Bacterium Bradyrhizobium Japonicum Research Articles

Impact of biological fungicides on the formation and functioning of symbiotic system soybean–Bradyrhizobium japonicum

Background. In order to reduce the negative impact of mineral fertilizers and pesticides on agroecosystems, environmentally friendly plant protection systems are increasingly being utilized in soybean cultivation technologies. These systems aim to provide essential nutrients to plants while minimizing ecological harm. Materials and Methods. In vegetative studies, the processes of formation and functioning of symbiotic systems in soybeans with active strains of nodulating bacteria Bradyrhizobium japonicum (РC07, PC09, B78, B144) were investigated under the influence of pre-sowing seed treatment with the biofungicides Mycosan-N and Phytocide-r. Microbiological, physiological, statistical methods, and gas chromatography were employed in the research. Results. The impact of biological fungicides on the nodulation activity of nodule bacteria has been identified. It was determined that with comprehensive treatment of soybean seeds with B. japonicum РC07 and Mycosan-N, the mass of nodules formed on the roots exceeded that of the control plants by 7‒22 %, and under the influence of Phytocide-r, the mass was lower by 6‒20 % throughout the vegetation period. With the combined application of B. japonicum PC09 and Mycosan-N, a decrease in the number of root nodules by 8‒25 % and a reduction in their mass by 18‒35 % was observed throughout the vegetation period. Under the influence of Phytocide-r, with the inoculation of rhizobia strain PC09, the number of nodules was lower compared to the control plants only in the full flowering stage. At the same time, the reduction in their mass was 6‒20 % throughout the entire observation period. A stimulating effect of both biofungicides on the formation of the symbiotic apparatus involving strains B. japonicum B78 and B144 has been identified. With the combined application of B. japonicum РC07 and Phytocide-r, nitrogenase activity of symbiotic systems decreased by 7 % only in the stage of three true leaves compared to plants whose seeds were treated only with rhizobia. The nitrogenase activity of symbiotic systems formed with the participation of B. japonicum B78 and under the influence of Mycosan-N increased by 28 %, 15 %, and 12 % in the stages of three true leaves, budding, and full flowering, respectively. The action of Phytocide-r, with the inoculation by the nodule bacteria B78, resulted in an increase in nitrogenase activity by 14 % only in the budding stage. Under the comprehensive treatment of soybean seeds with involved biofungicides and B. japonicum B144, an increase in the intensity of N2 assimilation was noted by 29‒34 % in the stage of three true leaves and by 10‒16 % in the budding stage. Conclusions. The application of a scientifically justified selection of rhizobial strains and biological plant protection agents for pre-sowing treatment of soybean seeds will enable a more complete realization of their nitrogen-fixing potential and phytoprotective effects.

Bionanomaterial composed of Bradyrhizobium japonicum and graphene oxide for determination of mercury in water and fruit juice samples

In the present work, the synthesis of a novel hybrid bionanomaterial formed by the bacteria Bradyrhizobium japonicum (BJ) and the graphene oxide (GO) nanomaterial implemented in the miniaturized dispersive microsolid phase extraction technique (D-μ-SPE) was reported for the first time for the preconcentration and determination of Hg in water and fruit juice samples after a multivariate optimization. Selective Hg extraction was achieved by forming a complex with ammonium pyrrolidinedithiocarbamate (APDC) reagent and a multivariate study was performed to optimize the experimental conditions. The optimal Hg extraction conditions for 25 mL of sample were 3 mg of BJ@GO, NaNO3 at 1% (w/v), 6 min of vortex agitation and 9 min of centrifugation. For back extraction, 500 µL of 14 mol L−1 HNO3, 6 min of vortex agitation and 6 min of centrifugation were used followed by detection by cold vapor atomic fluorescence spectrophotometry (CV-AFS). Under optimized experimental conditions, an extraction efficiency of 98%, a detection limit of 0.08 µg L−1, a relative standard deviation of 3.5% (at 1 µg L−1 of Hg, n = 10), and a linear range of 0.26 - 4.0 µg L−1 were obtained. In addition, the sustainability of the method developed through the use of the Analytical GREEnness calculator (AGREE) software was evaluated, obtaining a greenness degree of 0.59. This work reports the first application of BJ@GO along with the D-μ-SPE technique for trace Hg determination water and fruit juice samples. In this way, this research work allows the exact and precise determination of low Hg concentrations in water and juice samples, which allows the evaluation of the contamination degree by this toxic metal in beverages widely consumed by the population.

The Bradyrhizobium japonicum exporter ExsFGH is involved in efflux of ferric xenosiderophores from the periplasm.

