Nodule Bacteria Research Articles

Isolation and characterization of non-rhizobial bacteria and arbuscular mycorrhizal fungi from legumes

This study investigates non-rhizobial endophytic bacteria in the root nodules of chickpea (Cicer arietinum L), faba bean (Vicia faba), and cowpea (Vigna unguiculata L. Walp), as well as arbuscular mycorrhizal fungi in the rhizospheric soil of chickpea and faba bean. Out of the 34 endophytic bacterial populations examined, 31 strains were identified as non-rhizobial based on nodulation tests. All strains were assessed for their plant growth-promoting (PGP) activities in vitro. The results revealed that most isolates exhibited multiple PGP activities, such as nitrogen fixation, indole-3-acetic acid (IAA) and ammonia (NH3) production, phosphate solubilization, and exopolysaccharide production. The most effective PGP bacteria were selected for 16S rRNA analysis. Additionally, a total of 36 species of native arbuscular mycorrhizal fungi (AMF) were identified. Acaulospora (100%) and Scutellospora (91.66%) were the most prevalent genera in Cicer arietinum L. and Vicia faba L. plants, respectively. Acaulospora also exhibited the highest spore density and relative abundance in both plants. Moreover, the root colonization of Cicer arietinum L. and Vicia faba L. plants by hyphae, vesicles, and arbuscules (HVA) was significant. The findings of this study provide valuable insights into non-rhizobial endophytic bacteria associated with legume root nodules and the diversity of AMF. These organisms have great potential for PGP and can be manipulated by co-inoculation with rhizobia to enhance their biofertilizer effectiveness. This manipulation is crucial for promoting sustainable agriculture, improving crop growth, and advancing biofertilizer technology.

Caracterización fenotípica y simbiótica de las bacterias que nodulan Genista saharae en la región árida de Argelia

Twenty bacterial strains had been isolated from root nodules of Genista saharae that grew wild in Biskra and El Oued city (Northeastern Algerian Sahara). This study focused on obtaining isolates of legume nodule bacteria (LNB) from the plant G. saharae and evaluated their effectiveness in forming a symbiotic relationship with the legume species Vigna unguiculata through cross-inoculation. Additionally, the study aimed to identify the successful cross-inoculation group of LNB strains based on their phenotypic characteristics. The growth capacity of isolates under varying salinity conditions [NaCl] and pH levels was investigated using a spectrophotometer (96-microplate reader). The API 20NE and API 20E systems were used to identify the biochemical characteristics of the isolates. In addition, the rhizospheric soil samples from the two study sites were analyzed using standard analytical techniques of soil. All isolates established effective symbioses with Vigna unguiculata, were Gram-negative rods, and were fast-growing. The optimal growth temperature was between 28 °C and 37 °C; some isolates were thermophiles and specifically withstood extreme heat between 45-50 °C. Furthermore, they demonstrated a wide tolerance range to pH (5–10) with salt tolerance ranging from 100 mM to 500 mM. Biochemical results revealed that the isolates assimilated various sources of carbon and nitrogen and displayed numerous enzyme activities. Physicochemical analysis revealed that all the soils were deficient in nutrients and had an alkaline pH. This study enabled us to identify the effective stress-tolerant strains, which could be used in the future to inoculate plants for environmental applications. Keywords: Rhizobia, morpho-physiological and biochemical characterization.

The Effect of Rhizobium Inoculation on the Nutritional Value of Crops in the Legume–Cereal Intercropping System in Northern Kazakhstan

