Nodules Of Plants Research Articles

Coinoculação com Bradyrhizobium ssp. e Azospirillum brasilense em feijão caupi sob estresse salino

ABSTRACT Plants grown under salinity are subject to many morphological and physiological disorders. In this context, practices that can enable the use of saline water are essential. The deleterious effects of salinity can be mitigated by using beneficial microorganisms, especially diazotrophic bacteria. In this context, this study aimed to evaluate the effect of bacterial inoculation and co-inoculation using Bradyrhizobium spp. and Azospirillum brasilense on the growth, nodulation and production of cowpea under salt stress. The experiment was conducted using a randomized block design, in a 4 × 5 factorial arrangement, corresponding to four nitrogen sources (N1 - without nitrogen and without inoculant; N2 - fertilization with mineral nitrogen and without inoculant; N3 - inoculation of Bradyrhizobium spp.; and N4 - co-inoculation of Bradyrhizobium spp. and Azospirillum brasilense) and five values of electrical conductivity of irrigation water - ECw (0.4, 1.9, 3.4, 4.9 and 6.4 dS m-1), with five replicates. Co-inoculation with Azospirillum brasilense and Bradyrhizobium spp. favors the growth, production and nodulation of cowpea plants up to ECw of 0.4 dS m-1. Increase in the electrical conductivity of irrigation water negatively affects the growth, production and nodulation of cowpea plants in Corujinha variety, regardless of the nitrogen source used.

Comparative study of symbiotic activity of legumes when using Risotorphin and Epin-extra.

In a vegetation experiment with soybean plants of the Svapa and Mageva varieties and in a field experiment with bean plants of the Geliada and Shokoladnitsa varieties, we studied the effect of pre-sowing treatment of the seeds of these plants with Rizotorfin and Epin-extra on the nitrogenase activity of the nodules of these plants and their ultrastructure. Analysis of the ultrastructure of the nodule tissue of beans and soybeans was carried out in the flowering phase. It was found that the highest indices of the mass and number of nodules and the activity of nitrogenase in them were found in bean plants of the Heliada cultivar when the seeds were treated with Epin-extra against the background of inoculation with Rizotorfin, where the largest area of symbiosomes, volutin and their number was noted in the nodules. Beans of the Shokoladnitsa variety showed the protective effect of Rizotorfin. In the nodules of soybean plants of the Svapa variety, the seeds of which were treated with Epin-extra against the background of inoculation with Rizotorfin, the presence of a large number of symbiosomes, bacteroids, volutin inclusions with a larger area and a minimum number of inclusions of poly-β-hydroxybutyric acid (PHB) was noted, and the highest indicators of symbiotic activity. Soybean plants of the Mageva variety showed the protective effect of Rizotorfin. The efficiency of the symbiotic system was determined by the number and weight of nodules and the activity of the nitrogenase enzyme.

Morpho-Physiological and Biochemical Characterization of Soybean-Associated Rhizobia and Effect of Their Liquid Inoculant Formulation on Nodulation of Host Plants in the Cameroon Cotton Fields Zone

Morpho-Physiological and Biochemical Characterization of Soybean-Associated Rhizobia and Effect of Their Liquid Inoculant Formulation on Nodulation of Host Plants in the Cameroon Cotton Fields Zone

Indigenous diazotrophs and their effective properties for organic agriculture

Nitrogen-fixing microorganisms (diazotrophs) converting the atmospheric N2 into usable form NH4 are considered the key players in the nitrogen cycle, chiefly responsible for the enriching nitrogen content in the soils. Globally, biological fixation of N2 greatly contributes to plant growth, lessens the need for chemical fertilizers, and thus contributes to the mitigation of greenhouse gases NOx. In this study, diazotrophic bacterial strains were isolated from rhizosphere soils and root nodules of legume and non-legume plants in Vietnam. Quantitative analyzes by the acetylene reduction assay showed that the isolates have high nitrogen fixation activity compared with that of reference strain Azospirillum vinelandii KCTC 2426. In addition, other effective capabilities of the isolated strains toward supporting agriculture were investigated, i.e. synthesizing IAA and siderophore for promoting plant growth, or producing exopolysaccharides for maintaining soil moisture. Taxonomic positions of the isolated strains were identified based on the comparative analyses of sequences of the 16S rDNA and gene related to nitrogen fixation (nifH), revealing a high taxonomic diversity among free-living and symbiotic diazotrophic isolates. Direct support of the selected isolates to plant growth was proven in experiments with mung beans under laboratory conditions. Thus, the native diazotrophic strains obtained in this study would be good microbial sources for application in organic agriculture and soil amendment.

