The common bean is of great importance for human nutrition worldwide as its grain is a cheap source of protein, but it has a high-cost production system due to the use of synthetic inputs such as nitrogen fertilizers and pest and disease control products. An alternative to promote the sustainability of the production system and increase crop productivity is the use of co-inoculation of plant growth-promoting rhizobacteria (PGPR) to reduce and/or replace the use of nitrogen fertilizers and pest and disease control products. Since 2010, Brazil has stood out on the international stage with research focused on the application of rhizobia combined with PGPR in bean crops. Currently, the use of microorganisms that effectively promote plant growth through different mechanisms of action is already a consolidated reality in Brazil, with experimental results proven by years of research. Despite these advances, there are still challenges to overcome to maximize the potential of co-inoculation in bean crops. These include the need to develop more efficient formulations containing elite microbial strains and optimized microbial consortia, as well as the search for inoculation methods that optimize root colonization by the bacteria and maximize their beneficial effects on the plant. In this review, we discuss the history of common-bean production and the types and doses of inoculants marketed, as well as the challenges for the efficiency of co-inoculation and for the development of inoculants of rhizobial consortia with PGPRs for the common-bean cultivation.

Entomopathogenic fungi are increasingly recognized as multifunctional bioinputs, but robust methodological approaches are needed to evaluate their compatibility with established microbial inoculants and their effects on plant performance. Our study proposes an integrative framework to assess Metarhizium-Bradyrhizobium interactions in soybean (Glycine max), combining laboratory compatibility screening with greenhouse assessment. First, Metarhizium anisopliae LCM S04 and Metarhizium brunneum LCM S11 were tested against Bradyrhizobium diazoefficiens BR 85 (=SEMIA 5080) and Bradyrhizobium japonicum BR 86 (=SEMIA 5079) using dual-culture assays in a medium that supported the growth of both fungal and bacterial partners, allowing direct evaluation of intermicrobial compatibility. Subsequently, the microorganisms were evaluated in soybean under greenhouse conditions through direct seed inoculation. Each seed received bacterial culture, fungal suspension, or both (including absolute and nitrogen-fertilized controls). Plants were maintained under controlled conditions for 47 days. Plant height, leaf production, branching, nodulation, biomass, and root morphology were assessed. No inhibition zones were observed in vitro, and co-inoculation did not impair nodulation. The combination BR 86 + LCM S04 improved fine root number in soybean. This framework provides a reproducible approach for evaluating microbial compatibility and functional bioinput benefits and can be adapted to crops other than soybean.

Species of the genus Stylosanthes show great potential for use in mixed pastures and as green manure due to their symbiotic potential in symbiosis with diazotrophic bacteria, particularly Bradyrhizobium. These herbaceous legumes can improve pasture quality and contribute to sustainability. This study evaluated performance of Stylosanthes cv. Campo Grande when inoculated with Bradyrhizobium strains isolated from Oxisols in Roraima, Brazil, versus standard strains. A greenhouse experiment tested 18 new Bradyrhizobium strains isolated from soil alongside two recommended strains, including uninoculated and nitrogen-fertilized controls. Also, two field experiments were conducted using the best-performing greenhouse strain, the recommended strains, and absolute control and nitrogen fertilization (30 kg ha- 1 of N). Variables measured included nodule number and dry nodule mass, root and shoot biomass, and total nitrogen in shoot biomass. The proportion of nitrogen derived from biological nitrogen fixation (percentage of N derived from the atmosphere) was estimated by the 1 5N natural abundance method, with Urochloa brizantha as the reference plant. In the greenhouse, strains ERR 917, ERR 922, ERR 942, ERR 1110, and ERR 1173 present great nodulation and dry matter production, identifying ERR 917 as optimal for field testing. Inoculation significantly increased both shoot and root biomass of cv. Campo Grande. Standard strain BR 446T and ERR 917 enabled plants to obtain 38%-44% of their total nitrogen from biological nitrogen fixation, with inoculated plants accumulating about twice as much nitrogen and biomass versus the uninoculated control (averaging 1900 and 3900 kg ha- 1 of dry biomass for control and inoculated treatments, respectively).

