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In this study, the effectiveness of an inoculant containing a nitrogen (N)-fixing microorganism (Methylobacterium symbioticum) was evaluated on maize (Zea mays L.) grown both in the field (silage maize) and in pots over two years (2021 and 2022). The field trial included the following two treatments: with (Yes) and without (No) the inoculant. The pot experiment was designed as a factorial arrangement with two factors: the application of the inoculant (Yes and No) and N applied to the soil (0, 0.4, 0.8, and 1.6 g pot−1). In the field, total dry matter yield (DMY) did not differ significantly between treatments, although the average DMY was higher in the inoculant treatment. In pots, the total DMY varied significantly across all N rates but was only significantly affected by the inoculant application in 2022. N fixation estimates in the field were 58.8 and 14.5 kg ha−1 for 2021 and 2022, respectively, representing 23.7% and 9.1% of the N recovered in the aboveground plant parts. In pots, the estimated fixed N values were −49.2 and 199.2 mg pot−1 in 2021 and 2022, respectively, which corresponded to −5.2% and 18.5% of the N found in the aboveground plant parts. Considering the average values obtained across the four cultivation conditions, there was a positive outcome for the treated plants. However, these values cannot be considered significant when compared to nitrogen removal in maize crops. A commercial product should provide an unequivocal and quantitatively relevant contribution to plant nutrition, which did not appear to be the case. Thus, for this inoculant to provide reliable guarantees of positive outcomes for farmers and become a useful tool in promoting more sustainable agriculture, further studies appear necessary. These studies should aim to determine in which crops and under what cultivation conditions the application of the inoculant is truly effective in enhancing N fixation and improving crop productivity.

Most Earth system models (ESMs) do not explicitly represent the carbon (C) costs of plant nutrient acquisition, which leads to uncertainty in predictions of the current and future constraints to the land C sink. We integrate a plant productivity-optimizing nitrogen (N) and phosphorus (P) acquisition model (fixation & uptake of nutrients, FUN) into the energy exascale Earth system (E3SM) land model (ELM). Global plant N and P uptake are dynamically simulated by ELM-FUN based on the C costs of nutrient acquisition from mycorrhizae, direct root uptake, retranslocation from senescing leaves, and biological N fixation. We benchmarked ELM-FUN with three classes of products: ILAMB, a remotely sensed nutrient limitation product, and CMIP6 models; we found significant improvements in C cycle variables, although the lack of more observed nutrient data prevents a comprehensive level of benchmarking. Overall, we found N and P co-limitation for 80% of land area, with the remaining 20% being either predominantly N or P limited. Globally, the new model predicts that plants invested 4.1 Pg C yr-1 to acquire 841.8 Tg N yr-1 and 48.1 Tg P yr-1 (1994-2005), leading to significant downregulation of global net primary production (NPP). Global NPP is reduced by 20% with C costs of N and 50% with C costs of NP. Modeled and observed nutrient limitation agreement increases when N and P are considered together (r 2 from 0.73 to 0.83).

The adoption of sustainable technologies that optimize biological nitrogen fixation (BNF) by peanut crops is essential to improve plant growth and ensure high grain yield potential. A study was conducted to investigate the efficiency of inoculation with Bradyrhizobium spp. and foliar application of molybdenum (Mo) in improving the productive response of peanut crop (Arachis hypogaea L., cv. IAC Tatu ST) in moderately acidic sandy soil (pH 5.0) of the Cerrado region, Brazil during two growing seasons (2016/2017 and 2017/2018). Treatments consisted of uninoculated seeds (control) and inoculated with Bradyrhizobium spp. and foliar application of Mo at five rates (0, 25, 50, 100, and 150 g Mo ha−1). These treatments were arranged in a randomized block design in split plots with four replicates. Nodulation, shoot dry matter, leaf N concentration, and peanut grain yield were significantly high in plants inoculated with Bradyrhizobium spp. in both growing seasons. The grain yield increased by 13% and 12% with the inoculation of Bradyrhizobium spp. compared with non-inoculated plants in the 2016/2017 and 2017/2018 seasons, respectively. Plants fertilized with Mo showed higher root nodulation, leaf N concentration, and shoot dry matter production than control plants in both growing seasons. However, the crop grain yield was significantly high (24%) only in the 2016/2017 season in plants fertilized with Mo. The optimal rate of foliar Mo application in peanut plants cultivated in sandy soils varies from 80 to 100 g of Mo ha−1. Due to improved root nodulation, N uptake, plant growth, and higher crop grain yield, rhizobium inoculation and foliar application of Mo are sustainable agricultural practices that should be recommended for peanut growers. [ABSTRACT FROM AUTHOR]

Free‐living N2 fixation (FNF) is an important source of bioavailable nitrogen for forests. Though increasing plant species diversity (PSD) benefits soil nitrogen accumulation, how it impacts FNF rate has not been explored.Forty‐five plots covering a PSD gradient were selected in a subtropical karst forest, southwest China. FNF rates in leaf litter and soil were measured using acetylene reduction assay calibrated with a 15N2 fixation method. Diazotrophic community, litter and soil properties were determined as well.The nifH gene abundance was not significantly altered in litter but was significantly decreased in soil by higher PSD. Diazotrophic community's Shannon diversity was significantly increased by higher PSD in both litter and soil. Increasing PSD inhibited litter FNF rate via aggravating phosphorus limitation and decreasing the relative abundance of Betaproteobacteria and Deltaproteobacteria. For the mineral soil horizon, increasing PSD suppressed FNF rate via decreasing nifH gene abundance caused by elevated N availability and lowered phosphorus and iron availability or through decreasing the relative abundance of Deltaproteobacteria but increasing that of Betaproteobacteria.Our findings, for the first time, reveal the microbial and abiotic controls on FNF in litter and soil in response to PSD, and hence benefit the prediction of nitrogen input via biological N2 fixation under changes in PSD. Read the free Plain Language Summary for this article on the Journal blog. [ABSTRACT FROM AUTHOR]

The interest in growing pulse crops and oilseeds together as intercrop combinations is increasing. However, little is known about nutrient dynamics in pulse–oilseed intercropping systems in western Canada, particularly the contributions from biological nitrogen fixation (BNF). Kabuli chickpea and brown flax were grown without added fertilizer as monocrops and as intercrops in mixed and alternate seed row configurations at sites located in the Brown (Central Butte, SK) and Black (Redvers, SK) soil zones in 2019 and 2020. Comparison was made of grain and straw yields, nitrogen (N) and phosphorus (P) uptake of grain and straw, and proportion and amount of biologically fixed N contributed from the chickpea and transferred to the flax. The intercrop grain yield land equivalent ratio (LER) and N and P uptake LER values were at or above 1 for the four site-years, indicating benefit from intercropping in increasing total yield and crop N and P uptake from the land area. The proportion of N derived from BNF was not enhanced in this intercrop combination, but considerable biologically fixed N (8%–22%) was transferred from the chickpea to the flax during the growing season. This was associated with reduced depletion of soil N compared to monocrop flax in part due to the contribution of biologically fixed N in the chickpea–flax intercrop system. [ABSTRACT FROM AUTHOR]

Nitrogen is essential for all organisms, but biological nitrogen fixation (BNF) occurs only in a small fraction of prokaryotes. Previous studies divided nitrogenase-gene-carrying prokaryotes into Groups I to IV and provided evidence that BNF first evolved in bacteria. This study constructed a timetree of the evolution of nitrogen-fixation genes and estimated that archaea evolved BNF much later than bacteria and that nitrogen-fixing cyanobacteria evolved later than 1,900 MYA, considerably younger than the previous estimate of 2,200 MYA. Moreover, Groups III and II/I diverged ∼2,280 MYA, after the Kenorland supercontinent breakup (∼2,500–2,100 MYA) and the Great Oxidation Event (∼2,400–2,100 MYA); Groups III and Vnf/Anf diverged ∼2,086 MYA, after the Yarrabubba impact (∼2,229 MYA); and Groups II and I diverged ∼1,920 MYA, after the Vredefort impact (∼2,023 MYA). In summary, this study provided a timescale of BNF events and discussed the possible effects of geological events on BNF evolution. [ABSTRACT FROM AUTHOR]