The gram-negative bacterium Bradyrhizobium japonicum can take up structurally dissimilar ferric siderophores from the environment (xenosiderophores) to meet its nutritional iron requirements. Siderophore-bound iron transported into the periplasm is reduced to the ferrous form by FsrB, dissociated from the siderophore and the free ion is then transported into the cytoplasm by the ferrous iron transporter FeoAB. Here, we identified the RND family exporter genes exsFG and exsH in a selection for secondary site suppressor mutants that restore growth of an fsrB mutant on the siderophores ferrichrome or ferrioxamine. The low level of radiolabel accumulation from 55Fe-labeled ferrichrome or ferrioxamine observed in the fsrB mutant was restored to wild type levels in the fsrB exsG mutant. Moreover, the exsG mutant accumulated more radiolabel from the 55Fe-labeled siderophores than the wild type, but radiolabel accumulation from inorganic 55Fe was similar in the two strains. Thus, ExsFGH exports siderophore-bound iron, but not inorganic iron. The rescued fsrB exsG mutant required feoB for growth, indicating that ExsFGH acts on those siderophores in the periplasm. The exsG mutant was more sensitive to the siderophore antibiotic albomycin than the wild type, whereas the fsrB mutant was more resistant. This suggests ExsFGH normally exports ferrated albomycin. B. japonicum is naturally resistant to many antibiotics. The exsG strain was very sensitive to tetracycline, but not to six other antibiotics tested. We conclude that ExsFGH is a broad substrate exporter that is needed to maintain siderophore homeostasis in the periplasm.

The divalent metal ion exporter IhpABC is required to maintain iron homeostasis under low to moderate environmental iron conditions in the bacterium Bradyrhizobium japonicum.

Bacterial iron export mitigates high iron stress, but a role for it under lower iron conditions has not been established. MbfA is the high iron stress exporter in Bradyrhizobium japonicum. Here, we identify the ihpABC genes in a selection for secondary site mutations that suppress the poor growth phenotype of feoAB mutants defective in iron acquisition. IhpABC belongs to the RND tripartite efflux pump family. High iron conditions that derepress the mbfA gene partially rescued the growth of an ihpC mutant but reverted the feoB ihpC mutant to the feoB growth phenotype. The ihpA mutant grown under low iron conditions accumulated higher levels of iron compared to the wild type, and it displayed aberrant iron-responsive gene expression. The mbfA mutant was more sensitive than the wild type to H2 O2 , but the ihpA mutant was not sensitive. The ihpA mutant accumulated more Zn, Co and Cd than was found in the wild type, and growth of the mutant was more sensitive to inhibition by ZnCl2 , CoCl2 and CdCl2 . The findings suggest that IhpABC is a divalent metal ion exporter that helps maintain iron homeostasis under low to moderate environmental iron levels. Thus, iron export is not limited to managing high iron stress.

Оценка комплексной обработки семян сои фунгицидами и инокулянтом

The Russian and foreign scientists have already proved an important role of the symbiosis of rhizobia and soybean through which plants obtain the neces-sary nitrogen amount from the air for effective growth and protein synthesis. At the same time, they stated the seed treatment with some fungicides sup-press the rhizobia growth and lowered the formation of nitrogen-fixing nodules in fields. Now, the search of fungicides effective against pathogens but not suppressing rhizobia at joint application is actual in an agrotechnological system. In 2020–2021, we treated soybean seeds with an inoculant Hycout su-per soya based on a bacterium Bradyrhizobium japonicum. It did not reduce soybean plants infes-tation with diseases but caused additional seed yield 0.37 (2020) and 0.27 t/hа (2021) through the symbi-otic activity. The combined application of fungicides and an inoculant showed their effectiveness against fusarium wilt. The soybean seed treatment with Max-im, CS, Delit Pro, CS, and Tebu 60, OE combined with the inoculant did not suppress the formation of nitrogen-fixing nodules on soybean plant roots and did not reduce their air-dry weight. The highest amount (33 and 83 pcs./plant) and weight (0.09 and 0.32 g/plant) of nodules are noted on plant roots in a variant when treated the seeds with a mix Maxim, CS + inoculant, L.

