Orchid seeds generally rely on mycorrhizal fungi for successful germination and development, but communication between fungal exudates and orchid seeds before contact is poorly understood. The researchers used G. conopsea and GS2, a strain originally isolated from the orchid’s roots, to investigate whether fungal products alone could initiate host responses. The fungus was grown in liquid culture, and its filtered exudates were freeze-dried, concentrated 60-fold, sterilized and applied to surface-disinfected seeds for 18 days. Sterile water served as the control.

The study combined RNA sequencing with four biological replicates, untargeted metabolomics of the exudates, hormone measurements with five replicates, fatty acid profiling with five replicates and transmission electron microscopy. RNA sequencing identified 5,172 significantly upregulated and 18,051 significantly downregulated genes in treated seeds relative to controls. Upregulated genes were enriched in starch and sucrose metabolism, fatty acid biosynthesis and elongation, plant hormone signal transduction, brassinosteroid biosynthesis and zeatin biosynthesis. Among the induced genes were fatty acid-associated genes including PKp, KASII, KAR, FatB and RAM2, several brassinosteroid-biosynthesis genes, and the cytokinin-related genes IPT and CYP735A.

Targeted hormone analysis found that brassinolide content in treated seeds reached 14 times the control level. Several bioactive cytokinins also increased, including trans-zeatin-riboside, cis-zeatin-riboside, cis-zeatin and N6-(Δ2-isopentenyl)adenosine, with cis-zeatin-riboside measured at 4.7 times the control value. Three bioactive gibberellins—GA3, GA4 and GA7—were detected, with GA7 accounting for more than 99.9% of total gibberellins and increasing significantly after treatment. Gibberellin regulatory genes were induced and abscisic-acid signaling was repressed, patterns the authors associated with possible release of seed dormancy. These measurements show associations with exudate exposure; they do not prove that every altered pathway was required for fungal colonization.

Separate experiments using GS2 during symbiotic germination provided functional evidence for a role for brassinosteroids and cytokinins. Exogenous 24-epibrassinolide at 5 µmol L-1 increased seed germination by 3.0 percentage points, whereas the brassinosteroid-biosynthesis inhibitor brassinazole reduced it by 4.7 points and significantly reduced the number of colonized cells in protocorms. Exogenous 6-benzylaminopurine at 10 µmol L-1 increased germination by 5.7 points, while zeatin at 1 and 5 µmol L-1 increased it by 4.6 and 6.7 points, respectively. Lovastatin did not significantly alter germination rate but reduced protocorm dimensions in a concentration-dependent manner; at 50 µmol L-1, protocorm length and width fell by 29.5% and 28.8%, respectively.

Exudate exposure was also associated with increased lipid accumulation. Transmission electron microscopy showed a greater area ratio of lipid droplets in treated seeds, and quantitative gas chromatography-mass spectrometry confirmed higher fatty acid content than in controls. The induction of RAM2 and FatB occurred alongside this increase. During symbiotic germination, fatty acid accumulation was observed before GS2 colonization, which began at 12 days post-symbiosis, while key components of the common symbiosis signaling pathway—CASTOR, POLLUX/DMI1, CCaMK/DMI3 and CYCLOPS/IPD3—were induced by day 3 and remained highly expressed after colonization. The authors therefore proposed that the pathway may mediate some exudate-induced changes, describing this as a potential mechanism rather than a demonstrated complete signaling chain.

Evolutionary comparisons showed that the orchid retained RAM2 and other components of the common symbiosis signaling pathway, whereas the fully mycoheterotrophic orchids Neottia fugongensis and Gastrodia elata lacked those genes in the analysis. The gene FatM and the arbuscular-mycorrhiza-specific genes RAM1, STR and STR2 were not detected in orchid-mycorrhizal hosts. GcRAM2 expression was approximately 25 times higher in fungus-colonized adult orchid roots than in non-colonized roots, and in situ hybridization localized its expression to GS2-colonized protocorm cells. In a cross-species functional test, expression of GcRAM2 restored substantial colonization by Rhizophagus irregularis in roots of the Medicago truncatula ram2 mutant, supporting conservation of its fatty acid-related function.