Despite its enormous benefits, mining is responsible for intense changes to vegetation and soil properties. Thus, after extraction, it is necessary to rehabilitate the mined areas, creating better conditions for the establishment of plant species which is challenging. This study evaluated mineral and organic fertilization on the growth, and carbon and nitrogen (N) metabolism of two Crotalaria species [Crotalaria spectabilis (exotic species) and Crotalaria maypurensis (native species from Carajás Mineral Province (CMP)] established on a waste pile from an iron mine in CMP. A control (without fertilizer application) and six fertilization mixtures were tested (i = NPK; ii = NPK + micronutrients; iii = NPK + micronutrients + organic compost; iv = PK; v = PK + micronutrients; vi = PK + micronutrients + organic compost). Fertilization contributed to increased growth of both species, and treatments with NPK and micronutrients had the best results (up to 257% cf. controls), while organic fertilization did not show differences. Exotic Crotalaria had a greater number of nodules, higher nodule dry mass, chlorophyll a and b contents and showed free ammonium as the predominant N form, reflecting greater increments in biomass compared to native species. Although having lower growth, the use of this native species in the rehabilitation of mining areas should be considered, mainly because it has good development and meets current government legislation as an opportunity to restore local biodiversity. [ABSTRACT FROM AUTHOR]

Symbiotic diazotrophs form associations with legumes and substantially fix nitrogen into soils. However, grasslands on the Qinghai-Tibet Plateau are dominated by non-legume plants, such as Kobresia tibetica. Herein, we investigated the diazotrophic abundance, composition, and community structure in the soils and roots of three plants, non-legume K. tibetica and Kobresia humilis and the legume Oxytropis ochrocephala, using molecular methods targeting nifH gene. Diazotrophs were abundantly observed in both bulk and rhizosphere soils, as well as in roots of all three plants, but their abundance varied with plant type and soil. In both bulk and rhizosphere soils, K. tibetica showed the highest diazotroph abundance, whereas K. humilis had the lowest. In roots, O. ochrocephala and K. humilis showed the highest and the lowest diazotroph abundance, respectively. The bulk and rhizosphere soils exhibited similar diazotrophic community structure in both O. ochrocephala and K. tibetica, but were substantially distinct from the roots in both plants. Interestingly, the root diazotrophic community structures in legume O. ochrocephala and non-legume K. tibetica were similar. Diazotrophs in bulk and rhizosphere soils were more diverse than those in the roots of three plants. Rhizosphere soils of K. humilis were dominated by Actinobacteria, while rhizosphere soils and roots of K. tibetica were dominated by Verrumicrobia and Proteobacteria. The O. ochrocephala root diazotrophs were dominated by Alphaproteobacteria. These findings indicate that free-living diazotrophs abundantly and diversely occur in grassland soils dominated by non-legume plants, suggesting that these diazotrophs may play important roles in fixing nitrogen into soils on the plateau. [ABSTRACT FROM AUTHOR]

Inadequate management of nitrogen fertilization, due to high costs, compromises the nutrition and productivity of winter forage plants, highlighting the need to adopt sustainable alternatives, such as using nitrogen-fixing bacteria. This research evaluated the growth promotion of black oat (Avena strigosa) and ryegrass (Lolium multiflorum) plants inoculated with Azospirillum brasilense rhizobacteria. The experiments were carried out over two consecutive harvests in Santa Maria, Brazil. The sowing density was 300 viable seeds per square meter for black oats and 100 viable seeds per square meter for ryegrass. A dose of 5 ml per kg of seeds from the consortium was used as an inoculant. Following sowing, the seeds were incorporated into the soil through light harrowing. At the beginning of tillering, 50 kg of N ha-1 was applied to create treatments with and without nitrogen. Plant emergence, plant height, number of leaves and tillers were evaluated. In the first year, an inoculation response was observed for the number of black oat plants emerged m-2. In the second year (corn residue), an inoculation response was observed for the number of leaves and height of black oats. Under soybean straw for oat leaf number and ryegrass tiller number. The current research highlights the effectiveness of inoculation with Azospirillum brasilense in promoting significant improvements in the morphological components of forage crops. Following corn, an increase in the number of leaves and tillers was observed, as well as in the height of black oats. Similarly, following soybeans, there was an increase in the number of black oat tillers and the number of ryegrass leaves. The results emphasize the importance of inoculation as an effective practice to enhance crop development and performance in agricultural systems, improving sustainability and productivity under soybean and corn residues. [ABSTRACT FROM AUTHOR]

Nitrogenase is a complicated enzyme that actives the ATP-dependent reduction of dinitrogen (N2) to ammonia (NH3). The aim of this manuscript is to review the nitrogenase evolution with considering nitrogenase, structure of nitrogenase, action mechanism of nitrogenase and oxygen sensitive mechanism of nitrogenase. The searches focused on publications from 1980 to February 2023, using PubMed, Google Scholar, Science Direct, and Scopus databases. In the term of evolution, the nitrogen cycle has experienced highly changes; at the beginning of life and suggested the exact anoxic scenario, the comparatively sufficient ammonium was possibly used in an assimilation/mineralization cycle by protocellular organisms. The main nif gene products which are active in nitrogen fixation are nifH, nifD, nifK, nifT, nifY/nafY, nifE, nifN, nifX, nifU, nifS, nifV, nifW, nifZ, nifM, nifF, nifL, nifA, nifB, fdxN, nifQ, and nifJ. The main vnf gene products which are active in nitrogen fixation are vnfA, vnfE, vnfN, vnfX, vnfH, vnfFd, vnfD, vnfG, vnfK, and vnfY. Oxygen can be either detrimental or beneficial for diazotrophs in organisms suitable for an aerobic catabolism, and it supports the production of a substrate for nitrogenase (ATP), but it can also impede the activity and suppress the synthesis of this enzyme. [ABSTRACT FROM AUTHOR]

Plants perceive N-acetyl-d-glucosamine-containing oligosaccharides that play a role in the interaction with bacteria and fungi, through cell-surface receptors containing a tight bundle of three LysM domains in their extracellular region. However, the identification of LysM domains of receptor-like kinases (RLK)/receptor-like proteins (RLP) using sequence based methods has led to some ambiguity, as some proteins have been annotated with only one or two LysM domains. This missing annotation was likely produced by the failure of the LysM hidden Markov model (HMM) from the Pfam database to correctly identify some LysM domains in proteins of plant origin. In this work, we provide improved HMMs for LysM domain detection in plants, that were built from the structural alignment of manually curated LysM domain structures from the Protein Data Bank and AlphaFold Protein Structure Database. Furthermore, we evaluated different sets of ligand-specific HMMs that were able to correctly classify a limited set of fully characterised RLK/Ps by their ligand specificity. In contrast, the phylogenetic analysis of the extracellular region of RLK/Ps, or of their individual LysM domains, was unable to discriminate these proteins by their ligand specificity. The HMMs reported here will allow a more sensitive detection of plant proteins containing LysM domains and help improve their characterisation. [ABSTRACT FROM AUTHOR]

The legume family includes approximately 19,300 species across three large subfamilies, of which Papilionoideae stands out with 13,800 species. Lentils were one of the first crops to be domesticated by humans, approximately 11,000 BP. They are diploid legumes that belong to the Papilionoidea subfamily and are of agricultural importance because of their resistance to drought and the fact that they grow in soil with a pH range of 5.5–9; therefore, they are cultivated in various types of soil, and so they have an important role in sustainable food and feed systems in many countries. In addition to their agricultural importance, lentils are a rich source of protein, carbohydrates, fiber, vitamins, and minerals. They are key to human nutrition since they are an alternative to animal proteins, decreasing meat consumption. Another characteristic of legumes, including lentils, is their ability to form nodules, which gives them a growth advantage in nitrogen-deficient soils because they enable the plant to fix atmospheric nitrogen, thus contributing nitrogen to the soil and facilitating the nutrition of other plants during intercropping. Lentils have also been applied for protection against various human diseases, as well as for phytoremediation, and they also have been applied as environmental bioindicators to identify cytotoxicity. This review addresses the importance of lentils in agriculture and human health. [ABSTRACT FROM AUTHOR]

