Root nodule symbiosis allows legumes to meet their nitrogen requirements. However, the causes of symbiosis failure at high temperatures remain unknown. We demonstrated that elevated temperatures affec...

“Few researchers bridge fundamental science and applied innovation as seamlessly as Prof. Handelsman. Her work consistently yields ideas that are not only publishable but patentable. From a patent strategy standpoint, Handelsman’s research is a goldmine—her insights into soil microbiomes and antibiotic resistance offer clear pathways to novel compounds and methods with strong commercial viability. It was no surprise when one of her projects was selected as part of our Plant Sciences Challenge grant program, supported jointly by WARF and Bayer.”

John Yeley has spent the last 26 years working on his farm, which his family has owned since 1852.

Arbuscular mycorrhizal fungi (AMF) and soil microbiome form synergistic interactions that drive nutrient mobilisation, carbon sequestration, and climate resilience in terrestrial ecosystems, yet their integrated roles in the global carbon-energy interactions remain underexplored.​ The present study aims to present a comprehensive understanding of the interactions between microbes in soil ecosystems, emphasising how they can improve carbon sequestration and establish mitigation and adaptation strategies regarding climate change. This review, adhering to PRISMA guidelines, synthesised 155 studies from the Scopus database (2000-2026) and Quantitative synthesis shows AMF channel 4-20% of plant photosynthates into hyphal networks and glomalin-related soil proteins (GRSP), up to 27% of soil organic carbon (SOC), enhancing aggregation and long-term Carbon (C) storage, while soil microbes regulate 2.15 × 1021 J yr⁻1 of heterotrophic respiration, while microbial carbon use efficiency (0.3-0.6) controls C stabilization versus CO2 loss. AMF-microbiome synergies ampify nutrient mobilisation, microbial biomass carbon and belowground carbon fluxes. Ecosystem- specific analyses reveal consistent AMF functions, SOC accumulation in forests; plant diversity and aggregate stability in grasslands; improved nutrient use efficiency in agriculture; stress tolerance in alpine systems, and accelerated restoration in degraded lands. Climate stressors (warming, elevated CO2, drought, salinity) modulate these interactions, often enhancing short term AMF biomass but risking SOC losses via reduced carbon use efficiency. Critical gaps remain in multi-stressor field trials, molecular signalling, and bioinoculants scalability. Integrating AMF with soil microbiome offers a natural climate solution to stabilise SOC, reduce greenhouse gas emissions, and bolster food-energy security under climate change.

It has been shown that cold long-term storage of Entrophospora etunicata inoculum may decrease the concentration of spore proteins. However, no studies have been carried out to understand how long-term storage under cold conditions may interfere with glomalin-related soil proteins (GRSP) concentration, thermostable glycoproteins released by arbuscular mycorrhizal fungi (AMF). To elucidate how this shelf-life factor may modulate GRSP concentration, the same isolate of E. etunicata was cultivated in 2000 (Ee2000) and 2023 (Ee2023) and stored since then under cold conditions (4–6ºC). The easily extractable fraction of GRSP (EE-GRSP) was extracted from these inocula using 0.25g of soil-inoculum, equivalent weight for 50 spores in the soil-inoculum, and a spore suspension with 50 spores. The results were expressed as the concentration in µg for each approach evaluated and as the Carbon of GRSP (C-GRSP). In the treatment of equivalent weight for 50 spores, the EE-GRSP and C-GRSP concentrations were reduced by 35% in Ee2000 in comparison to Ee2023, indicating that such a reduced concentration of GRSP and C-GRSP may occur in stored inocula at a slow rate due to cold storage. A slight decrease (9.5%) in EE-GRSP and C-GRSP was observed in Ee2000 soil-inoculum from 0.25g treatment, and no difference in protein concentration was shown in the spore suspension. It is concluded that E. etunicata inoculum has its EE-GRSP and C-GRSP negatively affected by long-term storage in cold conditions. This is the first study to report the behaviour of GRSP in cold long-term stored AMF inoculum.

Root nodules are distinct organs assembled from ancient developmental components. A new study shows that rhizobial Nod factors activate a conserved auxin module controlling lateral root formation, revealing how symbiotic signals can recruit pre-existing programmes without reproducing lateral root development.

What are Frankia root nodules? Frankia root nodules are specialized nitrogen-fixing structures that form on the roots of woody angiosperms following colonization by soil bacteria of the genus Frankia (Figure 1). Within these nodules, Frankia converts atmospheric dinitrogen (N2) into ammonia — a form of nitrogen the plant can assimilate — in exchange for photosynthetically derived carbon. The result is a mutualism with considerable ecological consequence: actinorhizal symbioses contribute fixed nitrogen to soils at rates comparable to the well-studied legume–rhizobia system, yet they occur in ecosystems ranging from boreal forests, riparian systems to coastal dunes, where legumes are largely absent.

Root system architecture (RSA) is pivotal to plant nutrient acquisition and environmental adaptation. In recent years, peptide hormones—intercellular signaling molecules that act locally or systemically—have emerged as critical regulators of RSA. These hormones are recognized by specific receptor kinases, which transduce peptide signals by activating downstream pathways involving calcium fluxes, reactive oxygen species bursts, and mitogen-activated protein kinase cascades, or by directly modulating core signaling components. These signaling networks integrate endogenous developmental cues with exogenous environmental stimuli to fine-tune root growth and development, thereby shaping RSA plasticity. This review provides a systematic analysis of the essential roles of peptide hormones in regulating RSA plasticity, elucidates their associated molecular pathways, and critically assesses their potential to optimize crop RSA, improve nutrient use efficiency, and enhance stress resistance, thereby contributing to sustainable agriculture.

Strains of the arbuscular mycorrhizal (AM) fungus Rhizophagus irregularis belong to one of two nuclear organizations: homokaryons, which carry genetically similar nuclei, and dikaryons, which harbour two genetically distinct nuclear populations. Although traits tend to be conserved within each group, it is unclear whether their capacity to adapt or exhibit phenotypic plasticity in response to environmental change differs. Here, we tested whether dikaryotic strains have greater potential than homokaryotic strains for rapid adaptation or plasticity in response to shifts in soil phosphorus (P) availability. The experiment first included a conditioning phase in which we measured the growth responses of Allium ampeloprasum L. inoculated with four homokaryotic or four dikaryotic strains under contrasting soil P levels. Next, in the adaptive potential phase, we grew the conditioned strains under both low and high P to assess whether prior P exposure influenced subsequent mutualistic functioning. During conditioning, host biomass was greater under high than low P for both nuclear groups, but increased roughly 3.2-fold for dikaryons and 1.9-fold for homokaryons. In the adaptive potential phase, strains conditioned under low P enhanced host biomass significantly more than those conditioned under high P, and by a similar amount in both P environments. This effect did not differ between dikaryons and homokaryons, but its magnitude varied markedly among individual strains, from no benefit to a roughly two-fold increase in host biomass. Together, these results provide the first experimental evidence that conditioning AM fungi under low soil P can enhance their subsequent benefits to host plants, even when P availability increases. Soil P conditioning strategies for AM fungi, therefore, merit consideration in agriculture and other managed ecosystems, although the strong strain dependence of the effect implies that strain choice and conditioning regime would need to be optimized together.

