Symbiosome-like intracellular colonization of cereals and other crop plants by nitrogen-fixing bacteria for reduced inputs of synthetic nitrogen fertilizers

It has been forecast that the challenge of meeting increased food demand and protecting environmental quality will be won or lost in maize, rice and wheat cropping systems, and that the problem of environmental nitrogen enrichment is most likely to be solved by substituting synthetic nitrogen fertilizers by the creation of cereal crops that are able to fix nitrogen symbiotically as legumes do. In legumes, rhizobia present intracellularly in membrane-bound vesicular compartments in the cytoplasm of nodule cells fix nitrogen endosymbiotically. Within these symbiosomes, membrane-bound vesicular compartments, rhizobia are supplied with energy derived from plant photosynthates and in return supply the plant with biologically fixed nitrogen, usually as ammonia. This minimizes or eliminates the need for inputs of synthetic nitrogen fertilizers. Recently we have demonstrated, using novel inoculation conditions with very low numbers of bacteria, that cells of root meristems of maize, rice, wheat and other major non-legume crops, such as oilseed rape and tomato, can be intracellularly colonized by the non-rhizobial, non-nodulating, nitrogen fixing bacterium, Gluconacetobacter diazotrophicus that naturally occurs in sugarcane. G. diazotrophicus expressing nitrogen fixing (nifH) genes is present in symbiosome-like compartments in the cytoplasm of cells of the root meristems of the target cereals and non-legume crop species, somewhat similar to the intracellular symbiosome colonization of legume nodule cells by rhizobia. To obtain an indication of the likelihood of adequate growth and yield, of maize for example, with reduced inputs of synthetic nitrogen fertilizers, we are currently determining the extent to which nitrogen fixation, as assessed using various methods, is correlated with the extent of systemic intracellular colonization by G. diazotrophicus, with minimal or zero inputs.     

OBPC Symposium: Maize 2004 &; beyond—Intracellular colonization of cereals and other crop plants by nitrogen-fixing bacteria for reduced inputs of synthetic nitrogen fertilizers

Summary The problem of environmental nitrogen enrichment is most likely to be solved by reducing the inputs of synthetic nitrogen fertilizers through the creation of cereals that, like legumes, are able to fix nitrogen. In legumes, rhizobia present intracellularly in vesicles in the cytoplasm of nodule cells fix nitrogen endosymbiotically. Rhizobia within these membrane-bounded compartments are supplied with energy from plant photosynthates and, in return, the bacteria provide the plant with biologically fixed nitrogen. Recently, we have demonstrated, using novel inoculation conditions with very low numbers of bacteria, that cells of the root meristems of maize, rice, wheat, and other major non-legume crops can be colonized intracellularly by the non-rhizobial, non-nodulating, nitrogen-fixing bacterium, Gluconacetobacter diazotrophicus, that occurs naturally in sugarcane. G. diazotrophicus expressing nitrogen-fixing genes is present in membrane-bounded compartments in the cytoplasm of cells of the root meristems of the target cereals and non-legume species, similar to the intracellular colonization of legume root nodule cells by rhizobia. In order to obtain an indication of the likelihood of adequate growth and yield of maize, for example, with reduced inputs of synthetic nitrogen fertilizers, we are determining the extent to which nitrogen fixation is correlated with systemic intracellular colonization by G. diazotrophicus, with minimal or zero inputs of synthetic nitrogen fertilizer.     

Symbiosome-like intracellular colonization of cereals and other crop plants by nitrogen-fixing bacteria for reduced inputs of synthetic nitrogen fertilizers

It has been forecast that the challenge of meeting increased food demand and protecting environmental quality will be won or lost in maize, rice and wheat cropping systems,and that the problem of environmental nitrogen enrichment is most likely to be solved by substituting synthetic nitrogen fertilizers by the creation of cereal crops that are able to fix nitrogen symbiotically as legumes do. In legumes, rhizobia present intraceliularly in membrane-bound vesicular compartments in the cytoplasm of nodule cells fix nitrogen endosymbiotically. Within these symbiosomes, membrane-bound vesicular compartments, rhizobia are supplied with energy derived from plant photosynthates and in return supply the plant with biologically fixed nitrogen, usually as ammonia. This minimizes or eliminates the need for inputs of synthetic nitrogen fertilizers. Recently we have demonstrated, using novel inoculation conditions with very low numbers of bacteria, that cells of root meristems of maize, rice, wheat and other major non-legume crops, such as oilseed rape and tomato, can be intracellularly colonized by the non-rhizobial, non-nodulating, nitrogen fixing bacterium, Gluconacetobacter diazotrophicus that naturally occurs in sugarcane. G. diazotrophicus expressing nitrogen fixing (nifH) genes is present in symbiosome-like compartments in the cytoplasm of cells of the root meristems of the target cereals and non-legume crop species, somewhat similar to the intracellular symbiosome colonization of legume nodule cells by rhizobia. To obtain an indication of the likelihood of adequate growth and yield, of maize for example, with reduced inputs of synthetic nitrogen fertilizers,we are currently determining the extent to which nitrogen fixation, as assessed using various methods, is correlated with the extent of systemic intracellular colonization by G. diazotrophicus,with minimal or zero inputs.     