Lime and gypsum are widely used to correct soil acidity and improve grain yields in Brazilian agricultural systems. However, limited information is available on their effectiveness and application practices in degraded sandy soils typical of older agricultural frontiers, such as those in Rio de Janeiro State. This study evaluated the effects of surface application versus the incorporation of lime and gypsum into the soil through tillage operations on soil chemical properties, nodulation, and grain yield of soybean cultivars grown in low-fertility Fluvisols. The experiment was conducted during the 2021/2022 growing season in Campos dos Goytacazes, Rio de Janeiro, using a strip-plot design with four soybean cultivars and two soil amendment placement strategies: surface application without tillage and incorporation through tillage. Soil chemical attributes, nodulation, nutrient uptake, and yield components were assessed. Incorporated application significantly increased soil pH, reduced Al3+ toxicity, and enhanced Ca2+, Mg2+, P, and K+ availability compared to surface application. Nodulation responses varied among cultivars, with incorporated treatments promoting up to 40% greater nodule biomass. Although primary root length was not affected, incorporation stimulated secondary root development and nutrient uptake, leading to approximately 50% higher pod number and grain yield. Overall, incorporating lime and gypsum through soil tillage was more effective than surface application in improving soil fertility, enhancing nodulation, and increasing soybean productivity under the conditions evaluated in this study. These findings suggest that lime and gypsum incorporation can represent an important management strategy for improving soybean production in degraded sandy soils.

Abstract The objective of this work was to evaluate the productive performance of four chickpea (Cicer arietinum) cultivars and their seed inoculation with Mesorhizobium strains, in low-altitude areas in the state of Rio de Janeiro, Brazil. The chickpea cultivars used were BRS Aleppo, BRS Cícero, BRS Cristalino, and BRS Toro. Five experiments were carried out in a greenhouse and in the field. Chickpea inoculation experiments with Mesorhizobium were performed in sterilized substrate, soil, in a greenhouse and in the field, in the municipality of Seropédica. Cultivar production was evaluated in the municipalities of Seropédica and Campos dos Goytacazes. Yields above 1,400 kg ha-1 were observed for BRS Aleppo, BRS Cristalino, and BRS Toro cultivars, in Seropédica, being lower in Campos dos Goytacazes. However, the presence of caterpillar infestations (Chloridea virescens and Helicoverpa armigera) reduced grain yield and quality in Seropédica. Despite their high-nodulation rates in sterilized conditions, the tested Mesorhizobium strains failed to establish an effective nodulation under field conditions. Cultivars BRS Aleppo, BRS Cristalino, and BRS Toro show potential for commercial cultivation in low-altitude areas in Rio de Janeiro. However, the available Mesorhizobium strains do not establish nodulation under field conditions.

Bradyrhizobium is a genetically diverse genus that forms symbioses with numerous legumes, including major crops such as cowpea (Vigna unguiculata) and soybean (Glycine max). Understanding the genetic and symbiotic diversity of native strains is essential for improving inoculant technologies and enhancing biological nitrogen fixation in tropical agricultural systems. This study investigated Bradyrhizobium strains associated with these two legumes grown in adjacent tropical soils in Brazil to elucidate their genetic relationships, taxonomic placement, and host compatibility. A total of 34 Bradyrhizobium strains isolated from cowpea and soybean nodules were characterized using multilocus phylogenetic analyses (16S rRNA, gyrB, recA, and nodC). Selected strains underwent whole-genome sequencing for comparative analyses based on average nucleotide identity (ANI) and digital DNA–DNA hybridization (dDDH). Cross-inoculation assays were performed to evaluate nodulation capacity and symbiotic efficiency on both hosts. The strains displayed high genetic diversity, forming multiple phylogenetic clusters. Most grouped within the B. elkanii superclade, whereas several occupied divergent lineages, some potentially representing new taxa. Genome-based analyses supported these findings, showing intracluster ANI values above 95–96% and intercluster values below 94%. A distinct group of cowpea-derived strains exhibited high symbiotic efficiency but low genomic similarity to known type strains, suggesting the presence of a novel species with potential use in inoculants. In contrast, some soybean-derived strains were genetically identical to commercial inoculants, indicating persistence or re-isolation from previously inoculated soils. Notably, strain BR 13971, isolated from soybean, nodulated both hosts efficiently, demonstrating a broad host range and suggesting a unique symbiovar. Cross-inoculation assays showed that soybean-derived strains effectively nodulated cowpea, whereas cowpea-derived strains did not nodulate soybean, indicating asymmetrical host compatibility. Particularly for cowpea, strains BR 10926 and BR 10750 demonstrated higher symbiotic efficiency than the strains currently recommended for this crop. Overall, these findings enhance the understanding of Bradyrhizobium diversity in tropical soils and highlight promising native strains for future inoculant development.

