Bradyrhizobium Strains Research Articles

Native Bradyrhizobium Strains From Ghana Can Enhance Grain Yields of Field-Grown Cowpea and Groundnut

The existence of large population of ineffective native rhizobia and inconsistent performance of exotic strains in Ghanaian soils necessitate the need to identify effective and locally adapted elite strains for enhanced legume-rhizobium symbiosis. This study was designed to test the suitability of two previously selected potential elite Bradyrhizobium strains for use as inoculants on cowpea (Vigna unguiculata L. Walp) and groundnut (Arachis hypogaea L.) in multi-location experiments. Field experiments were set up at 26 locations (12 planted with cowpea and 14 planted with groundnut) in the Northern region of Ghana. Four treatments were applied at each location: inoculation with Bradyrhizobium strains KNUST 1002 and KNUST 1006, a positive nitrogen (+N) control and a negative control (without nitrogen or inoculation) arranged in randomized complete blocks with four replications. The results showed that inoculation with strains KNUST 1002 and KNUST 1006 promoted significant increases in grain yields of both cowpea and groundnut. On average, inoculating cowpea with strains KNUST 1002 and KNUST 1006 resulted in 63 and 52% increases in grain yield when compared to the negative control. Pod yields of groundnut on the other hand, were significantly (p< 0.05) increased at 57% of the study location with one or both test strains. Responses to inoculation were, however, highly variable across the different study locations (i.e. significant Treatment ⸼ Location Interaction, TLI). A detailed analysis of this significant TLI based on the genotype main effect (G) plus genotype-by-environment (GE) interaction (GGE) biplot analysis revealed that location contributed 71 and 88% of the variation observed in cowpea and groundnut, respectively, and grouped the locations into mega-environments. These results demonstrate that native elite Bradyrhizobium strains KNUST 1002 and KNUST 1006 have potential for use as inoculants to increase cowpea and groundnut production in Northern Ghana.

Soybean seed chemical composition as influenced by Bradyrhizobium inoculation in soils with elevated nickel concentrations

Symbiotic nitrogen-fixing bacteria, rhizobia, besides their intrinsic ability to fix atmospheric nitrogen (N), may influence other element concentrations in leguminous plants. Adequate nickel (Ni) supply is of special importance for growth and N status of soybean, due to the role Ni holds as an enzyme component involved in N metabolism. However, high soil Ni availability also represents risks. Therefore, the objective of the study was to evaluate the effects of rhizobium inoculation on yield and the chemical composition (N, C, S, P, K, Ca, Mg, Cu, Fe, Mn, Zn, Ni) of soybean seeds produced on two soils with elevated total Ni concentrations (59 and 106 mg kg−1 of soil). The genetic characterisation of investigated rhizobial strains and screening for plant growth promoting traits were also performed. The field experiments included 9 treatments where soybean seeds were inoculated with different single rhizobial strains, a treatment with mineral fertilizer application, as well as a non-inoculated control (Ø). Most of the strains belonged to the genus Bradyrhizobium and produce the plant hormone indole-3-acetic acid. Bradyrhizobium inoculations significantly increased the soybean seed yield (up to 125%) and N%, and decreased Ni (up to 48%), P, Fe, Cu and Zn seed concentrations compared to the control Ø, depending on the strain and location. Soybean seed Ni concentrations were significantly influenced by treatment, location and their interaction and varied from 6.28 to 22.25 mg kg−1. There were no significant differences in C, S, Ca, Mg concentrations between treatments. Uptake of all nutrients by seeds in inoculated treatments was generally increased or similar, compared to the control Ø. Concentrations and uptake of Ni in inoculated treatments in both locations were significantly decreased compared to mineral fertilization. The study showed that inoculation with Bradyrhizobium strains effective in nitrogen fixation, together with yield increase may reduce the Ni concentrations in soybean seeds in organic cultivation systems of soybean.