Molecular diversity and phylogeny of indigenous <Emphasis Type="Italic">Rhizobium leguminosarum</Emphasis> strains associated with <Emphasis Type="Italic">Trifolium repens</Emphasis> plants in Romania

The symbiotic nitrogen fixing legumes play an essential role in sustainable agriculture. White clover (Trifolium repens L.) is one of the most valuable perennial legumes in pastures and meadows of temperate regions. Despite its great agriculture and economic importance, there is no detailed available information on phylogenetic assignation and characterization of rhizobia associated with native white clover plants in South-Eastern Europe. In the present work, the diversity of indigenous white clover rhizobia originating in 11 different natural ecosystems in North-Eastern Romania were assessed by a polyphasic approach. Initial grouping showed that, 73 rhizobial isolates, representing seven distinct phenons were distributed into 12 genotypes, indicating a wide phenotypic and genotypic diversity among the isolates. To clarify their phylogeny, 44 representative strains were used in sequence analysis of 16S rRNA gene and IGS fragments, three housekeeping genes (atpD, glnII and recA) and two symbiosis-related genes (nodA and nifH). Multilocus sequence analysis (MLSA) phylogeny based on concatenated housekeeping genes delineated the clover isolates into five putative genospecies. Despite their diverse chromosomal backgrounds, test strains shared highly similar symbiotic genes closely related to Rhizobium leguminosarum biovar trifolii. Phylogenies inferred from housekeeping genes were incongruent with those of symbiotic genes, probably due to occurrence of lateral transfer events among native strains. This is the first polyphasic taxonomic study to report on the MLSA-based phylogenetic diversity of indigenous rhizobia nodulating white clover plants grown in various soil types in South-Eastern Europe. Our results provide valuable taxonomic data on native clover rhizobia and may increase the pool of genetic material to be used as biofertilizers.     

Polyphasic characterization of rhizobia microsymbionts of common bean [<Emphasis Type="Italic">Phaseolus vulgaris</Emphasis> (L.)] isolated in Mato Grosso do Sul,a hotspot of Brazilian biodiversity

Common bean [Phaseolus vulgaris (Linnaeus)] is the key source of protein, carbohydrates and micronutrients for over 300 million people in the tropics. Like many legumes, P. vulgaris can fix atmospheric nitrogen in symbiosis with rhizobia, alleviating the need for the expensive and polluting N-fertilizers. The crop is known to nodulate with a wide range of rhizobia and, although Brazil is not a center of genetic origin/domestication of P. vulgaris, a variety of rhizobial species have been found as symbionts of the legume. Mato Grosso do Sul (MS) is one of the largest common bean producer states in Brazil, with reports of high yields and abundant natural nodulation. The objective of this study was to evaluate the diversity of 73 indigenous rhizobia isolated from common bean grown in 22 municipalities of MS. Great morphophysiological and genetic diversity was found, as indicated by the six and 35 clusters formed, considering the similarity level of 75 and 70%, respectively, for the phenotypic and rep-PCR dendrograms. Eleven representative isolates were selected for detailed genetic characterization using 16S rRNA and three protein-coding housekeeping genes, glnII, gyrB and recA. We identified species originated from the centers of origin/domestication of the legume, R. etli and R. phaseoli, species probably indigenous of Brazil, R. leucaenae and others of the Rhizobium/Agrobacterium clade, in addition to putative new species. The results highlight the great rhizobial diversity of the region.     

Genetic markers for search of rhizobia based on symbiotic genes

The possible application of rhizobial symbiotic genes as markers for the search and primary identification of rhizobia from temperate-zone legumes was studied. It was shown that conservative sym genes nifH and nifD could be used as markers for rapid search of rhizobia among the analyzed isolates, while more variable genes nifK and nodC could be used for their primary identification. Efficiency of the proposed method was shown in analysis of bacterial isolates obtained from Onobrychis arenaria and Astragalus cicer root nodules.     

