Common Bean Nodules Research Articles

Diversity and Phylogeny of Symbiotic Bacteria Nodulating Common Bean (Phaseolus vulgaris L.) in Côte d’Ivoire

Phaseolus vulgaris L. (common bean) is an agriculturally important legume that benefits from a symbiosis with bacteria belonging to the genus Rhizobium. Growing interest in the use of rhizobia as biofertilizers has led to the identification of a large number of rhizobia strains and studies of their diversity. Although much research has been carried out on rhizobia, there is little data on the diversity of rhizobia associated with common bean in Côte d'Ivoire. This study assessed the species diversity of common bean nodulating bacteria in ivorian soils. This diversity was assessed based on 16S rRNA gene sequencing. Ten high-performance bacterial isolates extracted from common bean nodules were used for genetic analysis. The 16S rRNA gene sequences of the native isolates were closely affiliated with members of the genera Rhizobium, Bradyrhizobium, Allorhizobium and Sinorhizobium demonstrating the presence of a diversity of native bean nodule bacteria. This study also reports for the first time the presence of Allorhizobium taibaishanense in common bean nodules. These results constitute an important step in the development of an effective microbial inoculum and sustainable food production.

Influence of native rhizobacteria co‐inoculation and formulation of bacterial inoculants on the growth and yield of common bean (Phaseolus vulgaris L.)

AbstractIntroductionIncorporation of inoculum in different carrier materials may increase the efficacy of bacterial inocula.Materials and MethodsField experiments were conducted using two strains of rhizobium and phosphate‐solubilizing bacteria (PSB) and their respective combinations using different carrier materials in common bean‐growing regions in three soil types in Kenya. The field experiment was laid out in a split‐plot arrangement with the strain inoculations as the main plot while the subplots consisted of the carrier materials (filter mud, peat moss and yeast extract mannitol broth [YEMB]). Each main plot included two controls: uninoculated negative control and uninoculated controls that received N and P fertilizer. The experiment was conducted for two cropping seasons. Data were collected on the nodulation, shoot and root biomass and yield.ResultsCo‐inoculation of the common bean with Rhizobium phaseoli + Bacillus aryabhattai strains had significantly higher number of nodules (55 nodules per plant) compared to single R. phaseoli inoculation (38 nodules). The co‐inoculation of the rhizobia and the PSB yielded statistically at par with the application of diammonium phosphate (18:46:0) across the soil types and seasons. The use of filter mud as a carrier material led to a higher number of nodules for most of the rhizobia strains inoculation and their respective co‐inoculation with the bacillus strains. Significantly higher yield was obtained with the filter mud (1.64 Mg ha−1) while there was no significant difference in the yield of common bean between peat moss and YEMB as carrier materials for the bacterial strains.ConclusionThe solid carrier material, specifically filter mud, showed potential for use in the formulation of inoculants. Specific co‐inoculation of rhizobia (R. phaseoli and Rhizobium pusense strains) and PSB (Paenibacillus polymyxa and B. aryabhattai) increased the growth, nodulation and yield of common bean more efficiently than the control.

Persulfidation of plant and bacteroid proteins is involved in legume nodule development and senescence.

Legumes establish symbiosis with rhizobia, forming nitrogen-fixing nodules. The central role of reactive oxygen species (ROS) and reactive nitrogen species (RNS) in nodule biology has been clearly established. Recently, hydrogen sulfide (H2S) and other reactive sulfur species (RSS) have emerged as novel signaling molecules in animals and plants. A major mechanism by which ROS, RNS, and RSS fulfil their signaling role is the post-translational modification of proteins. To identify possible functions of H2S in nodule development and senescence, we used the tag-switch method to quantify changes in the persulfidation profile of common bean (Phaseolus vulgaris) nodules at different developmental stages. Proteomic analyses indicate that persulfidation plays a regulatory role in plant and bacteroid metabolism and senescence. The effect of a H2S donor on nodule functioning and on several proteins involved in ROS and RNS homeostasis was also investigated. Our results using recombinant proteins and nodulated plants support a crosstalk among H2S, ROS, and RNS, a protective function of persulfidation on redox-sensitive enzymes, and a beneficial effect of H2S on symbiotic nitrogen fixation. We conclude that the general decrease of persulfidation levels observed in plant proteins of aging nodules is one of the mechanisms that disrupt redox homeostasis leading to senescence.