The application of nitrogen fertilizer is crucial to the growth and biological nitrogen fixation of peanut, especially in the seedling stage where nodules have not yet formed. However, it is still uncertain how much initial nitrogen fertilizer should be applied to promote peanut root growth, nodule formation, and biological nitrogen fixation (BNF). There, a 2-year pot experiment was conducted using Huayu 22 (HY22, large-grain cultivar) and Huayu 39 (HY39, small-grain cultivar) as experimental materials to research the effects of different initial nitrogen fertilizer application rates on peanut root growth (root weight, root length, root mean diameter, root activity) and biological nitrogen fixation capacity (nodule number, nodule weight, biological nitrogen fixation, and nitrogen fixation potential per plant). N0, as control, four initial nitrogen fertilizer application rates were established: 15 kg·hm−2 (N15), 30 kg·hm−2 (N30), 45 kg·hm−2 (N45), and 60 kg·hm−2 (N60). The present results showed that the nodule number, nodule dry weight, nitrogenase activity, and biological nitrogen fixation of the HY22 cultivar under the N15 treatment were higher compared to those under other treatments over the two growing seasons. In addition, the cultivar of HY39 treated with the N15 treatment also increased the nitrogen fixation potential per plant and BNF relative to other treatments. Although the application of 60 kg·hm−2 nitrogen increased the root surface area and root volume, it decreased the nitrogenase activity, nodule dry weight, and nitrogen fixation potential per plant of HY22 and HY39 varieties in both growing seasons. Above all, an initial nitrogen application of 15 kg·hm−2 may be the optimal treatment for promoting peanut nodule formation and biological nitrogen fixation. [ABSTRACT FROM AUTHOR]

Besides being a significant part of the Brazilian national economy, the common bean is a legume whose grains are part of the daily diet of the population of developing nations globally. Combining N-fixing bacteria and plant phytohormone producers through the use of inoculants is an alternative to improve plant growth and increase grain yield in the common bean production system. This study aimed to evaluate the effect of co-inoculation doses with Rhizobium tropici and Azospirillum brasilense, applied in the planting groove, on nodulation, growth and productivity parameters of the common bean. Five field experiments were conducted in a randomized block design with four replications. Restrictions were formulated, namely: absolute control, without fertilization and without co-inoculation (AC); nitrogen fertilization (NfT); single inoculation of R. tropici (Rt); R. tropici + one dose of A. brasiliense (Rt + Ab1f); R. tropici + two doses of A. brasiliense (Rt + Ab2f); R. tropici + three doses of A. brasiliense (Rt + Ab3f); R. tropici + four doses of A. brasiliense (Rt + Ab4f) and R. tropici + three doses of A. brasiliense (RP). The number of nodules (NN), nodule dry mass (NDM), aerial dry mass (ADM), root dry mass (RDM), pod number (NP), number of grains (NG), mass of 100 grain (100G) and grain yield (GY) were taken. The Rt + Ab4f co-inoculation provided a significant increase in NN, NDM, ADM and RDM values than the Rt and AC treatments. In addition, it also presented the highest GY stability, with an average production of 3499 kg ha− 1 from the five sites, which represents an increase of 15.4%, 14.8%, 11.0%, and 1.0% as compared to Rt, AC, RP, and NfT treatments, respectively. The results indicate the feasibility of the use of co-inoculation with R. tropici and A. brasilense in commercial production farms and for small producers in several countries of the world with similar edaphoclimatic conditions, reducing the use of nitrogen fertilizers, production costs and the prevention of negative environmental impacts. [ABSTRACT FROM AUTHOR]

Biological nitrogen fixation is a key process balancing the loss of combined nitrogen in the marine nitrogen cycle. Its relevance in upwelling or high nutrient regions is still unclear, with the few available studies in these regions of the ocean reporting rates that vary widely from below detection limit to > 100 nmol N L−1 d−1. In the eastern tropical Atlantic Ocean, two open ocean upwelling systems are active in boreal summer. One is the seasonal equatorial upwelling, where the residual phosphorus associated with aged upwelled waters is suggested to enhance nitrogen fixation in this season. The other is the Guinea Dome, a thermal upwelling dome. We conducted two surveys along 23° W across the Guinea Dome and the Equator from 15° N to 5° S in September 2015 and August–September 2016 with high latitudinal resolution (20–60 nm between stations). The abundance of Trichodesmium colonies was characterized by an Underwater Vision Profiler 5 and the total biological nitrogen fixation in the euphotic layer was measured using the 15N2 technique. The highest abundances of Trichodesmium colonies were found in the area of the Guinea Dome (9°–15° N) with a maximum of 3 colonies L−1 near the surface. By contrast, colonies were almost absent in the Equatorial band between 2° N and 5° S. The highest nitrogen fixation rate was measured at the northern edge of the Guinea Dome in 2016 (ca. 31 nmol N L−1 d−1). In this region, where diazotrophs thrived on a sufficient supply of both phosphorus and iron, a patchy distribution was unveiled by our increased spatial resolution scheme. In the Equatorial band, rates were considerably lower, ranging from below detection limit to ca. 4 nmol N L−1 d−1, with a clear difference in magnitude between 2015 (rates close to zero) and 2016 (average rates around 2 nmol N L−1 d−1). This difference seemed triggered by a contrasting supply of phosphorus between years. Our study stresses the importance of surveys with sampling at fine-scale spatial resolution, and shows unexpected high variability in the rates of nitrogen fixation in the Guinea Dome, a region where diazotrophy is a significant process supplying new nitrogen into the euphotic layer. [ABSTRACT FROM AUTHOR]

The common bean is known to require nitrogen, with mineral fertilizer being the main form of supply to the plants. However, more efficient inoculants containing strains of Rhizobium tropici that are potentially more efficient in fixing atmospheric N2 could replace the use of nitrogen fertilizers as long as some of the existing challenges in the application methodology are overcome (for example, ensuring the existence of active nodules in the most demanding crop phases: flowering and grain filling). This study aimed to verify the responses of common bean plants subjected to reinoculation using Rhizobium tropici in topdressing applied at different stages in a greenhouse and in a field between March and April 2020 and in the spring–summer harvest of 2020/2021, respectively. A randomized block design was used, along with four replications: T1 = uninoculated control; T2 = inoculation via seed (VS); T3 = VS + reinoculation at the V4 stage; T4 = VS + reinoculation at the R5 stage; T5 = VS + reinoculation at the R6 stage; T6 = VS + reinoculation at the V4 and R5 stages; T7 = VS + reinoculation at the V4 and R6 stages; T8 = VS + reinoculation at the R5 and R6 stages; T9 = VS + reinoculation at the V4, R5, and R6 stages; and T10 = mineral nitrogen fertilization. Nodulation and morphological variables were evaluated at the R7 stage, and agronomic variables were evaluated at the R9 stage at harvest. The use of reinoculation, in addition to inoculation, positively influenced the bean nodulation process. The morphological characteristics of the plants showed good development with the supply of N (carried out via inoculation and reinoculation), equivalent to the development that could be achieved via the addition of a mineral nitrogen fertilizer. Reinoculation from the R5 to R7 stages proved ineffective in supplying N to the common bean crop. Inoculation in the seed associated with reinoculation in coverage at the V4 stage provided higher yields (3.271 kg ha−1), reinforcing the potential for mineral nitrogen fertilizers to be replaced. The use of the seed inoculation technique with subsequent reinoculation under cover can completely replace the use of nitrogenous mineral fertilizers in fertilizing bean plants, making the cultivation of this fabacea more sustainable in terms of the associated costs and environmental problems. [ABSTRACT FROM AUTHOR]