Relationships of Bradyrhizobium strains nodulating three Algerian Genista species

The Mediterranean world is the cradle for the diversification of a large number of plant species, including legumes belonging to the Tribe Genisteae. Nodule bacteria from three species of Genista legumes indigenous to northwestern Africa (G. ferox, G. numidica, G. tricuspidata) were sampled across a 150km region of Algeria in order to investigate symbiotic relationships. Partial 23S rRNA sequences from 107 isolates indicated that Bradyrhizobium was the predominant symbiont genus (96% of isolates), with the remainder belonging to Rhizobium or Mesorhizobium. A multilocus sequence analysis on 46 Bradyrhizobium strains using seven housekeeping (HK) genes showed that strains were differentiated into multiple clades with affinities to seven species: B. canariense (17 isolates), B. japonicum (2), B. ottawaense (2), B. cytisi/B. rifense (9), ‘B. valentinum’ (5), and B. algeriense (11). Extensive discordance between the HK gene phylogeny and a tree for four loci in the symbiosis island (SI) region implied that horizontal transfer of SI loci has been common. Cases of close symbiont relationship across pairs of legumes hosts were evident, with 33% of isolates having as their closest relative a strain sampled from a different Genista species. Nevertheless, tree permutation tests also showed that there was substantial host-related phylogenetic clustering. Thus, each of the three Genista hosts utilized a measurably different array of bacterial lineages.

Multiphase characterization of wild Vigna associated root nodule bacteria from Japanese subtropical islands unveiled novel high temperature resistant Bradyrhizobium strains having high symbiotic compatibility with soybean and mungbean

ABSTRACT Vigna riukiuensis plant – a rare type of vigna, found only in Taiwan and the islands of Okinawa prefecture, Japan – possesses intrinsic property of high level of salt and heat tolerance. To understand the diversity and identify suitable rhizobia, multiphase characterization of root nodule bacteria associated with V. riukiuensis grown in Ishigaki and Iriomote Islands of Okinawa prefecture was performed. Multigene phylogenetic analysis of housekeeping genes based on 16S rRNA gene sequences, 16S-23S rRNA gene internal transcribed spacer (ITS) and 23S rRNA gene sequences identified three main groups closely similar to Bradyrhizobium japonicum, B. elkanii and B. jicamae family. However, analysis of symbiotic nifH and nodD1 genes and their phylogenetic trees showed similar topology, having only few discrepancies in comparison to the housekeeping gene phylogeny. Interestingly, for some of the isolates having similarity with B. elkanii, growth was observed at 40°C, which exceed the highest record for B. elkanii to the best of our knowledge. All the isolates were observed to have the capability of forming root nodules and fix nitrogen in their original host plant V. riukiuensis and two other crops: soybean and mungbean. Most of the isolates showed similar or higher nitrogen-fixing capability in comparison with B. diazoefficiens USDA110 in V. riukiuensis and V. radiata (mungbean), and Iri 5/6 in V. riukiuensis, Iri 5/12 in soybean and Ishi 7/2 in mungbean showed highest acetylene reduction assay (in µmol/h/gm nodule dry weight) activity, which was significantly higher than B. diazoefficiens USDA110. In addition, six isolates attained higher soybean biomass production compared with B. diazoefficiens USDA110, suggesting high symbiotic compatibility with soybean. Among them, Iri 5/7 of B. elkanii group contributed 29% higher soybean biomass production than B. diazoefficiens USDA110 and could grow at 40°C, hence it could be a promising soybean inoculant in the tropics. Abbreviations ARA: acetylene reduction assay, ISR: intergenic spacer regions, ITS: internal transcribed spacer

English

Effective nodulation of legume is usually solid carriers dependent. Peat, the most effective solid carrier is not readily available in Nigeria, hence, alternative solid carriers were evaluated with Bradyrhizobium strains using cowpea as a test crop. Mutant strains (USDA 3384 and USDA 3451) were evaluated in the laboratory using solid carriers and pot experiments. The survival (multiplication), infectivity and nodulating ability of the Bradyrhizobium mutant strains using cowpea as a test crop were assayed in peat, cowdung and composted maize cob. Mutant strains inoculated in the three solid carriers got to their peak of growth at 16th day of incubation. Peat significantly (p<0.05) increased the population of both USDA 3384 and USDA 3451 compared to cowdung and composted maize cob. USDA 3451 inoculated in cowdung significantly (p<0.05) increased multiplication of Bradyrhizobium mutant strain and nodulation of cowpea. Investigation shows it can be used as peat substitute when preparing inoculant. USDA 3384 inoculated in both composted maize cob and cowdung, respectively had consistent and significant positive effect on nodulation of cowpea and nodule dry weight. These results reveal their ability to subtitute peat. Hence, blind use of alternative solid carriers for inoculant preparations may hinder high infectivity and optimal nodulation which could facilitate positive effects of nitrogen fixation in low N tropical soil. Key words: Solid carriers, Bradyrhizobium,survival, infectivity, nodulation.