Viability of Nodule Bacteria Bradyrhizobium japonicummon Soybean Seeds by Treatment With Fungicides During Extended Storage

Background. With the increase in soybean cultivation areas, inoculants are becoming increasingly sought after. They are not only compatible with the original preparations for seed treatment of soybean but also offer the possibility of applying them for several days or even months before sowing into the soil. Objective. The viability of new strains of nodule bacteria Bradyrhizobium japonicum (strain PC07 and strain B78) was investigated on the surface of soybean seeds treated with fungicides Fever and Maxim XL during the extended storage of inoculated seeds. Additionally, their ability for nitrogen fixation under symbiotic conditions was evaluated. Methods. Сultivation, serial dilution method, determination of bacterial titer of inoculants, quantification of colony forming units (CFU), gas chromatography. Results. In laboratory conditions, on the soybean seeds of Almaz and Vasylkivska varieties, the viability of the nodule bacteria B. japonicum PC07 and B78, which are more resistant to the active substances of these fungicides in pure culture, decreased. This reduction was dependent on the storage period, the fungicide used, and the strain of inoculant. One day after inoculation on the seeds without the use of fungicide, 68.6–75.4% CFU of the initial number was retained. Seeds treated with the fungicide Fever contained 36.5–38.3% CFU, and those treated with Maxim XL contained 26.2–27.0% CFU. The use of inoculants based on fungicide-resistant strains of B. japonicum PC07 and B78 also provided high cell viability – 2.41–2.8×106 (on the seeds without treatment), 1.40–1.70×106 (with Fever) and 0.8–1.17×106 (with Maxim XL) CFU/seed after 5 days of storage. The nitrogen-fixing activity of root nodules in soybean plants of Almaz and Vasylkivska varieties, whose seeds were treated with Fever, decreased on both varieties by 18.4–22.4% and 32.1–39.5%, respectively and for treatment with Maxim XL, the reduction was 24.5–33.7% and 47.7–75.2%, respectively, during the storage of seeds for 5 and 7 days before sowing, compared to control variants (seeds without fungicide treatment). Conclusions. The utilization of fungicide-resistant strains of B. japonicum PC07 and B78 for bacterization of soybean seeds treated with fungicides Fever and Maxim XL provides a high inoculation titer during 5 days of storage. Viability of microbial cells on seeds treated with fungicides significantly diminishes during storage for more than 7–14 days. Insufficient titer of CFU of B. japonicum strains PC07 and B78 on the seed surface after 14 days of storage can considerably impede the efficacy of biopreparations. Therefore, it is crucial to seek substances that can enhance the resistance of rhizobia on seeds to the adverse effects of fungicide seed treatment, ensuring a longer period of their viability.

Soybean Response to N Fertilization Compared with Co-Inoculation of Bradyrhizobium japonicum and Azospirillum brasilense

The soybean [Glycine max (L.) Merrill] relationship with the bacteria Bradyrhizobium japonicum is responsible for providing around 60% of the nitrogen (N) required for the crop and the remaining N comes from the soil or supplemental fertilization. To investigate if higher yields are possible, supplemental N studies and co-inoculation of Rhizobium with Azospirillum are necessary. This N rate (0, 30, 56, 112, 336 kg N ha−1) and inoculation study was conducted across eight environments in eastern North Dakota, USA, in 2021 and 2022. Also, the effect of supplemental N and co-inoculation on nodulation was evaluated. When N was applied at 112 kg N ha−1, nodulation was significantly inhibited. Co-inoculation increased the number of large nodules and the volume of nodules; however, the yield was not different from inoculation with B. japonicum. Nitrogen at 112 and 336 kg ha−1 increased grain yield, protein yield, and seed weight; however, the higher N rate decreased plant population. There were significant positive relationships between yield and protein content and seed weight, and negative relationships between oil and protein content, and yield and oil content. Based on a polynomial relationship, the highest yield (3711 kg ha−1) would be achieved at 273 kg N ha−1. The application of N resulted in a yield increase but using current prices may not be an economical choice. Additional research is necessary to verify if co-inoculation with efficient strains can improve biological N fixation.

Soybean Response to Seed Inoculation or Coating with Bradyrhizobium japonicum and Foliar Fertilization with Molybdenum.

Soybean is one of the most important legumes in the world, and its advantages and disadvantages are well known. As a result of symbiosis with the bacterium Bradyrhizobium japonicum, soybean can assimilate nitrogen from the air and is therefore not fertilized with this element, or if it is, only at small doses. In soybean agriculture practice, an important treatment is the inoculation of seeds with symbiotic bacteria and optimal fertilization with selected nutrients. Therefore, a three-year (2019-2021) field experiment was carried out to investigate the effects of soybean in the field to a seed Rhizobium inoculation or coating and molybdenum foliar fertilization. There were no significant interactions between the tested treatments over the years. It was demonstrated that the best variant was seed inoculation before sowing in combination with foliar molybdenum application. As a result of this treatment, a significant increase in nodulation, soil plant analysis development (SPAD) index, leaf area index (LAI) and seed yield (by 0.61 t·ha-1) was obtained compared to the control. In addition, the content of total protein in the seeds increased, while the content of crude fat decreased, which significantly modified the yield of both components. Sowing coated seeds in the Fix Fertig technology was less effective compared to inoculation, but it was significantly better than that in the control. Coating seeds with B. japonicum, in combination with foliar fertilization with molybdenum, could be recommended for agricultural practice, which was confirmed by economic calculations. Future experiments will assess the soybean's response to seed inoculation or coating and fertilization with other micronutrients.