IntroductionBiological nitrogen fixation (BNF), an unparalleled metabolic novelty among living microorganisms on earth, globally contributes ~88-101 Tg N year−1 to natural ecosystems, ~56% sourced from symbiotic BNF while ~22-45% derived from free-living nitrogen fixers (FLNF). The success of symbiotic BNF is largely dependent on its interaction with host-plant, however ubiquitous environmental heterotrophic FLNFs face many limitations in their immediate ecological niches to sustain unhindered BNF. The autotrophic FLNFs like cyanobacteria and oceanic heterotrophic diazotrophs have been well studied about their contrivances acclimated/adapted by these organisms to outwit the environmental constraints for functional diazotrophy. However, FLNF heterotrophs face more adversity in executing BNF under stressful estuarine/marine/aquatic habitats.MethodsIn this study a large-scale cultivation-dependent investigation was accomplished with 190 NCBI accessioned and 45 non-accessioned heterotrophic FLNF cultivable bacterial isolates (total 235) from halophilic estuarine intertidal mangrove niches of Indian Sundarbans, a Ramsar site and UNESCO proclaimed World Heritage Site. Assuming ~1% culturability of the microbial community, the respective niches were also studied for representing actual bacterial diversity via cultivation-independent next-generation sequencing of V3-V4 rRNA regions.ResultsBoth the studies revealed a higher abundance of culturable Gammaproteobacteria followed by Firmicutes, the majority of 235 FLNFs studied belonging to these two classes. The FLNFs displayed comparable selection potential in media for free nitrogen fixers and iron-oxidizing bacteria, linking diazotrophy with iron oxidation, siderophore production, phosphorus solubilization, phosphorus uptake and accumulation as well as denitrification.DiscussionThis observation validated the hypothesis that under extreme estuarine mangrove niches, diazotrophs are naturally selected as a specialized multidimensional entity, to expedite BNF and survive. Earlier metagenome data from mangrove niches demonstrated a microbial metabolic coupling among C, N, P, S, and Fe cycling in mangrove sediments, as an adaptive trait, evident with the co-abundant respective functional genes, which corroborates our findings in cultivation mode for multiple interrelated metabolic potential facilitating BNF in a challenging intertidal mangrove environment.

Assuming that common bean (Phaseolus vulgaris) cultivars differ in terms of the potential to respond to symbiosis with rhizobia, in this direction, this work aimed to identify those that are more efficient and responsive to biological nitrogen fixation. The interaction of common bean cultivars and rhizobia inoculation at greenhouse conditions was evaluated. Six common bean cultivars (IPR-Sabiá, IPR-Quero-Quero, IPR-Andorinha, IPR-Curió, IPR-Tangará, and IPR-Campos Gerais) were inoculated using a mixture of rhizobia strains (Rhizobium tropici and R. freirei). The completely randomized experimental design arranged in a 6 × 2 factorial scheme with five replications was used. Growth, nodulation, and yield performance of common bean cultivars with and without rhizobia inoculation were evaluated. Rhizobia strains increased nodulation, plant growth, and grain yield in all cultivars used. There was an influence of interaction between rhizobia inoculation and common bean cultivars on root volume, and on dry matter root and shoot. The cultivar IPR-Curió showed the highest number of nodules and nodule dry matter. It can be concluded that the inoculation was consistent for all cultivars used, but some differences in the response from the cultivars indicate a possible effect of the genotype.

Abstract Background Astragalus mongolicus Bunge is used in traditional Chinese medicine and is thus cultivated in bulk. The cultivation of A. mongolicus requires a large amount of nitrogen fertilizer, increasing the planting cost of medicinal materials and polluting the environment. Isolation and screening of plant growth-promoting rhizobacteria (PGPR) and exploring the nitrogen fixation potential of A. mongolicus rhizosphere microorganisms would effectively reduce the production cost of A. mongolicus. Results This study used A. mongolicus roots and rhizosphere soil samples from Longxi County of Gansu Province, Jingle County, and Hunyuan County of Shanxi Province, China, to isolate and identify nitrogen-fixing bacteria. Through nitrogen fixation efficiency test, single strain inoculation test, and plant growth-promoting characteristics, three strains, Bacillus sp. J1, Arthrobacter sp. J2, and Bacillus sp. G4 were selected from 86 strains of potential nitrogen-fixing bacteria, which were the most effective in promoting the A. mongolicus growth and increasing the nitrogen, phosphorus, and potassium content in plants. The antagonistic test showed that these bacteria could grow smoothly under the co-culture conditions. The J1, J2, and G4 strains were used in a mixed inoculum and found to enhance the biomass of A. mongolicus plants and the accumulation of the main medicinal components in the field experiment. Mixed bacterial agent inoculation also increased bacterial diversity and changed the structure of the bacterial community in rhizosphere soil. Meanwhile, the relative abundance of Proteobacteria increased significantly after inoculation, suggesting that Proteobacteria play an important role in plant growth promotion. Conclusions These findings indicate that specific and efficient PGPRs have a significant promoting effect on the growth of A. mongolicus, while also having a positive impact on the structure of the host rhizosphere bacteria community. This study provides a basis for developing a nitrogen-fixing bacterial fertilizer and improving the ecological planting efficiency of A. mongolicus.

IntroductionIntercropping as an approach for development of sustainable agriculture systems is pursuing the purposes such as creating ecological balance, further exploitation of resources, increase the quantity and quality of yield and reduce losses due to pests, diseases and weeds and reduce the dependence of farmers to pesticides, to maintain product quality and marketability. Jahani et al (2007) reported that intercropping of lentil and cumin has significantly increased dry weight of vegetative organs, biological yield, 1000-seed weight and number of seeds per cumin umbrella and seed yield. Legume/non-legume intercropping system due to the beneficial use of resources produces more yields. In review of about the fennel and fenugreek intercropping, intercropping of the two plants significantly increased the fennel seed yield compared with monoculture.Materials and MethodsThe experiment was conducted in 2014-2015 growing season based on randomized complete block design with three replications at Agricultural Research Station, Faculty of Agriculture, Ferdowsi University of Mashhad. Experimental treatments were 25% mung bean + 75% fennel, 50% mung bean + 50% fennel, 75% mung bean + 25% fennel, monoculture of mung bean and monoculture of fennel. Species were sown at the same time in 12/06/2015 in rows with a distance of 50 cm. At harvesting time, dry matter, seed yield and yield components of these crops were measured. Yield advantages of intercrops over monocultures were compared by land equivalent ratios. Seeds of fennel were analyzed for essential oil content. Results and DiscussionThe highest seed yield, biomass and essential oil of fennel for monoculture with 1460.4, 12286.4 and 50.4 kg per hectare respectively and the lowest values for 25% fennel + 75% mung bean treatment with 263.2, 5843.3 and 8.1 kg per hectare respectively were observed. Also, the highest seed yield and biomass of mung bean for monoculture with 2167.1 and 8900.5 kg per hectare respectively and the lowest values for 25% fennel + 75% mung bean treatment with 740.5 and 2540.5 kg per hectare respectively were observed. The highest essential oil contents of fennel with 3.56% was observed in ratio of 50% fennel + 50% mung bean and the lowest value was observed in 25% fennel + 75% mung bean with 3.1%. It seems that interspecific competition between fennel and mung bean due to the abiotic stresses (competition) led to increasing essential oil percent in intercrops. The highest essential oil yield of fennel was observed in monoculture with 50.4 kg/ha and the lowest value was observed in 25% fennel + 75% mung bean with 8.1 kg/ha. The highest mung bean harvest index was 28% in 25% mung bean + 75% fennel and the lowest was observed in 50% mung bean + 50% fennel with 23.5%. Also, the highest fennel harvest index was observed with 12% in monoculture and the lowest in 25% fennel + 75% mung bean with 4.5%. The highest value of total land equivalent ratio (1.07) was obtained in ratio of 50% mung bean + 50% fennel indicating 7 percent yield advantage of intercropping compared to pure stands of species of these two products compare to each other that represents of showing the more efficient use of land. The lowest value by intercrops of land equivalent ratio was observed in ratio of 25% mung bean + 75% fennel which was 9% lower than the monoculture.ConclusionIn general, intercropping of fennel with mung bean, had positive effect on biological and seed yield of two species as well as percent and yield of fennel essential oil. So, in order to reduce the consumption of chemical fertilizers in sustainable agriculture and as for nitrogen biological fixation ability of mung bean, intercropping this plant with fennel can be increased resources efficiency and improve of the quality and quantity yield of fennel.