Genomic insight into the origins and evolution of symbiosis genes in Phaseolus vulgaris microsymbionts

BackgroundPhaseolus vulgaris (common bean) microsymbionts belonging to the bacterial genera Rhizobium, Bradyrhizobium, and Ensifer (Sinorhizobium) have been isolated across the globe. Individual symbiosis genes (e.g., nodC) of these rhizobia can be different within each genus and among distinct genera. Little information is available about the symbiotic structure of indigenous Rhizobium strains nodulating introduced bean plants or the emergence of a symbiotic ability to associate with bean plants in Bradyrhizobium and Ensifer strains. Here, we sequenced the genomes of 29 representative bean microsymbionts (21 Rhizobium, four Ensifer, and four Bradyrhizobium) and compared them with closely related reference strains to estimate the origins of symbiosis genes among these Chinese bean microsymbionts.ResultsComparative genomics demonstrated horizontal gene transfer exclusively at the plasmid level, leading to expanded diversity of bean-nodulating Rhizobium strains. Analysis of vertically transferred genes uncovered 191 (out of the 2654) single-copy core genes with phylogenies strictly consistent with the taxonomic status of bacterial species, but none were found on symbiosis plasmids. A common symbiotic region was wholly conserved within the Rhizobium genus yet different from those of the other two genera. A single strain of Ensifer and two Bradyrhizobium strains shared similar gene content with soybean microsymbionts in both chromosomes and symbiotic regions.ConclusionsThe 19 native bean Rhizobium microsymbionts were assigned to four defined species and six putative novel species. The symbiosis genes of R. phaseoli, R. sophoriradicis, and R. esperanzae strains that originated from Mexican bean-nodulating strains were possibly introduced alongside bean seeds. R. anhuiense strains displayed distinct host ranges, indicating transition into bean microsymbionts. Among the six putative novel species exclusive to China, horizontal transfer of symbiosis genes suggested symbiosis with other indigenous legumes and loss of originally symbiotic regions or non-symbionts before the introduction of common bean into China. Genome data for Ensifer and Bradyrhizobium strains indicated symbiotic compatibility between microsymbionts of common bean and other hosts such as soybean.

Colonization by multi-potential Pseudomonas aeruginosa P4 stimulates peanut (Arachis hypogaea L.) growth, defence physiology and root system functioning to benefit the root-rhizobacterial interface

The beneficial associations between Arachis hypogaea L. (peanut) and fluorescent Pseudomonas species have been poorly explored despite their predominance in the peanut rhizosphere. The present study explores the mutually beneficial interactions between peanut roots and P. aeruginosa P4 (P4) in terms of their impact on plant growth, defence physiology and the root-rhizobacterial interface. The efficient phosphate solubilizer P4 exhibited biocontrol abilities, including the production of siderophores, pyocyanin, indole-3-acetic acid and hydrogen cyanide. The bacterization of peanut seeds with multi-potential P4 significantly enhanced in vitro seed germination and seedling vigour. Under sand-based gnotobiotic (10 days post-inoculation) and sterile soil-based cultivation systems (30 days post-inoculation), sustained P4 colonization enhanced the peanut root length and dry plant biomass. The subsequent increase in catalase, polyphenol oxidase and phenylalanine ammonia lyase activities with increased phenolic contents in the peanut roots and shoots suggested the systemic priming of defences. Consequently, the altered root exudate composition caused enhanced chemo-attraction towards P4 itself and the symbiotic N2-fixing Bradyrhizobium strain. Co-inoculating peanuts with P4 and Bradyrhizobium confirmed the improved total bacterial colonization (∼2 fold) of the root tip, with the successful co-localization of both, as substantiated by scanning electron microscopy. Collectively, the peanut-P4 association could potentially model the beneficial Pseudomonas-driven multi-trophic rhizosphere benefits, emphasizing the plausible role of non-rhizobium PGPR in promoting N2 fixation.

The intensity of ethylene release by soybean plants under the influence of fungicides in the early stages of legume-rhizobial symbiosis