Role of Phosphorus and Inoculation with Bradyrhizobium in Enhancing Soybean Production

Soybean (Glycine max L. Merril) is among the key oil seed crops worldwide, providing several benefits from human consumption to the enhancement of soil productivity. In Uganda, legumes are cultivated on roughly 1.5 million ha, with soybean being produced on a lower production area of 150,000 ha compared to beans (925,000 ha) and groundnuts (253,000 ha). In terms of achievable yield, soybean emerges the highest at 1.2 t·ha−1 as compared to beans (0.5 t·ha−1) and groundnuts (0.7 t·ha−1). Despite the smallest production coverage area, the crop’s feasible grain yield is projected at 4.6 t·ha−1 under optimal environmental conditions. The major bottleneck to the crop’s production is the decreasing soil fertility, mainly caused by low nitrogen (N) but also phosphorus (P) levels in the soil. There is a high potential for supplying N from the atmosphere through biological N fixation (BNF), a natural process mediated by the symbiotic bacteria Bradyrhizobium japonicum, which requires optimum P levels for effective N fixation and increased yield. The current work reviews the present status of soybean production in Uganda, highlights its ecological requirements, importance, and constraints, and proposes the use of inoculation and P application to boost its production.

Realization of Nodulation and Nitrogen-Fixing Activities of Bradyrhizobium japonicum and Rhizosphere Microbiota through Seed Treatment with Pesticide Standak Top and Spraying Plants with Soybean Seed Lectin

The idea of the study was to use soybean lectin as a biologically active compound with a broad spectrum of action to spray soybean plants for stabilization of the formation and functioning of the soybean-rhizobium symbiosis as well as the nitrogen-fixing activity of rhizosphere microbiota against the background of seed treatment with chemical plant protection product Standak Top — an innovative pesticide with fungicidal and insecticidal activity for the control of major diseases and pests of soybean plants. Aim. To study the peculiarities of formation and functioning of soybean-rhizobium symbiosis as well as the nitrogen-fixing activity of rhizosphere microbiota under spraying plants with specific soybean seed lectin on the background of seed treatment with Standak Top and inoculation with nodule bacteria Bradyrhizobium japonicum 634b on the sowing day in the conditions of pot experiments with soil as a substrate. Methods. Physiological, microbiological, gas chromatography, and statistical methods were used. Results. It was shown that after seed treatment with Standak Top (1.5 L/ton of seeds) on the sowing day, there was observed suppression of the process of nodule formation on the roots in the period of soybean vegetative growth. The nitrogen-fixing activity of the symbiotic system was at the control level, while the functional activity of soil diazotrophs was suppressed (by 1.2—2.2 times). Spraying plants in the phase of two trifoliate leaves (V2) with soybean seed lectin (without pesticide) led to an increase in the total mass of nodules on the plant (by 1.5 and 1.9 times as well as by 2.3 and 2.0 times compared to the control of inoculation in the phase of three trifoliate leaves (V3) and beginning of pod formation (R3), respectively). The increase in the total mass of the symbiotic apparatus on soybean roots in the phases V3 and R3 respectively was by 1.4 and 1.5 times in comparison with seed treatment with Standak Top, and the mass of one nodule was higher by 1.3 and 1.6 times, respectively. Soybean seed lectin led to a signifi cant increase in the actual nitrogenase activity of the soybean-rhizobium symbiosis. It was 2.9 and 1.9 times higher compared to control of inoculation and 2.1 and 1.8 times compared to the variant of inoculation + pesticide in the V3 and R3 phases, respectively. The functional activity of soil nitrogen-fixing microorganisms did not change significantly. The use of soybean seed lectin against the background of the seed treatment with Standak Top and inoculation contributed to the stabilization and increase in the rhizobia nodulation ability, the suppression of which was due to the infl uence of such an anthropogenic factor as pesticides. Th ere was observed an increase in the number (by 1.6 and 1.2 times) and mass of root nodules (by 2.2 and 1.5 times and 1.4 and 1.2 times, respectively, compared to the controls of inoculation and inoculation + pesticide). Soybean seed lectin significantly increased the nitrogenase activity of the symbiosis against the background of Standak Top (by 1.9 and 1.6 times and 1.4 and 1.5 times, respectively, in the V3 and R3 phases of soybean ontogenesis) compared to the control of inoculation and inoculation + Standak Top. Conclusions. The obtained results suggest the possibility of using the method of spraying plants with specific lectin as a means of leveling (or mitigating) the negative effect of pesticides used for the seed treatment on the formation and functioning of the symbiosis and rhizosphere diazotrophic microbiota. This indicates the prospects of studying the biological activity of phytolectins in spraying plants in order to regulate the formation and functioning of phytobacterial systems, as well as their responses to various environmental or anthropogenic stress factors, in particular, to the effect of chemical plant-protecting products used for the seed treatment.