“Few researchers bridge fundamental science and applied innovation as seamlessly as Prof. Handelsman. Her work consistently yields ideas that are not only publishable but patentable. From a patent strategy standpoint, Handelsman’s research is a goldmine—her insights into soil microbiomes and a...

Arbuscular mycorrhizal (AM) symbiosis is a widespread mutualism between plant roots and Glomeromycotina fungi that enables nutrient exchange through arbuscules. Although many transcriptional regulators of AM symbiosis have been identified, the role of the INDETERMINATE DOMAIN (IDD) proteins remain unknown. We show that rice IDD7 expression is strongly induced in arbuscule-containing cells, and its loss markedly reduces fungal colonization and disrupts arbuscule development. IDD7 is required for induction of genes associated with fatty acid biosynthesis, nutrient transport, and symbiotic signaling during AM symbiosis. IDD7 interacts with the transcription factors SLENDER RICE 1 (SLR1) and PHOSPHATE STARVATION RESPONSE 2 (PHR2) through its conserved TQDFLG domain. Electrophoretic mobility shift assays show that IDD7 binds multiple promoter motifs. Together with SLR1 and PHR2, IDD7 synergistically activates the promoters of PHOSPHATE TRANSPORTER 11 (PT11) and the AM-associated transcription factors PHR2, REQUIRED FOR ARBUSCULAR MYCORRHIZATION 1 (RAM1), WRINKLED 5a (WRI5a), and CYCLOPS. Moreover, IDD7 is required for PHR2 expression in arbuscule-containing cells. Here, we show that IDD7 is a central transcriptional regulator in arbuscule-containing cells.

Background Enhancing plant immunity often improves resistance to pathogens, yet its consequences for beneficial symbionts remain underexplored. Here, we assessed the effects of disrupting GmLOPP, a soybean type 2 C protein phosphatase, on both immune activation and rhizobial symbiosis. Results Loss-of-function Gmlopp mutants displayed enhanced resistance against both bacterial blight and pustule disease. Upon treatment with lipopolysaccharides (LPS), major components of the bacterial outer membrane, these mutants showed elevated reactive oxygen species (ROS) production and upregulated expression of GmOXI1 and GmWRKY33. Importantly, Gmlopp plants exhibited no detectable trade-offs in growth or nodulation compared with the wild type. Conclusion Collectively, our findings establish that GmLOPP knockout confers disease resistance without compromising rhizobial symbiosis, highlighting its potential as a gene-editing target for soybean breeding.

Under nitrogen limiting conditions, legume plants interact with nitrogen fixing bacteria known as rhizobia, resulting in the formation of a new organ, the nodule. This process is accompanied by dramatic changes in gene expression, which operate at different levels. A previous study revealed that histone methylation is differentially modulated during nodulation. However, the histone methyl transferases and demethylases involved in this modulation have not been characterized. In this study we report the identification of the Medicago truncatula putative histone lysine demethylase MtPKDM9B, which is subject to alternative splicing (AS), and the differential modulation of AS variants at translational level during nodule symbiosis. Knockdown of MtPKDM9B impaired infection by rhizobia, nodule development, bacterial viability and the expression of the leghemoglobin coding gene MtLHB1. MtPKDM9B is the putative ortholog of Arabidopsis EARLY FLOWERING 6 (ELF6/AtPKDM9B) gene involved in the removal of the repressive mark H2K27me3. A combination of ChIP-seq and RNA-seq experiments revealed that MtPKDM9B is required for demethylation of H3K27me3 in regions nearby or contained within gene bodies of symbiotic genes and the upregulation of the cognate mRNAs in response to rhizobia, including those encoding the putative ubiquitin ligase MtPUB2, the MYB transcription factor MtMYB040 and the auxin conjugating enzyme MtGH3 (Gretchen Hagen 3). Our findings illustrate how AS and translational regulation of this plant specific histone lysine demethylase contributes to the removal of the repressive mark H3K27me3, promoting transcriptional activation of symbiotic genes required for the formation of functional nitrogen fixing nodules.

The nitrogen-fixing symbiosis between legume plants and rhizobia represents one of the most remarkable examples of developmental plasticity in plants. Following the recognition of compatible rhizobia, legumes initiate the formation of root nodules. These specialized organs accommodate nitrogen-fixing bacteria and mediate the exchange of plant-derived carbon for bacterially fixed nitrogen (Roy et al. 2020). Nodule development has traditionally been viewed as a specific symbiosis process, but increasing evidence indicates that rhizobia-induced nodulation relies on pre-existing developmental programs controlling root growth, vascular organization, and organogenesis. How ancient developmental pathways have adapted to integrate and support symbiotic interactions remains one of the major unresolved questions in plant biology.

In order to identify the mechanisms underlying the formation of low-molecular-weight forms of acidic exopolysaccharide in the symbiotic bacteria Rhizobium leguminosarum bv. viciae, a comparative analysis of exo-oligosaccharide production was conducted in the wild-type strain and its derivatives with mutations in one (plyB), two (plyBC, pssWplyB), and three (pssWplyBC) glycanase genes. Based on viscometry data and quantitative analysis of polysaccharides, it was concluded that the mutants synthesized an exopolysaccharide with a higher degree of polymerization of repeating octasaccharide units compared to that in the wild-type strain. It was shown that knockout of glycanase genes led to a decrease in the amount of acidic oligosaccharides in the culture medium. It was established that the efficiency of polysaccharide depolymerization by extracellular polysaccharide lyases PlyBC depends on the presence of a functionally active periplasmic glycoside hydrolase PssW. Based on the results of mass spectrometric analysis of exo-oligosaccharides, a scheme for the location of polysaccharide cleavage sites by glycanases was proposed. It was shown that effectiveness of symbiosis with garden pea was reduced in the mutant strains plyBC and pssWplyBC.