The effect of pre-sowing treatment of soybean seeds with fungicides on the intensity of ethylene release, the processes of nodulation and nitrogen fixation in different symbiotic systems in the early stages of ontogenesis were investigated. The objects of the study were selected symbiotic systems formed with the participation of soybean (Glycine max (L.) Merr.) Diamond variety, strains Bradyrhizobium japonicum 634b (active, virulent) and 604k (inactive, highly virulent) and fungicides Maxim XL 035 PS (fludioxonil, 25 g/L, metalaxyl, 10 g/L), and Standak Top (fipronil, 250 g/L, thiophanate methyl, 225 g/L, piraclostrobin, 25 g/L). Before sowing, the seeds of soybean were treated with solutions of fungicides, calculated on the basis of one rate of expenditure of the active substance of each preparation indicated by the producer per ton of seed. One part of the seeds treated with fungicides was inoculated with rhizobium culture for 1 h (the titre of bacteria was 107 cells/mL). To conduct the research we used microbiological, physiological, biochemical methods, gas chromatography and spectrophotometry. It is found that, regardless of the effectiveness of soybean rhizobial symbiosis, the highest level of ethylene release by plants was observed in the stages of primordial leaf and first true leaf. This is due to the initial processes of nodulation – the laying of nodule primordia and the active formation of nodules on the roots of soybeans. The results show that with the participation of fungicides in different symbiotic systems, there are characteristic changes in phytohormone synthesis in the primordial leaf stage, when the nodule primordia are planted on the root system of plants. In particular, in the ineffective symbiotic system, the intensity of phytohormone release decreases, while in the effective symbiotic system it increases. At the same time, a decrease in the number of nodules on soybean roots inoculated with an inactive highly virulent rhizobia 604k strain due to the action of fungicides and an increase in their number in variants with co-treatment of fungicides and active virulent strain 634b into the stage of the second true leaf were revealed. It was shown that despite a decrease in the mass of root nodules, there is an increase in their nitrogen-fixing activity in an effective symbiotic system with the participation of fungicides in the stage of the second true leaf. The highest intensity of ethylene release in both symbiotic systems was recorded in the stage of the first true leaf, which decreased in the stage of the second true leaf and was independent of the nature of the action of the active substances of fungicides. The obtained data prove that the action of fungicides changes the synthesis of ethylene by soybean plants, as well as the processes of nodulation and nitrogen fixation, which depend on the efficiency of the formed soybean-rhizobial systems and their ability to realize their symbiotic potential under appropriate growing conditions.

Bradyrhizobium uaiense sp. nov., a new highly efficient cowpea symbiont.

This study describes two Bradyrhizobium strains, UFLA03-164T and UFLA03-153, which share more than 99% sequence similarity of the 16S rRNA with the type strains of 15 species in this genus. The concatenation of three housekeeping genes (recA, gyrB, and dnaK) indicated that both strains formed a single clade separate from known Bradyrhizobium species. B. viridifuturi, represented by SEMIA 690T, is the closest neighboring species (96.2%). Low (< 92%) average nucleotide identity (ANI) was observed between strain UFLA03-164T and any of the closest species on the phylogenetic trees based on concatenated housekeeping genes. The DNA G+C content of UFLA03-164T is 63.25%. Phenotypic characteristics were determined for both UFLA strains. Based on the data, the two strains represent a new species for which the name Bradyrhizobium uaiense is proposed, with UFLA03-164T (= LMG 31509T) as type strain.

Isolation, Characterization, and Complete Genome Sequence of a Bradyrhizobium Strain Lb8 From Nodules of Peanut Utilizing Crack Entry Infection.

In many legumes, the colonization of roots by rhizobia is via “root hair entry” and its molecular mechanisms have been extensively studied. However, the nodulation of peanuts (Arachis hypogaea L.) by Bradyrhizobium strains requires an intercellular colonization process called “crack entry,” which is understudied. To understand the intercellular crack entry process, it is critical to develop the tools and resources related to the rhizobium in addition to focus on investigating the mechanisms of the plant host. In this study, we isolated a Bradyrhizobium sp. strain, Lb8 from peanut root nodules and sequenced it using PacBio long reads. The complete genome sequence was a circular chromosome of 8,718,147 base-pair (bp) with an average GC content of 63.14%. No plasmid sequence was detected in the sequenced DNA sample. A total of 8,433 potential protein-encoding genes, one rRNA cluster, and 51 tRNA genes were annotated. Fifty-eight percent of the predicted genes showed similarity to genes of known functions and were classified into 27 subsystems representing various biological processes. The genome shared 92% of the gene families with B. diazoefficens USDA 110T. A presumptive symbiosis island of 778 Kb was detected, which included two clusters of nif and nod genes. A total of 711 putative protein-encoding genes were in this region, among which 455 genes have potential functions related to symbiotic nitrogen fixation and DNA transmission. Of 21 genes annotated as transposase, 16 were located in the symbiosis island. Lb8 possessed both Type III and Type IV protein secretion systems, and our work elucidated the association of flagellar Type III secretion systems in bradyrhizobia. These observations suggested that complex rearrangement, such as horizontal transfer and insertion of different DNA elements, might be responsible for the plasticity of the Bradyrhizobium genome.

