Plant and Soil

Microbial communities can shift under drought in ways that enhance plant performance during drought ("microbe-mediated acclimation"). However, it is also possible for microbial communities to shift in ways that worsen the effects of drought ("mal-acclimation"). It is unclear how and where microbe-mediated acclimation vs. mal-acclimation occurs, or if there are types of soils or microbial communities that are more likely to harbor microbes that enhance plant acclimation and limit mal-acclimation. We tested for microbe-mediated plant acclimation/mal-acclimation to drought in soils from 21 maize farms in the midwestern United States, spanning a range of climate, soil types, and management practices. We first conditioned soil microbial communities to drought vs. well-watered conditions in a greenhouse and then tested for microbe-mediated acclimation by growing maize in soils inoculated with the conditioned microbial communities under drought and well-watered conditions. Drought-conditioned soils did not enhance plant performance under drought. In fact, one third of the farms exhibited mal-acclimation, especially under well-watered conditions where wet-conditioned soils reduced plant performance in well-watered contemporary conditions. Farm management practices, climate, soil texture, and microbial diversity generally did not predict when this microbe-mediated mal-acclimation occurred. Overall, these results suggest that in agricultural soils, microbes may frequently impede-rather than facilitate-plant acclimation to soil moisture levels. Open research statementThe plant and soil data used in this study are available via the Environmental Data Initiative repository at https://doi.org/10.6073/pasta/f4a0db3a076cf6d8cef908947f82736e. The bacterial and fungal amplicon sequence data are available via the European Nucleotide Archive under accessions PRJEB110071 and PRJEB109827, respectively.

O_LIThe causal factors shaping plant-associated microbiota are incompletely known. Elevated concentrations of the micronutrients zinc (Zn), manganese (Mn) and copper (Cu), and exposure to non-essential trace elements including cadmium (Cd) and arsenic (As), can be toxic. Here we explored whether differences in metal(loid) sensitivity between plants and bacteria influence phyllosphere bacterial community composition. C_LIO_LI224 representative Arabidopsis thaliana phyllosphere bacterial strains were screened on metal(loid) concentration series in synthetic media. We obtained leaf apoplastic fluid ionomes for comparisons with bacteriotoxicity profiles, and tested for relationships between strain-wise metal(loid) tolerances, phylogeny and gene content. C_LIO_LILeaf apoplastic Zn2+ and Cd2+ concentrations were the most likely to arrest growth of metal-sensitive bacteria in planta. Soil bacterial strains were several-fold more sensitive to both these metals than leaf strains, consistent with selection for increased bacterial Zn and Cd tolerance in the phyllosphere. Strains known to govern bacterial community structure were metal-sensitive, with only minor influences of between-metal and between-strain interactions. Bacterial genus explained considerable proportions of the variances in metal(loid)-related gene content and tolerance phenotypes. Bacterial Cd tolerance correlated with the presence and copy number of known Cd-related genes. C_LIO_LIOur results suggest that plant metal homeostasis contributes to structuring bacterial communities in the leaf endosphere. C_LI

O_LISymbiosis between legumes and rhizobia is beneficial on nutrient-poor soils, as it enables the fixation of atmospheric N2. To establish this symbiosis, gene expression in both the host plant and the symbiont has to be regulated. To understand the underlying RNA-mediated regulation of host gene expression, we designed experiments to identify competing endogenous networks involving circular RNA, microRNA, and linear transcripts during symbiosis, using wt and symbiosis-deficient Lotus japonicus mutants with the rhizobium Mesorhizobium loti (M. loti). C_LIO_LICircRNA, miRNA, and linear transcripts were identified from Lotus japonicus wildtype and CCamK mutant (ccamk-13; snf-1) seedlings without inoculation or with M. loti inoculation using deep short-read sequencing with rRNA-depletion and random primers. C_LIO_LIDifferentially expressed miRNAs showed negative correlations to predicted target genes and may regulate symbiotic processes. The symbiosis essential iron-sensor LjnsRING/BRUTUS expresses a circRNA which was upregulated in symbiotic treatments. This circRNA may act as a target mimic and contribute to nodule longevity. CircRNAs are predicted to act predominantly as trans-regulatory molecules with similar frequencies in Arabidopsis thaliania, Oryza sativa, and Lotus japonicus. C_LIO_LIWe identified novel miRNAs, long noncoding RNAs, and circRNAs, and nominated several as potential new regulatory non-coding RNAs that may act as target mimics to stabilize genes and support symbiosis. C_LI SummarySymbiosis between Lotus japonicus and Mesorhizobium loti involves treatment-specific regulation of competing endogenous RNA networks involving circular RNA, miRNA, and linear transcripts.