Rhizobium etli is able to emit nitrous oxide by connecting assimilatory nitrate reduction with nitrite respiration in the bacteroids of common bean nodules

ABSTRACT Legumes can contribute to emissions of the potent greenhouse gas nitrous oxide (N2O) directly by some rhizobia species that are able to denitrify under free-living conditions and in symbiotic association with the plant. In this study, the capacity of Phaseolus vulgaris-Rhizobium etli symbiosis to emit N2O in response to nitrate (NO3 -) has been demonstrated for the first time. We found that bacteroidal assimilatory nitrate reductase (NarB) as well as nitrite reductase (NirK) and nitric oxide reductase (cNor) denitrifying enzymes contribute to nitric oxide (NO) and N2O formation in nodules. We also show that R. etli NarK is involved in NO2 - extrusion and links NO3 - reduction by NarB in the cytoplasm with NirK and cNor denitrification activities in the periplasm. The knowledge generated in this work will be instrumental for exploring strategies and sustainable practices in agricultural soil management to increase legume crop yield and mitigate greenhouse gas emissions.

The outstanding diversity of rhizobia microsymbionts of common bean (Phaseolus vulgaris L.) in Mato Grosso do Sul, central-western Brazil, revealing new Rhizobium species.

Common bean is considered a legume of great socioeconomic importance, capable of establishing symbioses with a wide variety of rhizobial species. However, the legume has also been recognized for its low efficiency in fixing atmospheric nitrogen. Brazil is a hotspot of biodiversity, and in a previous study, we identified 13 strains isolated from common bean (Phaseolus vulgaris) nodules in three biomes of Mato Grosso do Sul state, central-western Brazil, that might represent new phylogenetic groups, deserving further polyphasic characterization. The phylogenetic tree of the 16S rRNA gene split the 13 strains into two large clades, seven in the R. etli and six in the R. tropici clade. The MLSA with four housekeeping genes (glnII, gyrB, recA, and rpoA) confirmed the phylogenetic allocation. Genomic comparisons indicated eight strains in five putative new species and the remaining five as R. phaseoli. The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) comparing the putative new species and the closest neighbors ranged from 81.84 to 92.50% and 24.0 to 50.7%, respectively. Other phenotypic, genotypic, and symbiotic features were evaluated. Interestingly, some strains of both R. etli and R. tropici clades lost their nodulation capacity. The data support the description of the new species Rhizobium cerradonense sp. nov. (CNPSo 3464T), Rhizobium atlanticum sp. nov. (CNPSo 3490T), Rhizobium aureum sp. nov. (CNPSo 3968T), Rhizobium pantanalense sp. nov. (CNPSo 4039T), and Rhizobium centroccidentale sp. nov. (CNPSo 4062T).

Biofertilizer and NPSB fertilizer application effects on nodulation and productivity of common bean (Phaseolus vulgaris L.) at Sodo Zuria, Southern Ethiopia.

Common bean is among the vital legumes cultivated for nutrition, foreign currency earnings, and income generation. Common bean production is constrained mainly by nutrient deficiencies. A field experiment was conducted at Sodo Zuria Woreda to assess the effects of blended N-P-S-B fertilizer rates and Rhizobium strain on yield and yield traits of common bean and income of farmers. Treatments were five different rates of blended NPSB fertilizer (0, 50, 100, 150, and 200 kg ha-1) and two types of Rhizobium strains (HB-A15 and HB-429 (Rhizobia etli)) and control (without inoculation). Randomized complete block design in factorial arrangement with three replications were used. Results showed that interaction of blended NPSB rate and Rhizobium strain type had significant effect on the number of effective nodule, grain yield, and above ground dry biomass. Application of 150 kg NPSB ha-1 along with inoculation of Rhizobium etli resulted in the highest grain yield (3017.7 tons ha-1). Similarly, economic analysis indicated that the highest net return of 38298.545 ETB ha-1 was obtained from combined application of 150 kg NPSB ha-1 along with Rhizobium etli. Thus, it could be concluded that using Rhizobium etli strain with 150 kg NPSB ha-1 was found to be appropriate for common bean production in the area.

Microbiome of Nodules and Roots of Soybean and Common Bean: Searching for Differences Associated with Contrasting Performances in Symbiotic Nitrogen Fixation.