Anthyllis vulneraria and Lotus corniculatus on calamine heaps form nodules with Bradyrhizobium liaoningense-related strains harboring novel in Europe symbiotic nifD haplotypes

Calamine tailings, a habitat known for high concentrations of zinc and lead, are inhabited by a limited number of plants. The objective of this project was a phylogenetic characterization of Bradyrhizobium strains that form effective symbiosis with Anthyllis vulneraria L. and Lotus corniculatus L. (Fabaceae) spontaneously growing on old calamine tailings in Southern Poland using a multilocus sequence analysis (MLSA) approach based on amplification of selected housekeeping dnaK, glnII, gyrB, recA, and rpoB, and symbiotic nifD genes. The isolates were also tested for their metal tolerance and nitrogenase activity. The analyses have shown enhanced growth of Bradyrhizobium strains in the presence of lead (Pb) or zinc (Zn), unlike uninoculated plants which died within three weeks. In housekeeping gene trees, all heavy metal-resistant bradyrhizobia clustered close to B. liaoningense, a species which until now has not been recorded among Bradyrhizobium symbionts of Loteae tribe. In nifD gene tree, Bradyrhizobium isolates clustered within a group referred to as Clade XVII, encompassing Fabaceae symbionts that originate from different geographical areas but also non-nodulating Bradyrhizobium diazotrophs. Summing up, this study revealed that in these heavy metal-contaminated soils L. corniculatus and A. vulneraria are infected by Bradyrhizobium strains harboring symbiotic nifD gene haplotypes phylogenetically unrelated to Clade II strains that predominate in Europe among bradyrhizobia nodulating Genisteae and Loteae spp.

Characterization of Bradyrhizobium strains indigenous to Western Australia and South Africa indicates remarkable genetic diversity and reveals putative new species

Bradyrhizobium are N2-fixing microsymbionts of legumes with relevant applications in agricultural sustainability, and we investigated the phylogenetic relationships of conserved and symbiotic genes of 21 bradyrhizobial strains. The study included strains from Western Australia (WA), isolated from nodules of Glycine spp. the country is one genetic center for the genus and from nodules of other indigenous legumes grown in WA, and strains isolated from forage Glycine sp. grown in South Africa. The 16S rRNA phylogeny divided the strains in two superclades, of B. japonicum and B. elkanii, but with low discrimination among the species. The multilocus sequence analysis (MLSA) with four protein-coding housekeeping genes (dnaK, glnII, gyrB and recA) pointed out seven groups as putative new species, two within the B. japonicum, and five within the B. elkanii superclades. The remaining eleven strains showed higher similarity with six species, B. lupini, B. liaoningense, B. yuanmingense, B. subterraneum, B. brasilense and B. retamae. Phylogenetic analysis of the nodC symbiotic gene clustered 13 strains in three different symbiovars (sv. vignae, sv. genistearum and sv. retamae), while seven others might compose new symbiovars. The genetic profiles of the strains evaluated by BOX-PCR revealed high intra- and interspecific diversity. The results point out the high level of diversity still to be explored within the Bradyrhizobium genus, and further studies might confirm new species and symbiovars.

Temporal biological nitrogen fixation pattern in soybean inoculated with Bradyrhizobium

AbstractBiological nitrogen fixation (BNF) in soybean [Glycine max (L.) Merr.] plays an important role in sustainable agriculture, reducing the limitations associated with other sources of N such as fertilizers and soils. Our major objective was to evaluate the weekly pattern of BNF in soybean influenced by nodule formation, using three different laboratory‐cultured Bradyrhizobium strains. Plants were grown in 1‐m polyvinyl chloride (PVC) columns for 17 week in a greenhouse and BNF was determined using an integrated approach by assessing nodule formation, stem ureide‐N, and N partitioning in plant parts. Bradyrhizobium strains showed overall similar plant performance and N2−fixation capacity. During the beginning of flowering/full bloom (R1/R2) growth stages, nodule formation significantly increased and reached a maximum at pod‐filling (R4) stage. Stem ureide‐N was detected at early growth stages even with fewer small nodules, which significantly increased after the beginning of pod formation (R3). Peak N2−fixing rate (g N kg−1 d−1) started to decline after the onset of seed filling (R5.5). Relationship between BNF and nodule parameters (nodule number [R2 = .65] and nodule weight [R2 = .62]) suggested that they can be used as predictors of BNF.