Advances in understanding the evolutionary ecology of the rhizobia-legume mutualism have been constrained by methodological limitations in efficiently measuring relative strain frequencies alongside measurements of absolute population sizes of rhizobia living in nodules. To examine strain competition in natural and agricultural ecosystems that harbor multiple strains of rhizobia, an increasing number of manipulative and observational studies have recently begun to examine dozens or hundreds of strains simultaneously. Assessing the competitive fitness of multiple strains in legume nodules requires, first, processing pools of dozens to hundreds of nodules to overcome the stochasticity of nodule formation; second, focusing on the reproductively viable rhizobial population, since this trait represents rhizobia's reproductive success in nodules and is pivotal for evolutionary interpretations. Our approach has been optimized in the Medicago truncatula–Sinorhizobium meliloti system, where rhizobia induce the formation of indeterminate nodules that harbor two subpopulations: terminally-differentiated bacteroids and undifferentiated rhizobia that retain reproductive viability. This protocol has also been used for other legumes with terminally-differentiated bacteroids, such as pea and vetch, as well as for those with non-terminally differentiated bacteroids, such as soybean and cowpeas. The protocol we present enables rapid and reproducible homogenization of pools containing hundreds of nodules using a tissue homogenizer. We also enrich for undifferentiated rhizobia using two centrifugation steps: first, a low-speed centrifugation to deplete nodule debris and large, endoreduplicated, terminally-differentiated bacteroids, followed by a high-speed centrifugation to pellet the remaining undifferentiated rhizobia. The pellet can later be used for DNA extraction, followed by whole-genome or amplicon sequencing, and then downstream analysis to estimate strain fitness. Finally, we include an optional step for a reliable, reproducible system for nodule imaging, which is especially useful for quantifying nodule abundance and studying morphological variation. ​

Symbiotic interactions between plants and nitrogen-fixing microorganisms are essential for sustainable agriculture, yet the molecular mechanisms underlying plant–cyanobacterium symbiosis remain poorly understood. In particular, the nature of the signalling mechanisms mediating partner recognition in associations involving Nostoc species is largely unknown. Recent proteomic analyses have identified proteins homologous to rhizobial Nod factors biosynthetic enzymes in Nostoc punctiforme, suggesting the existence of a Nod-like signalling system. However, the functional role of these components has not been experimentally validated. Here, we investigate the contribution of nod-like biosynthetic and regulatory genes to symbiosis by analysing mutants of N. punctiforme affected in genes with homology to nodB and nodD. Phenotypic characterization revealed that disruption of nodB-like genes does not impair free-living growth but affects early stages of plant association and colonization. Specifically, the nodB1 mutant is impaired in plant association and shows a mild defect in colonization, whereas the nodB3 mutant exhibits a severe defect in colonization. In contrast, nodD-like mutants exhibited altered symbiotic phenotypes, with specific regulators differentially affecting interaction and colonization efficiency in rice (Oryza sativa). In particular, mutation of nodD2 and nodD3 reduced plant association and severely compromised colonization in Oryza sativa, with a more pronounced phenotype in nodD3 mutant. Altogether, our results provide genetic evidence supporting the involvement of Nod-like components in cyanobacterial symbiosis and suggest the existence of a regulatory and biosynthetic module contributing to plant colonization. These findings shed new light on the evolution and diversity of symbiotic signalling mechanisms across plant–microbe interactions.

Comparative genomic analyses provide insight into the mechanisms underlying gene-family evolution and crop adaptation. Here, we used the legume phenylalanine ammonia-lyase (PAL) gene family as a model and integrated pan-genomic, phylogenetic, molecular evolutionary, duplication-mode, and transcriptomic analyses, while developing GFtool for gene family identification. Across 45 genomes, we identified 302 PAL genes and classified them into five Groups. Groups 1–3 represented ancient lineages shared with outgroups, whereas Groups 4 and 5 were legume-specific. Molecular-clock analyses placed the divergence of Group 2 near the Paleocene-Eocene transition, while Groups 4 and 5 diversified from the middle Eocene to the early Oligocene. WGD/segmental duplication broadly contributed to PAL copy-number expansion, whereas tandem duplication was enriched in Group 5 of Papilionoideae. Group 2 genes showed drought-induced expression, whereas Group 5 genes were associated with early root nodule development. GFtool provides a scalable framework for gene-family studies.

Food production must rise while its environmental footprint falls, an imperative sharpened by pressure to use fewer external inputs. Meeting this challenge requires gains from aspects that crop breeding has so far overlooked. One such source is the soil, because every crop transforms the environment in which it grows, shaping the conditions experienced by the next crop. We propose that these legacy effects, expressed through altered soil nutrients, water, structure and microbial communities, could offer new breeding targets. Many traits that modulate these effects vary within major crop species, and this variation is heritable and therefore selectable. We advocate breeding for farming systems to exploit this untapped genetic dimension. We outline the mechanisms underpinning this relationship, the frameworks needed to quantify it and the practical challenges of integrating legacy-aware selection into breeding pipelines. Every crop leaves a legacy, and understanding this provides a pathway to a more sustainable agricultural future.

Arbuscular mycorrhizal fungi (AMF) are an early-diverging subphylum of fungi that inhabit plant roots and provide their host with phosphorous and sometimes nitrogen in exchange for carbon. Often, this exchange benefits the plant leading to interest in developing AMF as natural fertilizers. This goal proved incredibly challenging due to the complexity of the system. Despite often being treated as a monolith, individual species of AMF have been demonstrated to play different roles in their plant association, with some species behaving more like parasites than mutualists in certain environments. Additionally, AMF harbor diverse internal and external microbiomes that impact their association with the plant host. In this dissertation I utilize amplicon and whole genome shotgun sequencing of individual AMF spores isolated from Georgia and Arizona derived pot cultures to characterize different genera of AMF and their microbiomes. Through the amplicon sequencing effort, I obtained ITS-OTUs from 211 Entrophosphora spores and 2 Racocetra spores. Each Entrophosphora spore contained multiple divergent OTUs, with the Georgia derived samples being more diverse than the Arizona derived samples. Of these 213 spores, 22 contained ITS-OTUs from non-AMF fungi. Through the genome sequencing portion of the project, I obtained genome assemblies of 15 Entrophospora samples, 10 Racocetra/Cetraspora samples, and one each of Scutellospora and Funneliformis. Burkholderia-related endobacteria metagenome assembled genomes (MAGs) were recovered from 9 spores in the Gigasporales and 32 Mollicutes-related endobacteria MAGs were recovered from 27 AMF spores, with some spores containing multiple distinct MRE. Two well-supported novel BRE clades were identified from my samples. Co-phylogenies of the endobacteria and their host revealed primarily vertical transmission for the BRE and mixed transmission for the MRE. Together, these complementary approaches establish a methodological framework for single-spore AMF research and reinforce the view that the AMF symbiosis is best understood not as a bipartite plant–fungus association but as a multi-kingdom system spanning fungi, bacteria, and plants. The genomic assemblies, reference data, and bioinformatic workflows developed here provide a foundation for future AMF research and contribute to the long-term goal of reducing synthetic fertilizer inputs by harnessing the natural functional capacities of AMF and their microbiomes.