Exudates from GS2, five additional Ceratobasidium strains and a Tulasnella strain induced root-hair branching in M. truncatula. This assay is consistent with the presence of bioactive mycorrhizal factors, potentially lipo-chitooligosaccharides, but the researchers did not identify or structurally characterize those compounds. Untargeted profiling found that GS2 exudates contained 979 lipid or lipid-like metabolites, 673 organoheterocyclic compounds and 591 organic acids and derivatives, with fatty acyls the largest lipid category at 39.63%.

The findings remain limited to one orchid species, selected fungal strains and controlled laboratory seed treatments. Treated seeds entered a pre-germination stage but did not fully germinate, possibly because starch, free sugars or a continuous supply of fungal metabolites was insufficient. The specific fungal molecules that trigger the observed responses, their receptors and their interaction with hormone signaling remain unresolved. The study therefore provides evidence that fungal exudates can alter orchid seed transcription, hormone levels and fatty acid content before physical fungal contact, while presenting hypotheses rather than demonstrated mechanisms for later symbiotic germination and conservation applications.

Subject of Research: Pre-symbiotic responses of Gymnadenia conopsea seeds to Ceratobasidium sp. GS2 exudates

Article Title: Ceratobasidium sp. GS2 exudates elicit downstream transcriptional and physiological responses in seeds of Gymnadenia conopsea ( Orchidaceae )

Article References: Wang, Y., Qian, X., Li, J., Wang, H., Yang, A., Yang, L., Liu, J., & Xing, X. (2026). Ceratobasidium sp. GS2 exudates elicit downstream transcriptional and physiological responses in seeds of Gymnadenia conopsea (Orchidaceae). IMA Fungus, 17, Article e191381. https://doi.org/10.3897/imafungus.17.191381

Image Credits: AI Generated

DOI: 10.3897/imafungus.17.191381

Keywords: Gymnadenia conopsea, Ceratobasidium, orchid seed germination, fungal exudates, mycorrhizal symbiosis, gene expression, phytohormones, fatty acid biosynthesis, common symbiosis signaling pathway, plant–fungus communication, exudates, elicit

The study was conducted at the MycoPhylo experimental field in southeastern Estonia, a site specifically designed to integrate plant phylogenetic diversity with various mycorrhizal types. The researchers sampled leaf tissue from 158 plots representing 110 distinct plant species, ensuring that all plants were of similar age and exposed to comparable edaphic and microclimatic conditions. This controlled environment allowed the scientists to test specific hypotheses regarding the influence of host phylogeny, mycorrhizal associations, and leaf phenotypic characters on foliar fungal assemblages. The experimental setup included trees, shrubs, and herbaceous plants, providing a broad representation of plant growth forms.

Using a metabarcoding approach targeting the Internal Transcribed Spacer (ITS) region of the fungal ribosomal RNA, the team generated a high-resolution profile of the fungal communities present on the leaves. The sequencing data revealed that the fungal communities were dominated by the classes Dothideomycetes, Tremellomycetes, and Taphrinomycetes. Functional guild analysis indicated that plant pathogens and saprotrophs were the most abundant ecological groups within these assemblages. The high level of taxonomic diversity observed underscores the complexity of the phyllosphere and the importance of understanding the mechanisms that regulate microbial colonization in this exposed habitat.

The statistical analysis revealed that plant growth form and mycorrhizal association were significant drivers of foliar fungal alpha diversity and community composition. Plant phylogenetic eigenvectors accounted for up to 25.8% of the variation in fungal richness, indicating that evolutionary history plays a substantial role in determining which fungi can successfully colonize a host. Specifically, plants belonging to certain families, such as Grossulariaceae and Cistaceae, harbored relatively higher fungal richness compared to hosts from families like Lauraceae and Betulaceae. This phylogenetic signal suggests that closely related plant species may share similar microbial communities due to conserved leaf traits or shared evolutionary pressures.