Recent changes to the classification of symbiotic,nitrogen-fixing,legume-associating bacteria: a review

The Rhizobia are collectively comprised of gram negative soil bacteria that have the ability to form symbiotic nitrogen-fixing root and/or stem nodules in association with leguminous plants. The taxonomy of these bacteria is continually in a state of flux, in large part due to rapid development of refined molecular biology techniques. The isolation and characterization of new, and often different, legumes-nodulating bacteria on a variety of plant hosts has resulted in the naming of many new rhizobial species. Here we update the taxonomy of the legume-nodulating bacteria and describe newly identified rhizobia capable of nodulating edible legumes and legume trees. In 1990, there was only one bacterial species that was known to nodulate common bean worldwide (Rhizobium leguminosarum sv. phaseoli), one species that nodulated faba bean (Rhizobium leguminosarum sv. viciae), and two species that nodulated soybean (Bradyrhizobium japonicum and Rhizobium fredii). Today, nearly 14, 11, 6, 5, 5, 4, 3 and 2 species have been defined that are capable of nodulating common bean, soybean, cowpea, chickpea, peanut, lentils, faba bean and pea, respectively. The recent use of whole genome based taxonomy (genomotaxonomy) will surely change how we define this important group of bacteria. The identification of several rhizobial species that are able to nodulate and fix nitrogen with edible legumes may enhance the production of these crops and can compensate for worldwide deficiencies in human nutritional needs in the future.     

Evolution of <Emphasis Type="Italic">fixNOQP</Emphasis> genes encoding cytochrome oxidase with high affinity to oxygen in rhizobia and related bacteria

Many bacteria belonging to the order Rhizobiales have fixNOQP genes which encode cytochrome oxidase with high affinity to oxygen required for oxidative phosphorylation in microaerophilic conditions. There is one copy of the identified fixNOQP operon in ancestral forms of rhizobia (Bradyrhizobium), as well as in their putative evolutionary predecessors (bacteria related to Rhodopseudomonas). At the same time, forms deeply specialized in symbiosis (Rhizobium leguminosarum, Sinorhizobium meliloti) have multiple (2–3) copies, some of them have a high similarity (>90%) to fixNOQP genes of Bradyrhizobium and Rhodopseudomonas, and others have only 30–50% similarity. Two divergent copies fixNOQP are detected in Tardiphaga, which is a representative of the Bradyrhizobiaceae family, lacking the ability to fix N₂ (lack of nif genes encoding the synthesis of nitrogenase) and to induce the formation of nodules on legumes roots (lack of nod genes encoding the synthesis of signal Nod factors activating symbiosis development). The presence of Tardiphaga in nodule bacterial communities from a range of legumes, including Vavilovia formosa (relic representative of the tribe Fabeae, for which R. leguminosarum bv. viciae is the main microsymbiont), suggests that the ancestral gene duplication and subsequent divergence of fixNOQP operon in bacteria related to Tardiphaga opened the possibility of wide dissemination of functionally different copies of this cluster among symbiotically active forms of Rhizobiales. It is possible that the acquisition of fixNOQP genes determines adaptation of bacteria to microaerophilic niches not only in plants nodules but also in their environment (the rhizosphere, rhizoplane, internal portions of soil aggregates).     

Nodulation ability in different genotypes of <Emphasis Type="Italic">Phaseolus lunatus</Emphasis> by rhizobia from California agricultural soils

Phaseolus lunatus is the second economically most important species of the genus Phaseolus. It carries out N fixation through symbiosis with rhizobia. However, it is unclear whether P. lunatus can nodulate with native rhizobia from soils where this legume is not native or was not cultivated previously. Thus, this study assessed the ability of 14 geographically distant lima bean genotypes to nodulate with rhizobia from three California agricultural soils: without a history of legumes or P. lunatus cultivation, with a history of legumes as a cover crop, and with a history of P. lunatus cultivation. Nodulation only occurred on genotypes grown in the soil with a history of P. lunatus planting. The analysis of variance of nodulation traits showed that the genotype effect was highly significant in all the traits measured. Shoot biomass had a higher correlation with nodule size and nodule weight than with nodule number. In addition, shoot biomass and leaf N content were positively correlated with nodule coloration and with nodule position close to the main root of the plant. This study suggests that agricultural soils from California do not appear to have native rhizobia able to nodulate P. lunatus, which suggests the need to inoculate, at least initially, the seeds at planting in order to establish the population of rhizobia. Also, geographically distant lima bean genotypes have different responses to nodulating bacteria and it suggests that future studies to test these genotypes across different environments should be pursued.     

