In root nodule symbiosis, symbiosome compartments accommodate nitrogen-fixing rhizobia inside the plant cell. Differentiated into bacteroids, the rhizobia are surrounded by a peribacteroid space and a plant-derived peribacteroid membrane, which separates them from the plant cytoplasm but allows signal and nutrient exchange between host and microbe. The morphological features of symbiosomes are primarily determined by ultrastructural single focal plane imaging, with limited information about spatial details. This study combines 2D and 3D imaging, using transmission electron microscopy and focused ion beam scanning electron microscopy as complementary techniques to analyse the symbiosome ultrastructure and organisation in Lotus japonicus wild-type plants. The 3D model of a mature colonised root nodule cell region demonstrates a dense, puzzle-like arrangement of symbiosomes relative to one another and adjacent plant organelles. The symbiosome shape and size depends on the orientation and number of bacteroids within the compartment and features connective tubular structures. Furthermore, vesicular structures, some likely of bacterial origin, were present at the interface. The study presents a multi-angled analysis of symbiosome-related structures, highlighting their volumes, spatial distribution, and pronounced compactness. Interface associated vesicles, protrusions and connective structures hint towards a dynamic and flexible system that contributes to the plant-microbe crosstalk.

Plant-associated microbiomes are central to crop productivity, nutrient efficiency, and stress resilience, yet conventional microbiome manipulatio

Root nodule symbiosis and pathogen fungi infection represent contrasting biotic interactions that affect plant growth and demand precise cellular reprogramming in Phaseolus vulgaris. Inoculation with Rhizobium tropici promoted progressive nodule organogenesis, reaching their maximum nodule number and diameter at 21 and 30 days post-inoculation, respectively. This developmental progression was accompanied by significant reductions in plant height, leaf and root weight, alongside an increase in photosynthetic pigment content, consistent with the energetic requirements of symbiosis establishment. In contrast, infection by the necrotrophic fungus Sclerotinia sclerotiorum caused extensive foliar necrosis and high infection disease, confirming the susceptibility of P. vulgaris. To explore the molecular regulators associated with these processes, 11 ROP (Rho of Plants) GTPase genes were characterized in the P. vulgaris genome based on the Ras superfamily domain (PF00071) and the Rho-specific motif (PS51420). Structural and phylogenetic analyses grouped PvROPs into five conserved subfamilies, while promoter inspection revealed cis-regulatory elements associated with nodulation (OSE1ROOTNODULE and OSE2ROOTNODULE) and immune signaling (WRKY710S), suggesting context-dependent transcriptional control. Expression analyses demonstrated that PvROPs respond dynamically to both interactions. During early symbiosis, PvROP5, PvROP8, and PvROP10 were significantly upregulated at 7 dpi, indicating roles in infection thread formation and initial nodulation signaling, whereas PvROP1, PvROP7, and PvROP9 were upregulated at 21 dpi, likely contributing to nodule maturation and functionality. Conversely, PvROP1, PvROP3, PvROP7, and PvROP10 were strongly induced in leaves infected with S. sclerotiorum, suggesting a potential role in the plant defense response. Our results provide valuable insight into the role of PvROPs to understand how legumes balance both pathogenic and symbiotic interactions in P. vulgaris.

More than 80% of land plants form symbiotic relationships with arbuscular mycorrhizal (AM) fungi for nutrient uptake. As emerging soil pollutants, nanoplastics (NPs) accumulate in both crop and AM fungal tissue, posing non-negligible toxicity and health risks. However, whether and how NPs can impact plant-AM fungal partnership throughout the symbiotic process remains poorly understood. Here, using axenic root-fungal culture, fluorescence NP tracking, and real-time symbiotic signal monitoring, we show that during pre-colonization phase, NPs reduced spore germination rates (−48%) due to the NP accumulation on spore surface, hindering symbiotic signal perception. During the colonization phase, NPs entered fungal cells, disrupted organelles, and accelerated hyphal senescence, consequently reducing hyphal branching length (−22%) and secondary spore production (−32%). In real-world soil, inoculation of secondary spores (reproduced under NPs) formed fewer arbuscule structures (−46%) within maize roots with reduced carbon allocation to AM fungus, leading to lower hyphal length density (HLD) (−24%). During the post-colonization phase, lower HLD impaired the well-known function of phosphorus (P) mineralization by hyphae-enriched bacteria, reduced soil available P (−5.7%) and maize shoot P (−20%), eventually resulting in compromised plant performance. Our findings reveal an integrated yet largely underexplored mechanism of how NPs hinder plant performance by disrupting the dynamic relationship between plants and their symbiotic partners. In a broader context, understanding the alteration of plant-microbial interaction (rather than separately) under emerging stress can provide ecologically relevant implications for sustaining agricultural and terrestrial ecosystems.