16S rDNA is the historical gold standard for bacterial identification, particularly in metabarcoding approaches reliant on sequence similarity thresholds. We analyzed 6,660 Rhizobium 16S rRNA gene sequences from GenBank to examine the relationship between sequence identity and three metadata: species name, host plant, and geographic origin. Using an iterative BLAST-based pipeline, we detected 116,069 pairwise matches and assessed concordance among sequences (average length 1,328 bp) sharing 100% identity. For those in which the organism name, host plant and country of isolation were present in the record, surprisingly, 66.59% of identical sequence pairs showed full discordance across all three metadata, while only 1.40% shared the same name, host, and country. The most widespread sequence, detected 371 times, was associated with over 56 different host plants across 25 countries and bore multiple species name designations. These results highlight a striking mismatch between the 16S barcode and the taxonomic, ecological, and phenotypic variability it is assumed to reflect, likely arising from the slow evolution of rRNA genes contrasted with the mobility of ecologically relevant genes via horizontal transfer on plasmids, transposons, and phages. Our findings further challenge the limitations of relying on 16S rRNA alone for fine-scale taxonomic and metadata-based inference in capturing the true functional and ecological diversity of bacteria, endorsing the critical importance of polyphasic taxonomic approaches that integrate genomic, phenotypic, and ecological data. An interesting byproduct of the analysis was to realize the possibility of treating these data as if they were ‘citations.’ The more one finds the same query sequence, the more that sequence can be considered biologically ‘cited’, i.e., re-proposed elsewhere in the world. Thus, one can also analyze the h-index of such a ranking. In our Rhizobium dataset, we calculated an h-index = 201, meaning the sequence ranked 201st had 202 identical homologues in GenBank. Although the research effort on given species is directly connected with it, this number provides a quantitative indicator of a taxon’s sequence recurrence and distribution within public databases, independent of nomenclatural inconsistencies, offering a novel framework for assessing bacterial representation across global datasets.

Cut the Chemicals: Tell the FDA to limit dangerous additives in our food now

The symbiosome, a temporary plant organelle enabling nitrogen fixation in legume-rhizobia symbiosis, consists of a plant-derived symbiosome membrane (SM), symbiosome space (SS), and enclosed bacteroid. Here, we isolate and purify symbiosomes from Medicago truncatula-Sinorhizobium meliloti root nodules and perform label-free quantitative mass spectrometry to profile protein abundances in the symbiosomes. We identify 1,018 M. truncatula proteins, including 829 in the SM and 457 in the SS. Combined with transport assays, our data reveal multiple dicarboxylate transporters in the SM that potentially deliver carbon sources to bacteroids. The SM is enriched in membrane trafficking proteins, lipid raft-associated components, and receptor-like proteins, together with numerous cell wall-associated proteins, highlighting the extracellular properties of the symbiosome. Proteomic and metabolomic analyses reveal the SS as a metabolically active compartment enriched in both plant and rhizobial proteins involved in carbon and amino acid metabolism. These findings offer insights into the molecular basis of symbiotic nitrogen fixation.

Arbuscular Mycorrhizal fungi are widespread symbionts that support plant nutrition, health and resilience while shaping soil microbial communities. This review synthesizes current evidence, with emphasis on arbuscular mycorrhizal (AM) fungi, on their roles in nutrient acquisition, carbon cycling, soil aggregation, disease suppression and tolerance to abiotic stress. The mycorrhizosphere is considered a dynamic interaction hub in which AM fungi, bacteria and nematodes influence nutrient fluxes and multi-trophic ecosystem processes. Recent findings on common mycorrhizal networks (CMNs) are also discussed, particularly their potential roles in resource redistribution, defense signaling and plant competition, while acknowledging continuing debate regarding their prevalence and ecological significance. Using an integrative systems-level perspective, this review identifies methodological inconsistencies, geographic and taxonomic biases, and underexplored relationships between AM fungal community composition and ecosystem service delivery. Applications of mycorrhizae in sustainable agriculture, phytoremediation and ecosystem restoration are highlighted, together with constraints such as fungal antagonism, environmental variability and invasive species. Finally, the review outlines key research priorities, including long-term multi-site studies and trait-based approaches, to improve translation of mycorrhizal ecology into scalable management strategies for resilient land-use systems and enhanced ecosystem health.

Plants have evolved a signalling pathway in which, when one root senses local nitrogen (N) deficiency, nitrate uptake by other roots is enhanced in a complementary manner. This long-range communication, known as systemic N-demand signalling, is triggered when the root-to-shoot mobile signal, C-TERMINALLY ENCODED PEPTIDE (CEP), which is induced in roots under N starvation, is perceived by CEP RECEPTOR 1 (CEPR1) expressed in the leaf phloem. However, the molecular components required for CEP-dependent CEPR1 activation remain unknown. Here we identified a leucine-rich repeat receptor kinase that interacts with CEPR1 in a CEP-dependent manner, which we named CEP RECEPTOR INTERACTOR (CERI). CERI belongs to the last functionally uncharacterized clade within the Arabidopsis leucine-rich repeat receptor kinase subgroup II. Loss of CERI impairs systemic N-demand signalling but does not affect CEPR1-mediated regulation of root system architecture. CERI functions as a co-receptor that confers signalling specificity on CEPR1 by selectively mediating systemic N-demand signalling.