Forms of natural selection controlling the genomic evolution in nodule bacteria

The role of different forms of natural selection in the evolution of genomes in root nodule bacteria (rhizobia) is analyzed for the first time. In these nitrogen-fixing symbionts of leguminous plants, two types of genome organization are revealed: (i) unitary type, where over 95% of genetic information is encoded by chromosomes (5.3–5.5 Mb in Azorhizobium, 7.0–7.8 Mb in Mesorhizobium, 7.3–10.1 Mb in Bradyrhizobium); (ii) multipartite type, where up to 50% of genetic information is allocated to plasmids or chromids which may exceed 2 Mb in size and usually control the symbiotic properties (pSyms) in fast-growing rhizobia (Rhizobium, Sinorhizobium, Neorhizobium). Emergence of fast-growing species with narrow host ranges are correlated to the extension of extrachromosomal parts of genomes, including the increase in pSyms sizes (in Sinorhizobium). An important role in this evolution is implemented by diversifying selection since the genomic diversity evolved in rhizobia owing to symbiotic interactions with highly divergent legumes. However, analysis of polymorphism in nod genes (encoding synthesis of lipo-chitooligosaccharide signaling Nod factors) suggests that the impacts of diversifying selection are restricted to the bacterial divergence for host specificity and do not influence the overall genome organization. Since the extension of rhizobia genome diversity results from the horizontal sym gene transfer occurring with low frequencies, we suggest that this extension is due to the frequency-dependent selection anchoring the rare genotypes in bacterial populations. It is implemented during the rhizobia competition for nodulation encoded by the functionally diverse cmp genes. Their location in different parts of bacterial genomes may be considered as an important factor of their adaptive diversification implemented in the host-associated microbial communities.     

Plasmids impact on rhizobia-legumes symbiosis in diverse environments

Rhizobia are a well-known group of soil bacteria that establish symbiotic relationship with leguminous plants, fix atmospheric nitrogen, and improve soil fertility. To fulfill multiple duties in soil, rhizobia are elaborated with a large and complex multipartite genome composed of several replicons. The genetic material is divided among various replicons, in a way to cope with, and satisfy the diverse functions of rhizobia. In addition to the main chromosome, which is carrying the essential (core) genes required for sustaining cell life, the rhizobia genomes contain several extra-chromosomal plasmids, carrying the nonessential (accessory) genes. Occasionally, some mega-plasmids, denoted as secondary chromosomes or chromids, carry some essential (core) genes. Furthermore, specific accessory gene sequences (the symbiotic chromosomal islands) are incorporated in the main chromosome of some rhizobia species in Bradyrhizobium and Mesorhizobium genera. Plasmids in rhizobia are of variable sizes. All of the plasmids in a Rhizobium cell constitute about 30–50% of the genome. Rhizobia plasmids have specific characters such as miscellaneous genes, independent replication system, self-transmissibility, and instability. The plasmids regulate several cellular metabolic functions and enable the host rhizobia to survive in diverse habitats and even under stress conditions. Symbiotic plasmids in rhizobia are receiving increased attention because of their significance in the symbiotic nitrogen fixation process. They carry the symbiotic (nod, nif and fix) genes, and some non-symbiotic genes. Symbiotic plasmids are conjugally-transferred by the aid of the non-symbiotic, self-transmissible plasmids, and hence, brings about major changes in the symbiotic interactions and host specificity of rhizobia. Besides, the rhizobia cells harbor one or more accessory, non-symbiotic plasmids, carrying genes regulating various metabolic functions, rhizosphere colonization, and nodulation competitiveness. The entire rhizobia-plasmid pool interacting in harmony and provides rhizobia with substantial abilities to fulfill their complex symbiotic and non-symbiotic functions in variable environments. The above concepts are extensively reviewed and fairly discussed.     

Functional analysis of the two cyclophilin isoforms of <Emphasis Type="Italic">Sinorhizobium meliloti</Emphasis>

The nitrogen fixing Sinorhizobium meliloti possesses two genes, ppiA and ppiB, encoding two cyclophilin isoforms which belong to the superfamily of peptidyl prolyl cis/trans isomerases (PPIase, EC: 5.2.1.8). Here, we functionally characterize the two proteins and we demonstrate that both recombinant cyclophilins are able to isomerise the Suc-AAPF-pNA synthetic peptide but neither of them displays chaperone function in the citrate synthase thermal aggregation assay. Furthermore, we observe that the expression of both enzymes increases the viability of E. coli BL21 in the presence of abiotic stress conditions such as increased heat and salt concentration. Our results support and strengthen previous high-throughput studies implicating S. meliloti cyclophilins in various stress conditions.     