The effect of infection with soybean mosaic virus on the growth and formation of bacterial nodules of soybean plants

دلت نتائج الدراسة على ان الاصابة بموزائيك فول الصويا ادت الى اختزال معنوي في عدد ووزن العقد البكتيرية اضافة الى اختزال الوزن الجاف للبذور وقد كانت نسب الاختزال محسوبة على اساس معاملة المقارنة .

Systemic control of symbiotic nodulation in legume plants: genetic engineering in functional studies of key regulators

Legumes are important suppliers of vegetable protein. In symbiotic interactions with soil bacteria rhizobia, legume plants form nitrogen-fixing nodules on their roots, where molecular nitrogen is fixed and incorporated into organic compounds. A host plant controls the number of symbiotic nodules to meet its nitrogen demands. The presence of high amount of nitrate in the soil suppresses the formation of nodules. CLE (CLAVATA3/EMBRYO SURROUNDING REGION) peptides produced in the root in response to rhizobial inoculation and/or nitrate were shown to control the number of symbiotic nodules. Previously, we have identified the MtCLE35 gene upregulated by the rhizobia and the nitrate treatment in Medicago truncatula, which systemically inhibited nodulation when overexpressed. Using genetic engineering approaches we increased its transcriptional activity in transgenic roots, which almost completely prevented the formation of symbiotic nodules. Moreover, we obtained stable transgenic lines overproducing the MtCLE35 peptide, and found that their had lower shoot and root biomass in comparison to the wild-type plants. The study of metabolome of MtCLE35-overproducing plants revealed the increased level of amino acids in the roots, suggesting the stimulating effect of MtCLE35 on amino acid synthetic pathways.
 In addition, we obtained several knock-out lines, where the MtCLE35 gene was edited using the CRISPR/Cas9-mediated system. The detailed analysis of these plants will allow us to understand the mechanisms of MtCLE35 action in the regulation of plant nitrogen status and symbiotic interaction with rhizobia.
 This work was supported by the grant from Saint Petersburg State University ID 93020341 and by the Russian Foundation for Basic Research project 20-016-00129.

Response of groundnut to phosphorus and bio-organics in Alfisols of Red and Laterite zone of India

Present study was carried out for three consecutive kharif season of 2018, 2019 and 2020 to evaluate the performance of different levels of chemical phosphatic fertilizers and FYM in combination with PSB inoculant on groundnut growth, yield, quality and nutrient uptake in an acidic Red and Laterite soil. Application of higher dose of phosphatic fertilizer along with PSB inoculant yielded better results with respect to groundnut plant height, nodulation, pegs and pods plant−1, weight of mature pods plant−1, kernel and haulm yield, protein and oil content, and nutrient uptake. Treatment combination T7 (60 kg ha−1 of P2O5 + PSB inoculant) was observed as the best treatment combination for deriving maximum growth, yield and nutrients uptake of kharif groundnut crop. Treatment T6 (application of 40 kg ha−1 of P2O5 + PSB inoculant) reported similar results. However, maximum profit ($862), benefit cost ratio (2.7) and net return per unit cost of input was significantly higher in T7 compared to T6. Thus, we recommend application of 60 kg ha−1 of P2O5 with PSB inoculant (T7) for the light textured acid soils of Red and Laterite zone of India for higher production and profitability.

Genetic and Physiological Characterization of Soybean-Nodule-Derived Isolates from Bangladeshi Soils Revealed Diverse Array of Bacteria with Potential Bradyrhizobia for Biofertilizers.