Decomposition of organic matter is a key process in soils contributing to carbon and nutrient cycling. To identify management strategies for agroecosystems that reduce nutrient losses while maximizing plant growth, it is important to understand which parameters determine decomposition rates. This study therefore investigated how the presence of winter wheat (Triticum aestivum var. Asory) affects decomposition in a controlled Ecotron setup with two soil types with varying organic matter content across three simulated climates (2013, 2068, 2085). Using the tea bag index, interstitial soil pore water analyses, microbial biomass quantification, bacterial and fungal gene abundance, and soil respiration measurements, we tested the hypotheses that plant exudates would enhance decomposition rate and microbial biomass, while plant nitrogen uptake would deplete soil nitrate, potentially mitigated by fertilization. Contrary to expectations, decomposition rates were lower in planted than in unplanted soils, suggesting resource competition between plants and microbes. No significant differences were observed in microbial biomass or respiration due to plant presence, and fertilization effects on nitrate or microbial mineralization were undetectable, likely due to rapid turnover of organic molecules including uptake by plants and microbes. Mechanistically, fungi and soil humidity were more important for decomposition than bacteria or temperature. The findings corroborate climate impacts on decomposition but also indicate microbial resilience and highlight the potential of management strategies like cover crops, adjusted planting dates and crop residual management which can contribute to healthy soils by sustaining carbon and nutrient cycling.

O_LIPhyllosphere microbiomes are subject to microbial import from various sources and undergo substantial changes during phenological changes of plants. However, these processes are still poorly understood for forest canopies. We propose that phenology-driven changes in host properties, and rainwater-mediated, within-canopy transport shape the phyllosphere microbiome in temperate forests. Leaves and throughfall samples were collected from oak, ash and linden trees at top, mid, and bottom canopy positions at the Leipzig canopy crane facility (Germany) at time points representing early, mid and late phenological stages. Bacterial community composition was assessed by 16S rRNA gene amplicon sequencing. C_LIO_LIPhenological stages explained 19% of phyllosphere bacterial community variation, followed by tree species identity (12%) and canopy position (2%). Later phenological stages exhibited more homogeneous and functionally redundant phyllosphere communities along with a strong decline of plant pathogens and increasing potential for microbially mediated biocontrol mechanisms. Throughfall transported up to 1011 bacterial cells per litre with maximum bacterial fluxes at the canopy top. C_LIO_LIOur findings demonstrate that in temperate forests, phenology-driven effects on the phyllosphere microbiome are far more important than tree species specific effects. Extent and selectivity of throughfall-mediated mobilization may play a crucial role for the spatial heterogeneity of microbial communities in tree crowns. C_LI

O_LIA productivity-driven higher nutrient demand of trees in diverse mixtures is frequently reported. Yet, it remains unclear how tree diversity influences microorganisms-plants interactions, in which microbes facilitate tree nutrient acquisition in exchange for carbon (C) to meet the resource demand of both. C_LIO_LIUsing a long-term tree diversity experiment in the subtropics, we assessed microbial investment in C-, nitrogen (N)-, and phosphorus (P)-acquiring enzymes in litter and mineral soil, testing the effects of tree species richness and mycorrhizal type (arbuscular (AM)- vs. ectomycorrhizal (EcM)-associated tree species). C_LIO_LIWith increasing tree species richness, microbial investment in C acquisition decreased, while investment in N and/or P acquisition increased in litter and in mineral soil. In mineral soil of AM-associated tree mixtures, ecoenzymatic stoichiometry revealed a shift from microbial investment in C toward P acquisition as tree species richness increased. C_LIO_LIOur findings suggest that tree diversity strengthens microbe-tree interactions in terms of C-for-nutrient exchange. This highlights the key role of soil microorganisms, particularly in AM symbiosis, shaping tree diversity-biogeochemical feedbacks. C_LI

