Nodule Bacteria Research Articles

Unveiling root nodulation in Tribulus terrestris and Roystonea regia via metagenomics analysis.

Root nodule symbiosis is traditionally recognized in the Fabales, Fagales, Cucurbitales, and Rosales orders within the Rosid I clade of angiosperms. However, ambiguous root nodule formation has been reported in Zygophyllaceae and Roystonea regia (Arecaceae), although a detailed analysis has yet to be conducted. We aimed to perform morphological analyses of root structures in these plants and utilize metagenomic techniques to identify and characterize the bacterial populations within the nodule-like structures. We collected root samples of Tribulus terrestris (Zygophyllaceae) and Roystonea regia from West Asia and the Caribbean, respectively. We conducted detailed morphological analyses of nodule-like structures, isolated and genome-sequenced the endophytes, and employed metagenomic techniques to identify the bacterial populations within these formations. We observed nodule-like structures in both plant species. Symbiosomes, which are hallmark structures of nodulating plants, were not detected. Metagenome sequence data analysis revealed potential nodulating and nitrogen-fixing bacteria in the nodule-like structures of both species. Canonical nodulation and nitrogen-fixation genes were identified in microbes associated with the nodules. However, the phylogenomic analysis showed that the bacteria isolated from T. terrestris and R. regia are within Gammaproteobacteria and Bacilli, which are not typically known as nodulating bacteria. The observed structures differ significantly from traditional nodules found in legumes and actinorhizal plants, suggesting unique characteristics with hosting nitrogen-fixing bacteria. Although bacteria identified through in silico analysis or culture are well-known nitrogen-fixers, their specific role in root nodule formation remains to be investigated.

Morphological and Molecular Characterization of Endophytic Bacteria Isolated from Bambara Groundnut (Vigna subterranea) Nodules in Daloa, Côte d’Ivoire

A thorough understanding of the diversity, functions and specific interactions of endophytic bacteria present in legumes is essential for improving crop production. This study aimed to identify roots nodule endophytes associated to Bambara groundnut by analyzing their morphological diversity and genetic diversity. A total of 34 bacterial isolates were sampled from different cultivation sites and associated with 5 local varieties of Bambara groundnut. Morphological analysis using macroscopic and microscopic observation revealed that the bacterial isolates were morphologically diverse in terms of colony appearance, shape, colony size, color, opacity and Gram stain result. These bacteria exhibited characters different from those of the symbiotic bacteria. In addition, molecular identification based on sequencing the 16S-rRNA gene and 1081 bp analysis showed the existence of non-symbiotic bacteria in Bambara groundnut nodule. The community of roots nodule endophyte isolated from Bambara groundnut belonged to the genera Pseudomonas, Bacillus, Bacterium, microbacterium, Rahnella, Paenibacillus, Lysobacter with 98 to 99.5% similarity, classified under the Firmicutes, Actinobacteria and Proteobacteria phyla. In Bambara groundnut, the predominant nodule endophytes were Bacillus (56%) and Pseudomonas (17%). Future research could further investigate the ecological implications of these bacterial interactions, and their potential applications in biotechnology and agronomy, particularly in biofertilization and host plant growth promotion.

Relation of soybean productivity to the functioning of the symbiotic and photosynthetic apparatuses