Genetic and physiological characterization of bacteria derived from nodules of leguminous plants in the exploration of biofertilizer is of paramount importance from agricultural and environmental perspectives. Phylogenetic analysis of the 16S rRNA gene of 84 isolates derived from Bangladeshi soils revealed an unpredictably diverse array of nodule-forming and endosymbiotic bacteria-mostly belonging to the genus Bradyrhizobium. A sequence analysis of the symbiotic genes (nifH and nodD1) revealed similarities with the 16S rRNA gene tree, with few discrepancies. A phylogenetic analysis of the partial rrn operon (16S-ITS-23S) and multi-locus sequence analysis of atpD, glnII, and gyrB identified that the Bradyrhizobium isolates belonged to Bradyrhizobium diazoefficiens, Bradyrhizobium elkanii, Bradyrhizobium liaoningense and Bradyrhizobium yuanmingense species. In the pot experiment, several isolates showed better activity than B. diazoefficiens USDA110, and the Bho-P2-B2-S1-51 isolate of B. liaoningense showed significantly higher acetylene reduction activity in both Glycine max cv. Enrei and Binasoybean-3 varieties and biomass production increased by 9% in the Binasoybean-3 variety. Tha-P2-B1-S1-68 isolate of B. diazoefficiens significantly enhanced shoot length and induced 10% biomass production in Binasoybean-3. These isolates grew at 1-4% NaCl concentration and pH 4.5-10 and survived at 45 °C, making the isolates potential candidates for eco-friendly soybean biofertilizers in salty and tropical regions.

The Formation of Symbiotic Potential and Yields of Soybean Depending on Elements of Growing Technology

The study of the nitrogen-fixing activity of nodule bacteria of soybean plants and features of the formation of the number and mass of nodules depending on variety, sowing method and seeding rate influenced by water regime of the soil and different lighting intensity.
 Methods: Field and laboratory experiments, evaluation and comparison. A series of laboratory experiments in sand culture have explored the influence of water logging and dehydration of the root system and light intensity on soybean nitrogenase activity and the release of CO2 by nodules of plants. According to the results of field research it was found that the intensity of nodule formation was increased from the phase of the third trifoliate leaf prior to flowering. At a sowing rate of 500 seeds it made from 20,0 PCs. for the variety Ustya to 27,4 in the variety of Romance. The increase in seeding rate contributed to the increase in the number of nodules to 28,1 per piece (Romance), 27,5 pieces (Ustya) and 25,5 units (Vorskla). The mass of active nodules was within 75% of the total number. Seed rate more than method of sowing influenced the amount of soybean yield. Increase the seeding rate to 800 thousand/ha of viable seeds, especially of sowing on later dates, have not contributed to a significant increase in productivity. Therefore it is most expedient to sow soybean normal line (15 cm) or in wide (45 cm) methods with a seeding rate of 700 thousand/ha of viable seeds. In the line way of sowing increasing the seeding rate contributed to the increase in the number of nodules to 28,1 per piece (Romance), 27,5 pieces (Ustya) and 25,5 units (Vorskla). In wide way of sowing these figures were less than 4,5% to 10,6%. The mass of active nodules was within 75% of the total number. It is advisable to sow soybean normal line (15 cm) or in wide (45 cm) methods with a seeding rate of 700 thousand/ha of viable seeds.

Isolation, Identification and Evaluation of the Plant Growth Promoting Activities of Endophytic Stenotrophomonas maltophilia to Stimulate Growth of Clover Plants under Salt Stress

Two endophytic bacterial isolates were obtained from root nodules of clover plants grown in salt affected clay soil of Egypt. The isolates were closely linked to Stenotrophomonas maltophilia strains IPR-Pv696 and 262XG2 based on the sequencing and phylogenetic analysis of 16S rRNA genes, and deposited in GenBank with accession numbers OM980221.1 (AM1) and OM980223.1 (AM2) respectively. The isolates were evaluated for their potential to promote plant growth. The results revealed that the two isolates of S. maltophilia strains (IPR-Pv696 and 262XG2) respectively exhibited production for indole-3- acetic acid (30.26 & 31.15 µg/ml), exopolysaccharides (13.57 & 13.68 g/l), nitrogen fixation activity and they solubilize the phosphate (278 & 208 mg/l) and potassium (33.5 & 32.9 µg/ml). In a field trial, these two isolates increased clover plant growth, chlorophyll, carbohydrates content and nutrients uptake while lowering proline levels. Hence this highlights its application to be exploited as biofertilizer by leading to sustainable agriculture. This could be a promising inoculant for many other crops.