Efficient control of the productivity of agrophytocenosis of soybean (Glicyne max (L.) Merill)

Around the world, there is a surging demand for soy, the ‘king of beans’. Soy is a globally traded foodstuff produced in both temperate and tropical regions and is a major source of protein and vegetable oils. In the conditions of the Kaliningrad region, the duration of soybean vegetation in soddy-weakly podzolic gleyic light loamy soil, with the medium reaction close to neutral, high and very high content of mobile phosphorus, and exchangeable potassium attains 144 days. The agrophytocenosis productivity is controlled through application of different mineral and bacterial fertilizers. Analysis of variance of the experimental data obtained in the study showed a pronounced relationship of 0.71 for the effect of the fertilizer system on quantitative indicators of the biological yield of soybean seeds. The effectiveness of the microbial-plant system of soybean cultivar Merlin and strains of nodule bacteria Bradyrhizobium japonicum, under optimal conditions, increased the crop yield to 4.43 t/ha, which is 2.11 t/ha higher compared to the control and 1.01 t/ha higher relative to the experimental option with carbamide fertilizer.

Symbiotic and physiological indicators of soybean inoculated of Bradyrhizobium japonicum single-strain in 7 days before sowing

Results of investigation of soybean of the Almaz variety in inoculation with preparations based on nodule bacteria Bradyrhizobium japonicum (Kirchner, 1896), Jordan, 1982 B78, B157, D37, D87 are presented. Different periods of the soybean seeds inoculation were used - on the sowing day (control) and in 7 days before sowing (experimental variants). The differences between control and experimental plants in the formation and functioning of the symbiotic apparatus and its functional activity, depending on the period between from seed inoculation to sowing were analysed. It was determined that the number of root nodules in the control plants was higher. The mass of nodules at the stage of 3 true leaves exceeded the control by 1.5–2.0 times in plants inoculated in 7 days before sowing, and the intensity of nitrogen fixation by 1.7–6.6 times. At the budding-beginning of flowering stage, the mass and intensity of N2 fixation by the nodules of control plants increased. As a result, the difference between the nitrogen fixing activity of control and experimental plants decreased significantly. Stimulating effect on aboveground mass of Bradyrhizobium japonicum strains with increased nitrogen fixing activity was noted. Optimal conditions for the formation and functioning of bean-rhizobial symbiosis were provided at the use of both of these terms of soybean inoculation. This reveals the possibility of effective application of early inoculation of soybean seeds with preparations based on nodule bacteria Bradyrhizobium japonicum active strains.

Transgenic Soybeans Expressing Phosphatidylinositol-3-Phosphate-Binding Proteins Show Enhanced Resistance Against the Oomycete Pathogen Phytophthora sojae

Oomycete and fungal pathogens cause billions of dollars of damage to crops worldwide annually. Therefore, there remains a need for broad-spectrum resistance genes, especially ones that target pathogens but do not interfere with colonization by beneficial microbes. Motivated by evidence suggesting that phosphatidylinositol-3-phosphate (PI3P) may be involved in the delivery of some oomycete and fungal virulence effector proteins, we created stable transgenic soybean plants that express and secrete two different PI3P-binding proteins, GmPH1 and VAM7, in an effort to interfere with effector delivery and confer resistance. Soybean plants expressing the two PI3P-binding proteins exhibited reduced infection by the oomycete pathogen Phytophthora sojae compared to control lines. Measurements of nodulation by nitrogen-fixing mutualistic bacterium Bradyrhizobium japonicum, which does not produce PI3P, revealed that the two lines with the highest levels of GmPH1 transcripts exhibited reductions in nodulation and in benefits from nodulation. Transcriptome and plant hormone measurements were made of soybean lines with the highest transcript levels of GmPH1 and VAM7, as well as controls, following P. sojae- or mock-inoculation. The results revealed increased levels of infection-associated transcripts in the transgenic lines, compared to controls, even prior to P. sojae infection, suggesting that the plants were primed for increased defense. The lines with reduced nodulation exhibited elevated levels of jasmonate-isoleucine and of transcripts of a JAR1 ortholog encoding jasmonate-isoleucine synthetase. However, lines expressing VAM7 transgenes exhibited normal nodulation and no increases in jasmonate-isoleucine. Overall, together with previously published data from cacao and from P. sojae transformants, the data suggest that secretion of PI3P-binding proteins may confer disease resistance through a variety of mechanisms.