Background. Increasing the yield of soybean necessitates the maintenance of a high protein level in seeds, and therefore the process of fixing atmospheric N2. Seed inoculation with soybean nitrogen-fixing bacteria is known to improve N2-fixation and soybean grain yield. At the same time, the introduction of new nodule bacteria strains into preparations for soybean inoculation requires the study of their influence on the main interconnected physiological processes that form the basis of leguminous plants productivity – N2-fixation and photosynthesis. The aim of the work was to study the relationship of vegetative growth and grain productivity of soybean inoculated with new nodule bacteria Bradyrhizobium japonicum strains of different functional activity with the plants’ symbiotic and photosynthetic apparatuses functioning. Materials and Methods. The research was carried out on symbiotic systems created with soybean plants (Glycine max (L.) Merr.) of the Almaz variety and nodule bacteria B. japonicum strains: analytically selected PC09, and recombinant strains B157, B201, D45, D52 (pSUP5011::Tn5mob) and C30 (pSUP2021::Tn5) from the N2-fixing microorganisms museum collection of the Institute of Plant Physiology and Genetics NAS of Ukraine. Research methods – microbiological, biochemical and physiological, statistical analysis. Results. It was found that the N2-fixing activity (NFA) of nodules formed by B. japoni­cum PC09, D45, D52, B157 and B201 strains at the stage of 3 true leaves exceeded the NFA of nodules formed by Tn5-mutant C30 by 1.6–4.0 times, and at the stage of budding–beginning of flowering – by 4.2–6.2 times. Highly active strains also differed from each other in NFA, although to a lesser extent than with strain C30. On the basis of a comparative analysis of the physiological indices of soybean inoculated with B. japonicum strains of different activity, close positive linear correlations were found between NFA, photosynthetic rate, and the biological and grain productivity of plants. Conclusions. The results obtained indicate that the higher the nodulating and NFA of rhizobia in the symbiotic system soybean–Bradyrhizobium japonicum, the higher the functional activity of photosynthetic apparatus formed by plants. This provides a more complete genetic potential release of soybean crop productivity.

Isolasi dan Karakterisasi Bakteri Rhizobium asal Bintil Akar Tanaman Kacang Tanah (Arachis hypogaea) dan Koro Rawe (Mucuna bracteata)

Nitrogen (N) is naturally available in the form of N2. Nitrogen fixation can be facilitated by microbes such as Rhizobium bacteria, which can establish symbiosis with the roots of legume plants to form root nodules that perform nitrogen fixation. This study aims to determine the characteristics of Rhizobium bacteria in the root nodules of peanuts (Arachis hypogaea) and koro rawe (Mucuna bracteata). The methods used in this research include collecting root nodules, isolating Rhizobium bacteria, and identifying and characterizing these bacteria for their nitrogen fixation and phosphate solubilization capabilities. Characterization results on YEMA + Congo red media indicated that 10 out of 16 isolates from peanut root nodules and 30 out of 55 isolates from koro rawe root nodules were positive for Rhizobium bacteria. The nitrogen fixation activity test revealed that only 4 isolates from peanut root nodules and 6 from koro rawe root nodules exhibited nitrogen fixation ability in Jensen's medium. The highest phosphate solubilization index was obtained from isolate KK.5.1.1 (0.5±0.08), while the lowest was from isolate KK.5.2.2 (0.09±0.06). This research underscores the significance of Rhizobium in enhancing nitrogen availability and phosphate solubilization for legume plants.

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, Pedersen, and Greening (HydDB: a web tool for hydrogenase classification and analysis. Sci Rep 2016;6:34212. https://doi.org/10.1038/srep34212.); 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.