Biological nitrogen fixation (BNF) is a key process for the N input in agriculture, with outstanding economic and environmental benefits from the replacement of chemical fertilizers. However, not all symbioses are equally effective in fixing N2, and a major example relies on the high contribution associated with the soybean (Glycine max), contrasting with the low rates reported with the common bean (Phaseolus vulgaris) crop worldwide. Understanding these differences represents a major challenge that can help to design strategies to increase the contribution of BNF, and next-generation sequencing (NGS) analyses of the nodule and root microbiomes may bring new insights to explain differential symbiotic performances. In this study, three treatments evaluated in non-sterile soil conditions were investigated in both legumes: (i) non-inoculated control; (ii) inoculated with host-compatible rhizobia; and (iii) co-inoculated with host-compatible rhizobia and Azospirillum brasilense. In the more efficient and specific symbiosis with soybean, Bradyrhizobium presented a high abundance in nodules, with further increases with inoculation. Contrarily, the abundance of the main Rhizobium symbiont was lower in common bean nodules and did not increase with inoculation, which may explain the often-reported lack of response of this legume to inoculation with elite strains. Co-inoculation with Azospirillum decreased the abundance of the host-compatible rhizobia in nodules, probably because of competitiveness among the species at the rhizosphere, but increased in root microbiomes. The results showed that several other bacteria compose the nodule microbiomes of both legumes, including nitrogen-fixing, growth-promoters, and biocontrol agents, whose contribution to plant growth deserves further investigation. Several genera of bacteria were detected in root microbiomes, and this microbial community might contribute to plant growth through a variety of microbial processes. However, massive inoculation with elite strains should be better investigated, as it may affect the root microbiome, verified by both relative abundance and diversity indices, that might impact the contribution of microbial processes to plant growth.

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.

Influence of common bean genotypes and rhizobia interaction for nodulation and nitrogen fixation

Bacteria from the rhizobia group are able to associate symbiotically with bean crop, forming nodules in the root, in which the biological nitrogen-fixing process occurs. However, the efficiency of this process has been low and it can be attributed to genetic and environmental factors. Thus, the objective of this study was to evaluate the capacity of nodulation of local varieties and commercial common bean cultivars inoculated with a Rhizobium tropici strain used in commercial inoculants and rhizobia isolates from common bean root nodules. The experiment was carried out in a factorial scheme (2x4), in a randomized block design with four replicates. It was tested two local varieties and two commercial cultivars, inoculated with the reference Rhizobium tropici strain CIAT899 and the RBZ14 strain isolated from common bean nodules grown in soils of Southern Brazil, in adapted Leonard-type pots. The CIAT899 strain promoted either higher mass of viable nodules and higher nitrogen accumulation in the aerial part. The black group local variety showed better response than the cultivar TAA Dama for nodule viability, suggesting more efficiency for nodulation. The interaction between genotypes (local varieties and commercial cultivars) and bacteria showed the specificity of the complex symbiotic relationship of biological nitrogen fixation in common bean, requiring further studies of these interactions.

English

Most of the soils are N and P-deficient and require high applications of chemical fertilizers to ensure optimal crop production. However, this practice poses in most cases serious environmental issues. In recent years, inoculation with beneficial bacteria has emerged as a safe and sustainable alternative to chemical fertilizers. In this context, we investigated the effect of two symbiotic Agrobacterium radiobacter strains (LMR670 and LMR676) and two plant growth-promoting rhizobacteria Bacillus sp. (M131) and Enterobacter sp. (P1S6), as single or combined inoculants, on common bean growth and yield under N and P-deficient conditions. In a first trial, Agrobacterium strains’ symbiotic efficiency with common bean was evaluated in a low phosphorus and nitrogen soil under greenhouse conditions. Strain LMR670 recorded the highest nodules number (53 nodules per plant) and shoot dry weight (0.553 g plant-1). This strain was then used in combination with the PGP rhizobacteria in a common bean co-inoculation assay under sufficient and deficient P levels (80 kg ha-1of P and No P added). Single inoculation with LMR670 recorded the highest shoot dry weight (82% increase compared to non-fertilized control) compared to combined inoculants (46-47% increase). To corroborate the obtained results, a field experiment was conducted using the same treatments. LMR670 as a single inoculant or mixed with M131 was consistently effective leading to common bean yields comparable to N and P fertilized plants (13.07 t ha-1 and 12.35 t ha-1 respectively). In addition, single inoculation with the multi-PGP strain M131 showed positive effects on all common bean growth parameters and yield value (14 t ha-1); exceeding even N and P fertilized control plants. These results suggest that the strains LMR670 and M131 can be used, in a single or combined inoculation, as effective biological fertilizers for common bean cultivation to replace phosphorus and nitrogen fertilizers. Globally our results highlight the potential of native phytobeneficial strains for successful nodulation, growth and yield of common bean under N and P-deficient conditions.