Crop root systems develop in biologically complex soils where beneficial symbionts and pathogenic organisms can jointly influence root architecture and, consequently, belowground function. In this work, we used X-ray computed tomography (CT) to assess how colonisation by the arbuscular mycorrhizal fungus Rhizophagus irregularis (AMF) and infection by the potato cyst nematode Globodera pallida (PCN) influence root system architecture in soil-grown tomato and potato plants. Root architectural traits, including root volume and root surface area, were quantified non-destructively from intact root systems to evaluate the individual and combined effects of AMF colonisation and PCN infection over time. AMF inoculation increased root volume and surface area, whereas PCN infection caused pronounced reductions in these traits, particularly during early development. AMF-associated increases in root system size were maintained in both PCN-free and PCN-infected plants, indicating largely additive effects of beneficial and pathogenic soil biota on root architectural outcomes. These findings show that soil organisms can independently reshape crop root development in ways likely to influence soil exploration and resource acquisition under biologically complex conditions. More broadly, the study highlights the value of X-ray CT as a non-destructive approach for linking belowground biotic interactions with functionally relevant root traits in sustainable agroecosystems.

Antibiotic use in agricultural systems can unintentionally disrupt beneficial rhizosphere microorganisms, yet the consequences of this dysbiosis for plant fitness remain insufficiently understood. Building on previous findings that application of streptomycin to the roots decreases cyanobacteria and increases tomato plant susceptibility to foliar Xanthomonas infection, this study aimed to determine whether this relationship reflects causation or correlation. We evaluated whether targeted inoculation with the filamentous nitrogen-fixing cyanobacterium Cylindrospermum sp. (CI) or a complex rhizosphere microbiome transplant (RMT) could mitigate antibiotic-induced dysbiosis. As expected, streptomycin treatment significantly increased bacterial spot disease severity and reduced microbial richness in the rhizosphere, marked by a pronounced decline in cyanobacterial and Cylindrospermum operational taxonomic units. Co-occurrence network analysis revealed that this dysbiotic state was defined by reduced community connectivity and increased negative associations, indicating a breakdown in cooperative microbial relationships. Notably, both CI and RMT reduced plant disease severity, though they caused distinct rhizosphere community reassembly outcomes. While RMT relied on microbial functional redundancy, the targeted CI approach achieved more robust colonization and effectively "patched" the functional gap left by dysbiosis. Microbiome restoration directly influenced host physiology, significantly reducing the overactivation of ethylene-mediated defense genes, such as ERF1, and partially reinstating auxin-responsive signaling pathways (IAA21) that were disrupted under dysbiosis. These findings suggest that targeted microbial inoculation could reverse dysbiosis and enhance plant resilience under pathogen pressure as effectively as complex microbial transplants. This work highlights a shift in microbiome management: from the complex rebuilding of communities to the strategic repair of specific functional gaps.

Belowground, plants are exposed to a wide range of biotic stresses that vary in severity and nature, including tissue damage, disruption of vascular connectivity, and depletion of assimilates. How plants adapt their root systems to cope with different types of belowground biotic stresses is not well known. In this paper we compare above- and belowground plant adaptations to three nematode species with distinct tissue migration and feeding behaviours to study mechanisms underlying tolerance to different types of biotic stresses. We monitored both green canopy growth and changes in root system architecture of Arabidopsis inoculated with Pratylenchus penetrans, Heterodera schachtii, and Meloidogyne incognita. This revealed three distinct phases in aboveground plant responses: (i) initial growth inhibition associated with host invasion and tissue damage, (ii) persistent growth reduction associated with nematode sedentarism, and (iii) late growth stimulus in more advanced stages of infection. Specific adaptations in the root systems further revealed fundamentally different stress coping strategies. Tissue damage and intermittent feeding by P. penetrans in the root cortex did not induce significant changes in root system architecture. Tissue damage to the root cortex and prolonged feeding on host vascular cells by H. schachtii induced secondary root formation compensating for primary root growth inhibition. Prolonged feeding on host vascular cell by M. incognita alone did not induce secondary root formation, but was accompanied by typical local tissue swelling instead. Our data suggest that local secondary root formation and tissue swelling are two distinct compensatory mechanisms underlying tolerance to sedentarism by root-feeding nematodes. HighlightHow plants utilize root system plasticity to cope with different types of biotic stresses by root feeding nematodes remains largely unknown. Here, we report on specific adaptive growth responses in Arabidopsis roots to three nematode species, Pratylenchus penetrans, Heterodera schachtii, and Meloidogyne incognita, with fundamentally different strategies for host invasion, subsequent migration through host tissue, and feeding on host cells.

