizal colonization was significantly higher in the wetlands for both plant species. Both upland and wetland habitats supported distinct assemblages of ericaceous root associated fungi including habitat specific members of the genus Serendipita. Habitat was a stronger driver of ericoid mycorrhizal colonization and ericaceous root associated community composition than host or sampling season, with differences related to soil water content, soil nutrient content, or both. Our results indicate that ericaceous plant roots in forested wetlands are heavily colonized by habitat specific symbionts.