Nitrous oxide is a potent greenhouse gas, with N fertilizers being one of its major sources. Plant growth-promoting bacteria have been used to mitigate N2O emissions by improving N use efficiency in plants. In addition, some of these microorganisms are capable of reducing N2O to N2, a process that could be further explored as a complementary mitigation strategy. This study aimed to test whether Azospirillum brasilense strains Ab-V5 and Ab-V6, and strain Wa3, as well as Nitrospirillum viridazoti strain BR 11145, already used in commercial inoculants for N-fertilized crops and known to carry the nosZ gene encoding nitrous oxide reductase, could act as biological sinks for this gas. In the pot experiment, soils fertilized with N and inoculated with N. viridazoti exhibited consistently lower N2O emissions, but not when A. brasilense strains were used. The mitigation effect was observed both in bare soil (72% emission reduction) and in the presence of millet plants (60% emission reduction), confirming the ability of N. viridazoti strain BR 11145 to consume N2O. In a field experiment conducted with maize, inoculation with N. viridazoti again reduced N2O fluxes during the first two weeks after fertilization compared with the urea-only treatment. However, no significant differences were detected comparing emission factors, whose calculation requires consideration of the entire monitoring period, thereby adding more variability. While N2O mitigation was observed, no significant effects of inoculation or N fertilization were found on maize growth or yield. Nonetheless, the consistent reduction in N2O emissions achieved with N. viridazoti strain BR 11145 suggests that inoculants with this bacterium represent a promising biological sink for N2O, offering a novel nature-based solution to enhance the sustainability of tropical crop management.

Brazil maintains a leading position in agricultural exports and stands as the world's foremost producer and user of bioinputs in agriculture. These bioinputs generate annual savings of billions of dollars that would otherwise be allocated to chemical fertilizers and pesticides. The nation's regulatory framework enables bioinput agriculture and serves as a model for countries transitioning toward regenerative agriculture. Brazilian legislation categorizes bioinputs into: 1) biofertilizers (extracts); 2) biostimulants (plant growth-promoting and biocontrol agents); and 3) inoculants (active ingredient comprises one or more living microorganisms). The inoculation of soybeans with Bradyrhizobium strains provides approximately 90% of the nitrogen accumulated by this crop. Brazil has registered over six hundred inoculants, with at least 60% specifically designated for soybean cultivation. The annual sales of inoculants in Brazil reach approximately 120 million doses. Although beans (Phaseolus vulgaris and Vigna unguiculata) represent an essential food crop in Brazil's staple diet and benefit from inoculation, inoculant supply remains insufficient. Regarding biocontrol, soy, corn, sugarcane, and coffee rank among the most protected crops, employing biocontrol agents against bacteria, fungi, nematodes, and insects. Bacillus, Pseudomonas, Streptomyces, Rhizobium, Azotobacter, and Paenibacillus strains were predominantly cited in the 5,000+ bioproduct patents filed between 2022 and 2024. Among fungal genera, Trichoderma, and Penicillium received the most citations. EMBRAPA's biobanks maintain over 10,000 strains of bacteria, fungi, and viruses for biocontrol, and 14,000 strains of nutrient-fixing and plant-growth promoters. Production challenges include quality control, particularly as on-farm production of inoculants becomes prevalent on larger farms, alongside product availability and supply limitations. Brazilian farmers maintain global competitiveness partly through reduced chemical fertilizer and pesticide costs enabled by bioinput usage. As components of regenerative agriculture, bioinputs enhance soil quality, decrease carbon footprints, and support Sustainable Development Goals. Brazil's leadership in microbial bioinput utilization stems from its extensive agricultural sector, rich microbial biodiversity, and progressive regulatory framework.