Legumes convert atmospheric nitrogen into ammonium through symbiotic bacteria housed in root nodules, yet the molecular interactions between rhizobial and host proteins inside nodules remain poorly understood. Here we employed AlphaFold3 to construct a cross-kingdom interactome between Medicago truncatula and its symbiont Sinorhizobium meliloti. Screening more than 217,000 protein pairs yielded 7,137 putative interactions, providing a valuable resource for the broader symbiosis community. Within this network, we focused on DEFECTIVE IN NITROGEN FIXATION 2 (DNF2), a host protein required for rhizobial persistence within nodules. We showed that DNF2 localizes to the peribacteroid space and associates with previously uncharacterized secreted rhizobial proteins (SRPs), suggesting it may function as a hub for host-symbiont communication. Notably, knockout of two DNF2-interacting proteins, SRP86 and SRP485, results in white, nitrogen-fixation-deficient nodules with abnormal symbiosomes and elevated expression of senescence-associated genes, closely phenocopying the dnf2 loss-of-function mutant. Together, our findings define a DNF2-SRP molecular framework underlying symbiotic accommodation, and illustrate the potential of AI-guided interactome mapping to uncover molecular mechanisms of plant-microbe interactions with relevance to sustainable agriculture.

Large networks are particularly beneficial to researchers in under-represented groups.

Key Message We mapped key regions of the maize genome that influence the formation of aboveground (aerial) roots, which in some varieties have been associated with symbiosis with nitrogen-fixing bacteria. Abstract Modern agriculture relies heavily on chemically synthesized nitrogen fertilizers, which ensure high yields but also carry high economic and environmental costs. Biological nitrogen fixation (BNF) supplies high amounts of nitrogen to legumes, and several avenues of research are underway to extend it to cereal crops. In maize, aerial roots formed in Sierra Mixe landraces have been associated with BNF. However, much of the genetics underlying aerial root morphology remains unknown. Here, we evaluate aerial root morphology traits associated with BNF in three segregating populations derived from crosses between two Midwest-adapted inbred lines and three landraces. Inclusive composite interval mapping (iCIM) with flowering time as a covariate identified 37 quantitative trait loci (QTL) for three aerial root traits (nodes with roots, root size, and roots per node) which exhibit moderately high heritability (H2 = 0.65 to 0.83). The combined proportion of phenotypic variance explained by the detected QTL ranged from 23 to 51%, depending on the trait and population, and potential candidate genes were identified through literature searches, macrosynteny, and gene expression analyses. Introgressing the most relevant aerial root-associated QTL into elite genotypes may provide a path toward achieving meaningful levels of BNF-associated traits in maize, but further work is needed to assess this approach's viability under field conditions.

Nitrogen is often a limiting nutrient for agricultural crops. Most microbes also require an external fixed-nitrogen source for optimal growth. Even for microbes capable of biological nitrogen fixation (BNF), it is generally the case that they will regulate their metabolism to prioritize assimilation of fixed-nitrogen. Microbes employ various strategies and pathways to take advantage of available fixed-nitrogen compounds found in their natural environments. Azotobacter vinelandii is a model microbe for the study of BNF. Due to the high energetic cost of the process, BNF in A. vinelandii is repressed in the presence of ammonium, urea and nitrate. Prior studies indicated that strong inhibition of nitrogen fixation by urea and nitrate in A. vinelandii is actually the result of intracellular conversions of these metabolites into ammonium. In this study, we demonstrate a strategy to eliminate BNF inhibition by both urea and nitrate at concentrations ranging as high as 15 mM in a strain lacking the genes for urease and nitrate reductase, resulting in continued nitrogenase activity in the presence of these common fertilizer inputs. In contrast to the properties of A. vinelandii, the diazotroph Gluconacetobacter diazotrophicus naturally lacks these pathways, prompting the question of whether urea or nitrate inhibit BNF or support growth in G. diazotrophicus. To probe this observation, we developed experiments to demonstrate that while the presence of urea and nitrate delay the initial growth rate in G. diazotrophicus, nitrogenase activity and ammonium accumulation occurs at a similar rate in the presence of these metabolites. These results indicate that biological nitrogen fixation in G. diazotrophicus is somewhat insensitive to these nitrogen sources. This illustrates that alternative pathways in diazotrophic strains should be carefully considered in any efforts to optimize extracellular nitrogen production for biofertilizer applications and strain optimization, and that additional design strategies can be effective to assure that diazotrophs continue to fix nitrogen in the presence of specific nitrogen compounds common to industrial fertilizers.

Nitrate impairs both symbiotic and free-living biological nitrogen fixation (BNF). While nitrate-induced phosphorylation signalling has been implicated in the inhibition of symbiotic BNF, the suppression of free-living BNF has generally been attributed to ammonium generated during nitrate assimilation. However, whether nitrate can inhibit free-living BNF independently of ammonium feedback regulation remains unclear. Here, an ammonium-deregulated mutant of Azotobacter chroococcum (A4) was used to investigate whether nitrate inhibits nitrogen fixation independently of ammonium regulation. Despite the loss of ammonium-mediated inhibition, nitrate significantly suppressed nitrogen fixation. Nitrate at concentrations above 2 mM reduced extracellular ammonium accumulation, with 10 mM nitrate decreasing ammonium production to 61% of that observed under nitrogen-free conditions. Integrated multi-omics analyses revealed that nitrate triggered extensive regulatory reprogramming across multiple molecular layers, with both coordinated and layer-specific responses across transcriptomic, proteomic and phosphoproteomic levels. These responses differed from the typical ammonium-mediated feedback regulation characterized by substantial repression of nitrogen fixation-related genes or proteins, but nevertheless resulted in reduced ammonium excretion, accompanied by enhanced biomass accumulation and extracellular polymeric substance (EPS) production in A4. Together, these findings indicate that, rather than directly repressing the nitrogen fixation machinery, prolonged nitrate exposure suppresses nitrogen fixation output through global regulatory reprogramming that redirects cellular metabolism and resource allocation away from nitrogen fixation. This study provides new insights into nitrate-mediated regulation of free-living diazotrophs and has implications for optimising nitrogen management and improving the application of nitrogen-fixing microorganisms.