FUNCTIONING OF SOYBEAN-RHIZOBIA SYMBIOSIS AFTER VARIOUS METHODS APPLYING TRACE ELEMENTS COMPLEX

Objective. Investigate the effect of inoculation of soybean seeds with an active strain of nodule bacteria Bradyrhizobium japonicum T21-2 and treatment of seed or vegetative plants with a complex of trace elements Avatar-2 on the functioning of the symbiotic apparatus and grain productivity. Methods. Physiological, microbiological, gas chromatographic, vegetation experiment, statistical. Results. As a result of joint treatment of seeds with nodule bacteria and Avatar-2, inhibition of nodulation activity of nitrogen-fixing microorganisms in the stage of three true leaves was revealed in comparison with symbiotic systems formed as a result of standard inoculation of seed with rhizobia of Bradyrhizobium japonicum T21-2 strain. In the stage of budding / beginning of flowering and bean formation on the soybean roots of this variant, the number of nodules exceeded the corresponding figure in plants, bacterized without the use of trace elements by 16.6 % and 24.1 %, respectively. The combination of inoculation of seeds and feeding of plants during the vegetation with Avatar-2 did not have a significant effect on the activity of symbiotic systems in the stage of three true leaves and budding / beginning of flowering. However, in the stage of bean formation, a 38 % increase in nitrogen-fixing activity was detected compared to plants grown from seeds of rhizobia inoculated with Bradyrhizobium japonicum T21-2 strain (without the use of trace elements). Activation of soybean growth processes at all stages of plant development when exposed of seed bacterization and use of Avatar-2 by different methods was noted. Inoculation of seeds and using microfertilizers, both as separate elements of soybean cultivation technology and complex treatments in the conditions of vegetation experiment, resulted in a decrease in grain content of copper, zinc, iron and nickel. It was found that pre-sowing treatment of soybean seeds with a complex of chelated nutrients without bacterization caused an increase in grain weight per plant by 17.8 %, and when feeding non-inoculated growing plants with this fertilizer — by 27.5 % compared to the control. The combination of both factors — inoculation of seeds with a bacterial preparation and different using of trace element complex also had a positive effect on individual productivity of soybean plants, as evidenced by an increase of 14.4 % and 30.2 % compared to plants whose seeds were treated with rhizobia only. Conclusion. The combination of intensification factors in soybean cultivation technology through the use of different methods of complex microfertilizer and seed treatment with highly active strain of nodule bacteria has a significant impact on plant growth, content of individual trace elements in grain and creates a significant reserve of grain productivity of this crop.

Composition of pigment complex in leaves of soybean plants, inoculated by Bradyrhizobium japonicum, subject to metal nanocarboxylates and various-levels of water supply

A distinctive feature of legumes is the ability to combine two most important processes: photosynthesis and nitrogen fixation. However, the course of those processes, and therefore seed potential of those crops depend on a number of biotic and abiotic factors, the commonest being drought. Therefore, interest in physical-biochemical resistance of the plant organism to abiotic stress factors is increasing, as well as search for optimum ways to increase its adaptability. Success of adaptation of a plant’s organism to unfavourable environmental factors is known to largely depend on optimal functioning of assimilative apparatus. Some indicators of the condition of the apparatus are the content and ratio of photosynthesis pigments. Therefore, we aimed at determining the reaction of the pigment complex of Glycine max (L.) Merr. plants, grown against the background of optimal and insufficient watering, to inoculation of seeds with rhizobia bacteria Bradyrhizobium japonicum, cultivated using nanocarboxylates of chromium, cobalt, iron, copper and germanium. Research has shown that utilization of germanium nanocarboxylate as a component of inoculative suspension led to the highest content of chlorophylls in leaves of soybean of the studied variants in the blossoming phase during optimal watering, as well as significant increase in the content of carotenoids compared with the control plants regardless of the level of watering. At the same time, this element caused no significant effect on the chlorophyll content in plants grown in drought. It was confirmed that among soybean plants that were in stress conditions (blossoming phase) for two weeks, the highest content of chlorophylls was in leaves of plants grown from seeds inoculated with rhizobial suspension with addition of chromium and copper nanocarboxylates, which caused 25.3% and 22.8% increase in chlorophyll а, 29.4% and 32.3% in chlorophyll b and 26.4%% and 23.8% in them respectively, compared with the control. Furthermore, chromium and copper nanocarboxylates stimulated the content of carotenoids in the same plants, though it was less expressed than after adding germanium nanocarboxylate. The highest content of photosynthetic pigments in plants after the watering was resumed (phase of bean formation) was in cases of applying chromium and germanium nanocarboxylates. It was confirmed that the most efficient way to protect the pigment complex of soybean plants during drought was using chromium and germanium nanocarboxylates as components of inoculation suspension. The results we obtained indicate the possibility of applying chromium nanocarboxylate in the technology of cultivating soybean in the conditions of water deficiency as an effective way to improve biosynthesis of chlorophylls, as well as using germanium nanocarboxyllate as a component that provides a high level of activity of protective mechanisms of the pigment system of soybean, associated with resisting stress caused by water deficiency.