Native Rhizobia Improve Plant Growth, Fix N2, and Reduce Greenhouse Emissions of Sunnhemp More than Commercial Rhizobia Inoculants in Florida Citrus Orchards.

Sunnhemp (Crotalaria juncea L.) is an important legume cover crop used in tree cropping systems, where there is increased interest by growers to identify rhizobia to maximize soil nitrogen (N) inputs. We aimed to isolate and identify native rhizobia and compare their capabilities with non-native rhizobia from commercial inoculants to fix atmospheric dinitrogen (N2), produce and reduce nitrous oxide (N2O), and improve plant growth. Phylogenetic analyses of sequences of the 16S rRNA and recA, atpD, and glnII genes showed native rhizobial strains belonged to Rhizobium tropici and the non-native strain to Bradyrhizobium japonicum. Plant nodulation tests, sequencing of nodC and nifH genes, and the acetylene-dependent ethylene production assay confirmed the capacity of all strains to nodulate sunnhemp and fix N2. Inoculation with native rhizobial strains resulted in significant increases in root and shoot weight and total C and N contents in the shoots, and showed greater N2-fixation rates and lower emissions of N2O compared to the non-native rhizobium. Our results suggest that native rhizobia improve plant growth, fix N2, and reduce greenhouse emissions of sunnhemp more than commercial rhizobia inoculants in Florida citrus orchards.

RopB protein of Rhizobium leguminosarum bv. viciae adopts amyloid state during symbiotic interactions with pea (Pisum sativum L.).

Amyloids represent protein aggregates with highly ordered fibrillar structure associated with the development of various disorders in humans and animals and involved in implementation of different vital functions in all three domains of life. In prokaryotes, amyloids perform a wide repertoire of functions mostly attributed to their interactions with other organisms including interspecies interactions within bacterial communities and host-pathogen interactions. Recently, we demonstrated that free-living cells of Rhizobium leguminosarum, a nitrogen-fixing symbiont of legumes, produce RopA and RopB which form amyloid fibrils at cell surface during the stationary growth phase thus connecting amyloid formation and host-symbiont interactions. Here we focused on a more detailed analysis of the RopB amyloid state in vitro and in vivo, during the symbiotic interaction between R. leguminosarum bv. viciae with its macrosymbiont, garden pea (Pisum sativum L.). We confirmed that RopB is the bona fide amyloid protein since its fibrils exhibit circular x-ray reflections indicating its cross-β structure specific for amyloids. We found that fibrils containing RopB and exhibiting amyloid properties are formed in vivo at the surface of bacteroids of R. leguminosarum extracted from pea nodules. Moreover, using pea sym31 mutant we demonstrated that formation of extracellular RopB amyloid state occurs at different stages of bacteroid development but is enhanced in juvenile symbiosomes. Proteomic screening of potentially amyloidogenic proteins in the nodules revealed the presence of detergent-resistant aggregates of different plant and bacterial proteins including pea amyloid vicilin. We demonstrated that preformed vicilin amyloids can cross-seed RopB amyloid formation suggesting for probable interaction between bacterial and plant amyloidogenic proteins in the nodules. Taken together, we demonstrate that R. leguminosarum bacteroids produce extracellular RopB amyloids in pea nodules in vivo and these nodules also contain aggregates of pea vicilin amyloid protein, which is able to cross-seed RopB fibrillogenesis in vitro. Thus, we hypothesize that plant nodules contain a complex amyloid network consisting of plant and bacterial amyloids and probably modulating host-symbiont interactions.

Strains of Bradyrhizobium barranii sp. nov. associated with legumes native to Canada are symbionts of soybeans and belong to different subspecies (subsp. barranii subsp. nov. and subsp. apii subsp. nov.) and symbiovars (sv. glycinearum and sv. septentrionale).