Mycorrhizal type emerged as another key factor influencing fungal community structure, although the effect sizes were modest. Ectomycorrhizal plants were notably richer in Taphrinomycetes and Pucciniomycetes compared to arbuscular mycorrhizal and ericoid mycorrhizal hosts. This finding suggests that the belowground symbiotic relationship between the plant and mycorrhizal fungi may indirectly affect the aboveground microbial community, possibly through changes in leaf nutrient allocation or defense chemistry. However, the study noted that mycorrhizal type is often correlated with other host traits, making it difficult to disentangle the direct effects of the symbiosis from the effects of plant taxonomy and growth form.

Leaf functional traits, such as thickness and hairiness, showed a weaker but still significant influence on fungal richness. Thinner-leaved species generally harbored more fungal operational taxonomic units than thick-leaved species, a pattern consistent with previous studies suggesting that tougher leaf structures may limit fungal proliferation. Leaf hairiness, or the presence of trichomes, also had a minor effect on fungal richness, potentially due to the production of chemical compounds by the hairs that inhibit fungal colonization. Despite these correlations, the overall impact of these morphological traits on community composition was less pronounced than that of plant phylogeny and growth form.

The relationship between host phylogeny and fungal community dissimilarity was further examined using Mantel tests, which revealed weak but guild-dependent associations. While total fungal and saprotroph communities showed a positive relationship with host phylogenetic distance, plant pathogenic fungi did not exhibit a significant phylogenetic signal. This distinction highlights the different ecological strategies employed by various fungal groups; pathogens may be more influenced by specific host species identity and immune responses, whereas saprotrophs may be more responsive to broader phylogenetic and structural traits of the host plant. These findings suggest that the assembly of foliar fungi is a complex process involving multiple interacting factors.

The study also found that plant deciduousness significantly affected fungal community composition, even though it did not influence fungal richness. Deciduous plants harbored higher proportions of certain fungal classes, such as Tremellomycetes and Taphrinomycetes, compared to evergreen plants. This difference may be attributed to the structural and chemical properties of deciduous leaves, which are often thinner and shorter-lived, making them more susceptible to fungal colonization. Evergreen leaves, with their thicker cuticles and longer lifespan, may provide a more stable but less permissive environment for many fungal taxa. These results emphasize the importance of considering leaf phenology when studying phyllosphere microbial communities.

Overall, the findings indicate that host phylogeny and plant functional traits contribute to, but do not solely determine, the structure of foliar fungal assemblages under field conditions. The integration of host evolution with functional ecology provides a robust framework for understanding phyllosphere mycobiome variation. By identifying the specific plant characteristics that drive fungal diversity, this research offers valuable insights for managing plant health and resilience in managed ecosystems. Future studies should focus on the temporal dynamics of these communities and the causal mechanisms linking belowground mycorrhizal interactions to aboveground microbial assembly.

Subject of Research: Drivers of foliar fungal community composition in a common garden experiment

Article Title: Impact of plant species, mycorrhizal type, and leaf traits on foliar fungal communities (in a common garden experiment)

Article References: Ariyan, M., Mikryukov, V., Khalil, H., Gohar, D., Hosseyni Moghaddam, M. S., Drenkhan, R., & Tedersoo, L. (2026). Impact of plant species, mycorrhizal type, and leaf traits on foliar fungal communities (in a common garden experiment). IMA Fungus, 17, Article e173358. https://doi.org/10.3897/imafungus.17.173358

Image Credits: AI Generated

DOI: 10.3897/imafungus.17.173358

Keywords: foliar fungi, plant microbiome, mycorrhizal symbiosis, phyllosphere, plant phylogeny, leaf traits, metabarcoding, fungal diversity, common garden, ecological drivers, Impact, plant

