Bradyrhizobium Yuanmingense Research Articles

Isolation and characterization of mung bean (Vigna radiata L.) rhizobia in Myanmar

We collected soil samples from six major mung bean cropping regions in Myanmar: Sagaing, Mandalay, Nay Pyi Taw, and Magway in the tropical savanna climate zone and Bago and Yangon in the tropical monsoon climate zone. All fields grew mung bean for at least 5 years and had no history of rhizobial inoculation. Mung bean ‘Yezin-11’, a popular cultivar in Myanmar, was inoculated with soil suspensions. From the nodules formed on the roots, we isolated 55 rhizobial strains. Identification of the isolates revealed the dominant species of indigenous rhizobia in each region. We identified 53 Bradyrhizobium strains and 2 Ensifer strains. Bradyrhizobium yuanmingense was dominant in the tropical savanna zone and Bradyrhizobium sp. (B. liaoningense or B. diversitatis) and B. centrosematis were dominant in the tropical monsoon zone. Principal component analysis indicates that the dominance of B. yuanmingense in the tropical savanna zone might be due to high concentration of NO3-N and P2O5 in the soil. It also indicates that the dominance of B. centrosematis in the tropical monsoon zone might be caused by drastically low pH and high concentration of NH4 in the soil. Bradyrhizobium centrosematis YGN-M9, B. yuanmingense SGG-M3, and Bradyrhizobium sp. BGO-M5 significantly increased nodulation (nodule number and nodule dry weight), acetylene reduction activity, and shoot dry weight, respectively, relative to Ensifer terangae MDY-M6. Co-inoculation with these three strains increased nodulation significantly compared with single inoculation of BGO-M5. The characterization of mung bean rhizobia and selection of microbial inoculant candidates will be useful for the development of microbial inoculants in Myanmar.

Synergistic application of Bradyrhizobium yuanmingense and Trichoderma species improves tolerance of cowpea to aluminum stress by enhancing the antioxidant defense system

The positive effects of Trichoderma on the antioxidant responses of nodulated cowpea plants under aluminum (Al) stress are not well documented. This study aims to evaluate the effects of different microbial consortia of Bradyrhizobium yuanmingense (bradyrhizobia) and Trichoderma species (T. asperelloides T02, T. asperellum T77, or T. harzianum T78) on the growth and antioxidant responses of cowpea under Al stress. Coinoculation with Bradyrhizobia and Trichoderma resulted in greater growth (plant height and root length) and biomass (dry weight of shoots and roots) compared to single inoculations. The dual application of bradyrhizobia and T. asperellum T77 to Al-treated plants increased chlorophyll-a (36 %), chlorophyll-b (210 %), and carotenoid (92 %) contents. Under Al stress, plants inoculated with bradyrhizobia exhibited a 53 % increase in anthocyanin content compared to the unstressed control. The highest levels of reduced ascorbate and ascorbate redox state were observed in plants coinoculated with bradyrhizobia and Trichoderma, whereas oxidized ascorbate accumulated more in plants with single inoculations (bradyrhizobia or Trichoderma). Lipid peroxidation and hydrogen peroxide levels were 50 % lower in plants coinoculated with bradyrhizobia and Trichoderma. Additionally, these plants showed increased activities of superoxide dismutase, catalase, ascorbate peroxidase, and peroxidases under Al stress, particularly in those coinoculated with bradyrhizobia and T. asperelloides T02 and T. harzianum T78. Our results confirm that the presence of bradyrhizobia and Trichoderma in the cowpea rhizosphere mitigates oxidative damage caused by reactive oxygen species under Al stress and positively influences cowpea growth and development under stressful conditions.

Genetic diversity assessment and selection of Bradyrhizobium strains for Inceptisols based on symbiotic performance

The symbiotic association of pigeonpea (Cajanus cajan (L.) Millsp.) with rhizobia plays a pivotal role in plants’ growth and development. The present study was carried out at Institute of Agricultural Sciences, Banaras Hindu University, Varanasi, Uttar Pradesh during 2018–2020 to explore the diversity of rhizobia present in the Cajanus cajan root under Inceptisols and identify an elite strain that exhibits exceptional plant growth promotion and of nitrogenase activity for efficient nitrogen fixation. The bacterial identification using 16s rDNA sequencing revealed bacteria strains Bradyrhizobium japonicum (S12), Bradyrhizobium subterraneum (S1, S7, S8, S10, S13), Bradyrhizobium yuanmingense (S2, S3, S4, S6, S9, S11) while two bacteria were the endophytes identified as Pseudomonas azotoformans (S5) and Paenibacillus seodonensis (S15). Genetic diversity using UPGMA clustering revealed remarkable genetic variations, classifying 14 strains into three primary clusters and four secondary clusters. Strains S1, S3, S6, S9, and S13 exhibited elevated nitrogenase activity, indicative of their proficient nitrogen fixation capability. In plant growth experiments, all strains demonstrated significant growth of the plants compared to control. The notable growth enhancement by these five strains can be attributed to their remarkable efficiency in nitrogen fixation, as indicated by significantly higher nodule formation and nitrogenase activity. These findings provide valuable insights for identifying potential rhizobial inoculants to enhance pigeonpea productivity.