Ectomycorrhizal (EcM) fungi are well-recognized symbionts impacting tree health and ecosystem functioning globally, yet understanding of their timing of proliferation in soils across seasons and years remains limited. We analyzed monthly patterns of EcM fungal abundance and community structure over two years in five temperate monodominant forest plots via quantitative PCR and Illumina sequencing. We found that the phenological dynamics of EcM fungi differed significantly by host tree leaf habit, fungal exploration type, fungal genus, and soil moisture. Overall, total EcM fungal abundances based on qPCR consistently peaked in autumn, and were more dynamic in evergreen than deciduous plots, supporting ideas of surplus carbon and asymmetric above-belowground dynamics. Longer-distance exploration types peaked earlier and were more stable than shorter-distance types, suggesting an independent and supportive role in releasing spring nutrients. About half of 20 focal taxa consistently peaked in either autumn, summer, or spring, while others were either host- and/or year-dependent. Our findings highlight that phenology is a key EcM fungal trait best explained by both host and fungal contributions, and future studies across biomes should consider seasonal shifts and sampling to elucidate phenological traits. Summary- The timing of belowground production and seasonal community dynamics remain poorly understood for ectomycorrhizal (EcM) fungi. - We collected soils monthly for two years from five temperate monodominant forest plots. - Fungal production peaked in autumn, shorter-distance and evergreen-associated spanned wider ranges, and half of focal fungal genera showed seasonal preference, emphasizing autumn surplus carbon and spring nutrients from long-distance types. - Future studies should consider seasonal shifts when sampling EcM fungal communities, and forest carbon models should include asymmetric above-belowground phenology. Translated Summary (Spanish)- La fenologia de la produccion y composicion de comunidades de hongos ectomicorrizicos (EcM) es poco estudiada. - Recolectamos suelos mensualmente por dos anos de cinco parcelas mono-dominantes templados. - Produccion maxima de hongos ocurrio en otono, hongos asociados con arboles siempreverdes y de exploracion de corta-distancia observaron rangos mas amplios, y la mitad de generos de hongos focales observaron preferencia estacional, enfatizando extra carbono en otono y nutrientes en primavera de tipos larga-distancia. - Estudios deben considerar cambios estacionales para el muestreo de hongos EcM, y modelos de carbono deben incluir fenologia asimetrica entre hojas y hongos. Plain language summaryEctomycorrhizal fungi are critical for the global carbon cycle, but their seasonal and inter-annual growth patterns remain unclear. We sample soil DNA monthly over two years across five different monodominant temperate forest stands. We find an overall belowground peak in autumn, with significantly later growth under wetter conditions, more dynamism with evergreen trees, and distinct spring growth by longer-distance fungi.

O_LIPlants take up nutrients from the soil while investing in absorptive root surface or mycorrhizal partners. Root hairs - a major structure for nutrient uptake and cheap to build - increase the absorptive root surface. As such they are an important component of plant resource economics but largely neglected in root economic concepts so far. C_LIO_LIThis is mainly due to data scarcity, which we set out to overcome by measuring root-hair traits on 82 European grassland species in a greenhouse experiment. Using fluorescence and light microscopy, root-hair length and incidence was measured along with mycorrhizal colonization. C_LIO_LIWe found a phylogenetically conserved trade-off between plant investment into root hairs and mycorrhiza. A similar trade-off between root-hair incidence and mycorrhiza occurred at the intraspecific level, while patterns were heterogeneous among species. Plant species with high colonization rates showed the highest variability in root-hair incidence. C_LIO_LIWe conclude that plants vary along a gradient ranging from investment into root hairs as part of a "do-it-yourself" strategy to collaboration with mycorrhizal fungi while showing intraspecific variation in root-hair incidence. These findings demonstrate that root hairs play a fundamental role in fine-root trait variation and need to be considered when studying belowground plant economic strategies. C_LI