Mycorrhizal symbioses are among the oldest alliances in biology. Nearly all land-forming trees and plants rely on fungi that colonize their roots, extending the reach of the root system into the soil and trading minerals and water for sugars produced by photosynthesis. Ecologists divide these partnerships into broad functional groups. Arbuscular mycorrhizal fungi, often abbreviated AM, penetrate the cells of the root cortex and are ancient partners of flowering plants. Ectomycorrhizal fungi, by contrast, wrap themselves around root tips without penetrating cell walls and dominate among conifers and many trees of the oak, birch, and beech families. The two symbioses come with different nutritional economies: AM fungi are generally thought to be less selective foragers that deliver nitrogen-rich nutrients quickly, while ectomycorrhizal fungi excel at mining organic nitrogen directly from litter and soil organic matter. Which symbiosis dominates a forest therefore shapes how fast carbon cycles, how much carbon stays locked in soils, and which seedlings can establish beneath the canopy.

Because of these links to carbon and nutrient cycling, ecologists have long wanted to know whether the balance between mycorrhizal types is stable or whether it shifts with environmental change. Individual plots have shown responses to nitrogen pollution, drought, and warming, but evidence for a coherent, archipelago-wide reorganization has been scarce. The new analysis of forests across Japan provides exactly that: a broad-scale signal that the relative dominance of arbuscular mycorrhizal trees is changing along the length of the country, and that the change is not random.

The study’s central finding is that AM-associated trees are expanding their hold on Japanese forests, and that two explanatory threads run through the pattern. The first is disease. Tree diseases, driven by fungi, oomycetes, and insect vectors that thrive in a warming world, do not strike all tree species equally. Species that harbor ectomycorrhizal partnerships and species that harbor AM partnerships differ in their susceptibility, their rates of recovery, and their competitive ability following damage. When pathogens and pests remove or weaken particular canopy trees, the species that recruit into the gaps may disproportionately belong to the AM group, tipping the local balance and, cumulatively, the regional one. The second thread is climate warming itself. As temperatures rise, the climatic envelopes that once favored cool-adapted, ectomycorrhizal-rich forests in northern and montane Japan are shifting toward conditions that favor warm-adapted, AM-rich communities. Warming thus acts both directly, by altering which species can tolerate the local climate, and indirectly, by amplifying the diseases that thin the canopy.

What makes the result scientifically important is its breadth. Surveys and forest inventories distributed along the Japanese archipelago, which spans a remarkable gradient of climate from subtropical Ryukyu islands to boreal-influenced Hokkaido, revealed a coherent geographic pattern rather than a scatter of local anomalies. The archipelago functions in the study as a natural laboratory: because it is elongated along a latitudinal and thermal gradient, it allows researchers to ask whether the composition of forests is tracking climate in the way theory predicts. The answer is that it is, but with an added twist. The shift in mycorrhizal dominance is not simply a by-product of species moving poleward; it is entangled with the dynamics of disease, which can accelerate, redirect, or amplify the compositional change that warming alone would produce.

The mechanism by which disease and mycorrhizal type interact is an active area of research, and the Japanese findings add an important macroecological perspective to it. Mycorrhizal fungi do more than feed their hosts; they influence host defense, drought tolerance, and seedling survival. Ectomycorrhizal networks can support seedling establishment under parent trees, while AM associations often favor rapid growth and fast nutrient acquisition. A forest in which pathogens selectively remove ectomycorrhizal trees may therefore experience a cascade: fewer ectomycorrhizal adults mean fewer ectomycorrhizal propagules in the soil, less supportive fungal networks for the remaining regeneration, and an increasingly favorable environment for AM seedlings that thrive on disturbed, nutrient-flushed soils. Disease, in other words, can act as a ratchet, converting temporary losses into durable shifts in symbiotic identity.

Climate warming feeds this ratchet in several ways. Warmer winters fail to kill off insects and pathogens that cold once suppressed, extending their active seasons and geographic ranges. Warmer, sometimes drier summers stress trees, making them more vulnerable to attack. Extreme events such as typhoons, which regularly strike Japan, create large areas of disturbed forest in which fast-growing, disturbance-adapted species, many of them AM-associated, gain a foothold. Each of these processes has been documented in local studies; what the new research contributes is evidence that their combined effect is visible at the scale of the entire archipelago, registered in the shifting balance between mycorrhizal types.