The use of the nodule bacteria as a remedy for expanding adaptive possibilities of soybean under drought conditions

Symbiotic systems created by soybean variety Almaz (Glycine max (L.) Merr.) and strains of nodule bacteria Bradyrhizobium japonicum 646, РС09, В157 with were investigated under different water supply are investigated (30 and 60 % field capacity, FC). It was shown that drought inhibited the process of nodulation, depending on the functional and adaptive capacity of each of the microsymbionts. The number and mass of root nodules on plants decreased compared to control plants under insufficient watering. Insufficient watering (30 % FC) negatively impacted the functioning of symbiotic systems Glycine max (L.) Merr.—Bradyrhizobium japonicum depending on the drought duration. Strains of Bradyrhizobium japonicum PC09 and B157 formed more drought-tolerant symbiotic systems. They had higher nitrogen fixing activity under reduced moisture supply (30 % FC) and during the recovery period compared to the Bradyrhizobium japonicum 646 strain. It was shown that with increasing drought duration (from the 3rd to the 10th day) the difference in physiological and symbiotic parameters between treated and control plants increased, but with the resumption of watering they partially leveled depending on the inoculant strain.

Effects of Constant Electric Field on Biodegradation of Phenol by Free and Immobilized Cells of Bradyrhizobium japonicum 273

It is shown that bacteria Bradyrhizobium japonicum 273 were capable of degrading phenol at moderate concentrations either in a free cell culture or by immobilized cells on granulated activated carbon particles. The amount of degraded phenol was greater in an immobilized cell preparation than in a free culture. The application of a constant electric field during cultivation led to enhanced phenol biodegradation in a free culture and in immobilized cells on granulated activated carbon. The highest phenol removal efficiency was observed for an anode potential of 1.0 V/S.H.E. The effect was better pronounced in a free culture. The enzyme activities of free cells for phenol oxidation and benzene ring cleavage were very sensitive to the anode potential in the first two steps of the metabolic pathway of phenol biodegradation catalyzed by phenol hydroxylase—catechol-1,2-dioxygenase and catechol-2,3-dioxygenase. It was observed that at an anode potential of 0.8 V/S.H.E., the meta-pathway of cleavage of the benzene ring catalyzed by catechol-2,3-dioxygenase became competitive with the ortho-pathway, catalyzed by catechol-1,2-dioxygenase. The obtained results showed that the positive effect of constant electric field on phenol biodegradation was rather due to electric stimulation of enzyme activity than electrochemical anode oxidation.

Assessing life cycle environmental impacts of inoculating soybeans in Argentina with Bradyrhizobium japonicum