Two novel symbiovars of Bradyrhizobium yuanmingense, americaense and caribense, the symbiovar tropici of Bradyrhizobium pachyrhizi and the symbiovar cajani of Bradyrhizobium cajani are microsymbionts of the legume Cajanus cajan in Dominican Republic

Cajanus cajan L. (guandul) is commonly cultivated in Dominican Republic where this legume is a subsistence crop. Here we identified through MALDI-TOF MS several rhizobial strains nodulating C. cajan in two Dominican locations as Bradyrhizobium yuanmingense. The phylogenetic analysis of recA and glnII housekeeping genes showed that these strains belong to a wide cluster together with the type strain of B. yuanmingense and other C. cajan nodulating strains previously isolated in Dominican Republic. The comparison of genomes from strains representative of different lineages within this cluster support the existence of several genospecies within B. yuanmingense, which is the major microsymbiont of C. cajan in Dominican Republic where it is also nodulated by Bradyrhizobium cajani and Bradyrhizobium pachyrhizi. The analysis of the symbiotic nodC gene showed that the C. cajan nodulating strains from the B. yuanmingense complex belong to two clusters with less than 90% similarity between them. The strains from these two clusters showed nodC gene similarity values lower than 90% with respect to the remaining Bradyrhizobium symbiovars and then they correspond to two new symbiovars for which we propose the names americaense and caribense. The results of the nodC gene analysis also showed that C. cajan is nodulated by the symbiovar tropici, which has been found by first time in this work within the species Bradyrhizobium pachyrhizi. These results confirmed the high promiscuity degree of C. cajan, which is also nodulated by the symbiovar cajani of Bradyrhizobium cajani in Dominican Republic.

Genetic and Physiological Characterization of Soybean-Nodule-Derived Isolates from Bangladeshi Soils Revealed Diverse Array of Bacteria with Potential Bradyrhizobia for Biofertilizers.

Genetic and physiological characterization of bacteria derived from nodules of leguminous plants in the exploration of biofertilizer is of paramount importance from agricultural and environmental perspectives. Phylogenetic analysis of the 16S rRNA gene of 84 isolates derived from Bangladeshi soils revealed an unpredictably diverse array of nodule-forming and endosymbiotic bacteria-mostly belonging to the genus Bradyrhizobium. A sequence analysis of the symbiotic genes (nifH and nodD1) revealed similarities with the 16S rRNA gene tree, with few discrepancies. A phylogenetic analysis of the partial rrn operon (16S-ITS-23S) and multi-locus sequence analysis of atpD, glnII, and gyrB identified that the Bradyrhizobium isolates belonged to Bradyrhizobium diazoefficiens, Bradyrhizobium elkanii, Bradyrhizobium liaoningense and Bradyrhizobium yuanmingense species. In the pot experiment, several isolates showed better activity than B. diazoefficiens USDA110, and the Bho-P2-B2-S1-51 isolate of B. liaoningense showed significantly higher acetylene reduction activity in both Glycine max cv. Enrei and Binasoybean-3 varieties and biomass production increased by 9% in the Binasoybean-3 variety. Tha-P2-B1-S1-68 isolate of B. diazoefficiens significantly enhanced shoot length and induced 10% biomass production in Binasoybean-3. These isolates grew at 1-4% NaCl concentration and pH 4.5-10 and survived at 45 °C, making the isolates potential candidates for eco-friendly soybean biofertilizers in salty and tropical regions.

Genomic Diversity of Pigeon Pea (Cajanus cajan L. Millsp.) Endosymbionts in India and Selection of Potential Strains for Use as Agricultural Inoculants.

Pigeon pea (Cajanus cajan L. Millsp. ) is a legume crop resilient to climate change due to its tolerance to drought. It is grown by millions of resource-poor farmers in semiarid and tropical subregions of Asia and Africa and is a major contributor to their nutritional food security. Pigeon pea is the sixth most important legume in the world, with India contributing more than 70% of the total production and harbouring a wide variety of cultivars. Nevertheless, the low yield of pigeon pea grown under dry land conditions and its yield instability need to be improved. This may be done by enhancing crop nodulation and, hence, biological nitrogen fixation (BNF) by supplying effective symbiotic rhizobia through the application of “elite” inoculants. Therefore, the main aim in this study was the isolation and genomic analysis of effective rhizobial strains potentially adapted to drought conditions. Accordingly, pigeon pea endosymbionts were isolated from different soil types in Southern, Central, and Northern India. After functional characterisation of the isolated strains in terms of their ability to nodulate and promote the growth of pigeon pea, 19 were selected for full genome sequencing, along with eight commercial inoculant strains obtained from the ICRISAT culture collection. The phylogenomic analysis [Average nucleotide identity MUMmer (ANIm)] revealed that the pigeon pea endosymbionts were members of the genera Bradyrhizobium and Ensifer. Based on nodC phylogeny and nod cluster synteny, Bradyrhizobium yuanmingense was revealed as the most common endosymbiont, harbouring nod genes similar to those of Bradyrhizobium cajani and Bradyrhizobium zhanjiangense. This symbiont type (e.g., strain BRP05 from Madhya Pradesh) also outperformed all other strains tested on pigeon pea, with the notable exception of an Ensifer alkalisoli strain from North India (NBAIM29). The results provide the basis for the development of pigeon pea inoculants to increase the yield of this legume through the use of effective nitrogen-fixing rhizobia, tailored for the different agroclimatic regions of India.