Four bacterial strains isolated from root nodules of soybean plants that had been inoculated with root-zone soils of legumes native to Canada were previously identified as a novel Bradyrhizobium lineage consisting of symbiovars (sv.) glycinearum and septentrionale. Our purpose was to verify the taxonomic status of these strains using phylogenetic, genomic and phenotypic analyses. Multiple phylogenetic analyses including analysis of 51 full-length ribosome protein subunit (rps) gene sequences confirmed placement of the novel strains in a highly supported lineage distinct from named Bradyrhizobium species with B. japonicum USDA 6T as the closest relative. The results of genomic and phylogenomic analyses based on digital DNA–DNA hybridization and genome blast distance phylogeny showed that novel strains in comparisons with type strains of closest relatives were below the established threshold (70 %) for species delineation. Moreover, the novel strains were divided into two subspecies clusters based on the established threshold of 79 %. The genomes of strains 144S4T, 323S2, 1S5 and 38S5T have sizes of 11 399 526, 11 474 152, 10580853 and 10 530 141 bp with DNA G+C contents of 63.1, 63.0, 63.4 and 63.3 mol%, respectively. These strains possess symbiosis islands harbouring key nodulation, nitrogen-fixation and type III secretion system genes as well as abundant insertion sequences and between two and four putative plasmids. Strains 144S4T and 323S2 (sv. glycinearum) are effective with regard to nitrogen fixation in symbiotic association with soybeans whereas strains 1S5 and 38S5T (sv. septentrionale) are ineffective. Data for morphological, physiological and symbiotic characteristics complement the sequence-based results. The data presented here support the description of a new species and two new subspecies for which the names Bradyrhizobium barranii sp. nov. subsp. barranii subsp. nov. (sv. glycinearum) and Bradyrhizobium barranii sp. nov. subsp. apii subsp. nov. (sv. septentrionale) are proposed with strain 144S4T (=LMG 31552T=HAMBI 3722T) as the species type strain and type strain of subsp. barranii subsp. nov., and strain 38S5T (=LMG 31556T=HAMBI 3721T) as the type strain of subsp. apii subsp. nov.

Genotypic variability in nutrient uptake and use efficiency in chickpea grown under low phosphorus availability in a Mediterranean climate

Low phosphorus (P) availability is a major constraint for chickpea production. Consequently, P-efficient genotypes can improve productivity under conditions where the higher application of P is not economical. This study was conducted to characterise four chickpea genotypes for nutrient uptake and use efficiency under low-P conditions over two growing seasons. At flowering stage, plants were harvested and analysed for their nodulation, growth, P content and yield. Results indicate that low P availability significantly limited plant growth, nodulation and yield for all genotypes with the greatest effect on for Flip 84-92C and Flip 01-29C. The genotypes Flip 90-13C and ILC 32-79 showed the highest P uptake and use efficiency. The genotypes with high nutrient uptake had better efficiency in use of rhizobial symbiosis. It is concluded that nutrient uptake and use efficiency may be an important functional trait that may contribute to the selection of cultivars able to produce high quality seeds and efficiently fix nitrogen under conditions of low soil P.

Enhanced legume growth and adaptation to degraded estuarine soils using Pseudomonas sp. nodule endophytes.

The joint estuary of Tinto and Odiel rivers (SW Spain) is one of the most degraded and polluted areas in the world and its recovery is mandatory. Legumes and their associated bacteria are recommended sustainable tools to fight against soils degradation and loss of fertility due to their known positive impacts on soils. The aim of this work was to isolate and characterize plant growth promoting nodule endophytes (PGPNE) from inside nodules of Medicago spp. naturally growing in the estuary of the Tinto and Odiel Rivers and evaluate their ability to promote legume adaptation in degraded soils. The best rhizobia and non-rhizobia among 33 endophytes were selected based on their plant growth promoting properties and bacterial enzymatic activities. These strains, identified as Pseudomonas sp. N4, Pseudomonas sp. N8, Ensifer sp. N10 and Ensifer sp. N12, were used for in vitro studies using Medicago sativa plants. The effects of individual or combined inoculation on seed germination, plant growth and nodulation were studied, both on plates and pots containing nutrient-poor soils and moderately contaminated with metals/loids from the estuary. In general, inoculation with combinations of rhizobia and Pseudomonas increased plant biomass (up to 1.5-fold) and nodules number (up to 2-fold) compared to single inoculation with rhizobia, ameliorating the physiological state of the plants and helping to regulate plant stress mechanisms. The greatest benefits were observed in plants inoculated with the consortium containing the four strains. In addition, combined inoculation with Ensifer and Pseudomonas increased As and metals accumulation in plant roots, without significant differences in shoot metal accumulation. These results suggest that PGPNE are useful biotools to promote legume growth and phytostabilization potential in nutrient-poor and/or metals contaminated estuarine soils.