Phyllosphere microbiomes are increasingly recognized as key regulators of plant health and stress responses, although they are also known to change considerably over both space and time. In the phyllosphere, members of the genus Methylobacterium are often abundant and ecologically important as plant growth promoting bacteria. However, knowledge about the temporal abundances and community dynamics of Methylobacterium in agricultural systems remains limited. To address this gap, we characterized seasonal shifts in Methylobacterium-specific and total phyllosphere bacterial loads and community structure on two common summer crops and one overwintering cover crop. Leaf samples of Zea mays (corn), Glycine max (soybean), and Thlaspi arvense L. (pennycress) plants were collected over one year in Minnesota, USA and analyzed with host-associated microbial PCR (hamPCR). Microbial loads and community composition varied strongly among hosts and across growing seasons. Corn supported the highest Methylobacterium and total bacterial loads, increasing towards senescence, while pennycress exhibited the lowest loads and the most distinct communities. While there were strong host-specific patterns, a group of most abundant genera were shared across all crops (Methylobacterium, Sphingomonas, Pseudomonas, and Massilia) and the most abundant Methylobacterium amplicon sequence variants were present on all three hosts. Our findings highlight how microbial loads and community composition change during phyllosphere assembly across diverse summer and overwintering crops, with a small core of versatile taxa dominating multiple agricultural hosts. Understanding these host and season-linked patterns provides a foundation of harnessing Methylobacterium strains to enhance crop productivity and resilience.

1.Structured Abstract1.1 AbstractSoybean red crown rot, caused by the soil-borne fungus Calonectria ilicicola, causes substantial yield losses, but the response of the root-associated bacterial microbiome remains poorly understood. Here, we combined 16S rRNA gene sequencing, shotgun metagenomics, and single-cell genomics to characterize bacterial communities in soybean root-associated soils. 16S rRNA gene sequencing showed that diseased plants had rhizosphere and, more strikingly, rhizoplane microbiomes distinct from those of healthy plants, often with increased Enterobacterales. Shotgun metagenomics further revealed enrichment of genes associated with antibiotic resistance, particularly cationic antimicrobial peptide resistance, in diseased rhizoplane samples. Single-cell genomics recovered seven nonredundant Enterobacterales genomes and showed that plant pathogenicity-related genes were broadly distributed across these lineages. In contrast, dlt genes, which are associated with cationic antimicrobial peptide resistance, were detected only in the Enterobacterales lineages enriched in diseased rhizoplane soils. These results support a model in which soybean red crown rot is accompanied by microbiome restructuring and opportunistic enrichment of specific Enterobacterales lineages carrying putative cationic antimicrobial peptide resistance genes. More broadly, this study highlights the value of strain-resolved single-cell genomics for linking disease-associated community shifts to specific bacterial traits. 1.2 ImportanceUnderstanding crop disease requires resolving not only the primary pathogen but also the root-associated bacteria that respond to infection. Here, we used 16S rRNA gene sequencing, shotgun metagenomics, and single-cell genomics to examine the soybean rhizoplane microbiome under red crown rot. Diseased plants showed reproducible shifts in bacterial composition, including frequent enrichment of Enterobacterales and antimicrobial resistance-related functions. Strain-resolved genomes further revealed that the Enterobacterales lineages enriched in diseased rhizoplane soils specifically carried putative dlt-mediated resistance to cationic antimicrobial peptides, whereas general pathogenicity-related genes were broadly shared. These findings suggest that host defense-associated selection, rather than pathogenicity genes alone, may help shape disease-associated root microbiomes. This study demonstrates how single-cell genomics can uncover strain-level traits hidden within bulk community data and thereby clarify plant-pathogen-microbiome interactions.