Cowpea production potential often falls short, despite the crop's efficiency as a legume. This underperformance is primarily attributed to widespread deficiencies of phosphorus (P), cobalt (Co), and molybdenum (Mo) in Brazilian soils, especially in the Cerrado region. This study aimed to determine the optimal doses of P, Co, and Mo to enhance cowpea nodulation, biological nitrogen fixation, and overall plant growth. Two greenhouse exper-iments were conducted using a randomized complete block design in a 2 × 5 factorial scheme (two soils and five doses of each nutrient, inoculated with strain BR 3262), with four replicates. In Trial I, P₂O₅ doses of 0, 100, 200, 300, and 400 mg pot⁻¹ were tested, while in Trial II, Co:Mo ratios (w:w) were evaluated: 0:0, 2:8, 3:16, 4:32, and 6:64 (mg pot⁻¹). The variables analyzed included phytotechnical parameters. The application of 200 mg pot⁻¹ of P₂O₅ (200 kg ha⁻¹) resulted in the highest nodulation, nitrogen accumulation, and increased cowpea biomass. Furthermore, medium-high micronutrient levels (Mo at 32 g ha-1 and Co at 4 g ha-1) provided superior nodulation, biomass, and nitrogen accumu-lation. Adequate P and micronutrient fertilization is essential for plant development, rein-forcing its pivotal role in maximizing cowpea productivity under Cerrado soil conditions.

In this study, we characterized 86 plant growth-promoting bacterial strains belonging to the genus Nitrospirillum, isolated from diverse host plants and geographic regions. We investigated their evolutionary relationships through phylogenetic analyses of the 16S rRNA and recA genes, complemented by phylogenomic approaches incorporating genomic similarity metrics, such as ANI and dDDH. The classification of type strains was further supported by in silico analyses of chemotaxonomic markers, particularly genes involved in fatty acid biosynthesis and elongation, phospholipid and quinone production, and nitrogen fixation (nifHDK operon). Phenotypic and chemotaxonomic characterization was performed using matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry, fatty acid methyl ester (FAME) profiling, and physiological assays. These included evaluations of nitrogen fixation capacity, antibiotic resistance, carbon source utilization, and enzymatic activities. This integrative approach provided detailed insight into the characteristics and diversity of the studied strains. Phylogenetic and genomic analyses revealed six novel taxa within the genus Nitrospirillum, in addition to the previously described species N. amazonense, N. iridis, and N. viridazoti. The distinctiveness of these new lineages was supported by both genomic metrics and phenotypic traits. All novel strains also exhibited activity of nitrogenase enzyme, confirming their nitrogen-fixing ability under in vitro conditions. Based on these findings, we propose the formal description of six novel species: Nitrospirillum bahiense sp. nov. (= BR 11865T, = UCCCB 233T), Nitrospirillum guanabarense sp. nov. (= BR 11163T, = UCCCB 228T), Nitrospirillum guaranorum sp. nov. (= BR 11164T, = UCCCB 229T), Nitrospirillum karajorum sp. nov. (= BR 11752T, = UCCCB 231T), Nitrospirillum goiasense sp. nov. (= BR 11828T, = UCCCB 232T), and Nitrospirillum pindoramense sp. nov. (= BR 11622T, = UCCCB 230T).

The genus Bradyrhizobium comprises phylogenetically diverse bacteria that establish symbioses with numerous legumes, including economically important crops such as cowpea (Vigna unguiculata) and soybean (Glycine max). This study characterized 34 Bradyrhizobium strains isolated from nodules of cowpea and soybean cultivated in adjacent tropical soils in Brazil to elucidate the relationships between the symbionts and their hosts. Phylogenetic analyses of 16S rRNA, gyrB, recA, and nodC genes, combined with genome sequencing and comparative analyses (ANI and dDDH), revealed high genetic diversity and distinct taxonomic affiliations. Most strains clustered within the B. elkanii superclade, although several occupied divergent lineages, some potentially representing new taxons. Genome-based analyses confirmed these patterns, showing ANI values above 95–96% within groups and below 94% across distant strains. One group of cowpea-derived strains exhibited high symbiotic efficiency and low similarity to all known type strains, suggesting the presence of a novel species with potential for the crop. Conversely, some soybean-derived strains were genetically identical to commercial inoculants, indicating persistence or re-isolation from inoculated soils. Remarkably, strain BR 13971, isolated from soybean, nodulated both hosts efficiently, suggesting a distinct symbiovar with broad host compatibility. Cross-inoculation assays demonstrated that soybean-derived strains nodulated cowpea, but the reverse was not observed. These findings expand the understanding of Bradyrhizobium diversity in tropical soils and highlight the potential of native strains for inoculant development.