Isolation and Characterization of High-Efficiency Rhizobia From Western Kenya Nodulating With Common Bean.

Common bean is one of the primary protein sources in third-world countries. They form nodules with nitrogen-fixing rhizobia, which have to be adapted to the local soils. Commercial rhizobial strains such as Rhizobium tropici CIAT899 are often used in agriculture. However, this strain failed to significantly increase the common bean yield in many places, including Kenya, due to the local soils’ low pH. We isolated two indigenous rhizobial strains from the nodules of common bean from two fields in Western Kenya that have never been exposed to commercial inocula. We then determined their ability to fix nitrogen in common beans, solubilize phosphorus, and produce indole acetic acid. In greenhouse experiments, common bean plants inoculated with two isolates, B3 and S2 in sterile vermiculite, performed better than those inoculated with CIAT899 or plants grown with nitrogen fertilizer alone. In contrast to CIAT899, both isolates grew in the media with pH 4.8. Furthermore, isolate B3 had higher phosphate solubilization ability and produced more indole acetic acid than the other two rhizobia. Genome analyses revealed that B3 and S2 are different strains of Rhizobium phaseoli. We recommend fieldwork studies in Kenyan soils to test the efficacy of the two isolates in the natural environment in an effort to produce inoculants specific for these soils.

Effects of Rhizobium tropici azide-resistant mutants on growth, nitrogen nutrition and nodulation of common bean (Phaseolus vulgaris L.)

Inoculation with Rhizobium tropici and Rhizobium freirei can partially supply the nitrogen demand required for normal development and high productivity of common bean (Phaseolus vulgaris L.). Here, a simple and low-cost approach was used to select R. tropici sodium azide-resistant mutants with improved efficiency as microsymbiont of common bean. We mutagenized R. tropici CIAT889 by treatment with methyl methanesulfonate (MMS) and selected azide-resistant mutants. Six mutants (AzR14, AzR15, AzR16, AzR17, AzR18, and AzR19) with varying levels of resistance were obtained and characterized for symbiotic traits and performance upon inoculation of common bean. Mutants grew and produced exopolysaccharides similar to the CIAT899 wildtype; however, swarming motility, biofilm, and auxin production were altered. About nodulation, shoot biomass, and nitrogen content, the performance of mutant strains was improved or similar to that of CIAT899. All mutants enhanced nodulation of common bean; furthermore, AzR14, AzR18, and AzR19 increased nodule biomass by 25%–36%. AzR18 and AzR19 increased shoot biomass by up to 13% compared to CIAT899, whereas AzR14, AzR15, AzR16, and AzR17 reduced biomass accumulation by 9%–19%. In addition, AzR18 and AzR19 showed higher motility or biofilm production than the other mutant strains, suggesting that these traits are involved in bacterial symbiotic effectiveness. Shoot N content do not differ between plants inoculated with the CIAT899 or mutant strains, although plants inoculated with AzR18, AzR19 and CIAT899 had increased the shoot N as compared to uninoculated plants grown at N-limiting conditions. Our results demonstrated the possibility to improve the performance of the R. tropici microsymbiont through induced mutagenesis followed by the selection of azide resistant mutants. This approach allowed to identify the AzR18 and AzR19 strains, which showed superior nodulation and symbiotic effectiveness than CIAT899 while providing equivalent amounts of N to the inoculated plants. We propose the AzR18 and AzR19 as promising candidates for developing new and more efficient inoculants for common bean, nevertheless further trials under field conditions are required to confirm their superior performance to the wild strain.