Pseudomonas spp. make up 1.6% of the bacteria in the soil and are found throughout the world. More than 140 species of this genus have been identified, some beneficial to the plant. Several species in the family Pseudomonadaceae, including Azotobacter vinelandii AvOP, Pseudomonas stutzeri A1501, Pseudomonas stutzeri DSM4166, Pseudomonas szotifigens 6HT33bT, and Pseudomonas sp. strain K1 can fix nitrogen from the air. The genes required for these reactions are organized in a nitrogen fixation island, obtained via horizontal gene transfer from Klebsiella pneumoniae, Pseudomonas stutzeri, and Azotobacter vinelandii. Today, this island is conserved in Pseudomonas spp. from different geographical locations, which, in turn, have evolved to deal with different geo-climatic conditions. Here, we summarize the molecular mechanisms behind Pseudomonas-driven plant growth promotion, with particular focus on improving plant performance at limiting nitrogen (N) and improving plant N content. We describe Pseudomonas-plant interaction strategies in the soil, noting that the mechanisms of denitrification, ammonification, and secondary metabolite signaling are only marginally explored. Plant growth promotion is dependent on the abiotic conditions and differs at sufficient and deficient N. The molecular controls behind different plant responses are not fully elucidated. We suggest that superposition of transcriptome, proteome, and metabolome data and their integration with plant phenotype development through time will help fill these gaps. The aim of this review is to summarize the knowledge behind Pseudomonas-driven nitrogen fixation and to point to possible agricultural solutions. [Graphic: see text] Copyright © 2023 The Author(s). This is an open access article distributed under the CC BY 4.0 International license.

Exploring natural diversity for biological nitrogen fixation in maize and its progenitors is a promising approach to reducing our dependence on synthetic fertilizer and enhancing the sustainability of our cropping systems. We have shown previously that maize accessions from the Sierra Mixe can support a nitrogen-fixing community in the mucilage produced by their abundant aerial roots and obtain a significant fraction of their nitrogen from the air through these associations. In this study, we demonstrate that mucilage production depends on root cap and border cells sensing water, as observed in underground roots. The diameter of aerial roots correlates with the volume of mucilage produced and the nitrogenase activity supported by each root. Young aerial roots produce more mucilage than older ones, probably due to their root cap's integrity and their ability to produce border cells. Transcriptome analysis on aerial roots at two different growth stages before and after mucilage production confirmed the expression of genes involved in polysaccharide synthesis and degradation. Genes related to nitrogen uptake and assimilation were up-regulated upon water exposure. Altogether, our findings suggest that in addition to the number of nodes with aerial roots reported previously, the diameter of aerial roots and abundance of border cells, polysaccharide synthesis and degradation, and nitrogen uptake are critical factors to ensure efficient nitrogen fixation in maize aerial roots.

Given the importance of biological nitrogen (N) fixation in agroecosystems, using inoculants with phyllosphere N fixers effective across various crops would revolutionize agriculture. In this study, the application of an inoculant prepared from Methylobacterium symbioticum was tested on young olive trees. The pot experiment was arranged in a factorial design, with inoculant (Yes and No) and mineral N applied to the soil [0 (N0), 25 (N25), 50 (N50), and 100 (N100) kg ha−1], and four replicates. The inoculant application did not increase plant dry matter yield (DMY), whereas the application of mineral N had a significant and pronounced effect. The inoculant also did not significantly increase N concentration in tissues, unlike the strong increase observed with N applied to the soil. The inoculant significantly increased plant N recovery, a cumulative effect resulting from small increases in DMY and N concentration in tissues. This increase represented 5.2% more N in plants receiving the inoculant compared to untreated ones. However, only treatments receiving mineral N recorded positive values of fixed N, with the highest value observed in the N50 treatment (12.4%), whereas a negative value (−7.7%) was observed in the N0 treatment. Overall, these low values of fixed N question the economic rationale of using this inoculant by farmers and especially render it unsuitable for organic farming systems, where plants tend to have lower N levels in tissues.

IntroductionIntercropping cultivation is an excellent method to increase agricultural productivity per unit area based on environmental mechanisms. In this planting system, it is necessary to select crops with different capabilities in uptake resources and establish a complementary relationship. However, legumes in intercropping cultivation can be used an alternative and sustainable way to increase efficiency. Besides, it is necessary to study the interactions in the above and belowground parts to achieve the mentioned goals. Therefore, this study aimed to investigate inter-specific interactions in millet and soybean intercropping cultures. Material and MethodsA field experiment was conducted for two years (2018-19) at the research farm of Sari agricultural science and natural resources university, Iran (33º, 36ʹ N, 53º, 03ʹ E with 43 m altitude). The experimental design was a randomized complete block with four replications. The experimental treatment was 75% Soybean + 25% Niger, 50% Soybean + 50% Niger, 25% Soybean+75%Niger based on the replacement method and monoculture. This study investigated the agronomic traits, LAI, TDW, nitrogen derived from the atmosphere, and total microbial activity in the soil in different planting ratios. Result and DiscussionThe results showed that soybean monoculture had the highest plant height during different stages of growth. Furthermore, with the increase in the share of planting soybean in intercropping cultivation, the shoot height of the millet also increased. Also, unlike the millet, the soybean succeeded in producing more leaf area indexes among different mixed cultivation ratios than monoculture. The leaf area index of millet showed a significant decrease as the proportion of soybean plantings increased. However, despite this decrease, the total dry matter of soybean exhibited a higher percentage increase at various growth stages compared to the decrease in millet yield. As a result, after 60 days of planting, the intercropping cultivation of millet and soybean demonstrated positive complementarity in terms of competition impact. Additionally, the intercropping cultivation showed a higher percentage of nitrogen derived from the atmosphere compared to soybean monoculture. During the study period, the planting ratio of 75% millet and 25% soybean exhibited the highest increase in nitrogen fixation (Ndfa), with average values of 93.84% and 83.85% at 90 days after planting, respectively. Increasing the number of millet sowing rows in the intercropping patterns played a crucial role in intensifying competition with soybeans during the early growth stages and had an impact on the biological activity of soybeans. Moreover, the microbial activity in the soil (at a depth of 0-30 cm) was higher in the different intercropping ratios compared to monocultures of soybean and millet. In addition, soybean and millet monoculture had the highest and lowest grain yields. Among the different intercropping ratios, the 75:25 in 1398 with an average of 4639.77 kg. ha-1 had the highest grain yield. Besides, the planting ratio of 25:75 and 50:50, with an average of 1.23 and 1.02, respectively, had an LER higher than one. With increasing the share of soybean planting in intercropping, the competitiveness of the plant increases, and the competition ratio of millet was higher than the unit only in the planting ratio of 25:75 (millet-soybean). Therefore, the competitiveness of millet in mixed cultivation is significantly reduced by reducing the share of planting row. Finally, increasing productivity in the 25:75 combinations reduced soybean's competitive dominance, increasing millet grain yield in comparison to expected values and improving total microbial activity. ConclusionDue to limited growth period, millet had competition played an essential role in increasing total microbial activity and the percentage of nitrogen derived from the atmosphere at a 25:75 planting ratio before the initial reproductive growth. In addition, 25:75 and 50:50 planting ratios reduce soybean's competitive dominance, resulting in enhanced productivity. Finally, the selection of complementary plants based on physiological, morphological characteristics, and competitiveness in intercropping cultivation can be effective in the above and belowground interactions.

ABSTRACT The objective of this work was to assess the effects of A. brasilense inoculation on seeds on the yield performance of maize and black oat as a function of nitrogen rate over a 3-year succession. The experiment was conducted from 2015 to 2018 under field conditions. Maize was grown for three growing seasons in rotation with two seasons of black oat at the same experimental site. A randomized block design with a split-plot arrangement was used. Main plots consisted of A. brasilense inoculation (inoculated and uninoculated plants) and subplots of four nitrogen rates (0, 40, 80, and 120 kg ha−1). Maize plants were evaluated for biometric parameters, yield, and yield components at flowering. Black oat plants were analyzed for dry matter yield at flowering. The inoculation of A. brasilense in maize and black oat seeds in succession does not promote productivity nor does it favor the nitrogen fertilization of these cultures.