ABSTRACT Dark septate fungi (DSF) are endophytic microorganisms characterized by the formation of detached and melanized hyphae that confer adaptive advantages, including benefits in plant development. This study aimed to evaluate the nutrient accumulations, growth, and production of cotton plants inoculated with different DSF accessions. The experiment was carried out in a greenhouse in a completely randomized design with eight replicates. Cotton plants were grown in plastic bags containing commercial substrate and watered daily. Data were collected over the cycle of plants. Promising results were found in all inoculants; however, those obtained from ERR 26 and ERR 42 accessions provided broad benefits for cotton plants, improving the accumulation of N and P and improving plant growth and production. It is worth highlighting the significant contribution of the ERR 42 inoculant to the accumulation of calcium, which brings additional benefits to the formation and resistance of the fruit cell wall. This information is unprecedented and elevates the usage prospects of this inoculant for the biofertilization of cotton plants. However, further studies should be deepened to attest to its biostimulant advantages in cotton management.

ABSTRACT Neotropical Beta-rhizobia have a particular affinity to the large legume ( Fabaceae ) genus Mimosa and some of its relatives in the tribe Mimosae of the Caesalpinioideae subfamily. However, little is still known about the ecology of this interaction, especially the relationship between the rhizobia of “widespread” pan-tropical Mimosa species like M. pudica and the rhizobia that nodulate endemic Mimosa species that are very restricted in their habitats. The objective of this study was to examine the microsymbionts of Mimosa spp. and some other mimosoids in climates ranging from tropical to subtropical, humid to semi-arid, with varied soil characteristics and altitudes, with the aim of testing the hypothesis that widespread species have more cosmopolitan symbiont preferences than endemic ones. Nodules were sampled from more than 40 Mimosa spp. and related taxa in eleven Brazilian states, many endemics or biome-restricted, but particular attention was paid to sample nodules from the widespread species M. pudica at all locations. The Mimosa symbionts comprised 19 potential 16S rRNA and recA groups at the species level, with 16 belonging to the genus Paraburkholderia , including six lineages that may represent new species. The remaining genotypes consisted of 14 strains in two lineages of Cupriavidus that were mainly isolated from M. pudica growing at low altitudes, plus a single lineage of Rhizobium also from M. pudica . It is concluded that a high diversity of Paraburkholderia strains dominate as symbionts of Mimosa in the acidic soils of its main center of radiation in Central Brazil but that Cupriavidus and Rhizobium comprise a significant minority of symbionts of widespread Mimosa spp., especially M. pudica , in lowland or disturbed areas with less acidic soils. Mimosa symbiont diversity is thus driven either by edapho-climatic characteristics for widespread species and/or by co-evolution of the symbiotic partners for endemic species.

Dark Septate Endophytic (DSE) fungi can benefit plants by optimizing nutrient uptake, biosynthesis of phytohormones-like compounds, and stress relief such as toxic metals. The objective was to characterize in vitro 57 strains of the DSE Periconia macrospinosa isolated from sugarcane roots, indicating the most promising in solubilizing phosphate sources, growing in different metal and vinasse contents, and producing molecules related to indoleacetic acid (IAA). The strains were from the Embrapa Agrobiology Fungi Collection. Over 35% of the strains solubilize calcium phosphate, highlighting A356 and A155. No strain solubilizes aluminum phosphate. Three strains did not grow in the presence of Cd (A333, A334, A163), but A226, A332, and A423 stand out showing high growth with Cd. All strains grew in the presence of Cu, Zn, and Vinasse. A163, A164, and A328 were even stimulated by Cu addition. A331 and A335 showed a marked growth decrease with Zn. All strains are highly adapted to grow in vinasse presence. A257 and A226 produced the highest amount of IAA. The most biotechnological potential strains are indicated by presenting high growth under Cd, Cu, Zn, and vinasse stress, associated with a high tolerance index to these pollutants, further calcium phosphate solubilization, or IAA production.