Arbuscular mycorrhizal (AM) symbiosis is a widespread mutualism between plant roots and Glomeromycotina fungi that enables nutrient exchange through arbuscules. Although many transcriptional regulators of AM symbiosis have been identified, the role of the INDETERMINATE DOMAIN (IDD) proteins remain unknown. We show that rice IDD7 expression is strongly induced in arbuscule-containing cells, and its loss markedly reduces fungal colonization and disrupts arbuscule development. IDD7 is required for induction of genes associated with fatty acid biosynthesis, nutrient transport, and symbiotic signaling during AM symbiosis. IDD7 interacts with the transcription factors SLENDER RICE 1 (SLR1) and PHOSPHATE STARVATION RESPONSE 2 (PHR2) through its conserved TQDFLG domain. Electrophoretic mobility shift assays show that IDD7 binds multiple promoter motifs. Together with SLR1 and PHR2, IDD7 synergistically activates the promoters of PHOSPHATE TRANSPORTER 11 (PT11) and the AM-associated transcription factors PHR2, REQUIRED FOR ARBUSCULAR MYCORRHIZATION 1 (RAM1), WRINKLED 5a (WRI5a), and CYCLOPS. Moreover, IDD7 is required for PHR2 expression in arbuscule-containing cells. Here, we show that IDD7 is a central transcriptional regulator in arbuscule-containing cells.

We report X-ray crystallographic structures of the Azotobacter vinelandii nitrogenase MoFe protein showing the 8Fe-7S electron-transfer P-cluster in four redox states. Using electrochemical poising of protein crystals, together with in crystallo EPR spectroscopic verification of the redox state, we obtain structures showing the P-cluster at PN, P1+, P2+, and P3+ levels. This provides a detailed structural characterization of P-cluster rearrangement between the catalytically relevant PN and P1+ levels and the first experimental confirmation that the S = 7/2 P3+ state is structurally similar to P2+. These studies pave the way for future understanding of the structure–function relationship in nitrogenase catalysis.

This study reveals extensive changes in A/B chromatin compartmentalization and enhancer promoter interactions during legume–rhizobium symbiosis through high-throughput chromosome conformation capture. Notably, a cytokinin (CK)-responsive distal enhancer forms a long-range chromatin loop with the promoter of NODULE INCEPTION through the CK-signaling transcription factor type-B RESPONSE REGULATOR 3, uncovering a mechanism by which hormonal signaling activates nodulation.

Nod factors (NFs) are microbial signals originally identified for their key role in the nitrogen-fixing root nodule symbiosis in legumes. Beyond symbiosis, NFs also possess a conserved capacity to induce lateral-root formation across diverse plant species, including non-legumes. It is now well established that the nodule organogenesis program has co-opted several molecular mechanisms involved in root development, which raises the question of the developmental pathway controlled by NFs to trigger lateral-root formation and how it overlaps with nodule organogenesis in legumes. In Medicago truncatula, NF stimulation of lateral-root formation is independent of the cytokinin receptor CYTOKININ RESPONSE 1 (CRE1), a negative regulator of lateral-root formation. Here, we show that this stimulation is also independent of the NODULE INCEPTION (NIN) transcription factor, a major regulator of nodule organogenesis acting downstream of cytokinin perception. Instead, NFs stimulate lateral-root formation by influencing auxin biosynthesis and modulating auxin signaling, notably through Auxin/INDOLE-3-ACETIC ACID 7 (Aux/IAA7) in M. truncatula. Using reverse genetics and cross-species complementation, we show that orthologs of MtIAA7, AtIAA29 in Arabidopsis thaliana and SlIAA29 in tomato share a conserved role in lateral-root formation. MtIAA7 also interacts with AUXIN RESPONSE FACTOR (ARF) orthologs of AtARF7 and AtARF19, which are known to control lateral-root formation in Arabidopsis. Altogether, our findings show that NFs control a true lateral-root formation pathway, independent of the nodule organogenesis pathway in M. truncatula, by acting through a conserved auxin signaling module.

Understanding the molecular mechanisms underlying plant–microbe interactions is essential for developing innovative microbe-based agrotechnologies. However, deciphering these mechanisms within the complexity of natural microbial communities remains challenging. Such challenges can be addressed by employing synthetic microbial communities (SynComs) derived from well characterized microbial culture collections. Despite their importance, plant-associated microbial collections from major agricultural crops remain scarce. To bridge this gap, we established TriMic, a taxonomically and functionally representative culture collection of wheat root–associated bacteria. Complementing this collection, we include high quality genome sequences and an overview of genes involved in plant colonization, nutrient cycling, and plant growth promotion. Furthermore, we expanded this experimental toolkit by designing a reduced complexity SynCom that enables controlled dissection of plant-microbe interactions. Together, these resources lay the groundwork for mechanistic studies of plant-microbe interactions to accelerate biostimulant development aimed at enhancing agricultural productivity and sustainability. The TriMic collection and whole genomes are publicly available at the DSMZ (https://www.dsmz.de/collection/catalogue/microorganisms/microbiota/trimic) and NCBI.

Legumes establish mutualistic symbiosis with nitrogen (N)-fixing bacteria, which allows them to utilize atmospheric N2. Because the maintenance of symbiosis requires abundant carbon (C) sources, legumes regulate the balance between carbon consumption and nitrogen acquisition by systemically controlling the nodule number through CLAVATA1 (CLV1)-like receptors. In Lotus japonicus, the CLV1-like receptor HYPERNODULATION ABERRANT ROOT FORMATION1 (HAR1) acts in shoots to regulate root nodulation and contributes to symbiotic C/N coordination. This raises the possibility that HAR1 may also influence plant growth and nitrogen utilization beyond symbiotic nodulation. In this study, we showed that HAR1 plays a critical role in regulating nitrogen use to enhance growth under conditions of high nitrate availability, even in non-symbiotic environments. Unlike the wild-type, the har1 mutant failed to increase its growth in response to higher nitrate availability. This lack of growth response was associated with a lower rate of net biomass production per unit leaf area and a reduced capacity for biomass production per unit plant nitrogen. We further found that nitrate-responsive TCA cycle-related organic acids were higher in har1 leaves than in wild-type leaves even under low nitrate conditions. Because the HAR1 mutation did not affect photosynthetic traits, we propose that HAR1 promotes growth under non-symbiotic conditions by coordinating nitrogen utilization with primary metabolism.

