Mycorrhizal and saprotrophic macromycetes contribute strongly to the carbon and nitrogen cycles of forest ecosystems, often studied by tracing stable isotope composition of carbon and nitrogen. The phenomenon of the saprotrophic-mycorrhizal divide highlights the difference in the stable isotope composition of fruiting bodies of mycorrhizal and saprotrophic fungi. Much less is known about the isotopic composition of the mycelium, which plays an important role in the formation of the soil organic matter and fuels the fungal trophic channel in soil food webs. In this study, we assessed whether the saprotrophic-mycorrhizal divide in the natural δ13С and δ15N values can be traced throughout entire fungal organisms. This hypothesis was tested using 16 species of ectomycorrhizal and six species of saprotrophic basidiomycetous fungi. We showed that not only fruiting bodies, but also the mycelium of ectomycorrhizal and saprotrophic fungi differs in the δ13C and δ15N values. In both ectomycorrhizal and saprotrophic fungi, the δ13C and δ15N values increased from mycelium to hymenophores and correlated positively with the total N content in the corresponding tissues. The differences between ectomycorrhizal and saprotrophic mycelium can be used to reconstruct the fungal-driven belowground carbon and nitrogen allocation, and the contribution of saprotrophic and mycorrhizal fungi to soil food webs.

Mycoheterotrophic plants, which depend entirely on mycorrhizal fungi for carbon acquisition, often exhibit high specificity toward their fungal partners. Members of Thismiaceae are generally recognized for their extreme mycorrhizal specialization and rarity. In this study, we examined the mycorrhizal associations of Relictithismia, a recently discovered monotypic genus within Thismiaceae, and Thismia abei, a Thismia species with a similar distribution in southern Japan, by employing high-throughput DNA sequencing of the 18S rRNA gene. Our analyses revealed that both R. kimotsukiensis and T. abei are predominantly associated with two specific virtual taxa (VTX00295 and VTX00106) of the genus Rhizophagus (Glomeraceae). These shared associations may reflect either phylogenetic niche conservatism, in which the common ancestor of R. kimotsukiensis and T. abei retained the same AM fungal partners, or convergent evolution, in which the AM fungal phylotypes were independently recruited due to their potential benefits for these mycoheterotrophic plants. Furthermore, BLAST searches demonstrated that VTX00295 and VTX00106 are widely distributed globally, suggesting that highly specialized mycorrhizal interactions are unlikely to be the primary drivers of the limited distribution and rarity of R. kimotsukiensis and T. abei. Overall, our findings enhance our understanding of high mycorrhizal specificity in Thismiaceae. However, broader investigations, combining extensive sampling of Thismiaceae species with ancestral state reconstruction, are needed to determine whether the shared associations detected here reflect phylogenetic niche conservatism or convergent evolution.

Mycorrhizal associations play a crucial role in afforestation efforts, as they enhance the acquisition of nutrients and water, thereby supporting seedling establishment. However, the influence of nitrogen (N) forms in the soil, particularly the organic N, on the formation of mycorrhizal associations and their subsequent effects on seedling morpho-physiology remains poorly understood. In this study, we examine the mycorrhizal colonization, along with morpho-physiological and functional traits, in Pinus cooperi seedlings following fertilization with organic N in controlled nursery conditions. A factorial experiment was performed with Pinus cooperi C. E. Blanco seedlings using two N sources: organic N (amino acids) and inorganic N (NH4NO3) and two N doses: low and high (60 vs 200 mg N seedling-1). Seedlings were inoculated with a mixture of native fungi, but the phylogenetic analysis showed that Suillus placidus (Bonord.) Singer was the only species colonizing roots. Organic N promoted similar morphology and nutritional status as inorganic N, though at a low N rate, it improved root growth and mycorrhizal colonization. High N fertilization improved seedling growth and nutritional status but reduced mycorrhizal colonization. Mycorrhizal colonization improved needle P concentration, delayed plant desiccation, and reduced root cellular damage when seedlings were subjected to desiccation, though it decreased plant growth and needle N concentration. We conclude that organic N fertilization improves mycorrhization of P. cooperi with S. placidus, but the fertilization dose should be adjusted to meet species-specific requirements in order to optimize plant quality and promote afforestation success.