In this study, the changes in yield, nutrient content, and amino acid levels in legume–cereal grass mixtures were qualitatively evaluated depending on the legume–cereal combination and inoculation with preparations based on Rhizobium. This study, taking into account the biological characteristics of legume forage crops, used inoculations with strains of nodule bacteria and associative nitrogen fixers to enhance the process of the nitrogen fixation of mixed crops of legumes and cereal. The aim of this study was to compare the yields and nutritional values of monocultures and mixed crops, as well as to determine the effects of preparations based on strains of nodule bacteria and the associated nitrogen fixer on the photosynthetic activity and yield of combined annual legume–grain crops. A comparative study of forage crop biomass was conducted to analyze crude protein, fiber, carotene, and amino acid content in monocultures and legume–cereal mixtures, with and without the use of nodule-bacteria-based preparations (Rhizotorphin, Mizorine, Flavobactrin, and Azolene). The combined effect of crop mixtures and biological products led to increased green mass yield, protein content, and feed productivity. Notably, two-component mixtures with Rhizotorphin inoculation increased green mass yield by 8.79%, while three-component mixtures saw a 16.49% increase. The oat–pea mixture showed the most significant amino acid improvements, with lysine increasing by 6.26% and tyrosine by 3.24%. The general conclusion reached by the two-year experiment of 2022–2023 in the hill–plain zone of northern Kazakhstan is that double grass mixtures treated with nodule bacteria are more productive than monoculture crops in this area. These results suggest that inoculation with bacterial strains can effectively enhance the productivity of forage crops in northern Kazakhstan, providing a basis for future recommendations on optimizing herbaceous crop combinations. It is recommended to grow annual forage crops in mixtures with legumes to produce highly nutritious feeds with high metabolic energy in terms of biochemical composition.

Frankia [NiFe] uptake hydrogenases and genome reduction: different lineages of loss.

Uptake hydrogenase (Hup) recycles H2 formed by nitrogenase during nitrogen fixation, thereby preserving energy. Among root nodule bacteria, most rhizobial strains examined are Hup-, while only one Hup- Frankia inoculum had been identified. Previous analyses had led to the identification of two different [NiFe] hydrogenase syntons. We analysed the distribution of different types of [NiFe] hydrogenase in the genomes of different Frankia species. Our results show that Frankia strains can contain four different [NiFe] hydrogenase syntons representing groups 1f, 1h, 2a and 3b according to Søndergaard et al. (2016); no more than three types were found in any individual genome. The phylogeny of the structural proteins of groups 1f, 1h and 2a follows Frankia phylogeny; the phylogeny of the accessory proteins does not consistently. An analysis of different [NiFe] hydrogenase types in Actinomycetia shows that under the most parsimonious assumption, all four types were present in the ancestral Frankia strain. Based on Hup activities analysed and the losses of syntons in different lineages of genome reduction, we can conclude that groups 1f and 2a are involved in recycling H2 formed by nitrogenase while group 1h and group 3b are not.

Hot Spots of Site-Specific Integration into the Sinorhizobium meliloti Chromosome.

The diversity of phage-related sequences (PRSs) and their site-specific integration into the genomes of nonpathogenic, agriculturally valuable, nitrogen-fixing root nodule bacteria, such as Sinorhizobium meliloti, were evaluated in this study. A total of 314 PRSs, ranging in size from 3.24 kb to 88.98 kb, were identified in the genomes of 27 S. meliloti strains. The amount of genetic information foreign to S. meliloti accumulated in all identified PRSs was 6.30 Mb. However, more than 53% of this information was contained in prophages (Phs) and genomic islands (GIs) integrated into genes encoding tRNAs (tRNA genes) located on the chromosomes of the rhizobial strains studied. It was found that phiLM21-like Phs were predominantly abundant in the genomes of S. meliloti strains of distant geographical origin, whereas RR1-A- and 16-3-like Phs were much less common. In addition, GIs predominantly contained fragments of phages infecting bacteria of distant taxa, while rhizobiophage-like sequences were unique. A site-specific integration analysis revealed that not all tRNA genes in S. meliloti are integration sites, but among those in which integration occurred, there were "hot spots" of integration into which either Phs or GIs were predominantly inserted. For the first time, it is shown that at these integration "hot spots", not only is the homology of attP and attB strictly preserved, but integrases in PRSs similar to those of phages infecting the Proteobacteria genera Azospirillum or Pseudomonas are also present. The data presented greatly expand the understanding of the fate of phage-related sequences in host bacterial genomes and also raise new questions about the role of phages in bacterial-phage coevolution.