Agroecological modeling of nitrogen and carbon transfers between decomposer micro-organisms, plant symbionts, soil and atmosphere in an intercropping system

The modeling of continuous transfers of carbon (C) and nitrogen (N) previously published in the literature has paid little attention to the functional role of micro-organisms. In general, only monoculture systems have been modeled. Furthermore, there have been few experiments under field conditions at farm scale, where clear evidence for the benefits of intercropping is lacking. This work focus on mechanistic modeling approaches based on the ecological functioning of the microbial biomass, to quantify the daily exchange of C and N between plant organs, micro-organisms, rhizobial symbionts, soil compartments and the atmosphere in an arable intercropping system. The MOMOS model was validated on C and N data collected from a common bean (Phaseolus vulgaris L. cv. El Djadida) and maize (Zea mays L. cv. Filou) intercropping system. The experiment was performed at two field sites that were chosen with farmers to represent both high and low soil P availability. The results show that all C and N exchanges were successfully predicted at 5% significance and that they depend on the phenological stage, especially the flowering stage. Increased C allocation from photosynthesis to roots contributed to increasing both grain yield and N grain for intercropped maize. C and N stocks in the common bean nodules were lower in intercropping than in monocultures, and this is associated with the decrease of total atmospheric nitrogen (N2) fixation by intercropped common beans, in particular with a high soil P. However, the rate of N2 fixation was higher in the intercrops than in the monoculture when the soil is P-deficient. Micro-organisms were responsible for most of the C losses from the soil to the atmosphere but intercropping significantly reduced the C losses by improving micro-organism C use efficiency. These results uncover the strong link between N and C stocks, confirming the robustness of the newly formulated MOMOS equations that are validated in this paper. This agroecological modeling experiment demonstrated the functional role of microbial biomass, in both the growth of the intercrops crop and their symbiosis, improving the prediction of the daily C and N flows between plant organs, soil compartments and the atmosphere.

Fe3O4 nanoparticles and Rhizobium inoculation enhance nodulation, nitrogen fixation and growth of common bean plants grown in soil

The effects of Fe3O4 nanoparticles (NPs) and Rhizobium inoculation on nodulation, nitrogen fixation and plant growth of common bean (cv. Red Guama, Phaseolus vulgaris) plants were investigated in growth chambers. Plants were exposed to: Fe3O4 NPs (2000 mg/L) (T1), Rhizobium inoculation (T2) and Fe3O4 NPs + Rhizobium inoculation (T3); non-treated plants were considered as controls. Harvested 35-day-old treated plants showed improved symbiotic performance including increased nitrogenase activity (51.2–90.7%), nodule leghaemoglobin (44.8–80.9%) and iron content (83.4–84.2%), number of active nodules per plant (58.7–122%) and nodule dry weight (40.2–70.6%). This resulted in enhanced symbiotic nitrogen fixation,and increased shoot (26.5–50.2%) and root (24.1–48.2%) total nitrogen content in treated plants in comparison with the controls. The best result was obtained using treatment T3. Furthermore, Fe3O4 NPs were taken up by bean plants in treatments T1 and T3, and these accumulated in their organs, including in nodules. All treatments led to an increase in root (51.9–79.8%) and shoot (27.5–52.7%) lengths, in leaf area (10.9–16.8%) and in root (10.1–17.8%), stem (9.8–12.7%) and leaf dry weight (8–17.3%) compared to control plants. Thus applied treatments have the potential to improve common bean plant growth through enhancement of nodulation and nitrogen fixation during vegetative growth.This study also provides strong evidence that the presence Fe3O4 NPs in nodules improves the symbiotic performance between Rhizobium (leguminosarum CF1 strain) and the common bean plant, due to enhanced nodulation and nitrogen fixation.

Comprehensive Identification and Expression Profiling of Circular RNAs During Nodule Development in Phaseolus vulgaris.

Symbiotic nitrogen fixation by legume nodules provides an abundant nitrogen source for plants, and understanding this process is key for developing green agriculture. Circular RNA (circRNA), a type of endogenous RNA produced by reverse splicing of mRNA precursors, plays important regulatory roles in plants at the transcriptional and post-transcriptional levels. However, the relationship between circRNAs and legume–rhizobium is unknown. Here, we performed comprehensive identification and expression profiling of circRNAs during nodulation in common bean (Phaseolus vulgaris) compared to uninoculated roots of corresponding ages by constructing circRNA-seq and mRNA-seq libraries. We identified 8,842 high-confident circRNAs, 3,448 of which were specifically produced during symbiosis, with the highest number at the nitrogen-fixing stage. Significantly, more circRNAs were derived from exons than from intergenic regions or introns in all samples. The lengths and GC contents of the circRNAs were similar in roots and nodules. However, circRNAs showed specific spatiotemporal expression patterns during nodule and root development. GO and other functional annotation of parental genes of differentially expressed circRNAs indicated their potential involvement in different biological processes. The expression of major circRNAs during symbiosis is independent of parental genes’ expression to a certain degree, while expression of the remaining minor circRNAs showed positive correlation to parental genes. Functional annotation of the targeted mRNAs in the circRNA–miRNA–mRNA network showed that circRNAs may be involved in transmembrane transport and positive regulation of kinase activity during nodulation and nitrogen fixation as miRNA sponges. Our comprehensive analysis of the expression profile of circRNAs and their potential functions suggests that circRNAs may function as new post-transcriptional regulators in legume–rhizobium symbiosis.