The normal development of mycorrhizal symbiosis is a dynamic process, requiring elaborately regulated interactions between plant roots and compatible fungi, mandatory for both partners´ survival. In the present study, we further elucidated the mycorrhizal development of the desert truffles Terfezia claveryi with the host plant Helianthemum almeriense as an ectendomycorrhizal symbiosis model under greenhouse conditions. To investigate this, we evaluated the morphology of mycorrhizal colonization, concomitantly with the dynamic expression of selected marker genes (6 fungal and 11 plant genes) measured every week until mycorrhiza maturation (three months). We were able to determine 3 main stages in the mycorrhization process, 1) pre-symbiosis stage where mycelium is growing in the soil with no direct interaction with roots, 2) early symbiosis stage when the fungus spreads along the roots intercellularly and plant-fungal signaling is proceeding, and 3) late symbiosis stage where the fungus consolidates and matures with intracellular hyphal colonization; this is characterized by the regulation of cell-wall remodeling processes.

Robusta coffee, grown by 25million farmers across more than 50 countries, plays an important role in smallholder farmers' livelihoods and the economies of many low-income countries. Coffee establishes a mutualistic symbiosis with arbuscular mycorrhizal fungi (AMF); however, the impact of agricultural practices and soil characteristics on AMF diversity and community composition is not well understood. To address this, we characterised the AMF community composition of robusta coffee in part of its region of origin, the Democratic Republic of Congo. AMF diversity and community composition were compared between coffee monoculture, agroforestry systems and wild robusta in its native rainforest habitat. Using Illumina sequencing on 304 root samples, we identified 307 AMF operational taxonomic units (OTUs), dominated by the genera Glomus and Acaulospora. OTU richness did not vary across the three studied systems, yet large differences in community composition were found. Many unique OTUs were only observed in the coffee in the rainforest. In general, lower available soil phosphorus (P) and lower soil bulk density increased AMF diversity, yet higher available soil P and pH increased AMF diversity in the wild forest coffee. Shifts in AMF community composition across coffee systems were driven by canopy closure, soil pH, available soil P and soil bulk density. Our study is the first to characterise mycorrhizal communities in wild robusta coffee in its region of origin and shows that even low-input agricultural practices result in major AMF community shifts as compared to a natural baseline.

Like other plant-microbe symbioses, the establishment of orchid mycorrhiza (ORM) is likely to require specific communication and metabolic adjustments between the two partners. However, while modulation of plant and fungal metabolism has been investigated in fully established mycorrhizal tissues, the molecular changes occurring during the pre-symbiotic stages of the interaction remain largely unexplored in ORM. In this study, we investigated the pre-symbiotic responses of the ORM fungus Tulasnella sp. SV6 to plantlets of the orchid host Serapias vomeracea in a dual in vitro cultivation system. The fungal mycelium was harvested prior to physical contact with the orchid roots and the fungal transcriptome and metabolome were analyzed using RNA-seq and untargeted metabolomics approaches. The results revealed distinct transcriptomic and metabolomic remodelling of the ORM fungus in the presence of orchid plantlets, as compared to the free-living condition. The ORM fungus responds to the presence of the host plant with a significant up-regulation of genes associated with protein synthesis, amino acid and lipid biosynthesis, indicating increased metabolic activity. Metabolomic analysis supported the RNA-seq data, showing increased levels of amino acids and phospholipids, suggesting a remodelling of cell structure and signalling during the pre-symbiotic interaction. In addition, we identified an increase of transcripts of a small secreted protein that may play a role in early symbiotic signalling. Taken together, our results suggest that Tulasnella sp. SV6 may perceive information from orchid roots, leading to a readjustment of its transcriptomic and metabolomic profiles.

The community structure of ectomycorrhizal (ECM) fungi typically displays temporal dynamics. However, heavy snow cover hinders belowground investigations in temperate-to-boreal forests where ECM trees dominate, and the dynamics of the ECM fungal community structure during winter have not been fully elucidated. Given that boreal conifer species start root production in response to snowmelt, studies on the response of the ECM fungal community to snowmelt are needed. In the present study, to infer the community dynamics during the snowmelt season and their susceptibility to host tree conditions, we investigated ECM fungi associated with saplings of the evergreen conifer Abies sachalinensis immediately after the start and end of snowmelt in a common garden experiment. Saplings derived from two sources of contrasting snowfall conditions (heavy vs. little) were grown under two different light conditions (open vs. shaded), and the ECM fungal community dynamics patterns were compared across these combinations. The response of the ECM fungal community structure varied across treatments; although significant loss of ECM fungal operational taxonomic units (OTUs) was observed when saplings from the heavy snowfall region were grown under shade conditions, no change in community structure across the snowmelt season was observed for the other combinations. The stability of community composition despite the change in abiotic conditions with snowmelt, together with the effects of host origin and light conditions on community dynamics patterns, would imply the importance of host-mediated community dynamics of ECM fungi during the snowmelt season.