Application of rice endophytic Bradyrhizobium strain SUTN9-2 containing modified ACC deaminase to rice cultivation under water deficit conditions

ABSTRACT Bacterial 1-aminocyclopropane-1-carboxylate (ACC) deaminase is a key factor for alleviating the plant ethylene biosynthesis, which is induced by stress. The ACC deaminase-improved strains of the rice endophytic Bradyrhizobium sp. SUTN9-2, SUTN9-2 (ACCDadap) and SUTN9-2:pMG103::acdRS, exhibit 1.4- and 8.9-fold higher ACC deaminase activity than the wild type, respectively (Sarapat S, Songwattana P, Longtonglang A, Umnajkitikorn K, Girdthai T, Tittabutr P, Boonkerd N, Teaumroong N. 2020. Effects of Increased 1-Aminocyclopropane-1-Carboxylate (ACC) Deaminase Activity in Bradyrhizobium sp. SUTN9-2 on Mung Bean Symbiosis under Water Deficit Conditions. Microbes Environ. 35). The effects of these on rice growth under water deficit conditions were evaluated. The bacterial inoculations reduced ethylene synthesis, leading to a reduction in membrane destruction and the chlorophyll content of rice. Furthermore, the bacterial inoculations improved the leaf relative water content, survival, recovery rates, and improved the crop yield in field conditions. Therefore, the water deficit tolerance of rice was improved by controlling ethylene biosynthesis by improving ACC deaminase activity with endophytic SUTN9-2. Moreover, the SUTN9-2 (ACCDadap) strain can be used as a bio-inoculant under field conditions to enhance rice growth, grain yield, and enhance drought tolerance.

Phylogenetic Diversity and Evaluation the Effectiveness of Indigenous &amp;lt;i&amp;gt;Bradyrhizobium&amp;lt;/i&amp;gt; Strains for Myanmar Black Gram (&amp;lt;i&amp;gt;Vigna mungo&amp;lt;/i&amp;gt; L. Hepper) Cultivars

Black gram (Vigna mungo L. Hepper) is one of the main leguminous crops that provide chief source of food. Several Bradyrhizobium species are able to induce effective nodules in black gram cultivars. In the present study, we characterized forty isolates of indigenous black gram bradyrhizobia from Myanmar based on the sequence analysis of the bacterial 16S rRNA gene. The sequence analysis confirmed that all isolates were categorized and identified as the genus Bradyrhizobium and they were conspecific with B. elkanii, B. sp., B. liaoningense, B. japonicum and B. yunamingense. Almost all the collected isolates from major black gram growing regions of Nyaunglebin Bago Regio, Chaungzon Mon State, Sittwe Rakhine State, Danubyu Ayeyarwady Region and Launglon Tanintharyi Region were identified as B. liaoningense. At Danubyu Ayeyarwady Region and Pyinmanar Nay Pyi Taw Region, most of the strains were identified as B. japonicum. On the other hand, more or less all the isolates from Launglon Tanintharyi Region and Hpa-an Kayin State were related to B. elkanii. However, all B. sp. strains were found in Salingyi Sagaing Region black gram growing region. This is the first report describing Bradyrhizobium strains that were isolated from soil samples of major black gram growing areas in Myanmar. Evaluation of the effectiveness of Myanmar Bradyrhizobim strains isolated from soil samples of major black gram growing areas of Myanmar for plant growth and nitrogen fixation was studied in pot experiments with completely randomized design and three replicates. The nodule dry weight, shoot dry weight and acetylene reduction activity of the plant inoculated with Bradyrhizobium elkanii LauBG38 were significantly higher in ARA per plant, nodule and shoot dry weights than the other tested isolates in both Yezin-4 and Yezin-7 black gram varieties. We expect that Myanmar Bradyrhizobium elkanii LauBG38 will be able to use as Biofertilizer for black gram cultivars.