Potential Nitrogen Fixing Rhizobia Isolated from Some Wild Legumes of Nagaland Based on RAPD with Nif-directed Primer and Their Biochemical Activities

Wild legumes are widely dispersed and can survive in challenging environments as bacteria dwell in their nodules and help each other. Although Nagaland is home to many wild legume varieties, research on the microbial diversity that goes along with them is still in its infancy. This work aimed to characterize several wild legume root nodules and distinguish possible rhizobial isolates using RAPD and nif-directed RPO1 primer. Nodule bacteria were isolated in Yeast extract culture media. Based on their colony morphology, 150 isolates were selected for performing RAPD with nif-directed RPO1 primer. Eighty-four isolates were bonded with RPO1 primer, and a few biochemical tests were conducted on RPO1-positive isolates. Activities that promoted plant development were also investigated for these isolates. Of all the isolates, 18 exhibited phosphate solubilization capacity, while 38 isolates were found to produce IAA. This study entails a large variety of rhizobia in the nodules, which were able to promote growth. Hence, these isolates promise to be bio-fertilizers that could improve agricultural operations.

Responsiveness of alfalfa varieties to inoculation by nodule bacteria

Responsiveness of alfalfa varieties to inoculation by nodule bacteria

Effect of Environmental Factors on Recombinant Activity of Root Nodule Bacteria

Effect of Environmental Factors on Recombinant Activity of Root Nodule Bacteria

Screening for drought-tolerant mungbean root nodule bacteria with multiple plant growth promoting traits in Aridisol

Mungbean (Vigna radiata L. Wilczek) is an economically important legume crop grown across India. The growth and yield of the crop have been declining over the recent years due to climate change which leads to decrease in soil moisture. Applications of drought-tolerant plant growth-promoting (PGP) rhizobial strains have been found to enhance crop productivity under water deficiency. Therefore, the objective of this study was to screen the bacterial strains from the mungbean root nodule that may help in improving the plant growth under drought condition. Bacteria were isolated from mungbean root nodules and screened for in vitro drought-tolerance using different concentrations of PEG 6000 at 10, 20, 30 and 40 %, and temperature-tolerance at 30, 35, 40 and 45 °C. In primary screening, out of 98 root nodule bacterial isolates tested, only 25 % showed drought-tolerance at 40 % polyethylene glycol (PEG)-6000 while 22 % of bacteria survived at 45 °C. During secondary screening on combined stress-tolerance, only 8 % of isolates showed tolerance of 40 % PEG-6000 at 45 °C. The in vitro drought-tolerance of bacteria varied significantly according to their sampling region/district. Various PGP traits, i.e., nitrogen fixation, phosphate solubilization, and production of indole acetic acid, ammonia, and 1-aminocyclopropane-1-carboxylate (ACC) deaminase, were analyzed in isolated stress-tolerant bacteria, which may contribute towards stress-tolerance and crop productivity in mungbean. The most promising drought-tolerant nodule bacterial isolates were identified as Rhizobium sp. and Pseudomonas indica, respectively by 16S rRNA sequencing. Moreover, bacterial isolates MuJs52b, MuJs53b, MuJs72a and MuBk32b, which exhibited drought-tolerance of 40 % PEG-6000 and different PGP activities, were used as bioinoculant on mungbean plants grown under moderate to severe drought at 50 and 25 % of field capacity (FC) in pot experiment. Bacteria inoculated plants showed maximum increase in nodule dry weight (56.5 %) and shoot dry weight (87.5 %) per plant even under severe drought at 25 % FC. The results indicated that application of four potential nodule bacterial isolates, which were able to tolerate drought stress of 40 % PEG-6000, and having multiple PGP traits, showed stimulation of the mungbean growth even up to 25 % FC. These plant growth-promoting nodule bacterial strains may be used for the enhancement of mungbean crop productivity under drought stress and could be used as biofertilizer.

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.

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.

Isolation, characterization and identification of root nodule bacteria from Macrotyloma uniflorum (Lam.) Verdc

The present study was conducted to characterize the native plant growth promoting rhizobacteria (PGPRs) from the nodules of Macrotyloma uniflorum (Lam.) Verdc. The seeds of Macrotyloma uniflorum was grown in two different soils i.e., sterile and non-sterile soil and four bacterial isolates were isolated. The bacterial isolates were characterised on the basis of morphological and different biochemical analysis. Phylogenetic identification was done on the basis of 16S rRNA analysis. Four isolates obtained belongs to Bacillus velezensis and Enterobacter cloacae.