Genetic diversity of Agrobacterium species isolated from nodules of common bean and soybean in Brazil, Mexico, Ecuador and Mozambique, and description of the new species Agrobacterium fabacearum sp. nov.

Agrobacterium strains are associated with soil, plants and animals, and known mainly by their pathogenicity. We studied 14 strains isolated from nodules of healthy soybean and common bean plants in Brazil, Mexico, Ecuador and Mozambique. Sequence analysis of the 16S rRNA gene positioned the strains as Agrobacterium, but with low phylogenetic resolution. Multilocus sequence analysis (MLSA) of three partial housekeeping genes (glnII, gyrB and recA) positioned the strains in four distinct clades, with Agrobacterium pusense, Agrobacterium deltaense, Agrobacterium radiobacter and Agrobacterium sp. genomospecies G1. Analysis by BOX-PCR revealed high intraspecies diversity. Genomic analysis of representative strains of the three clades indicated that they carry the protelomerase telA gene, and MLSA analysis with six complete housekeeping genes (atpD, glnII, gyrB, recA, rpoB and thrC), as well as average nucleotide identity (less than 90 % with closest species) and digital DNA-DNA hybridization (less than 41 % with closest species) revealed that strain CNPSo 675T and Agrobacterium sp. genomospecies G1 compose a new species. Other phenotypic and genotypic characteristics were determined for the new clade. Although not able to re-nodulate the host, we hypothesize that several strains of Agrobacterium are endophytes in legume nodules, where they might contribute to plant growth. Our data support the description of the CNPSo 675T and Agrobacterium sp. genomospecies G1 strains as a new species, for which the name Agrobacterium fabacearum is proposed. The type strain is CNPSo 675T (=UMR 1457T=LMG 31642T) and is also deposited in other culture collections.

Evaluation of beneficial and inhibitory effects of nitrate on nodulation and nitrogen fixation in common bean (Phaseolus vulgaris)

AbstractThe effects of applied nitrate on symbiotic nitrogen fixation in legumes are complex. Both inhibition and promotion of nodulation by nitrate have been observed in a dose‐dependent manner. The objectives of this study were to determine the effects of nitrate at different concentrations on root nodulation in different genotypes in common bean (Phaseolus vulgaris). Six genotypes were inoculated with the same rhizobial strain and grown hydroponically in growth pouches in a growth chamber and exposed to six nitrate concentrations, including 0, 2.5, 5, 10, 15, and 20 mM for 4 weeks. The tested genotypes included three recombinant inbred lines (RILs, 25, 46, and 70) that differed in their responses to nitrogen (based on observations of one field growing season), their parents (Mist and Sanilac—registered varieties), which are different in N‐fixing abilities, and one nonnodulating mutant (R99). Our results showed that small amounts of nitrate (2.5 and 5 mM) promoted nodule formation and increased nodule biomass, compared with plants in the 0 nitrate control treatment. In contrast, nitrate concentrations over 10 mM inhibited nodulation, resulting in reductions in nodule number and nodule biomass. Nodulation was completely inhibited by 15‐mM nitrate in all the genotypes. Regression analyses indicated that 5‐mM nitrate is the optimum concentration for promoting nodulation as measured by the total number of nodules formed, the number of effective nodules formed, and the nodule biomass formed. In contrast, nitrogen fixation was inhibited by all levels of nitrate. No genotypic differences were observed in nodulation among the three RILs and their parental cultivars, but all were significantly different than R99, a nonnodulating mutant.

Combined Effect of Organic and Mineral Fertilization on Nodulation and Yield of Common Bean (Phaseolus vulgaris L.) under Sandy Soil Conditions.