English

Soybean is an important crop in the Democratic Republic of Congo, a country faced with high levels of war induced malnutrition but its productivity is limited by poor soil fertility coupled with low access to nitrogen mineral fertilizers. Commercial rhizobia strains introduced in 2010 failed to adapt and increase soybeans yields at desired level. We studied the performance of six indigenous rhizobia strains in enhancing soybean productivity compared to two commercial strains USDA110 and SEMIA5019. The study was carried out in the greenhouse and field of Kalambo station of International Institute of Tropical Agriculture (IITA), D. R. Congo during 2016/2017 cropping season. The treatments included: (1) N-, control without inoculation and N-fertilizer; (2) N+, non-inoculated control with 80 kg of N ha−1; and inoculated with (3) commercial strain Bradyrhizobium diazoefficiens USDA110; (4) commercial strain Bradyrhizobium elkanii SEMIA5019; (5) local strains Bradyrhizobium japonicum NAC17; (6) NAC22; (7) NAC37, (8) NAC42 (9) NAC 46 and (10) NAC78. Greenhouse and field experiments were laid out as completely randomized design and randomized complete block design respectively. The best inoculation treatments across all experiments were the indigenous strains NAC46 and NAC17 which nodulated equally or better than the commercial strain USDA 110. In the field NAC46 and NAC17 increased soybean grain yield from 2.4 to 3.3 t ha-1 and 3.4 t ha-1; indicating the increase of 68.7 and 70.8% respectively, over the commercial strain USDA110. The results demonstrated that indigenous rhizobia NAC46 and NAC17 would thus be the silver bullet to enhanced BNF and soybean yields in South Kivu province of Democratic Republic of Congo. Key words: Inoculation, local rhizobia; soil fertility, USDA110.

Are Cowpea-Nodulating Bradyrhizobial Communities Influenced by Biochar Amendments in Soils? Genetic Diversity and Symbiotic Effectiveness Assessment of Two Agricultural Soils of Brazilian Drylands

The objective of this study was to evaluate the influence of biochar amendment on the bacterial genetic diversity and symbiosis in cowpea-nodulating rhizobial communities in two agricultural soils of Brazilian drylands. Vertisol and Ultisol surface layer soil samples were collected in the Brazilian semiarid region. The soils were mixed with biochar and cowpea seeds (BRS Marataoa) and sown in pots. Forty-five days after plant emergence, the nodule bacteria were isolated. The genetic variability of the nodule bacteria was evaluated by means of restriction fragment length polymorphism of 16S-23S rRNA intergenic spacer region (IGS-RFLP), and those that shared identical IGS-RFLP profiles were fingerprinted by BOX-PCR. By selecting of representative isolates in the genetic clusters, 27 bacteria were identified by 16S-23S IGS sequencing and symbiotically assessed. The IGS-RFLP dendrogram of the 73 isolates had 20 clusters; the multivariate correspondence analysis grouped both Vertisol treatments, and the Ultisol treatments were placed far away in the biplot. Ultisol showed higher bradyrhizobial diversity than the Vertisol. The fingerprinting indicated the presence of only three clonal colonies, showing high intraspecific diversity. The 16S-23S IGS sequences indicated the prevalence of bacteria related to Bradyrhizobium guangxiense in the Vertisol and Bradyrhizobium zhanjiangense in the Ultisol, in addition to the presence of three putative Microvirga spp. in the Ultisol. The symbiotic efficiency of 22 out of 27 bacteria was comparable with that of the inoculant strain Bradyrhizobium yuanmingense BR 3267. The genetic diversity of the cowpea-nodulating bradyrhizobia was not primarily affected by the biochar application in two agricultural soils of Brazilian drylands. Both soils harbored symbiotically efficient bacteria, without the influence of biochar amendment .

Host Range and Symbiotic Effectiveness of N2O Reducing Bradyrhizobium Strains.

Emissions of the potent greenhouse gas N2O is one of the environmental problems associated with intensive use of synthetic N fertilizers, and novel N2O mitigation strategies are needed to minimize fertilizer applications and N2O release without affecting agricultural efficiencies. Increased incorporation of legume crops in agricultural practices offers a sustainable alternative. Legumes, in their symbiosis with nitrogen fixing bacteria, rhizobia, reduce the need for fertilizers and also respond to the need for increased production of plant-based proteins. Not all combinations of rhizobia and legumes result in efficient nitrogen fixation, and legume crops therefore often need to be inoculated with compatible rhizobial strains. Recent research has demonstrated that some rhizobia are also very efficient N2O reducers. Several nutritionally and economically important legumes form root nodules in symbiosis with bacteria belonging to Bradyrhizobium. Here, the host-ranges of fourteen N2O reducing Bradyrhizobium strains were tested on six legume hosts; cowpea, groundnut, mung bean, haricot bean, soybean, and alfalfa. The plants were grown for 35 days in pots in sterile sand supplemented with N-free nutrient solution. Cowpea was the most promiscuous host nodulated by all test strains, followed by groundnut (11 strains) and mungbean (4 strains). Three test strains were able to nodulate all these three legumes, while none nodulated the other three hosts. For cowpea, five strains increased the shoot dry weight and ten strains the shoot nitrogen content (pairwise comparison; p < 0.05). For groundnut the corresponding results were three and nine strains. The symbiotic effectiveness for the different strains ranged from 45 to 98% in cowpea and 34 to 95% in groundnut, relative to fertilized controls. The N2O reduction capacity of detached nodules from cowpea plants inoculated with one of these strains confirmed active N2O reduction inside the nodules. When released from senescent nodules such strains are expected to also act as sinks for N2O produced by denitrifying organisms in the soil microbial community. Our strategy to search among known N2O-reducing Bradyrhizobium strains for their N2-fixation effectiveness successfully identified several strains which can potentially be used for the production of legume inoculants with the dual capacities of efficacious N2-fixation and N2O reduction.