Symbiotic activity and productivity of soybean in the Western Forest Steppe

Purpose. To determine the effect of seed inoculation and foliar application of fertilisers on the growth and development of soybean. Methods. Field research was carried out in 2019–2021 in the fields of the Kozatska Dolyna 2006 LLC (Khmelnytskyi Region). Experiments were carried out according to the following design: factor A: inoculation of seeds: 1) without inoculation, 2) inoculation with the biopreparation Rizoactive; factor B: foliar feeding: 1) without foliar feeding, 2) foliar feeding with Fulvohumin; factor C: soybean varieties: 'Samorodok' (control), 'Rohiznianka', 'Triada', 'Orfei', 'Evridika', 'Aratta', 'Azymut', and 'Avrora'. Results. Foliar application of Fulvohumin contributed to obtaining 4.09 more nodules per plant. At the same time, inoculation with Bradyrhizobium japonicum strains eko/001, eko/002, and eko/003 contributed to an increase in the number of nodules by 8.35 (29.5 nodules per plant in the control treatment). That is, in areas where soybean is grown quite often, a certain indigenous type of microorganisms is supported in the soil, capable of inoculating plants during soybean cultivation; however, the use of pure, dedicated strains of microorganisms is a more effective way of increasing the efficiency of symbiotic interactions. In the case of seed inoculation with Rizoactive and foliar application of Fulvohumin, the best conditions for the formation of nodules were formed by varieties 'Azymut' (1.30 g/plant), 'Orfei' (1.31 g/plant), and 'Rohiznianka' (1.32 g/plant). Therefore, the improvement of the physiological state of plants with foliar application of Fulvohumin contributed to a general increase in the weight of nodules by 0.20 g/plant. However, inoculation with strains of Bradyrhizobium japonicum eko/001, eko/002 and eko/003 was more effective, because, due to the displacement of indigenous strains of microorganisms an increase in the weight of nodules of 0.36 g/plant was obtained. Based on the analysis of the factors of influence, it was determined that seed inoculation is a driver of changes in the number of colonies of active nodule bacteria on soybean roots (50%), even under the strong influence of the weather conditions of the growing season (33%). The importance of inoculation in the beginning of photosynthesis with the use of Bradyrhizobium japonicum strains eko/001, eko/002 and eko/003 is obvious. The mass of active nodules on plant roots is even more dependent on seed inoculation (57%), which supports the assumption that local native species are not able to cooperate with soybean plants as effectively as specially selected strains of nodule bacteria. On average, according to the experiment, the yield of soybean seeds in 2019 was 3.41 t/ha, in 2020, it was 3.10, in 2021, it was 3.92, and in 2022, it was 3.20 t/ha. The all-year leader in grain yield was the 'Aurora' variety. However, in 2019, 'Triada' showed good results, in 2020 – 'Azymut' and 'Triada', in 2021 – 'Triada', and in 2022 – 'Orfei', 'Azymut' and 'Triada' showed good results. This indicates the individual varietal response to growing conditions and the importance of technology adaptation for certain varieties and their in-depth verification in the area of ​​distribution regarding the possibility of forming a high yield level. Conclusions. The correct choice of mineral nutrition for soybean provided general stimulation of plant development and the subsequent formation of the legume-rhizobial relationship and its functioning from the point of view of assimilation of nitrogen from the atmosphere. This emphasises the role of small doses of starter nitrogen fertilisers from the standpoint of better crop development, especially when combined with foliar feeding of plants during the growing season. Even with the maximum need of plants for nutrients at the time of the formation of beans and seeds, and filling grain, their good start at the beginning of the growing season provided an advantage due to better rooting, development, and work of the leguminous rhizobial apparatus, etc. And therefore, against the background of the main fertilizer N30Р60K60, foliar application of Fulvohumin in combination with soybean seed inoculation contributed to the formation of a high level of productivity. All varieties increased their productivity in these treatments: 3.70 t/ha in 'Evridika', 3.76 t/ha in 'Aratta', and 4.02 t/ha in 'Orfei', while the highest indicators over the years of research were recorded in 'Azymut' (4.11 t/ha), ‘Triada’ (4.12 t/ha), and ‘Avrora’ (4.16 t/ha).

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.