Improvement of cowpea productivity by rhizobial and mycorrhizal inoculation in Burkina Faso

Cowpea is one of the most important food legume crops in Burkina Faso. It is able to associate with arbuscular mycorrhizal fungi (AMF) and rhizobia. This dual symbiosis improves nitrogen and phosphorus nutrient uptake in cowpea. As the application of exotic inoculants frequently lacks positive responses in field experiments, this study set out to select well-adapted native symbiotic rhizobial and AMF strains. Soil samples were collected from six study sites in three different climatic zones of Burkina Faso to investigate their native symbiotic strains. Soil-extraction of native spores led to the identification of four AMF genera (Scutellospora, Gigaspora, Glomus and Entrophospora) by morpho-anatomical characterization. The two most effective cowpea fungal strains were selected after spore isolation from field-collected soils, multiplication on maize roots and inoculation on cowpea seedlings in a greenhouse experiment. Cowpea-nodulating rhizobial strains were trapped in the greenhouse by planting cowpea seeds in collected soil samples and the strains were characterized using molecular methods. This characterization led to the rhizobial isolates being classified in four clusters on the phylogenetic tree (using the Maximum-Likelihood Phylogenies method). All strains belonged to the Bradyrhizobium genus and most of them were included in the B. japonicum branch. Some groups were clearly distinct species already identified and may be new species. The two most effective strains for cowpea yield improvement in the field were selected after cowpea inoculation in a greenhouse experiment. The inoculation design in the field experiment consisted of four single inoculation treatments, either rhizobial or mycorrhizal, along with four dual inoculations, one treatment with chemical fertilizers, and one uninoculated control. The results showed that cowpea productivity was significatively improved by dual inoculation with native rhizobial and mycorrhizal strains, reaching the same level as the application of commonly used chemical fertilizers [Nitrogen, Phosphorus and Potassium fertilizers (NPK)]. In addition, dual inoculation resulted in the highest iron content in cowpea leaves.     

Crop rotation associating a legume and the nickel hyperaccumulator <Emphasis Type="Italic">Alyssum murale</Emphasis> improves the structure and biofunctioning of an ultramafic soil

Nickel (Ni) agromining aims to phytoextract heavy metals using hyperaccumulators whilst at the same time rehabilitating ultramafic soils. After removing the bioavailable metal, ultramafic soils are improved in terms of their agronomic properties with the aim of future agricultural uses. The low fertility of ultramafic soils can be compensated by integrating legumes already used in traditional agro-systems because of their importance in soil nitrogen enrichment. However, few studies have evaluated the potential profits of legumes on Ni agromining and their potential benefits on soil biological fertility. Here, we characterized the effect of a crop rotation with two plants, a legume (Vicia sativa) and a hyperaccumulator (Alyssum murale), on the phytoextraction efficiency and on soil structure and biofunctioning. A pot experiment was set up in controlled conditions to grow A. murale and four treatments were tested: rotation with V. sativa (Ro), fertilized mono-culture (FMo), non-fertilized mono-culture (NFMo) and bare soil without plants (BS). No significant difference was found between the Ro and NFMo treatments for the dry biomass yield. However, the Ro treatment showed the highest Ni concentrations ([Ni]) in A. murale shoots compared to FMo and NFMo treatments. The Ro treatment plants had more than twice as many leaves [Ni] compared to FMo. Soil physico-chemical analyses showed that the Ro treatment was better structured and showed the highest presence of bacterial micro-aggregates, as well as less non-aggregated particles. Legumes integration in Ni-agromining systems could be a pioneering strategy to reduce chemical inputs and to improve soil biofunctioning and thus fertility.     

Nitrogen deposition decreases the benefits of symbiosis in a native legume

Aims

Anthropogenic nitrogen deposition can provide legumes with a cheap source of nitrogen relative to symbiotic nitrogen fixation, leading to the potential breakdown of this critical symbiosis. Here, the effects of nitrogen deposition were tested on a native symbiosis between legumes and rhizobia.