Inoculation with native Bradyrhizobium strains formulated with biochar as carrier improves the performance of pigeonpea (Cajanus cajan L.)

Abstract Pigeonpea (Cajanus cajan) originated in India and is now a major source of protein in countries in tropical and subtropical regions worldwide. However, little is known about the effect on crop yields of inoculation with appropriate rhizobia strains. The objective of this study was to explore the possibility of improving pigeonpea yields in the Caribbean region, by inoculating with selected N-fixing symbionts. To achieve the study objective, we selected three elite strains of the genus Bradyrhizobium isolated from root nodules of pigeonpea in the Dominican Republic, we formulated them using innovative carriers, and finally we tested the formulated inoculant on eight farms located in the Dominican Republic. Three different carriers (sewage sludge, pine bark biochar and poultry litter) plus perlite as control were tested for the formulation. The first step was to test bacterial survival in the different carriers at different times up to one year (shelf-life). Pine bark biochar and the control resulted in the best shelf-life and were selected for the field experiment. The experimental design comprised two pigeonpea cultivars and five fertilisation strategies: Inoculation with three Bradyrhizobium strains without nitrogen (N) fertilisation, plus two non-inoculated controls one of them fertilised with 125 and 210 kg N ha−1 in non-irrigated and irrigated fields respectively. Nodule occupancy by the inoculated bacterial strain depended solely on the level of soil native nodulating bacteria and not on the bacterial strain or the carrier. Inoculation produced, on average, a significant yield increase compared to the non-inoculated non-N-fertilised control. Furthermore, the yield with the inoculated treatments did not significantly differ from the yield of the N-fertilised control. However, either inoculation or N-fertilisation were ineffective in two out of the eight fields with more than 104 nodulating bacteria g soil−1, compared to the non-inoculated non-N-fertilised control. Yield was neither significantly affected by the bacterial strain nor by the cultivar-strain interaction, and therefore, the two cultivars can be considered promiscuous. Neither carrier had a significant effect on yield. Thus, inoculants based on strains of Bradyrhizobia isolated from the root nodules of pigeonpea, formulated with pine bark biochar or perlite as carrier, are recommended as a good strategy to ensure food security in tropical agroecosystems with pigeonpea.

Genetic analysis for biological nitrogen fixation (BNF) in cowpea

This work aimed to estimate the genetic parameters of biological nitrogen fixation (BNF) in cowpea to guide the development of efficient cultivars in N2 fixation. Inoculation was performed with a mix of three Bradyrhizobium strains. Two soil inoculations were performed with 1mL of the inoculum, at sowing and at five days after seedlings emergence. Plants were harvested and evaluated at 40 days after sowing. The means and variances of parents F1 and F2 and the BCs IC-1 (inefficient) × BRS Marataoã (efficient) (cross 1) and IC-1 × BRS Pujante (efficient) (cross 2) were analyzed. Segregation analyses for efficient and non-efficient plants indicate that BNF is controlled by two genes with dominant-recessive epistatic effect. Narrow-sense heritability estimates for crosses 1 and 2 were, respectively, 90% and 78% for shoot dry matter (SDM); 78% and 58%, for number of nodules/plant (NN); 86% and 58%, for nodule fresh matter (NFM); and 85% and 71%, for accumulated nitrogen (AN). Gene number estimates for SDM, NN, NFM, AN ranged from two to nine by the Mather and Jinks’ method, and from one to two, by the Lande’s method, in both crosses. SDM x NFM and NFM x AN had significant and positive correlations, while NN showed significant negative correlation with AN and SDM. BNF efficiency in cowpea can be easily achieved using autogamous breeding methods.