This study was carried out on sandy soil of El- Maraka village, El- Tahreer region, Behara Governorate, Egypt to define the pest application time of organic (molase and vinase) and mineral (P + Micronutrients) to two cultivars (Karnack and Nebraska) of common bean plants (phaselus volgaris L) and their effect on plant growth, productivity and seeds quality. Fresh and dry matter yields as well as the content of N, P, K, Mn, Zn and Cu were determined for the plant samples at 45 and 70 days. Also, nodules number at growth period of 45 day. Seeds contents of N, P, K, Mn, Zn, Cu and protein were determined. The data of this study show the importance of both organic and mineral fertilizer to increase sandy soil productivity of common bean plants and improved its quality. Nebraska cultivar was more suitable and high productivity under sandy soil conditions compared with Karnack cultivated. The latter applications of both organic and mineral fertilization were more high efficiency than of earlier application.

Molecular and biochemical changes of aging-induced nodules senescence in common bean

In the common bean (Phaseolus vulgaris L.), premature nodule senescence affects biological nitrogen fixation (BNF), reducing crop yield. Here, we investigated the molecular and biochemical changes in common bean nodules undergoing aging senescence. The experiments were carried out with three common bean genotypes and two nodule groups: senescent and non-senescent nodules. The expression patterns of 12 genes were assessed using RT-qPCR. In addition, global activities of ascorbate peroxidase (APX) and catalase (CAT), lipid peroxidation and total sugar content of nodules were also determined. Our results show that PvLb gene expression was down-regulated and PvGS(n-1) and PvUriII, genes involved in N metabolism, also decreased in senescent nodules. Transcripts related to the ethylene, abscisic acid and cytokinin phytohormones were up-regulated upon nodule senescence. Interestingly, PvSnakin-2 (SNAKIN-like cysteine rich protein), a gene related to plant-pathogen interaction, was induced in senescent nodules. The biochemical assays showed that in nodules undergoing senescence, APX activity was altered in only one genotype but CAT activity decreased in all common bean genotypes. Lipid peroxidation increased but total sugar content was not altered in nodules upon senescence. All these findings suggest that a delayed plant-response to Rhizobium infection is activated in common bean nodules undergoing aging senescence and this mechanism seems to be regulated by phytohormones and cell signaling as well, whereas the antioxidant defenses mediated by CAT are repressed, suggesting oxidative damage in nodules upon senescence. Our results offer insights into understanding nodule senescence metabolism and provide potential senescence markers for common bean nodules.

Nodulation and Yields of Common Bean are Not Affected Either by Fungicides or by the Method of Inoculation

Core Ideas Captan and difenoconazole compromise in vitro survival of Rhizobium tropici CIAT 899. In the field, native rhizobia were clearly tolerant to most of the fungicides evaluated. Among the fungicide treatments assessed in the field, only difenoconazole is potentially damaging. When difenoconazole treatment is indispensable, in‐furrow inoculation may be the most alternative. Due to the effects observed in vitro, similar precautions should also be taken with captan in the field. Though some studies observed negative effects of fungicide seed treatment on yield of common bean (Phaseolus vulgaris L.), few studies have directly evaluated fungicide compatibility with seed inoculation treatments in field conditions. For fungicides that are incompatible, rhizobia inoculation in the planting furrow (in‐furrow inoculation) would permit combined use of both management practices since this would avoid direct contact between the chemical compound and the rhizobia. Thus, the aim of this study was to evaluate the compatibility of seed fungicide treatment and rhizobia inoculation techniques to improve N derived from rhizobia symbiosis in common bean. A preliminary in vitro experiment evaluated the survival of Rhizobium tropici after contact with fungicides compared with an untreated control. Two field experiments were conducted in a 6 × 4 factorial arrangement involving the same treatments used for the in vitro survival test, as well as four sources of N [two types of liquid inoculation (inoculation on the seed or in‐furrow inoculation), plus two controls without inoculation]. We concluded that captan and difenoconazole fungicides reduced in vitro survival of rhizobia inoculated on the seed. Yield measurements in the field studies indicated that symbiotic nitrogen fixation could replace N fertilization, regardless of the inoculation treatment method and environmental conditions. Difenoconazole is potentially harmful to native bacteria in the field; therefore, in‐furrow inoculation may be recommended when the use of difenoconazole is necessary. Moreover, based on the harmful effects observed in vitro, precautions similar to the use of difenoconazole should also be taken when using captan fungicide in field conditions.