Mutualism in the symbiosis between arbuscular mycorrhizal fungi and plants is based upon the exchange of carbon for soil minerals, with phosphate being of central importance. The exchange of nutrients occurs when the fungus transiently colonises root cells, producing hyphal structures called arbuscules. The movement of phosphate from fungus to plant is well established, however its coordination and regulation at the ephemeral arbuscules remains elusive. Here, non-invasive imaging captures the complete growth and collapse of the arbuscules in unprecedented resolution, revealing heterogeneity in arbuscule development. Tracking the dynamics of rice PHosphate Transporter 1;11 (OsPHT1;11/ PT11) as a proxy for symbiotic phosphate transport shows consistent localisation across diverse arbuscules. However, we uncover phosphate-responsive variability in PT11 abundance, representing an essential, cellular-level layer of nutrient regulation. Such plasticity in arbuscule phosphate uptake capacity evidences uncoupling of arbuscule presence and arbuscule function, thereby demonstrating that arbuscules are not identical units of nutrient exchange.

The search for life now extends beyond the traditional habitable zone to include the icy moons of Jupiter and Saturn. These moons feature ice-covered surfaces overlying substantial oceans formed primarily of liquid water and other potential constituents, such as ammonia. On Earth, ammonia supports biochemistry at low concentrations by providing nitrogen but becomes disruptive at higher concentrations. Ammonia could therefore influence the habitability of extraterrestrial oceans, yet this topic has received limited attention in the literature. This review synthesises current research on ammonia in Saturn’s icy moons, Enceladus and Titan, and its effects on terrestrial life. We summarize the celestial incorporation, speciation, and phase behaviour of ammonia and review data on its occurrence and concentration in icy moon oceans. We examine the role of ammonia in prebiotic chemistry, biochemistry, and toxicity. Focusing on bacteria, we compare known survival limits in ammonia to estimated ammonia concentrations on Enceladus and Titan. We find that bacterial survival limits exceed concentrations estimated on Enceladus, but are below those estimated on Titan, and propose that ammonia measurements are crucial for assessing extraterrestrial habitability. Finally, we highlight outstanding knowledge gaps and challenges that influence our understanding of how ammonia shapes the potential for life beyond Earth.

Quorum-sensing (QS) systems based on acyl-homoserine lactones (AHLs) regulate gene expression in response to cell density in many bacteria, including Rhizobium. These systems, typically composed of LuxI-like synthases and LuxR-like regulators, control processes such as plasmid conjugation, biofilm formation, and plant interactions. However, their evolutionary dynamics and genomic distribution in Rhizobium remain poorly understood. We analyzed 142 complete Rhizobium genomes using comparative genomics, phylogenetic reconstruction, and genomic context analysis. LuxI/LuxR homologs were identified based on sequence similarity and Pfam domain architecture, and their genomic contexts were examined. Phylogenetic relationships and coevolution between LuxI/LuxR pairs were assessed using cophylogenetic approaches. QS systems showed a highly heterogeneous distribution across Rhizobium genomes: some strains lacked canonical systems, whereas others encoded one or multiple systems in chromosomes and/or plasmids. Chromosomal QS systems were associated with multiple distinct genomic contexts, supporting at least seven independent acquisition events. In contrast, plasmid-encoded systems exhibited substantially greater diversity in both sequence and genomic organization. Phylogenetic and comparative analyses revealed dynamic gains and losses of QS systems, variable coevolution among LuxI/LuxR pairs, and evidence of partner recruitment. Notably, plasmids appear to act as major reservoirs of QS systems and likely sources of their transfer to chromosomes. These findings indicate that QS systems in Rhizobium evolve through a combination of horizontal gene transfer, genomic rearrangement, and differential retention across replicons. The higher diversity and mobility of plasmid-encoded systems highlight their central role in shaping QS evolution and functional innovation. Overall, this study provides a comprehensive framework for understanding the diversification and evolutionary trajectories of QS systems in complex multipartite bacterial genomes.

Arbuscular mycorrhizal fungi (AMF) represent a key biological strategy for enhancing agricultural resilience under extreme climatic events such as drought

Symbiotic nitrogen fixation (SNF) by legumes is essential for sustainable agriculture, providing plant-available nitrogen while reducing reliance on synthetic fertilizers. The establishment of legume-rhizobium symbiosis requires tightly regulated host signaling to coordinate rhizobia infection, nodule development, and nitrogen fixation, while preventing excessive colonization or immune activation. Accumulating evidence indicates that ubiquitination, mediated by E1, E2, E3 ubiquitin ligases and deubiquitinating enzymes, plays a central role in controlling multiple stages of this process. In this review, we summarize current knowledge on ubiquitination-mediated regulation of symbiotic nitrogen fixation, with a focus on early symbiotic signaling and nodule development. We highlight key E3 ligases that modulate Nod factor receptor homeostasis, receptor-associated kinases, transcription factors, and infection thread growth, and discuss how ubiquitination interfaces with nutrient and stress signaling pathways. Finally, we outline key knowledge gaps and discuss the potential of manipulating ubiquitination pathways to improve nodulation efficiency and nitrogen use efficiency in crops.