Nitrogenase is a complicated enzyme that actives the ATP-dependent reduction of dinitrogen (N2) to ammonia (NH3). The aim of this manuscript is to review the nitrogenase evolution with considering nitrogenase, structure of nitrogenase, action mechanism of nitrogenase and oxygen sensitive mechanism of nitrogenase. The searches focused on publications from 1980 to February 2023, using PubMed, Google Scholar, Science Direct, and Scopus databases. In the term of evolution, the nitrogen cycle has experienced highly changes; at the beginning of life and suggested the exact anoxic scenario, the comparatively sufficient ammonium was possibly used in an assimilation/mineralization cycle by protocellular organisms. The main nif gene products which are active in nitrogen fixation are nifH, nifD, nifK, nifT, nifY/nafY, nifE, nifN, nifX, nifU, nifS, nifV, nifW, nifZ, nifM, nifF, nifL, nifA, nifB, fdxN, nifQ, and nifJ. The main vnf gene products which are active in nitrogen fixation are vnfA, vnfE, vnfN, vnfX, vnfH, vnfFd, vnfD, vnfG, vnfK, and vnfY. Oxygen can be either detrimental or beneficial for diazotrophs in organisms suitable for an aerobic catabolism, and it supports the production of a substrate for nitrogenase (ATP), but it can also impede the activity and suppress the synthesis of this enzyme.

Symbiotic diazotrophs form associations with legumes and substantially fix nitrogen into soils. However, grasslands on the Qinghai-Tibet Plateau are dominated by non-legume plants, such as Kobresia tibetica. Herein, we investigated the diazotrophic abundance, composition, and community structure in the soils and roots of three plants, non-legume K. tibetica and Kobresia humilis and the legume Oxytropis ochrocephala, using molecular methods targeting nifH gene. Diazotrophs were abundantly observed in both bulk and rhizosphere soils, as well as in roots of all three plants, but their abundance varied with plant type and soil. In both bulk and rhizosphere soils, K. tibetica showed the highest diazotroph abundance, whereas K. humilis had the lowest. In roots, O. ochrocephala and K. humilis showed the highest and the lowest diazotroph abundance, respectively. The bulk and rhizosphere soils exhibited similar diazotrophic community structure in both O. ochrocephala and K. tibetica, but were substantially distinct from the roots in both plants. Interestingly, the root diazotrophic community structures in legume O. ochrocephala and non-legume K. tibetica were similar. Diazotrophs in bulk and rhizosphere soils were more diverse than those in the roots of three plants. Rhizosphere soils of K. humilis were dominated by Actinobacteria, while rhizosphere soils and roots of K. tibetica were dominated by Verrumicrobia and Proteobacteria. The O. ochrocephala root diazotrophs were dominated by Alphaproteobacteria. These findings indicate that free-living diazotrophs abundantly and diversely occur in grassland soils dominated by non-legume plants, suggesting that these diazotrophs may play important roles in fixing nitrogen into soils on the plateau.

ABSTRACT The objective of this work was to evaluate the agronomic performance of a second crop maize grown under different methods of inoculation with Azospirillum brasilense. The experiment was conducted in a randomized block design with 6 treatments and four replications. The treatments used were: no inoculation (T1); inoculation with A. brasilense via seed (T2); inoculation with A. brasilense via sowing furrow (T3); inoculation with A. brasilense via leaf (T4); two inoculations with A. brasilense via leaf (T5); and combination of inoculations via seed, sowing furrow, and leaf (T6). Physiological, morphological, production, and yield components of the second crop maize were evaluated. The data were subjected to analysis of variance at 0.05 probability level and when significant, the means were subjected to the Scott-Knott test at 0.05 probability level, using the R-bio statistical program. Inoculation via seed results in higher nitrate reductase enzyme activity. The inoculation with A. brasilense does not result in gains in grain yield for second crop maize; however, it improves the maize quality by increasing its gross protein contents.

Phosphorus (P) is a major limiting factor for legume and symbiotic nitrogen fixation (SNF). Although overall adaptations of legumes to P supplementation have been extensively studied in connection with inorganic P, little information is currently available regarding nodulation or SNF responses to organic P (Po) in hydroponics. We investigated the mineral and carbon metabolism of Po-induced nodules of two contrasting faba bean varieties grown hydroponically under inorganic P (Pi), viz., in P-deficient (2 µM KH2PO4, −Pi), sufficient-P (200 µM KH2PO4, +Pi), and phytic acid (200 µM, Po) conditions, and were inoculated with Rhizobium leguminosarum bv. viciae 3841 and grown for 30 days. The results consistently reveal similar growth and biomass partitioning patterns between +Pi and Po, with both varying substantially from −Pi. In comparison, +Pi and Po observed equivalent accumulations of overall elemental P concentrations, with both increasing by 114 and 119%, respectively, relative to −Pi. A principal component analysis on metabolites showed a clear separation of the −Pi treatment from the others, with +Pi and Po correlating closely together, highlighting the nonsignificant differences between them. Additionally, the δ15N abundance of shoots, roots, and nodules was not significantly different between treatments and varieties and exhibited negative δ15N signatures for all tissues. Our study provides a novel perspective on mineral and carbon metabolism and their regulation of the growth, functioning, and reprogramming of nodules upon phytate supply. [ABSTRACT FROM AUTHOR]

Inoculation with Bacillus subtilis is a promising approach to increase plant yield and nutrient acquisition. In this context, this study aimed to estimate the B. subtilis concentration that increases yield, gas exchange, and nutrition of lettuce plants in a hydroponic system. The research was carried out in a greenhouse in Ilha Solteira, Brazil. A randomized block design with five replications was adopted. The treatments consisted of B. subtilis concentrations in nutrient solution [0 mL "non-inoculated", 7.8 × 10³, 15.6 × 10³, 31.2 × 10³, and 62.4 × 10³ colony forming units (CFU) mL-1 of nutrient solution]. There was an increase of 20% and 19% in number of leaves and 22% and 25% in shoot fresh mass with B. subtilis concentrations of 15.6 × 10³ and 31.2 × 10³ CFU mL-1 as compared to the non-inoculated plants, respectively. Also, B. subtilis concentration at 31.2 × 10³ CFU mL-1 increased net photosynthesis rate by 95%, intercellular CO2 concentration by 30%, and water use efficiency by 67% as compared to the non-inoculated treatments. The concentration of 7.8 × 10³ CFU mL-1 improved shoot accumulation of Ca, Mg, and S by 109%, 74%, and 69%, when compared with non-inoculated plants, respectively. Inoculation with B. subtilis at 15.6 × 10³ CFU mL-1 provided the highest fresh leaves yield while inoculation at 15.6 × 10³ and 31.2 × 10³ CFU mL-1 increased shoot fresh mass and number of leaves. Concentrations of 7.8 × 10³ and 15.6 × 10³ increased shoot K accumulation. The concentrations of 7.8 × 10³, 15.6 × 10³, and 31.2 × 10³ CFU mL-1 increased shoot N accumulation in hydroponic lettuce plants. [ABSTRACT FROM AUTHOR]

Simple Summary: Azotobacter vinelandii is a model organism used to study biological nitrogen fixation, a process by which nitrogen gas is transformed into ammonia, a form of nitrogen that can be assimilated by most organisms. This requires the synthesis and transfer of specific iron-containing cofactors to enzymes involved in nitrogen fixation. There are large gaps in our knowledge of iron uptake and utilization by A. vinelandii. In this review, our goal is to summarize current knowledge, propose novel elements based on our current understanding of bacterial iron homeostasis, and highlight those areas requiring more detailed research. Iron is an essential nutrient for all life forms. Specialized mechanisms exist in bacteria to ensure iron uptake and its delivery to key enzymes within the cell, while preventing toxicity. Iron uptake and exchange networks must adapt to the different environmental conditions, particularly those that require the biosynthesis of multiple iron proteins, such as nitrogen fixation. In this review, we outline the mechanisms that the model diazotrophic bacterium Azotobacter vinelandii uses to ensure iron nutrition and how it adapts Fe metabolism to diazotrophic growth. [ABSTRACT FROM AUTHOR]