The consequences of such a shift extend well beyond the identity of the trees themselves. Because AM and ectomycorrhizal forests differ in how they process nitrogen and store carbon, a wholesale conversion of forest symbiosis has implications for ecosystem function. Ectomycorrhizal-dominated forests are often associated with slower decomposition and greater storage of carbon in soil organic matter, partly because their fungi produce compounds that slow the breakdown of litter and because their nitrogen-mining strategy can suppress decomposer microbes. AM-dominated forests, in contrast, tend toward faster nutrient cycling, faster decomposition, and soils in which carbon is more exposed to microbial attack. A broad-scale transition from ectomycorrhizal toward AM dominance could therefore reduce the capacity of forest soils to lock away carbon, creating a feedback that adds to, rather than offsets, the warming that triggered the shift in the first place. The researchers emphasize that this is a hypothesis grounded in the established functional differences between the two symbioses, and that verifying the magnitude of any carbon feedback will require long-term monitoring of soils alongside vegetation.

There are also implications for biodiversity and forest management. The species that make up the AM and ectomycorrhizal pools differ in their economic and cultural value, in the wildlife they support, and in their responses to silvicultural treatment. Foresters in Japan have long managed stands of sugi, hinoki cypress, and other conifers, many of which rely on ectomycorrhizal partnerships, while broadleaved evergreens of the warm-temperate forests are predominantly AM-associated. A shift toward AM dominance would alter regeneration dynamics, the incidence of certain pests, and the suitability of land for different management objectives. Understanding the disease component of the shift gives managers an actionable lever: reducing pathogen spread, diversifying plantations, and protecting resistant genotypes could slow the conversion and buy time for adaptation.

The study also speaks to a growing recognition that global change operates through interactions rather than single causes. Warming alone would move species ranges; disease alone would reshape forests locally; but together, as the Japanese data show, they can produce a coordinated, archipelago-scale reorganization of one of the fundamental functional axes of forest ecosystems. For scientists modeling the future of the biosphere, the lesson is that predicting vegetation change requires tracking not only temperature and rainfall but also the health of the trees and the hidden fungal partnerships beneath their roots. For the forests of Japan, the finding is a warning and an opportunity in equal measure: the symbiotic identity of the woods is changing, and the window for understanding, anticipating, and perhaps guiding that change is open now, while the process is still measurable and, possibly, still manageable.

As monitoring continues, the Japanese archipelago will remain a bellwether. Its steep environmental gradients, rich forest flora, and dense long-term observational infrastructure make it one of the best places on Earth to watch the interplay of climate, disease, and symbiosis unfold in real time. The evidence assembled in this study indicates that the shift already underway is broad, structured, and driven by identifiable forces, and it establishes a baseline against which the forests of the coming decades will be judged.

Subject of Research: Climate warming and tree diseases driving a broad-scale shift in mycorrhizal tree dominance across Japanese forests

Article Title: Broad-scale shift in dominance of arbuscular mycorrhizal trees along the Japanese archipelago associated with tree diseases and climate warming

Article References: Schaefer, H., Yamashita, N., Hashimoto, S., Inagaki, Y., Kawanishi, A., Chatani, S., Shimadera, H., Furusawa, H., & Imaya, A. (2026). Broad-scale shift in dominance of arbuscular mycorrhizal trees along the Japanese archipelago associated with tree diseases and climate warming. Nature Communications. https://doi.org/10.1038/s41467-026-77711-w

Image Credits: AI Generated

DOI: 10.1038/s41467-026-77711-w

Keywords: arbuscular mycorrhiza, ectomycorrhizal fungi, Japanese forests, climate warming, tree diseases, forest ecology, mycorrhizal symbiosis, carbon cycling, soil fungi, forest composition, biogeography, Nature Communications