It has been suggested that invasive plant species are more generalist than non-invasive species in their interactions with arbuscular mycorrhizal fungi (AMF), allowing them to associate with novel AMF communities. There is emerging evidence suggesting that the flavonoid quercetin may play a role in regulating these interactions as a signaling compound. In this study, we experimentally grew three invasive alien and three non-invasive native woody species with AMF communities collected from within (though foreign to invasives) and outside their current distribution ranges. After 96days, we: (a) assessed mycorrhizal colonization rates; (b) evaluated the impact of these interactions on plant performance (growth and phosphorus nutrition); and (c) tested whether these responses were influenced by the addition of quercetin to the plant growth medium. Our findings reveal that the invasive species exhibited mycorrhizal colonization when grown with both novel AMF communities and benefited from them in terms of phosphorus (P) nutrition. In contrast, two of the three non- invasivenative species showed mycorrhizal colonization and enhanced P nutrition only with AMF from their current distribution range, but not with novel AMF from outside their range, suggesting selective behavior in their mycorrhizal interactions. The addition of quercetin did not have a strong effect on mycorrhizal colonization in either invasive or non-invasivenative species. However, quercetin promoted moderate increases in P nutrition in the two non-invasive native species when grown with the novel AMF communities. Overall, the results suggest that invasive species are more generalist in their AM symbiosis than two of the three non-invasive species, and that the addition of quercetin had a limited, moderate influence on their AM interactions.

Some heavy metal tolerant fungal isolates capable of forming ericoid mycorrhiza can also confer increased metal tolerance to the host plant. One of these fungal isolates, Oidiodendron maius Zn, has been characterized and a few molecular mechanisms underlying its metal tolerant phenotype have been identified. Here, we investigate the genomic divergences between the available genome of O. maius Zn and the genomes of metal tolerant and sensitive isolates of O. maius, with the aim of identifying genes or intergenic regions possibly involved in the display of the tolerance. The resequenced genomes of 8 tolerant and 10 sensitive isolates were mapped on the reference, O. maius Zn, yielding 357 gene models from the reference that were either missing or too polymorphic to be identified in the genomes of the sensitive isolates. These regions included genes with functions related to defense mechanisms and with unknown functions. One third of the predicted gene models turned out to be highly polymorphic, including many enriched GO terms, i.e. DNA/RNA metabolism and modification, chromosome/chromatin organization, protein biosynthesis, metabolism and function, energy consumption/transfer and mitochondrion. Overall, our findings indicate that the tolerant phenotype in O. maius likely arises from multiple genetic adaptations rather than a singular mechanism.

Panax quinquefolius L, a medicinal plant of the family Araliaceae, has been used in China for more than 300years. The quality of its medicinal materials is a significant concern. Our previous studies have shown that arbuscular mycorrhizal fungi (AMF) promote the growth of P. quinquefolius and facilitate the accumulation of the active ingredient ginsenosides. However, these beneficial effects are limited by the low AMF colonization rate in production settings, requiring interventions to improve the colonization rate. Biochar is considered an effective soil amendment. Our preliminary experiments indicate that biochar can enhance the inter-root microecology of P. quinquefolius, as well as increase the AMF colonization rate, but the mechanism was not clear. Therefore, we propose using biochar to increase the AMF colonization rate. In this study, we explore the use of biochar to promote the AMF infestation rate of P. quinquefolius and its potential mechanisms. The mechanism was explored by setting up eight treatments. The colonization rate and intensity of AMF in P. quinquefolius roots were assessed using a Trypan Blue solution. Rhizosphere soil microorganisms were analyzed by 16S and ITS sequencing, and secondary metabolites were identified via non-targeted metabolomics. The results showed that the AMF and 2% biochar combined (AMF + BC2) treatment significantly increased both the colonization rate and colonization intensity of AMF, which were 53.58% and 195.95% higher than that of AMF, respectively. The colonization and rhizosphere AMF data indicate that the application of biochar promotes AMF colonization from outside to inside the root. In addition, biochar attracted potentially beneficial microorganisms such as Sphingobium, Sphingomonas, and Novosphingobium, which are positively correlated with AMF and promote AMF colonization. These microorganisms are closely linked with active secondary metabolites, such as Sphingobium, which is positively correlated with L-malic acid. In conclusion, biochar can improve the quality of P. quinquefolius by promoting the formation of mycorrhizae. This finding provides a theoretical basis for the observed effect of the co-application of biochar and AMF on the growth and active ingredient accumulation of P. quinquefolius.