INTERACTION BETWEEN LUPIN PLANTS AND STRAINS OF THE BRADYRHIZOBIUM GENUS

Objective. Study the peculiarities of the interaction between three species of lupin and nodule bacteria of the genus Bradyrhizobium, isolated from various leguminous plants: lupin, seradella, soybean and cowpea. Methods. Microbiological (isolation of rhizobia from nodules and their cultivation), serological (defining serological affiliation of strains), vegetation experiment (study of host specificity of bradyrhizobia, study of the formation and functioning of the symbiosis between lupin and nodule bacteria), gas chromatographic (determination of the activity of symbiotic nitrogen fixation), mathematical and statistical. Results. Under the conditions of vegetation experiments on sod-podzolic soil and sterile vermiculite, the peculiarities of the interaction between different species of lupin and specific and non-specific strains of nodule bacteria of the genus Bradyrhizobium were studied. It was established that rhizobia isolated from lupin nodules (B. lupini 367a, 30l, LD4 and Bradyrhizobium sp. LD8), seradella (Bradyrhizobium sp. СР1) and soybean (B. lupini 631) are able to form an active symbiosis with white and yellow lupin plants. Microsymbionts of soybean (B. japonicum КВ11) and cowpea (B. diazoefficiens В22) did not infect these two species of lupin. In the experiment with narrow-leaved lupin, all strains of B. lupini (367а, 30l, LD4, 631) and LD8 Bradyrhizobium sp. ensured a reliable increase in the above-ground mass of plants by 14.3–22.6 % compared to the control. The maximum values of this indicator were obtained in variants inoculated with B. lupini LD4 and B. lupine 631 strains (1.03 and 1.02 g/plant versus 0.84 g/plant in the control). With the use of non-specific lupin strains B. japonicum КВ11 and B. diazoefficiens В22, a significant increase in the nitrogenase activity of nodules was registered (1.5 and 1.9 times compared to the control), however, the increase in the above-ground mass of plants was insignificant — 7.1 % and 10.7 %, respectively. Conclusion. It has been established that lupin-specific bradyrhizobia of various origins (B. lupini and Bradyrhizobium sp.) have a positive effect on the formation of the symbiotic apparatus, the growth and development of white, yellow and narrowleaved lupin plants. Rhizobia of soybean and cowpea (B. japonicum КВ11 and B. diazoefficiens В22) non-specific for lupin, contribute to the activation of the interaction between narrowleaved lupin and representatives of the soil population of B. lupini and act as PGPR microorganisms. B. lupini LD4 and B. lupini 631 strains are the best in terms of most symbiotic indicators and efficiency of symbiosis with narrow-leaved lupin.

SELECTION OF NEW STRAINS OF NODULE BACTERIA-SYMBIONTS OF MELILOTUS ALBUS AND TRIGONELLA FOENUM-GRAECUM

Objective. Study the properties of effective strains of nodule bacteria-microsymbionts of white melilot (Melilotus albus) and fenugreek (Trigonella foenum-graecum). Methods. Microbiological, biochemical, vegetative experiment, statistical. Results. During 2016–2021, a search for active strains of nodule bacteria of rare types of leguminous plants was carried out. Analytical selection methods have found 11 strains of Ensifer sp. — microsymbionts of white melilot and 3 of fenugreek. It was shown that the inoculation of white melilot seeds with bacterial suspensions of new strains of Ensifer sp. BR-4 and BR-201 contribute to an increase in the formation of nodules on plant roots by 15 % and 24 % versus the positive control (inoculation with Ensifer meliloti ДН-15), as well as an increase in vegetative mass by 25 % and 39 %, respectively. It was also established that during seed inoculation with suspensions of Ensifer sp. G-6 and G-10, the number of nodules exceeded this indicator of the positive control by 52 % and 72 %, the increase in vegetative mass was 4 % and 25 %, respectively. The peculiarities of the growth of new effective strains of rhizobia on pea agar medium, meat-peptone and mannitol-yeast agar, litmus milk, the ability to assimilate various sources of carbon and nitrogen and resistance to antibiotics were studied. The differences in the nature of growth on MPA and assimilation of nitrogen and carbon sources for melilot and fenugreek rhizobia were shown. Based on the results of research into the cultural-morphological and physiological-biochemical properties of the new microsymbiont isolates of the white melilot and fenugreek, it was shown that they correspond to the properties of the reference strain and characteristics of E. meliloti species. Conclusion. Analytical selection methods were used to obtain new strains of microsymbionts of the white melilot and fenugreek. The positive effect of bacterization of the seeds of these cultures with bacterial suspensions of the obtained strains on the symbiotic activity and productivity of plants was established. According to the cultural-morphological and physiological-biochemical properties, the new isolates have been assigned to E. meliloti. Differences in the nature of growth on MPA and assimilation of carbon and nitrogen between the rhizobia of Melilotus albus and Trigonella foenum-graecum were revealed.

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.