This thesis explores how certain plants, such as Medicago truncatula (Medicago), form a close partnership with soil bacteria collectively called rhizobia. These rhizobia have the unique ability to convert atmospheric dinitrogen (N2) into a form that plants can use for growth. In return, the plant provides the bacteria with sugars and a protected space inside small structures on its roots known as nodules. This interaction is especially important because nitrogen is an essential nutrient for plant growth. However, plants cannot access it directly from the atmosphere, but require a reduced form, such as ammonium (NH4+), which rhizobia provide. This research focuses on how these nodules begin to form, particularly during the earliest stages. A key question is how specific root cells know when and where to change their identity and start forming a root nodule. Unsurprisingly, plant hormones play a central role in guiding this process. One of these hormones, auxin, is known to regulate plant growth. Part of this work demonstrates that auxin accumulates very early in specific root cells in the pericycle after bacterial infection, prior to the first cell divisions. From here, it is relocated towards the root cortex. This process is tightly controlled through both local production and polar transport within the root and is essential for initiating cell divisions and nodule formation. In contrast, the hormone ethylene acts as a repressor of nodulation. The detected ethylene readout clearly shifts from the inside to the outside root tissue, preventing excessive nodule initiation. Additionally, ethylene acts in positioning nodules towards the xylem of the plant. Although the role of ethylene in nodule positioning was known, our results show that the mechanism by which this occurs is more complex than was previously assumed. This demonstrates that the function of ethylene is probably more than just an inhibitor of nodulation. The examples of auxin and ethylene emphasized how important the regulation of hormones and gene expression is in time and space during nodulation. Therefore, the next step in this thesis was to generate a near cell-type specific resolution atlas of gene expression during nodule initiation and development. This was reached by developing single-cell and single-nuclei RNA sequencing (scRNA-seq and snRNA-seq) for Medicago. Using this dataset, transcription factors were identified, that showed expression in the pericycle, which was validated in planta. Although the expression of these genes correlated with nodule initiation, their direct involvement in regulating nodulation could not reproducibly be validated. This either indicates limitations of our genetic tools, more complex regulatory mechanisms, or the need for stringent candidate selection. Together, this thesis provides insights into how Medicago controls nodule formation, both in space and time. Furthermore, it contains the first, robust, single-cell atlas of the root susceptible zone during nodule initiation, and describes how to generate such high-resolution atlases, which can be applied in other, legume, species.    

Every cell must decide what enters the nucleus and what stays out. The nuclear pore complex (NPC), built from proteins called nucleoporins (NUPs), spans the double membrane of the nuclear envelope and shuttles molecules between the cytoplasm and the nucleus. Most NPC components serve this universal housekeeping role, so it was striking when a handful turned out to matter specifically for symbiosis. The NUP107-160 subcomplex, which forms the outer ring of the pore (Tamura et al. 2010), was first linked to symbiosis in the model legume Lotus japonicus. Mutants in 3 of its members, NUP133, NUP85, and NENA, show impaired root nodulation and arbuscular mycorrhization. These mutants also lack the nuclear calcium spiking triggered by host perception of symbiotic microbes (Kanamori et al. 2006; Saito et al. 2007; Groth et al. 2010). This calcium spiking requires nuclear envelope-localized cation channels, CASTOR and POLLUX, in Lotus (Charpentier et al. 2008). The same pathway operates in the related model legume Medicago truncatula, where the Pollux ortholog DMI1 fills this role (Ané et al. 2004). The function of these channels likely depends on reaching the inner nuclear membrane, but how these NUPs influence symbiotic signaling has remained unclear for nearly 2 decades.

Beneficial plant–microbe associations (BMAs) offer a valuable opportunity to reduce dependence on synthetic inputs. However, traditional breeding has rarely targeted traits that enhance beneficial interactions, and conventional agricultural practices have often degraded soil health and microbial diversity. We present a framework that combines breeding for traits that facilitate BMA with soil management practices that enrich BMA. This approach integrates advanced breeding technologies with strategies for precise production and inoculation of microbes. Strengthening these complementary plant- and microbe-centered approaches and encouraging their adoption by farmers can foster more resilient and productive agricultural systems with reduced dependence on pesticides and fertilizers.

Nitrogenase plays a critical role in biological nitrogen fixation. However, current methods for detecting nitrogenase activity are labor-intensive and unsuitable for large-scale applications. To address this issue, we developed a real-time visual detection system for nitrogen-fixation ability in soybean (Glycine max) nodules using reporter gene-tagged rhizobial strains. Among the four reporters tested, the LUX reporter gene provided the most promising results for deep-tissue imaging because, unlike GFP and RFP, it suffers no autofluorescence interference, and unlike GUS, it does not require exogenous substrate. Further screening identified the nitrogenase-correlative promoter pc16350, which drove LUX expression with high sensitivity at levels strongly correlated to nitrogenase activity. This system enables the rapid, non-invasive monitoring of nitrogen-fixation activity in soybean nodules.

Plants preferentially absorb phosphorus (P) as orthophosphate/ inorganic Phosphate (Pi), which has low solubility and is readily fixed in soil. This generates the P limitation condition in many soils, for which plants have evolved a number of responses collectively known as the Phosphate Starvation Response (PSR). Significant advancements have been achieved in identifying PSR, and a holistic viewpoint is presented in this review to summarize responses to Pi starvation and their regulation. Plant response to Pi starvation is mediated by a plethora of players, including MYB transcription factors, strigolactone, malate, and brassinosteroids. The involvement of Arbuscular Mycorrhizal Fungi (AMF) in the process adds another dimension for enhancing PSR. Strigolactone secretion from the root increases AMF colonization and nodulation for enhancing Pi uptake. Pi uptake can be achieved directly by the plant or indirectly by the Pi transporter and AMF. Root system architecture changes according to the Pi present in the environment. Increased auxin signalling is associated with Pi starvation, playing a part in lateral root development. Overall, several unanswered questions remain in the phosphate starvation response of plants, especially from the perspective of AMF symbiosis. Research focused on enhancing PSR with symbiotic association with AMF using advanced biotechnological approaches would pave way for increasing crop productivity.

Mycorrhizal fungi form essential symbiotic relationships with plant roots, facilitating nutrient exchange and promoting plant health. Understanding their interactions can benefit from advanced imaging techniques capable of visualizing nutrient exchange and structural colonization at subcellular resolution across large sample sizes. This review explores novel imaging approaches that are revolutionizing our understanding of root–mycorrhizal fungal symbioses. Several techniques can now visualize and characterize mycorrhizal fungi and associated root structures non-destructively and in three dimensions, for example X-ray computed tomography (micro-CT), X-ray fluorescence (XRF), and X-ray absorption near edge structure (XANES) spectroscopy. Metabolic processes and nutrient exchange can be tracked through positron emission tomography (PET), fluorescent nanoparticles (FNPs), and the monitoring of electrical signalling. Artificial intelligence (AI)-powered image processing software is enabling high-throughput analysis of complex images generated from a range of sources. Mycorrhiza systems are also able to be tracked in-field at multiple scales: hyperspectral imaging can detect mycorrhizal associations at the kilometre scale, while portable MRI imagers can detect changes at the tissue scale. These converging technologies enable the direct, continuous measurement of structural and metabolic root–mycorrhizal fungi interactions, paving the way for a mechanistic understanding of these vital symbiotic partnerships and their impact on plant health and ecosystem functioning.