Phenotypic and genotypic diversity of root nodule bacteria from wild Lathyrus and Vicia species in Gaziantep, Turkey.

This study identified the phenotypic and genotypic characteristics of the bacteria that nodulate wild Lathyrus and Vicia species natural distribution in the Gaziantep province of Turkey. Principle component analysis of phenotypic features revealed that rhizobial isolates were highly resistant to stress factors such as high salt, pH and temperature. They were found to be highly sensitive to the concentrations (mg/mL) of the antibiotics neomycin 10, kanamycin, and tetracycline 5, as well as the heavy metals Ni 10, and Cu 10, and 5. As a result of REP-PCR analysis, it was determined that the rhizobial isolates were quite diverse, and 5 main groups and many subgroups being found. All of the isolates nodulating wild Vicia species were found to be related to Rhizobium sp., and these isolates were found to be in Clades II, III, IV, and V of the phylogenetic tree based on 16S rRNA. The isolates that nodulated wild Lathyrus species were in Clades I, II, IV, V, VI, VII, and VIII, and they were closely related to Rhizobium leguminasorum, Rhizobium sp., Phyllobacterium sp., Serratia sp., and Pseudomonas sp. According to the genetic analyses, the isolates could not be classified at the species level, the similarity ratio was low, they formed a distinct group that was supported by strong bootstrap values in the phylogenetic tree, and the differences discovered in the network analysis revealed the diversity among the isolates and gave important findings that these isolates may be new species.

ВЛИЯНИЕ ПРИМЕНЕНИЯ СРЕДСТВ ИНТЕНСИФИКАЦИИ НА БИОЛОГИЧЕСКУЮ АКТИВНОСТЬ И ФИТОСАНИТАРНОЕ СОСТОЯНИЕ ПОЧВЫ ПОД ПОСЕВОМ ГОРОХА

The aim of research is to determine the parameters of biological activity of meadow-chernozem soil and its phytoimmunity in grain-fallow crop rotation at different levels of cultivation intensification and the use of chemicals. The studies were carried out in a long-term (launched in 1973) stationary experiment in grain-fallow crop rotation (fallow – wheat – peas – wheat – barley) under pea crop in the Federal State Budgetary Scientific Institution Omsk Agrarian Scientific Center. In certain periods of 2022 and 2023, with favorable soil moisture, the presence of a significant amount of plant residues in the variant with minimum-zero tillage and the use of complex chemicals, the number of soil micromycetes and nodule bacteria increased to 93.0 thousand and 13.9 million CFU/g, respectively. The total number of soil microflora in the arable layer under different soil cultivation systems for peas without the use of complex chemicals was 51.4–61.2 million CFU/g with a difference of 19.0 % in the ascending series: No-till < moldboard < combined. The use of mineral fertilizers in combination with plant protection products in moldboard and combined tillage did not have a significant effect on the amount of soil microflora, a slight decrease to 17.0 % within the experimental error was noted. In the variants with No-till tillage in combination with intensification products, stimulation of the growth of the number of microorganisms to 22.0 % in relation to the control was noted. On average, according to the soil tillage factor, the greatest development of pea root rot was observed in the No-till variant (3.8 %), a slight decrease (3.2–3.6 %) – in moldboard and combined tillage. The use of plant protection products during the growing season of peas made it possible to reduce the development of root rot by 47.0 % relative to the control, depending on the soil tillage method. With combined treatment, the greatest prevalence of pea root system disease was noted (37.8 %), the least – with No-till treatment (30.8 %). In laboratory conditions, using the wet chamber method, a complex of root rot pathogens was determined, among which fungi of the genus Fusarium spp. prevailed.