Effect of Different Cover Crops on Suppression of the Weed Oxalis pes-caprae L., Soil Nutrient Availability, and the Performance of Table Olive Trees ‘Kalamon’ cv. in Crete, Greece

Winter cover crops are used in organic olive production to increase N-supply and yields, and to reduce weed competition. However, there is limited information on the effect of different cover crops on weed suppression, soil fertility and productivity of organic olive orchards. Here, we compared the relative effect of four contrasting cover crops established from (i) untreated vetch seed, (ii) vetch seed inoculated with a commercial Rhizobium seed inoculum, (iii) an untreated vetch/barley/pea seed mixture and (iv) untreated seed of Medicago polymorpha L. (a native legume species which establishes naturally in olive orchards in Crete) in a 35-year-old experimental table olive orchard. The use of a vetch/barley/pea mixture resulted in the greatest suppression of the dominant weed species Oxalis pes-caprae. Rhizobium inoculation of vetch seed resulted in significantly lower vetch establishment and significantly higher Oxalis suppression but had no significant effect on the root nodulation of vetch plants. There was no significant difference in fruit yield between cover crop treatments, but the fruit weight was significantly higher when cover crops were established from un-treated vetch seeds and the vetch/barley/pea seed mixture compared with the cover crops based on inoculated vetch or Medicago seed. However, although Medicago establishment was very low (<10 plants/m2), fruit yields were numerically 20% higher in the Medicago plots. These findings suggests that, overall, legume cover crops had no effect on fruit yields. This conclusion is supported by the results of the olive leaf analyses which detected no significant differences in nitrogen and other mineral macro- and micronutrient concentration between treatments, except for B (highest in olive leaves from Medicago and lowest in untreated vetch plots) and Mo (highest in olive leaves from Medicago and lowest in vetch/barley/pea mixture plots). Overall, our results suggest that the current recommendation to establish legume-based cover crops in organic olive orchards every year, may need to be revised and that establishing cover crops every 2–4 years may reduce costs without affecting olive fruit yields.

The Effects of Phyllosphere Bacteria on Plant Physiology and Growth of Soybean Infected with Pseudomonas syringae.

Phyllosphere bacteria are an important determinant of plant growth and resistance to pathogens. However, the efficacy of phyllosphere bacteria in regulating infection of Pseudomonas syringae pv. glycinea (Psg) and its influence on soybean growth and physiology is unknown. In a greenhouse study, we assessed the influence of a phyllosphere bacterial consortium (BC) of 13 species isolated from field-grown soybean leaves on uninfected and deliberately Psg infected soybean plants. We measured Psg density on infected leaves with and without the application of the BC. The BC application resulted in a significant reduction in Psg cells. We also measured plant biomass, nodule mass and number, gas exchange, and leaf chlorophyll and nitrogen in four treatment groups: control plants, plants with a BC and no infection (BC), plants with BC and infected with Psg (BC + Psg), and plants infected with Psg alone. For all variables, plants infected with Psg alone showed significant reduction in measured variables compared to both BC treatments. Therefore, the bacterial consortium was effective in controlling the negative effects of Psg on growth and physiology. The BC treatment sometimes resulted in increases in measured variables such as plant biomass, nodule numbers, and leaf chlorophyll as compared to control and BC + Psg treatments. Overall, the positive influence of BC treatment on plant growth and physiology highlights its potential applications to increase crop yield and control bacterial pathogens.

Can Organic P Inputs Alleviate P Limitation Effects on Nutrient Uptake and Biological N2-Fixing Capacity of Hairy Vetch (Vicia villosa)?