• Biological nitrogen fixation is a sustainable approach for boosting growth of rice and reducing the burden of chemical fertilizers • Azotobacter and Azospirillum spp. isolates (Az2 & As5) from native soil showed promising PGP functions • The isolates Az2 and As5 showed significantly higher N 2 -ase activity and fixed more nitrogen • Combined application of Azotobacter and Azospirillum spp. isolates (Az2 & As5) promoted the growth and development of rice (Oryza sativa L. var. Khandagiri) Biological nitrogen fixation is significantly effective and sustainable field procedure for the promotion of plant growth & development and reducing the burden of chemical fertilizers. In the present investigation, plant growth promoting activities of indigenous bio-fertilizer formulations of Azotobacter and Azospirillum spp. isolated from rice pots filled with local soil of Odisha was examined. The efficacy of Azotobacter and Azospirillum spp. were assessed by selecting five isolates from each genus that were analyzed by biochemical assay like utilization and fermentation of carbohydrate, citrate, nitrate reduction activity, oxidase activity, catalase activity and phosphatase activity. The isolates (Az2 and As5) showed maximum PGP activities among all the isolates of Azotobacter and Azospirillum spp. The analyzed isolates showed variation in production of ethylene between 63.11 ± 4.23 to 121.09 ± 4.56 nmole C 2 H 4 /mg bact./h. The isolates Az2 and As5 among Azotobacter and Azospirillum spp. respectively fixed more nitrogen in culture which is 101.66 ± 9.79 (Az2) nmole and 121.09 ± 4.56 (As5) nmole C 2 H 4 /mg bact./h. The treatments T 1 (Az2) and T 2 (As5) were used as individual application of Azotobacter and Azospirillum spp. and T 3 as combined application of both the strains which had the best response in all of the developmental metrics studied. Among all treatments, T 3 performed around 23-27% better than the T 1 and T 2 individually. [ABSTRACT FROM AUTHOR]

Background: Astragalus mongolicus Bunge is used in traditional Chinese medicine and is thus cultivated in bulk. The cultivation of A. mongolicus requires a large amount of nitrogen fertilizer, increasing the planting cost of medicinal materials and polluting the environment. Isolation and screening of plant growth-promoting rhizobacteria (PGPR) and exploring the nitrogen fixation potential of A. mongolicus rhizosphere microorganisms would effectively reduce the production cost of A. mongolicus. Results: This study used A. mongolicus roots and rhizosphere soil samples from Longxi County of Gansu Province, Jingle County, and Hunyuan County of Shanxi Province, China, to isolate and identify nitrogen-fixing bacteria. Through nitrogen fixation efficiency test, single strain inoculation test, and plant growth-promoting characteristics, three strains, Bacillus sp. J1, Arthrobacter sp. J2, and Bacillus sp. G4 were selected from 86 strains of potential nitrogen-fixing bacteria, which were the most effective in promoting the A. mongolicus growth and increasing the nitrogen, phosphorus, and potassium content in plants. The antagonistic test showed that these bacteria could grow smoothly under the co-culture conditions. The J1, J2, and G4 strains were used in a mixed inoculum and found to enhance the biomass of A. mongolicus plants and the accumulation of the main medicinal components in the field experiment. Mixed bacterial agent inoculation also increased bacterial diversity and changed the structure of the bacterial community in rhizosphere soil. Meanwhile, the relative abundance of Proteobacteria increased significantly after inoculation, suggesting that Proteobacteria play an important role in plant growth promotion. Conclusions: These findings indicate that specific and efficient PGPRs have a significant promoting effect on the growth of A. mongolicus, while also having a positive impact on the structure of the host rhizosphere bacteria community. This study provides a basis for developing a nitrogen-fixing bacterial fertilizer and improving the ecological planting efficiency of A. mongolicus. [ABSTRACT FROM AUTHOR]

Summary: Bacteria colonize plant roots and engage in reciprocal interactions with their hosts. However, the contribution of individual taxa or groups of bacteria to plant nutrition and fitness is not well characterized due to a lack of in situ evidence of bacterial activity.To address this knowledge gap, we developed an analytical approach that combines the identification and localization of individual bacteria on root surfaces via gold‐based in situ hybridization with correlative NanoSIMS imaging of incorporated stable isotopes, indicative of metabolic activity.We incubated Kosakonia strain DS‐1‐associated, gnotobiotically grown rice plants with 15N–N2 gas to detect in situ N2 fixation activity. Bacterial cells along the rhizoplane showed heterogeneous patterns of 15N enrichment, ranging from the natural isotope abundance levels up to 12.07 at% 15N (average and median of 3.36 and 2.85 at% 15N, respectively, n = 697 cells).The presented correlative optical and chemical imaging analysis is applicable to a broad range of studies investigating plant–microbe interactions. For example, it enables verification of the in situ metabolic activity of host‐associated commercialized strains or plant growth‐promoting bacteria, thereby disentangling their role in plant nutrition. Such data facilitate the design of plant–microbe combinations for improvement of crop management. [ABSTRACT FROM AUTHOR]

Wheat proteins provide around 20% of all human dietary protein, but their end-use qualities are determined by the form and quantity of nitrogen in the endosperm. In the developed world, there is a heavy reliance in grain production on nitrogen supplied from synthetic fertilisers, and this fertiliser can contribute up to 50% of the on-farm emissions of greenhouse gasses in agriculture. However, despite increasing rates of application of synthetic nitrogen to cereals, wheat grain protein levels, in developed nations, have been frequently failing to reach the premium grade required by the bread-making market. Here, for the first time, we report that biological nitrogen fixation from a new generation of hardseeded annual forage legumes, when grown in rotation with cereal crops, can replace fertiliser N without compromising grain protein. The forage legumes were grown in rotation with Triticum aestivum, and compared with rotations that included a fallow, or a cereal crop at three rainfed sites in Western Australia with differing soil types for 2–4 years. The wheat received low, medium and high rates of urea to indicate if forage legumes can provide sufficient nitrogen for sustainable wheat production. At all sites and years studied, we discovered that cereal grains produced following a year of forage legumes had significantly higher protein levels than when grown as part of a continuous cereal rotation. These results were achieved in combination with a reduction in on-farm emissions (by over 200 kg/ha of CO2) without compromising yield as indicated by emissions accounting. Including appropriate forage legumes in farming systems allows production of low emission intensity grain proteins in dryland farming. [ABSTRACT FROM AUTHOR]

The objective of this work was to evaluate the agronomic performance of a second crop maize grown under different methods of inoculation with Azospirillum brasilense. The experiment was conducted in a randomized block design with 6 treatments and four replications. The treatments used were: no inoculation (T1); inoculation with A. brasilense via seed (T2); inoculation with A. brasilense via sowing furrow (T3); inoculation with A. brasilense via leaf (T4); two inoculations with A. brasilense via leaf (T5); and combination of inoculations via seed, sowing furrow, and leaf (T6). Physiological, morphological, production, and yield components of the second crop maize were evaluated. The data were subjected to analysis of variance at 0.05 probability level and when significant, the means were subjected to the Scott-Knott test at 0.05 probability level, using the R-bio statistical program. Inoculation via seed results in higher nitrate reductase enzyme activity. The inoculation with A. brasilense does not result in gains in grain yield for second crop maize; however, it improves the maize quality by increasing its gross protein contents. [ABSTRACT FROM AUTHOR]

The use of ecological stoichiometry is quite effective for exploring the nutrient dynamics and relationships between plants and soils. However, the way that the plant and soil stoichiometry changes with soil remediation in mining ecosystems remains unclear. Biochar and vermicompost are generally applied to remediate contaminated soil. In this study, a pot experiment was conducted with a mine soil planted with alfalfa. Biochar (B) and vermicompost (V) were added to the soil separately in three different proportions, equivalent to application rates (w/w) of 0% (control, CT), 2.5% (low rate, l), and 5% (high rate, h). This resulted in nine treatments, including control (CT), Bl, Bh, Vl, Vh, BlVl, BlVh, BhVl, and BhVh. The carbon (C), nitrogen (N), and phosphorus (P) concentrations and stoichiometric characteristics of the alfalfa aboveground parts (plant) and soil were investigated. The results showed that biochar application significantly increased the concentrations of soil organic C (SOC), soil total N (TN), soil total P (TP), soil C:N, and plant P concentration, but decreased plant N concentration, and plant C:P and N:P ratios. The effects of vermicompost addition on SOC, soil TN, TP, and stoichiometric characteristics depended on the biochar addition rates, but it increased plant N concentration and N:P, and decreased plant C:N under the condition of low biochar addition. Additionally, the plant N concentration was negatively correlated with soil N and total manganese (Mn) concentrations, whereas there was a positive correlation between plant and soil P concentrations. The soil total and available cadmium (Cd) were positively correlated with plant N concentration but negatively correlated with plant P concentration. The results indicated that the stoichiometric characteristics of plants and soil had diverse responses to biochar and vermicompost additions, and different soil heavy metal elements. Biochar and vermicompost application improved external P and N utilization by plants, respectively. Vermicompost addition enhanced biological N fixation in alfalfa. These findings suggest that vermicompost addition could be an optimal method by which to promote vegetation restoration in mine soils with poor N levels, and that biochar could be applied to low-P soils. The effects of heavy metals on plant and soil stoichiometric characteristics should be taken into consideration. Consequently, this study will provide scientific references for biochar and vermicompost applications in alfalfa planting and management, and vegetation restoration in mining areas. [ABSTRACT FROM AUTHOR]