Bio-Efficacy of New Herbicide Molecules for Weed Management in Grain Legumes

Grain legumes including pigeon pea, chickpea, peanut, soybean, lentil, cowpea, common bean, faba bean, pea, and horse gram are extensively cultivated globally. Higher protein content, coupled with symbiotic nitrogen-fixing bacteria in root nodules enabling nitrogen fixation, highlights their importance in minimizing fertilizer use for agricultural production systems. Legume cultivation is limited by prolonged weed interference which is managed either through mechanical, cultural, chemical, or biological methods separately or in combinations. Nevertheless, herbicides remain and will persist as a crucial and economical element in global crop cultivation. While first-generation herbicides benefit agriculture, their long-term persistence and off-target toxicity have significant negative consequences, causing harm to both the environment and humans. Moreover, weed resistance and similar chemistry enforce the demand for new modes of action. Therefore, continuous research into new herbicides is important for combating herbicide resistance and ensuring crop output. In recent years, a few herbicides with novel mechanisms of action have occurred. Plants treated with new-generation herbicides resulted in increased WCE (weed control efficiency), decreased weed dry matter, and seed yield. There were no or minimal phytotoxicity symptoms observed on the existing and succeeding crops. This review emphasizes the importance of newly formulated herbicides signifying high selectivity, low toxicity, target specificity, and minimal application rates, contributing to an enhanced understanding of human and environmental health. Furthermore, it emphasizes the significance of economic viability and environmental friendliness in these formulations.

Morphological and physiological properties of lupine nodule bacteria (Bradyrhizobium lupini) when grown on a typical urban soil.

In the conducted studies, the moorphological and physiological properties of nodule bacteria of lupine were studied. Lupine plants were grown under the conditions of a microfield experiment on a typical medium loamy urban soil. In the study, a pure culture of Bradyrhizobium lupini was isolated. Then, the morphological properties of nodule bacteria cells and the chemical composition of cell membranes of nodule bacteria were determined. The acid resistance and physiological properties of lupine nodule bacteria were also determined, as well as the ratio of Bradyrhizobium lupini to antibiotics. All studies were carried out according to generally accepted methods. The results of the research showed that during the cultivation of lupine on a typical urban soil, nodule bacteria Bradyrhizobium lupini were isolated, which can be characterized as gram-negative, non-spore-forming rods that do not exhibit amylolytic activity. It was revealed that the rhizobia of nodule bacteria are not acid-resistant. Nodule bacteria turned out to be the least resistant to polymyxin, then to levomycetin, and Bradyrhizobium lupini showed the greatest resistance to tetracycline.

ВЛИЯНИЕ ПРЕДПОСЕВНОЙ ИНОКУЛЯЦИИ СЕМЯН И ВНЕКОРНЕВОГО ВНЕСЕНИЯ ПРЕПАРАТОВ НА ПРОДУКТИВНОСТЬ И КАЧЕСТВО ЗЕРНА СОИ В УСЛОВИЯХ ОРОШЕНИЯ ДОЖДЕВАНИЕМ В НИЖНЕМ ПОВОЛЖЬЕ

Purpose: to determine the effect of pre-sowing seed treatment and foliar application of drugs on the productivity and quality of the resulting soybean grain at various water availability under irrigation. The use of a biological preparation as an, inoculant which contributed to the growth of the necessary nodule bacteria on the root system of a given crop and, as a result, carried out the symbiosis of a legume plant, for the assimilation of nitrogen from the air and its conversion into a mineral compound in the soil. Materials and methods. Studies of this technique were conducted on light chestnut soils of the Volgograd region with a medium-ripened variety of own selection Volgogradka 2, bred by the All-Russian Research Institute of Irrigated Agriculture. The laying of experiments and related observations were carried out according to generally accepted methods for two years 2022–2023 on the lands of the All-Russian Research Institute of Irrigated Agriculture (experimental station “Irrigated”, Vodny settlement, Volgograd). Results. The obtained data prove the advantage of pre-sowing inoculation of seeds under different water regimes of this crop. At the same time, the increase in the yield of soybean grain, depending on such pre-sowing treatment, exceeded the control option by 0.40 t/ha under the water regime of 70-80-70 % the lowest moisture capacity and 0.54 t/ha not lower than 80% the lowest moisture capacity. Conclusions. In the conducted studies, the best option proved to be the option with a pre-watering soil moisture threshold of 80% the lowest moisture capacity in moisture layers of 0.4 and 0.6 m and pre-sowing inoculation of seeds in combination with leaf spraying with preparations of various origins, which contributed to a maximum yield of 3.91 t/ha and a profitability level of 242.10 %.