The rhizobia-legume symbiosis involves nodulation and nitrogen fixation, which are vital bioprocesses for the sustenance of agriculture, land and ecosystem, and the global nitrogen cycle. Legume plants and rhizobia interact intricately, resulting in the formation of specialized structures known as root nodules. Root nodules enhance soil fertility and promote more vigorous plant growth by converting atmospheric nitrogen into a form that plants can utilize. This symbiosis is regulated by complex and finely tuned molecular pathways that involve signal exchange during host recognition, nodulation, and nitrogen fixation, ensuring the successful initiation and maintenance of the symbiosis. This interaction comes with specific difficulties. This review explores the intricate dynamics of the symbiotic relationship between legumes and rhizobia. It involves the molecular, genetic, and ecological factors that govern this connection, providing a thorough understanding of its advantages and disadvantages. By examining these facets, this review aims to illuminate rhizobia’s potential to enhance legume yields and promote more environmentally conscious farming practices, while acknowledging the obstacles that must be overcome to maximize this mutualistic association.

Plant–microbe interactions shape key agronomic traits, yet breeding strategies to harness them remain unclear.
We propose distinguishing the varying population, which supplies heritable variation, from the carrying population, where the selected property is expressed.
In traditional breeding, the varying and carrying populations coincide within a single organism, whereas in novel strategies, they are distributed among plants and microorganisms, with one partner varying and the other carrying the selected property.
This distinction defines holobiont breeding as a strategy in which both plants and microorganisms are combined to form a varying population, enabling the selection of interspecific indirect genetic effects.
Our framework builds on the replicator–interactor distinction by localizing these roles within plant–microorganism associations, thereby making it operational for breeding program design.

In legumes, symbiotic root nodules undergo senescence in response to developmental or environmental cues. This process determines the maintenance and nitrogen-fixing capacity of the root nodules, but the molecular mechanisms underlying its initiation are poorly understood. The cysteine protease CYP35 is a positive regulator of nodule senescence in soybean (Glycine max), but its substrates remain unknown. Here, we demonstrate that CYP35 promotes nodule senescence by cleaving a subset of Nodule-Enriched Nodulin proteins (NENs). Sequence and phylogenetic analyses indicate that CYP35 is a cathepsin L-like cysteine protease, with Cys149 as a key catalytic residue. CYP35 physically interacts with a distinct subfamily of eight NENs, NEN1–8. Soybean quadruple and quintuple nen mutants obtained by multiplex gene editing develop nodules with accelerated senescence and reduced nitrogenase activity, whereas over-expression of NEN2 or NEN5 delays senescence and enhances nodule function. CYP35 proteolyzes NEN2, NEN5, NEN6, and NEN7 in vitro and cleaves NEN2 in vivo in a Cys149-dependent manner. Our findings establish a direct molecular link between cysteine protease–mediated Nodulin cleavage and the onset of nodule senescence in soybean, providing insights into the regulation of nodule lifespan and nitrogen fixation.

Symbiosis receptor kinase (SYMRK), a malectin-like-domain/leucine-rich-repeat receptor-like-kinase (MLD-LRR-RLK), is the upstream-most component in the common-symbiosis-signaling-pathway. We highlight 2 proline residues that were distinctly acquired by SYMRK orthologs in its hinge regions to constitute a signaling module for allowing progress of symbionts across transcellular barriers during rhizobia-legume symbiosis. Within the ectodomain hinge (EctoD-hinge), all MLD-LRR-RLKs have a conserved W1xnGDPCxnW2×4C motif, where SYMRK orthologs within legumes have a distinct signature with a proline preceding W2 enabling cleavage of SYMRK ectodomain for releasing MLD. Within the kinase hinge (KD-hinge) at gatekeeper + 1 position, a conserved glutamate in MLD-LRR-RLKs is replaced by proline in all SYMRK orthologs that enabled its dual-specific kinase activity for ensuring ectodomain cleavage. Substitution of either proline restricted cortical progression of symbionts, forming infection patches in the nodule apex without affecting epidermal invasion and nodule organogenesis. This halt was entirely overcome by ectopic expression of free MLD, demonstrating the released MLD has an active role in progress of symbionts. Overall, we show that conservation of distinct prolines in hinge regions of SYMRK orthologs in legumes generates a signaling module involving dimerization and optimal phosphorylation of SYMRK for releasing MLD as an active transducer of symbiosis signaling.

Plants exist as complex holobionts, whose health and productivity depend on dynamic interactions with diverse microbial partners. This chapter explores the ecological, molecular, and biotechnological dimensions of plant-microbe relationships, tracing the continuum from natural symbioses to their translation into microbe-based agricultural applications. It examines how plants actively structure their microbial communities across distinct yet interconnected compartments, such as the rhizosphere, phyllosphere, and endosphere, and how these microorganisms in turn modulate plant physiology, nutrient acquisition, and defense. Moving beyond the study of individual strains, we discuss the growing relevance of community-level approaches and synthetic microbial consortia as frameworks to decipher and replicate the ecological principles that sustain beneficial interactions. Finally, this chapter reflects on how the integration of ecological knowledge with emerging technologies such as synthetic biology, systems modeling, and artificial intelligence offers new opportunities to design resilient, predictable, and sustainable microbe-based solutions. By learning from these naturally optimized alliances, we can guide the transition to more adaptive and sustainable agricultural systems while sustaining plant health and productivity.