Common bean (Phaseolus vulgaris) is an important crop in Canada and globally. Like other legumes, common bean (Phaseolus vulgaris) establishes symbiotic interactions with nitrogen fixing bacteria called rhizobia. However, nitrogen fixation by rhizobia in association with common bean is often suboptimal, constraining its productivity and necessitating the application of nitrogen fertilizer. To support the development of high-performing, locally adapted rhizobial inoculants for Ontario common bean growers, we isolated 216 common bean-nodulating rhizobia from southern Ontario soils using a nodule trapping approach with four common bean cultivars. Whole genome sequencing followed by phylogenomic analyses of the 216 rhizobial isolates revealed substantial diversity, assigning them to 11 Rhizobium species, including two novel species. Nearly all isolates belong to the symbiovar phaseoli, spanning the nodC {gamma}-a, {gamma}-b, and alleles, with four isolates belonging to the symbiovar gallica. Soil origin had a significant impact on the species-level community composition recovered during the nodule trapping experiments, indicative of biogeographical structuring of common bean-nodulating rhizobia across southern Ontario. In contrast, host trapping cultivar had only a minor influence of the recovered Rhizobium population diversity. Greenhouse assays demonstrated that one of the novel Rhizobium species exhibited the highest average symbiotic effectiveness, although high-quality isolates were found across multiple species. Together, these results revealed a diverse and genomically variable Rhizobium community capable of forming effective symbioses with common bean in southern Ontario soils. Importantly, our genome-sequenced Rhizobium collection will serve as a valuable resource for identifying competitive and high-quality strains for the development of inoculants tailored to Ontario common bean production. IMPORTANCECommon bean is a globally important food crop, yet its productivity is often limited by suboptimal nitrogen fixation, forcing growers to rely on synthetic fertilizers. Consequently, identifying high-performing, locally adapted inoculant strains is essential for reducing dependence on synthetic nitrogen fertilizers and improving the sustainability of temperate agroecosystems. Our study provides a genome-sequenced collection of common bean-nodulating Rhizobium from southern Ontario, revealing substantial species and genomic diversity across sampling locations. Greenhouse studies allowed us to identify multiple isolates, including isolates from a novel Rhizobium species, that consistently fix nitrogen with, and enhance the growth of, common bean plants. Our findings highlight strong biogeographical structuring of rhizobial communities and demonstrate that Ontario soils already harbour strains with high symbiotic potential. In addition, our Rhizobium collection represents a foundational resource to support future inoculant development and enables future work on the ecology, evolution, and applied optimization of legume-rhizobium symbioses.

Studies of plant-associated microbial communities consistently indicate a role for classic assembly mechanisms, such as environmental and host filters, but often leave substantial unexplained variation. Biotic interactions within microbial communities may help to fill this gap, specifically cross-kingdom interactions between fungi and bacteria, as these are increasingly found to be important to both assembly and function. We hypothesized that direct interactions between bacteria and fungi are an important driver of composition in low-diversity leaf habitats, where pairwise interactions are more likely. In high-diversity root habitats, we expected diffuse, indirect interactions to be more relevant to composition. To test these hypotheses, we characterized bacterial and fungal communities of switchgrass (Panicum virgatum L.) leaves and roots at 14 sites spanning mountain to coastal ecoregions of North Carolina, USA. We analyzed putative direct and diffuse interactions using ecological network inference and partitioned variance explained in microbial community composition by spatial, environmental, and biotic interactions. We found that cross-kingdom biotic interactions contributed to microbial community structure. The largest improvements to variance explained (5-11%) were from direct interactions, except for root fungal communities where diffuse interactions (7.5%) explained more than double that of direct interactions (2.8%). These contributions were comparable to those from environmental and spatial factors. The joint effects of putative biotic interactions and environmental conditions also contributed to the explained variation, highlighting the importance of environmental tracking in microbes. These findings suggest that using network inference for identifying cross-kingdom ecological interactions can improve our fundamental understanding of how plant-associated microbiomes assemble, which is also directly relevant to applied efforts such as the effective development of synthetic communities.