A group of Gram-negative plant-associated diazotrophic bacteria belonging to the genus Nitrospirillum was investigated, including both previously characterized and newly isolated strains from diverse regions and biomes, predominantly in Brazil. Phylogenetic analysis of 16S rRNA and recA genes revealed the formation of a distinct clade consisting of thirteen strains, separate from the formally recognized species N. amazonense (the closest species) and N. iridis. Comprehensive taxonomic analyses using the whole genomes of four strains (BR 11140T = AM 18T = Y-2T = DSM 2788T = ATCC 35120T, BR 11142T = AM 14T = Y-1T = DSM 2787T = ATCC 35119T, BR 11145 = CBAmC, and BR 12005) supported the division of these strains into two species: N. amazonense (BR 11142 T and BR 12005) and a newly proposed species (BR 11140 T and BR 11145), distinct from N. iridis. The phylogenomic analysis further confirmed the presence of the new Nitrospirillum species. Additionally, MALDI-TOF MS analysis of whole-cell mass spectra provided further evidence for the differentiation of the proposed Nitrospirillum species, separate from N. amazonense. Analysis of chemotaxonomy markers (i.e., genes involved in fatty acid synthesis, metabolism and elongation, phospholipid synthesis, and quinone synthesis) revealed that the new species highlights high similarity and evolutionary convergence with other Nitrospirillum species. This new species exhibited nitrogen fixation ability in vitro, it has similar NifHDK protein phylogeny position with the closest species, lacked denitrification capability, but possessed the nosZ gene, enabling N2O reduction, distinguishing it from the closest species. Despite being isolated from diverse geographic regions, soil types, and ecological niches, no significant phenotypic or physiological differences were observed between the proposed new species and N. amazonense. Based on these findings, a new species, Nitrospirillum viridazoti sp. nov., was classified, with the strain BR 11140T (DSM 2788T, ATCC 35120T) designated as the type strain.

The use of nitrogen-fixing bacteria in leguminous plants is a widespread approach, and the exploration of symbiotic bacteria that are efficient in biological nitrogen fixation (BNF) continues to be explored. The aim of the present study was to evaluate the effect of inoculation and coinoculation of mung bean plants with different combinations of Bradyrhizobium (B. elkanii BR 2003, B. pachyrhizi BR 3262, B. yuanmingense BR 3267, B. paxllaeri BR 10398 and B. icense BR 10399), Azospirillum baldaniorum (Sp245) and Bacillus pumilus (UFPEDA 472) on the contribution of biomass, the concentration of nitrogen (N) compounds and BNF. The experiment was carried out under greenhouse conditions with pots containing washed and autoclaved sand. Mung bean seeds were inoculated or coinoculated (double or triple) with Bradyrhizobium, A. baldaniorum and/or Bacillus pumilus, and one absolute control (not inoculated) was used. The experimental design was a completely randomized design with 21 treatments harvested in two different periods (flowering and pod maturation). Inoculation and coinoculation positively influenced the number of nodules, shoot dry weight, N accumulated, total N content and inorganic and organic compounds (free ammonia, nitrate, ureides and leghemoglobin), indicating that there was efficiency in BNF and synergistic interaction between the bacteria used and the mung bean plants. Inoculation with Bradyrhizobium species and the combination of these strains with A. baldaniorum and Bacillus pumilus positively influenced N fixation and metabolism in mung bean plants, especially when B. elkanii BR 2003 and B. pachyrhizi BR 3262 were used.

Neotropical Beta-rhizobia have a particular affinity to the large legume (Fabaceae) genus Mimosa and some of its relatives in the tribe Mimosae of the Caesalpinioideae subfamily. However, little is still known about the ecology of this interaction, especially the relationship between the rhizobia of “widespread” pan-tropical Mimosa species like M. pudica and the rhizobia that nodulate endemic Mimosa species that are very restricted in their habitats. The objective of this study was to examine the microsymbionts of Mimosa spp. and other mimosoids in climates ranging from tropical to subtropical, humid to semi-arid, with varied soil characteristics and altitudes, with the aim of testing the hypothesis that widespread species have more cosmopolitan symbiont preferences than endemic ones. Nodules were sampled from >30 Mimosa spp. and related taxa in 13 Brazilian states covering all five national regions; many of the species were endemics or biome-restricted, but particular attention was also paid to sample nodules from the widespread species M. pudica at all locations. The Mimosa symbionts comprised 21 potential 16S rRNA and recA groups at the species level, with 17 belonging to the genus Paraburkholderia, including four lineages that may represent new species. The remaining genotypes consisted of 14 strains in two lineages of Cupriavidus that were mainly isolated from M. pudica growing at low altitudes, and a single lineage of Rhizobium also from M. pudica. In addition, a strain of Trinickia symbiotica was isolated from M. misera. It is concluded that diverse genotypes of Paraburkholderia dominate as symbionts of Mimosa in the acidic soils of its main center of radiation in Central Brazil but that Cupriavidus and Rhizobium comprise a significant minority of symbionts of widespread Mimosa spp., especially M. pudica, in lowland or disturbed areas with less acidic soils. Mimosa symbiont selection is thus driven either by edapho-climatic characteristics for widespread species and/or by co-evolution of the symbiotic partners for endemic species.