Influence of cultivation techniques on productivity and quality of meadow clover

Meadow clover (red clover) is the most important fodder plant in most parts of Russia. Positive characteristics of this plant are the presence of a significant amount of reserve nutrients (crude protein, protein, nitrogenfree extractive substances) in the composition of plant tissues, which provides high fodder value. As a legume, meadow clover accumulates in the soil organically bound nitrogen, a large amount of organic mass of crop root residues, enriching the soil with nutrients and contributing to the increase of its physical and mechanical properties. The disadvantage of this crop is its short productive life. In this regard, it is necessary to intensify the cultivation methods for a short period of use in order to obtain the greatest return from the crop in the fodder and agronomic aspect. Intensification of cultivation methods means everything that contributes to the increase of plant productivity: application of mineral and organic fertilizers to improve mineral nutrition, inoculation of seeds before sowing to mobilize and activate soil microflora and productive symbiosis between plants and microbiota, use of growth regulators to optimize photosynthesis processes. The study of the influence of these intensification methods was the purpose of our research. It was found that the use of estimated doses of mineral fertilizers contributed most optimally to the processes of yield formation and final productivity of clover grass. Among all the strains of nodule bacteria studied, inoculant strain 348a was the most effective because it is speciesspecific for meadow clover. The use of growth regulators Ribav Extra and Root Super is a positive technique that provides optimal growth and development of plants during vegetation. The methods studied also had a positive effect on the seed productivity of the clover plants, increasing yield and seed quality.

Genomic and Transcriptomic Analysis of Pea (Pisum sativum L.) Breeding Line 'Triumph' with High Symbiotic Responsivity.

Pea (Pisum sativum L.), like most legumes, forms mutualistic symbioses with nodule bacteria and arbuscular mycorrhizal (AM) fungi. The positive effect of inoculation is partially determined by the plant genotype; thus, pea varieties with high and low symbiotic responsivity have been described, but the molecular genetic basis of this trait remains unknown. Here, we compare the symbiotically responsive breeding line 'Triumph' of grain pea with its parental cultivars 'Vendevil' (a donor of high symbiotic responsivity) and 'Classic' (a donor of agriculturally valuable traits) using genome and transcriptome sequencing. We show that 'Triumph' inherited one-fourth of its genome from 'Vendevil', including the genes related to AM and nodule formation, and reveal that under combined inoculation with nodule bacteria and AM fungi, 'Triumph' and 'Vendevil', in contrast to 'Classic', demonstrate similar up-regulation of the genes related to solute transport, hormonal regulation and flavonoid biosynthesis in their roots. We also identify the gene PsGLP2, whose expression pattern distinguishing 'Triumph' and 'Vendevil' from 'Classic' correlates with difference within the promoter region sequence, making it a promising marker for the symbiotic responsivity trait. The results of this study may be helpful for future molecular breeding programs aimed at creation of symbiotically responsive cultivars of pea.

Genetically Modified Legume Plants as a Basis for Studying the Signal Regulation of Symbiosis with Nodule Bacteria

The development of legume–rhizobial symbiosis results in the formation of nitrogen-fixing root nodules. In response to rhizobial molecules, Nod factors, signal transduction is mediated by the interaction of activated receptors with downstream signaling proteins. Previously, some new regulators of the signal pathway, such as phospholipases D, which regulate the level of phosphatidic acid (PA), as well as mitogen-activated protein kinases (MAPKs), have been identified in legumes. Since PA is an important signal messenger, we tested the hypothesis that increasing the level of proteins involved in the reversible binding of PA in plant tissues may have a positive effect on symbiosis. Our findings showed that overexpression of MtSPHK1-PA, encoding the PA-binding domain of sphingosine kinase 1 (SPHK1), stimulated plant growth and nodule development in legume plants. Furthermore, the influence of MAPK6 on the development of symbiosis was studied. Using genetic engineering methods, we increased MAPK6 transcriptional activity in transgenic roots, leading to an increase in the number of nodules and the biomass of pea plants. Therefore, new approaches to obtain plants with an increased efficiency of symbiosis were tested. We report here that both genes that encode signaling proteins may be used as potential targets for future modification using biotechnological approaches.