Phosphorus (P) is a limiting nutrient in many agroecosystems and, apart from affecting plant growth, can also limit biological N2 fixation (BNF) by leguminous plants. Thus, increasing P supply can have a positive effect on BNF particularly in P-deficient soils. Here, we provide new insights into the response of hairy vetch (Vicia villosa), widely adopted as a legume cover crop, to P limitations, by comparing the effects of inorganic (Pi) and organic (Po) P supply on plant growth and BNF capacity. This was achieved by means of a greenhouse experiment in which rhizobia-inoculated hairy vetch was grown in a P-limited agricultural soil and changes in plant growth, nitrogen (N) and P uptake, BNF capacity, and soil phosphatases activities were evaluated as a function of Pi and Po inputs, in the form of orthophosphate or phytic acid, respectively. When compared to P-deficient conditions where BNF was primarily limited by plant growth rather than directly due to the high P costs of symbiotic N fixation, Pi addition substantially enhanced plant growth (threefold), nodule formation (16-fold), P acquisition (sixfold), and BNF efficiency (sevenfold). In contrast, even with the addition of the highest dose of Po, the increase in plant growth, nodule formation, P acquisition, and BNF capacity (1.7, 3.5, 2.4 and 2.1-fold, respectively) was much less expressed, indicating that hairy vetch could only minimally access Po sources over the growth period in order to alleviate the P limitation effect on N2 fixation in under P-deficient conditions. These findings suggest that hairy vetch will not be able to provide sufficient BNF for improving soil N inputs in low-fertility cropping systems that rely on organic inputs.

A novel function of the key nitrogen-fixation activator NifA in beta-rhizobia: Repression of bacterial auxin synthesis during symbiosis.

Rhizobia fix nitrogen within root nodules of host plants where nitrogenase expression is strictly controlled by its key regulator NifA. We recently discovered that in nodules infected by the beta-rhizobial strain Paraburkholderia phymatum STM815, NifA controls expression of two bacterial auxin synthesis genes. Both the iaaM and iaaH transcripts, as well as the metabolites indole-acetamide (IAM) and indole-3-acetic acid (IAA) showed increased abundance in nodules occupied by a nifA mutant compared to wild-type nodules. Here, we document the structural changes that a P. phymatum nifA mutant induces in common bean (Phaseolus vulgaris) nodules, eventually leading to hypernodulation. To investigate the role of the P. phymatum iaaMH genes during symbiosis, we monitored their expression in presence and absence of NifA over different stages of the symbiosis. The iaaMH genes were found to be under negative control of NifA in all symbiotic stages. While a P. phymatum iaaMH mutant produced the same number of nodules and nitrogenase activity as the wild-type strain, the nifA mutant produced more nodules than the wild-type that clustered into regularly-patterned root zones. Mutation of the iaaMH genes in a nifA mutant background reduced the presence of these nodule clusters on the root. We further show that the P. phymatum iaaMH genes are located in a region of the symbiotic plasmid with a significantly lower GC content and exhibit high similarity to two genes of the IAM pathway often used by bacterial phytopathogens to deploy IAA as a virulence factor. Overall, our data suggest that the increased abundance of rhizobial auxin in the non-fixing nifA mutant strain enables greater root infection rates and a role for bacterial auxin production in the control of early stage symbiotic interactions.

Symbiotic Nodule Development and Efficiency in the Medicago truncatula Mtefd-1 Mutant Is Highly Dependent on Sinorhizobium Strains.

Symbiotic nitrogen fixation (SNF) can play a key role in agroecosystems to reduce the negative impact of nitrogen fertilizers. Its efficiency is strongly affected by the combination of bacterial and plant genotypes, but the mechanisms responsible for the differences in the efficiency of rhizobium strains are not well documented. In Medicago truncatula, SNF has been mostly studied using model systems, such as M. truncatula A17 in interaction with Sinorhizobium meliloti Sm2011. Here we analyzed both the wild-type (wt) A17 and the Mtefd-1 mutant in interaction with five S. meliloti and two Sinorhizobium medicae strains. ETHYLENE RESPONSE FACTOR REQUIRED FOR NODULE DIFFERENTIATION (MtEFD) encodes a transcription factor, which contributes to the control of nodule number and differentiation in M. truncatula. We found that, in contrast to Sm2011, four strains induce functional (Fix+) nodules in Mtefd-1, although less efficient for SNF than in wt A17. In contrast, the Mtefd-1 hypernodulation phenotype is not strain-dependent. We compared the plant nodule transcriptomes in response to SmBL225C, a highly efficient strain with A17, versus Sm2011, in wt and Mtefd-1 backgrounds. This revealed faster nodule development with SmBL225C and early nodule senescence with Sm2011. These RNA sequencing analyses allowed us to identify candidate plant factors that could drive the differential nodule phenotype. In conclusion, this work shows the value of having a set of rhizobium strains to fully evaluate the biological importance of a plant symbiotic gene.