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Common bean is considered a legume of great socioeconomic importance, capable of establishing symbioses with a wide variety of rhizobial species. However, the legume has also been recognized for its low efficiency in fixing atmospheric nitrogen. Brazil is a hotspot of biodiversity, and in a previous study, we identified 13 strains isolated from common bean (Phaseolus vulgaris) nodules in three biomes of Mato Grosso do Sul state, central-western Brazil, that might represent new phylogenetic groups, deserving further polyphasic characterization. The phylogenetic tree of the 16S rRNA gene split the 13 strains into two large clades, seven in the R. etli and six in the R. tropici clade. The MLSA with four housekeeping genes (glnII, gyrB, recA, and rpoA) confirmed the phylogenetic allocation. Genomic comparisons indicated eight strains in five putative new species and the remaining five as R. phaseoli. The average nucleotide identity (ANI) and digital DNA–DNA hybridization (dDDH) comparing the putative new species and the closest neighbors ranged from 81.84 to 92.50% and 24.0 to 50.7%, respectively. Other phenotypic, genotypic, and symbiotic features were evaluated. Interestingly, some strains of both R. etli and R. tropici clades lost their nodulation capacity. The data support the description of the new species Rhizobium cerradonense sp. nov. (CNPSo 3464T), Rhizobium atlanticum sp. nov. (CNPSo 3490T), Rhizobium aureum sp. nov. (CNPSo 3968T), Rhizobium pantanalense sp. nov. (CNPSo 4039T), and Rhizobium centroccidentale sp. nov. (CNPSo 4062T). [ABSTRACT FROM AUTHOR]

Inoculants containing strains of bacteria that fix atmospheric N that are tolerant or resistant to chemical products commonly used in seed treatment are classified as long-life inoculants, which contribute to improving the efficiency of nodulating bacteria in soybean crops. The objective of this work was to evaluate the effects of applying commonly used pesticides in seed treatment and a long-life inoculant, alone or in combination, on the nodulation process and seed quality of soybeans during storage. The first experiment was carried out in a greenhouse using a completely randomized design, in an 8 × 2 factorial arrangement, with three replications. The treatments consisted of applications of industrial seed treatments: control, MaximAdvanced, Fortenza, long-life inoculant, MaximAdvanced + Fortenza, MaximAdvanced + long-life inoculant, Fortenza + long-life inoculant, and MaximAdvanced + Fortenza + long-life inoculant. The seeds were sown in pots containing soils from two crop areas. The second experiment was carried out in a laboratory, using a completely randomized design in an 8 × 7 factorial arrangement, with four replications. The treatments consisted of the same eight treatments used in the greenhouse experiment, which were applied before packaging and storing the seeds for two months. The industrial seed treatment with the mixture of fungicide, insecticide, and inoculants improved the soybean photosynthesis and nodulation processes, regardless of the history of the area. Soybean seeds can be stored for commercial purposes for up to 40 days, regardless of the seed treatment with fungicide, insecticide, and long-life inoculant applied alone or in combination. [ABSTRACT FROM AUTHOR]

Can farmers in sub-Saharan Africa (SSA) boost crop yields and improve food availability without using more mineral fertilizer? This question has been at the center of lively debates among the civil society, policy-makers, and in academic editorials. Proponents of the "yes" answer have put forward the "input reduction" principle of agroecology, i.e. by relying on agrobiodiversity, recycling and better efficiency, agroecological practices such as the use of legumes and manure can increase crop productivity without the need for more mineral fertilizer. We reviewed decades of scientific literature on nutrient balances in SSA, biological nitrogen fixation of tropical legumes, manure production and use in smallholder farming systems, and the environmental impact of mineral fertilizer. Our analyses show that more mineral fertilizer is needed in SSA for five reasons: (i) the starting point in SSA is that agricultural production is "agroecological" by default, that is, very low mineral fertilizer use, widespread mixed crop-livestock systems and large crop diversity including legumes, but leading to poor soil fertility as a result of widespread soil nutrient mining, (ii) the nitrogen needs of crops cannot be adequately met solely through biological nitrogen fixation by legumes and recycling of animal manure, (iii) other nutrients like phosphorus and potassium need to be replaced continuously, (iv) mineral fertilizers, if used appropriately, cause little harm to the environment, and (v) reducing the use of mineral fertilizers would hamper productivity gains and contribute indirectly to agricultural expansion and to deforestation. Yet, the agroecological principles directly related to soil fertility—recycling, efficiency, diversity—remain key in improving soil health and nutrient-use efficiency, and are critical to sustaining crop productivity in the long run. We argue for a nuanced position that acknowledges the critical need for more mineral fertilizers in SSA, in combination with the use of agroecological practices and adequate policy support. [ABSTRACT FROM AUTHOR]

Abstract Background Legumes can fix atmospheric nitrogen (N) and facilitate N availability to their companion plants in crop mixtures. However, biological nitrogen fixation (BNF) of legumes in intercrops varies largely with the identity of the legume species. The aim of our study was to understand whether BNF and concentration of plant nutrients by common bean is influenced by the identity of the companion plant species in crop mixtures. In this greenhouse pot study, common beans were cultivated with another legume (chickpea) and a cereal (Sorghum). We compared BNF, crop biomass and nutrient assimilation of all plant species grown in monocultures with plants grown in crop mixtures. Results We found beans to exhibit low levels of BNF, and to potentially compete with other species for available soil N in crop mixtures. The BNF of chickpeas however, was enhanced when grown in mixtures. Furthermore, biomass, phosphorous and potassium values of chickpea and Sorghum plants were higher in monocultures, compared to in mixtures with beans; suggesting competitive effects of beans on these plants. Concentration of calcium, magnesium and zinc in beans was higher when grown with chickpeas than with Sorghum. Conclusions It is generally assumed that legumes benefit their companion plant species. Our study highlights the contrary and shows that the specific benefits of cereal-legume mixtures are dependent on the growth rate of the species concerned. We further highlight that the potential of legume-legume mixtures is currently undervalued and may play a strong role in increasing N use efficiency of intercrop-based systems.

Abstract Background Biological nitrogen fixation converting atmospheric dinitrogen to ammonia is an important way to provide nitrogen for plants. Pseudomonas stutzeri DSM4166 is a diazotrophic Gram-negative bacterium isolated from the rhizosphere of cereal Sorghum nutans. Endogenous constitutive promoters are important for engineering of the nitrogen fixation pathway, however, they have not been systematically characterized in DSM4166. Results Twenty-six candidate promoters were identified from DSM4166 by RNA-seq analysis. These 26 promoters were cloned and characterized using the firefly luciferase gene. The strengths of nineteen promoters varied from 100 to 959% of the strength of the gentamicin resistance gene promoter. The strongest P12445 promoter was used to overexpress the biological nitrogen fixation pathway-specific positive regulator gene nifA. The transcription level of nitrogen fixation genes in DSM4166 were significantly increased and the nitrogenase activity was enhanced by 4.1 folds determined by the acetylene reduction method. The nifA overexpressed strain produced 359.1 µM of extracellular ammonium which was 25.6 times higher than that produced by the wild-type strain. Conclusions The endogenous strong constitutive promoters identified in this study will facilitate development of DSM4166 as a microbial cell factory for nitrogen fixation and production of other useful compounds.