Arbuscular mycorrhiza (AM) represents a key strategy for plants to overcome nutrient starvation. In legumes and rice, the development of this symbiosis requires the GRAS transcription factor DELLA, previously identified as central proteolytic target of gibberellin (GA) signaling. DELLA performs critical functions across multiple stages in AM development, including inner root cortex patterning, arbuscule initiation and degeneration, by regulating essential downstream genes driving these phenomena. Moreover, DELLA appears as a regulatory hub that integrates hormonal signals, environmental stimuli, and symbiosis formation. Here, we highlight recent advances in our understanding of DELLA-mediated regulation of AM development and provide state-of-the-art insights into how DELLA orchestrates these signaling pathways.

Glomalin-related soil proteins (GRSP) are operationally defined soil fractions associated with arbuscular mycorrhizal fungi (AMF) and are widely studied for their contributions to soil structure, carbon dynamics, and ecosystem functioning. Since its discovery, GRSP has attracted considerable attention because of its association with soil aggregation, carbon stabilization, and ecosystem sustainability. Glomalin, has been associated with various soil attributes, including the stability of soil aggregates, the size of soil carbon and nitrogen reservoirs, the sequestration of heavy metals, and the mitigation of diverse plant stresses. While GRSP concentrations in soil have often been correlated with AMF biomass measured through alternative (microscopic) methods, the chemical composition of GRSP extracted from soil remains intricate and not fully understood. This complexity arises from the nonspecific nature of its extraction and purification processes, as well as the diverse array of analytical techniques employed thus far to evaluate it. Current evidence suggests that GRSP contributes to soil organic carbon stabilization primarily through its association with soil aggregates. In this review, we endeavor to synthesize and explore various facets of glomalin, encompassing its composition, production mechanisms, soil-related functions, recalcitrant properties, and its potential role in the sequestration and stabilization of soil carbon.

Legumes acquire nitrogen via a symbiotic interaction with diazotrophic rhizobia bacteria. In return for getting fixed nitrogen, plants deliver high amount of photosynthate to the bacteria to support the nitrogen fixation process. Hence, biological nitrogen fixation in legume plants is a highly energy-demanding process that relies on the precise coordination of carbon allocation and metabolism between the host plant and its microbial symbiont. Although significant progress has been made in understanding carbon fluxes during nodulation, how these processes are spatially and functionally organized across different cell types and developmental stages within nodules remains poorly resolved. This limitation has hindered a comprehensive understanding of how carbon metabolism supports the establishment, maintenance, and termination of symbiosis. In this review, we explore the current understanding of carbon transport and metabolism throughout the nodulation process, from early allocation during rhizobial infection to the complex metabolic, transport, and regulatory networks in mature nitrogen-fixing and senescing nodules. We highlight key knowledge gaps, especially regarding cell-type-specific and spatial regulation of carbon metabolism. Finally, we discuss how emerging single-cell and spatial omics techniques offer powerful tools to resolve these gaps, enabling a deeper understanding of the metabolic and regulatory complexity that underpins legume–rhizobia symbiosis.

Root nodule organogenesis requires the activation of symbiotic developmental programs within a spatially restricted region of the root. Although many regulators of nodule initiation have been identified, the mechanisms that delimit the domain of organogenic cell proliferation remain poorly understood. In this study, we show that the AP2/ERF transcription factors PUCHI1 and PUCHI2 function in spatially restricting cortical cell division during nodulation in Lotus japonicus. The loss of PUCHI function increased infection thread formation and nodule primordium initiation; however, it did not increase the number of mature nodules. Instead, puchi1 and puchi2 mutants frequently formed clustered nodules accompanied by ectopic cortical cell divisions surrounding developing primordia. Constitutive activation of CCaMK induced broadened spontaneous proliferative structures in the mutant background even in the absence of rhizobia, indicating that this phenotype was not simply a consequence of enhanced infection. PUCHI1 expression was induced during early symbiotic signalling downstream of the NODULE INCEPTION (NIN)–associated transcriptional network; both PUCHI genes were preferentially expressed in the basal region of developing primordia, corresponding to sites of ectopic proliferation in the mutant. Transcriptome and reporter analyses suggested that PUCHI regulated auxin-response patterning, potentially via STY1-, YUCCA11-, TAR2-, and Forked1-like-associated pathways. Additionally, PUCHI1 promoted NIN expression via the cytokinin-responsive CE region of the distal promoter independently of LHK1-mediated cytokinin perception. These findings indicate that PUCHI genes define the spatial domain of organogenic cell proliferation during nodulation; they coordinate auxin-related patterning with the NIN transcriptional module to ensure localised nodule formation.

Plants are found in dynamic soil ecosystems, which are highly enriched with microorganisms that have significant impacts on the health and productivity of plants. Promotion of nutrient uptake, fixation of nitrogen, and tolerance to stress by arbuscular mycorrhizal fungi (AMF) and plant growth-promoting rhizobacteria occur via complex molecular communication with plant roots. Post-transcriptional gene silencing is mediated by MicroRNAs (miRNAs)-small non-coding RNAs that control gene networks in symbiosis and stress responses. Concurrently, phytohormones (auxin, cytokinin, ethylene, jasmonates, salicylic acid, abscisic acid, strigolactones, gibberellins, and brassinosteroids) control root development, defence, and microbial recruitment. Their combined regulatory functions in the establishment of root symbiosis are yet to be fully appreciated, although the integrated regulatory functions of miRNAs and phytohormones in regulating root symbiotic assembly have increasingly been recognised. The paper is a synthesis of the existing knowledge on root-associated beneficial symbioses and a critical assessment of the new roles of plant miRNAs and phytohormonal pathways in the establishment and maintenance of nodulation and arbuscular mycorrhizal associations. We suggest that miRNAs and phytohormones are linked in a complex regulatory network in which miRNAs can regulate hormone pathways and vice versa. We have also demonstrated crucial knowledge gaps by identifying molecular crosstalk between plants and their microbiota, functional validation of miRNA-target pairs, cross-kingdom small RNA trafficking and systemic shoot–root signalling that future studies can use to harness the full potential of plant–microbe interaction for achieving sustainable agriculture.

Soil microbiome studies often define ‘core microbiomes’ using different operational criteria, including occurrence thresholds, abundance thresholds, or other selection methods. Using 1813 soil sample...