Methods

Deposition rates, soil nitrogen concentration, and plant nitrogen isotopic composition were quantified along a predicted deposition gradient in California. Acmispon strigosus seedlings were exposed to fertilization spanning nitrogen concentrations observed in the plant’s California range. Both wild and experimental plants from pristine and nitrogen polluted sites were tested using rhizobial strains that varied in nitrogen fixation.

Results

Deposition intensity was tightly correlated with nitrogen concentration in soils. The growth benefits of rhizobial nodulation were dramatically reduced by even modest levels of mineral nitrogen, and all Acmispon lines failed to form root nodules at high nitrogen concentrations.

Conclusions

Our dataset suggests that anthropogenic deposition has greatly increased soil nitrogen concentrations in Southern California leading to significantly reduced benefits of rhizobial symbiosis. If nitrogen deposition increases continue, plant host mortality and a total collapse of the symbiosis could result.

     

The effects of gibberellins and mepiquat chloride on nitrogenase activity in <Emphasis Type="Italic">Bradyrhizobium japonicum</Emphasis>

Application of plant growth regulators (PGRs) to soybean plants is known to induce changes in nitrogenase activity in root nodules, and this led us to hypothesize that PGRs would affect nitrogenase activity in free-living rhizobia cultures. Little is known about the molecular basis of the effects of PGRs on nitrogenase activity in free-living rhizobia cultures. Therefore, a comparative study was conducted on the effects of gibberellins (GA₃) and mepiquat chloride (PIX), which regulate plant growth, on the nitrogenase activity of the nitrogen-fixing bacterium Bradyrhizobium japonicum. Fix and nif gene regulation and protein expression in free-living cultures of B. japonicum were investigated using real-time PCR and two-dimensional electrophoresis after treatment with GA₃ or PIX. GA₃ treatment decreased nitrogenase activity and the relative expression of nifA, nifH, and fixA genes, but these effects were reversed by PIX treatment. As expected, several proteins involved in nitrogenase synthesis were down-regulated in the GA₃-treated group. Conversely, several proteins involved in nitrogenase synthesis were up-regulated in the PIX-treated group, including bifunctional ornithine acetyltransferase/N-acetylglutamate synthase, transaldolase, ubiquinol-cytochrome C reductase iron-sulfur subunit, electron transfer flavoprotein subunit beta, and acyl-CoA dehydrogenase. Two-pot experiments were conducted to evaluate the effects of GA₃ and PIX on nodulation and nitrogenase activity in Rhizobium-treated legumes. Interestingly, GA₃ treatment increased nodulation and depressed nitrogenase activity, but PIX treatment decreased nodulation and enhanced nitrogenase activity. Our data show that the nif and fix genes, as well as several proteins involved in nitrogenase synthesis, are up-regulated by PIX and down-regulated by GA₃, respectively, in B. japonicum.     

Lima bean nodulates efficiently with <Emphasis Type="Italic">Bradyrhizobium</Emphasis> strains isolated from diverse legume species

Lima bean (Phaseolus lunatus L.) is an important legume species that establishes symbiosis with rhizobia, mainly of the Bradyrhizobium genus. The aim of this study was to evaluate the efficiency of rhizobia of the genus Bradyrhizobium in symbiosis with lima bean, in both Leonard jars and in pots with a Latossolo Amarelo distrófico (Oxisol). In the experiment in Leonard jars, 17 strains isolated from nodules of the three legume subfamilies, Papilionoideae (Vigna unguiculata, Pterocarpus sp., Macroptilium atropurpureum, Swartzia sp., and Glycine max), Mimosoideae (Inga sp.), and Caesalpinioideae (Campsiandra surinamensis) and two uninoculated controls, one with a low concentration (5.25 mg L^?1) and another with a high concentration (52.5 mg L^?1) of mineral nitrogen (N) were evaluated. The six strains that exhibited the highest efficiency in Leonard jars, isolated from nodules of Vigna unguiculata (UFLA 03–144, UFLA 03–84, and UFLA 03–150), Campsiandra surinamensis (INPA 104A), Inga sp. (INPA 54B), and Swartzia sp. (INPA 86A), were compared to two uninoculated controls, one without and another with 300 mg N dm^?3 (NH₄NO₃) applied to pots with samples of an Oxisol in the presence and absence of liming. In this experiment, liming did not affect nodulation and plant growth; the INPA 54B and INPA 86A strains stood out in terms of shoot dry matter production and provided increases of approximately 48% in shoot N accumulation compared to the native rhizobia populations. Our study is the first to indicate Bradyrhizobium strains isolated from the three legume subfamilies are able to promote lima bean growth via biological nitrogen fixation in soil conditions.