PurposeTo estimate life cycle impacts from introducing the yield-enhancing inoculant containing the nitrogen-fixing bacterium Bradyrhizobium japonicum and the signal molecule lipochitooligosaccharide (LCO) in Argentinian soybean production. The study focuses on soybeans grown in rotation with corn in the Buenos Aires province. We also provide the life cycle impact assessment for the inoculant production. The study represents a novel scope in terms of the studied crop, inoculant type, and location.MethodsConsequential LCA is used to assess the cradle-to-gate soybean production systems with and without inoculant use. Stepwise is used for quantification of 16 impacts at mid-point level. Also, the LCA-based guidance of Kløverpris et al. (2020) is followed, and we divide the change in impacts caused by the inoculant’s use into four effects. The field effect accounts for changes in field emissions. The yield effect accounts for additional soybean production in the inoculant system that displaces soybean production elsewhere (system expansion). The upstream effect covers the inoculant production and the downstream effect covers post-harvest changes such as soybean transport and drying. Small plot field-trials data is applied in the biogeochemical model DayCent to estimate field emissions, among others.Results and discussionThe use of this inoculant reduces environmental impacts from soybean production in all studied impact categories. The main contributing factor is the yield effect, i.e., reduced impacts via avoided soybean production elsewhere including reduced pressure on land and thereby avoided impacts in the form of indirect land-use-change (iLUC). The field effect is the second-largest contributor to the overall impact reduction. Upstream and downstream effects only had minor influence on results. The yield and field effects are closely tied to the yield change from the inoculant use, which was not fully captured in the DayCent modeling. Thereby, a potential underestimation of the environmental benefits of roughly 10% can be expected, corresponding to the difference of empiric yield data and the modeled yield data in DayCent.Conclusion and recommendationsThe use of this inoculant shows environmental benefits and no trade-offs for the 16 impacts assessed. Results depend primarily on avoided soybean production (the yield effect) which entails iLUC impacts in Brazil and USA, and to a lesser degree on field emissions modelled with DayCent. Better data and parametrization of DayCent, to better capture the change in yields and estimate field emissions, economic modelling for the system expansion assumptions, and accounting for uncertainty in iLUC modelling could improve the assessment.

Mechanistic insights into heme-mediated transcriptional regulation via a bacterial manganese-binding iron regulator, iron response regulator (Irr)

The transcription factor iron response regulator (Irr) is a key regulator of iron homeostasis in the nitrogen-fixating bacterium Bradyrhizobium japonicum Irr acts by binding to target genes, including the iron control element (ICE), and is degraded in response to heme binding. Here, we examined this binding activity using fluorescence anisotropy with a 6-carboxyfluorescein-labeled ICE-like oligomer (FAM-ICE). In the presence of Mn2+, Irr addition increased the fluorescence anisotropy, corresponding to formation of the Irr-ICE complex. The addition of EDTA to the Irr-ICE complex reduced fluorescence anisotropy, but fluorescence was recovered after Mn2+ addition, indicating that Mn2+ binding is a prerequisite for complex formation. Binding activity toward ICE was lost upon introduction of substitutions in a His-cluster region of Irr, revealing that Mn2+ binds to this region. We observed that the His-cluster region is also the heme binding site; results from fluorescence anisotropy and electrophoretic mobility shift analyses disclosed that the addition of a half-equivalent of heme dissociates Irr from ICE, likely because of Mn2+ release due to heme binding. We hypothesized that heme binding to another heme binding site, Cys-29, would also inhibit the formation of the Irr-ICE complex because it is proximal to the ICE binding site, which was supported by the loss of ICE binding activity in a Cys-29-mutated Irr. These results indicate that Irr requires Mn2+ binding to form the Irr-ICE complex and that the addition of heme dissociates Irr from ICE by replacing Mn2+ with heme or by heme binding to Cys-29.

EFFECT OF FUNGICIDES ON THE EFFICIENCY OF SOYBEAN INOCULATION WITH PESTICIDE-RESISTANT NODULE BACTERIA

Objective. Evaluate the efficiency of soybean inoculation with pesticide-resistant nodule bacteria Bradyrhizobium japonicum under the influence of pre-sowing seed dressing with Maxym XL,Standak Top and Fever and preventive spraying of plants with the fungicide Akanto Plus. Methods. Physiological, microbiological, gas chromatographic, vegetation experiment, statistical.Results. The number and weight of nodules formed on soybean roots during the growing seasonunder the action of the prothioconazole-based fungicide Fever were reported to be lower comparedwith the control and other variants of experiment. Pesticides Maxym XL and Standak Top had a lesspronounced toxic effect on the formation of legume-rhizobial symbiosis with the participation offungicide-resistant B. japonicum B78 Tn5 mutant compared to Fever. A decrease in the level of N2assimilation by 10.9–41.1 % in the three-leaf phase was found with the combined use of seed pesticides and nodule bacteria included in the study, relative to the variant with inoculation only. However, during the growing season, the negative effect of fungicidal preparations on the functioning oflegume-rhizobial symbiosis decreased. Conclusion. Under the conditions of the model vegetationexperiment with the concomitant use of Standak Top and Fever and bacterization, a decrease in individual plant productivity by 7.0 and 14.1 %, respectively, has been reported. The complex actionof the pesticide Maxym XL and inoculation with pesticide-resistant B. japonicum B78 transposonmutant, contributed to the maximum increase in yield by 27.2 %. The obtained data confirm the expediency of selection of fungicides and nitrogen-fixing microorganisms for concomitant use to ensure high economic and protective effects.