Beneficial interactions between plants and microorganisms strongly influence plant health and productivity, and root exudates play a central role in shaping these associations. This study analyzed the transcriptional responses of the bacterial endophytes Enterobacter asburiae RCA24 and Kosakonia sacchari RCA25 to root exudates from two commercial Italian rice accessions (Oryza sativa Baldo and Vialone Nano) and from an accession of the wild progenitor of tropical rice, Oryza rufipogon. Bacterial transcriptome analyses revealed that RCA24 responds differently to O. sativa varieties and that RCA25 was more stimulated by O. rufipogon. Changes in bacterial gene expression were mainly related to central metabolism, stress response, and signal transduction, highlighting a precise pattern of interaction. On the other hand, transcriptome analysis of inoculated rice revealed that RCA24 triggered broader transcriptional changes in plants than RCA25. Differentially expressed genes were related, especially in shoots, to defense responses, hormone-mediated signaling, and ribosome biogenesis, revealing that plants discriminate bacterial strains in a genotype-specific manner at the transcriptional level. Our findings suggest that traits beneficial to plant-soil microbiota interactions present in O. rufipogon and lost during domestication and diversification could be identified and reintroduced into modern rice varieties to improve sustainable field performance through beneficial microbial associations.

Climate change and increasing drought conditions significantly impedes citrus productivity in subtropical and tropical regions. This study explores the potential of combining arbuscular mycorrhizal fungi (AMF) Funneliformis mosseae and plant growth-promoting rhizobacteria (PGPR) Pseudomonas putida to mitigate drought resilience in Citrus reticulata (Red tangerine). AMF-mediated drought tolerance has been extensively documented; however, the collegial influence of PGPR and AMF on phytohormone signaling, photosynthetic efficiency, nutrient acquisition, and gene expression remains largely unexplored in citrus. We conducted a greenhouse experiment under both well water and drought stress conditions to assess the physiological and molecular responses to individual and co-inoculation with PGPR and AMF. Drought-stressed citrus plants, inoculated with AMF and PGPR, demonstrated significantly improved leaf water potential, stomatal conductance, carbon assimilation, and antioxidant defense. PGPR-AMF co-inoculation enhanced chlorophyll stability, osmotic adjustment, and nutrient uptake, while significantly reducing lipid peroxidation and ROS accumulation. The turquoise module emerged from transcriptomic and gene co-expression network analysis (WGCNA) as a potential key regulator of stress adaptation, revealed key regulatory transcription factors, e.g., CrMYB4, CrZFP8, CrSOS5, CrRGFR2, and CrQUA1, that were upregulated under combined inoculation, highlighting their potential role in stress adaptation. Our findings demonstrate that the synergistic PGPR-AMF interaction improves antioxidant enzyme activities and modulates gene expression to promote drought tolerance, providing new insights into the microbiomes role in plant resilience. These results offer a potential strategy to boost citrus growth and yield under water scarcity, with broad implications for agricultural resilience to climate change.

Microbial communities regulate carbon and nitrogen (N) cycling, yet their long-term responses to chronic global changes remain unclear. Using 12 years of grassland litter samples from the Loma Ridge Global Change Experiment in Irvine, California, we tested whether interactions between experimental drought and N deposition, and previously observed temporal variability are driven by background climatic conditions, including precipitation and temperature. Consistent with short-term studies, drought and N addition had relatively small effects on bacterial community composition compared to pronounced seasonal and interannual variability, with drought-by-year interactions explaining more variation than drought alone. Seasonal shifts were largely driven by short-term fluctuations in rainfall and temperature, whereas the substantial interannual variability in community composition was not captured by site-level climate metrics. Contrary to expectations, drought effects were influenced more by background temperature than precipitation, with the strongest effects observed in cooler years. Lastly, a bacterial taxons sensitivity to climate variability under ambient conditions did not predict its response to chronic drought. Together, our findings show that bacterial responses to drought are temporally dynamic and influenced by background temperature, underscoring the need for long-term longitudinal studies of soil microbial communities to better predict microbial responses under future global change. ImportanceMicrobial responses to global change, particularly drought and nitrogen addition, are often inferred from short-term studies (< 2 years), yet natural temporal variability may overshadow experimental effects. Using a 12-year dataset of grassland leaf litter communities, we show that temporal variability, both seasonal and interannual, exert a stronger influence on bacterial community composition than chronic drought or nitrogen deposition. These findings challenge assumptions about the magnitude of drought effects, particularly in naturally drought-affected ecosystem such as California grasslands and highlight the importance of long-term datasets for predicting microbial responses to climate change. By demonstrating that bacterial communities are strongly shaped by background climatic variability (baseline precipitation and temperature independent of imposed chronic treatments) and may be buffered to sustained drought, this work improves forecasts of ecosystem responses and informs the design of global change experiments and restoration strategies in future research studies.