The establishment of the rhizobium-legume symbiosis is generally based on plant perception of Nod factors (NFs) synthesized by the bacteria. However, some Bradyrhizobium strains can nodulate certain legume species, such as Aeschynomene spp. or Glycine max, independently of NFs, and via two different processes that are distinguished by the necessity or not of a type III secretion system (T3SS). ErnA is the first known type III effector (T3E) triggering nodulation in Aeschynomene indica. In this study, a collection of 196 sequenced Bradyrhizobium strains was tested on A. indica. Only strains belonging to the photosynthetic supergroup can develop a NF-T3SS-independent symbiosis, while the ability to use a T3SS-dependent process is found in multiple supergroups. Of these, 14 strains lacking ernA were tested by mutagenesis to identify new T3Es triggering nodulation. We discovered a novel T3E, Sup3, a putative SUMO-protease without similarity to ErnA. Its mutation in Bradyrhizobium strains NAS96.2 and WSM1744 abolishes nodulation and its introduction in an ernA mutant of strain ORS3257 restores nodulation. Moreover, ectopic expression of sup3 in A. indica roots led to the formation of spontaneous nodules. We also report three other new T3Es, Ubi1, Ubi2 and Ubi3, which each contribute to the nodulation capacity of strain LMTR13. These T3Es have no homology to known proteins but share with ErnA three motifs necessary for ErnA activity. Together, our results highlight an unsuspected distribution and diversity of T3Es within the Bradyrhizobium genus that may contribute to their symbiotic efficiency by participating in triggering legume nodulation.

As is customary in mycorrhizae, the interaction between plant and dark septate endophytic (DSE) fungi can result in physiological changes in the host plant, which are still poorly understood. This study aimed to evaluate the physiological changes in tomato plants colonized by DSE fungi. Four DSE isolates previously identified through ITS phylogeny were inoculated on tomato seeds and compared to non-inoculated plants (control). Kinetic parameters (Vmax and Km) were calculated measuring the nitrate content in the nutrient solution. The contents of NO3--N, NH4+-N, amino-N, soluble sugars in the root, petiole, stem and leaf, and the contents of macronutrients in the shoot were determined. The plants inoculated with A101 and A105 exhibited (i) significant increases in the soluble sugar contents; (ii) increases in the contents of P, K, Mg and S; and (iii) increased dry biomass compared to control. The A103 inoculation was antagonistic when compared to the other treatments, leading to a higher influx of NO3--N in the plants, resulting in a higher amino-N and the lower soluble sugar content in the shoot. The physiological parameters of tomato varied depending on the inoculation, and the changes ranged from positive to negative depending on each isolate involved in the interaction.

ABSTRACT This work aimed to evaluate the use of phosphorus (P) fertilizer, rhizobia inoculation, and seed enrichment with molybdenum (Mo) as tools for enhancing the cowpea yield in the Brazilian eastern Amazon. A set of field and greenhouse experiments were carried out in cerrado and upland forest environments. Four levels of P fertilization, eight rhizobia strains, and the application of mineral nitrogen (N) were evaluated for the cultivar BRS Tumucumaque (BR = Brasil e S = Sementes), and two levels of Mo seed enrichment were assessed for the cultivars BRS Tumucumaque and BRS Guariba. Grain yield increased linearly with the level of P applied. Inoculation with the rhizobia strain BR 3267 increased the shoot and nodule mass of the cultivar BRS Tumucumaque. There was no difference in grain yield and grain N accumulation between inoculation with seven rhizobia strains, N mineral fertilization and the control (without inoculation and N fertilization) in cerrado and upland forest environments. Cowpea grain yield and grain N accumulation were higher in the cerrado than in the upland forest environment and when the seeds were enriched with Mo. Phosphorus fertilization and seed enrichment with Mo are efficient tools to enhance cowpea yield in the Brazilian Amazon.