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.

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.

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.

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 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.

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.

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.

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.

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.

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. ​

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.

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.

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.

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.