BackgroundPotato is an important crop worldwide, yet its production is severely threatened by Phytophthora infestans, the causal agent of late blight. Alternatives to the current control strategies are needed, as these rely heavily on environmentally harmful treatments. The recruitment of beneficial microbes by plants upon stress ("cry-for-help" mechanism) may represent an opportunity to find new biocontrol agents but this has not yet been reported for potato. The aim of this study was to analyse whether foliar late blight infection induces shifts in the phyllosphere, rhizosphere and soil bacterial communities associated with two potato cultivars of differing sensitivity to late blight. Moreover, we aimed at isolating members of the plant microbiota to test whether bacteria putatively recruited upon infection would be particularly active in protecting the plant against late blight. ResultsControlled foliar infection triggered substantial, cultivar-specific shifts in the rhizosphere communities across two successive generations. Despite the number of differentially abundant ASVs detected being ten times higher in the second generation than in the first one, the same taxonomic groups were concerned by the shifts: Burkholderiales, Flavobacteriales, and Bacillales. Furthermore, the communities linked to the susceptible cultivar consistently shifted more strongly than the communities linked to the resistant cultivar. The obtained ASV sequences were used to identify 163 corresponding isolates. The inhibition potential of these strains against P. infestans spores was assessed through biological assays, which revealed the biocontrol potential of strains otherwise not yet known to inhibit phytopathogenic organisms, such as Advenella, Nocardioides and Phyllobacterium strains. Although we found no correlation between the relative abundance shift of the ASVs upon infection and the activity of the corresponding strains, we observed that the overall activity of strains isolated from the resistant cultivar was higher than that of the strains isolated from the susceptible one. ConclusionTaken together, the higher activity of the strains isolated from the resistant cultivar, along with its comparatively modest microbiome shifts upon infection suggest that the investigated resistant cultivar might harbour specific microbiota enriched in strains with efficient protective abilities against their host plants pathogens, which possibly contribute to its higher resistance against P. infestans.

Seed microbiota play a crucial role in plant health and development, yet remain understudied compared to other plant-associated microbial communities. This study aimed to characterize seed microbiota diversity across four major crops (common bean, rapeseed, tomato, and wheat) and establish a comprehensive strain collection of seed-borne microorganisms (bacteria and fungi). We employed a combination of culture-dependent and culture-independent approaches to analyze 68 seed samples representing diverse genotypes and production modes. Our results revealed highly variable seed microbiota, with bacterial colonization ranging from 10 to 100 million bacterial CFUs per gram of seeds, and microbial richness varying from 4 to 351 bacterial and 16 to 138 fungal amplicon sequence variants (ASVs) per sample. Both plant genotype and production mode significantly influenced microbiota composition, with each seed sample produced harboring a distinct microbial assemblage. Interestingly, seeds produced in confined environments exhibited lower bacterial colonization but higher microbial richness compared to field-produced seeds. We observed divergent ecological drivers shaping bacterial and fungal communities. Bacterial assemblages were more host-specific and variable, while fungal communities showed greater stability and a substantial core microbiome shared across plant species. Our culturomics approach yielded a collection of 2,510 bacterial and 837 fungal isolates, representing 10-21% of the seed microbiota diversity detected by metabarcoding and the majority of the prevalent and abundant taxa. Notably, 44-60% of cultured bacterial isolates were not detected by metabarcoding, highlighting the complementary nature of these approaches to detect rare or under amplified taxa in PCR. This study provides insights into the complexity and variability of seed microbiota across different crops and production conditions. Our findings emphasize the importance of combining culturomics and sequencing methods for comprehensive characterization of seed microbiota to uncover the potential of seed-borne microorganisms as bioinoculants for sustainable agriculture.