Influence of mixture of Arbuscular Mycorrhizae fungi and rhizobia on growth and yield of soybean at tidal swamp land

Tidal swamp land is a marginal land which has the potential for enhancement of many agricultural crop growths and yields. The objective of this study was to investigate the effect of a bio-fertilizer containing Arbuscular mycorrhizae fungi and a Rhizobium, Bradyrhizobium yuanmingense on productivity of soybean at the tidal swamp land. The field experiment was established in a completely randomized design with three treatments and three replications per treatments. The treatments include fertilization (P) consisting of a 100% of standard dose of NPK (P1) and the use of biological fertilizers combined with NPK; 100% of standard dose (P2) and 50% of the standard dose (P3) per hectare. The results showed that the yield of soybean treated with bio-fertilizer and combined with 50% standard dosages of NPK could increase production up to 44.7% compared to P1.

Characterization and nodulation capacity of native bacteria isolated from mung bean nodules used as a trap plant in Brazilian tropical soils

Mung bean [Vigna radiata (L.) Wilczek] is a legume of Asian origin with great importance worldwide, particularly in developing countries. It is known for its promiscuous association with soil rhizobia, which provides nitrogen via biological nitrogen fixation (BNF). From the prospective of increasing productivity, efforts are ongoing to identify bacteria that form more efficient symbiotic associations with mung bean. In this regard, we evaluated the nodulation of two mung bean cultivars with native rhizobia from 10 samples of tropical Brazilian soil and performed the isolation of bacterial strains present in the nodules. These isolates were characterized morphologically, and grouped based on sequencing of the 16S rRNA gene. The isolates were also evaluated for nodulation capacity in mung bean and BNF efficiency. We found differences in mung bean plant development depending on the soil for both cultivars. In general, the plants cultivated in soil samples from the Atlantic Forest region showed greater nodulation and shoot dry weight compared to the Brazilian Cerrado (tropical savanna). On the basis of an evaluation of nodulation by native rhizobia, we isolated 101 bacterial strains, which were classified according to 16S rRNA gene into nine genera: Bradyrhizobium (66), Rhizobium (19), Mesorhizobium (4), Ensifer (3), Leifsonia (3), Bacillus (3), Agrobacterium (1), Mycolicibacterium (1), and Kaistia (1). Bradyrhizobium was the most abundant genus, and its isolates formed 12 phylogenetic groups. Inoculation assessments revealed that only Bradyrhizobium isolates induced nodule formation in mung bean, with the BNF efficiency of isolates differing depending on phylogenetic group and soil of origin. Plants inoculated with isolates grouped with Bradyrhizobium yuanmingense B071 were found to show greater increases in shoot dry weight. Considering the soil origin, we observed that plants inoculated with Bradyrhizobium isolated from Mato Grosso state soil samples showed smaller increments in nodulation and shoot dry weight. These results suggest that the Bradyrhizobium is the predominant symbiont in the nodules of mung bean grown in Brazilian tropical soils, and its isolates have the potential for mung bean inoculation, especially when grown in soils that provide low nodulation by native rhizobia.

Root Nodule Rhizobia From Undomesticated Shrubs of the Dry Woodlands of Southern Africa Can Nodulate Angolan Teak Pterocarpus angolensis, an Important Source of Timber.

Pterocarpus angolensis, a leguminous tree native to the dry woodlands of Southern Africa, provides valuable timber, but is threatened by land conversion and overharvesting while showing limited natural regeneration. Nitrogen-fixing root nodule symbionts that could improve establishment of young seedlings have not yet been described. Therefore, we investigated the ability of P. angolensis to form nodules with a diverse range of rhizobia. In drought-prone areas under climate change with higher temperatures, inoculants that are heat-tolerant and adapted to these conditions are likely to be of advantage. Sources of bacterial isolates were roots of P. angolensis from nurseries in the Kavango region, other shrubs from this area growing near Pterocarpus such as Indigofera rautanenii, Desmodium barbatum, Chamaecrista sp., or shrubs from drought-prone areas in Namaqualand (Wiborgia monoptera, Leobordea digitata) or Kalahari (Indigofera alternans). Only slight protrusions were observed on P. angolensis roots, from which a non-nodulating Microbacterium sp. was isolated. Rhizobia that were isolated from nodules of other shrubs were affiliated to Bradyrhizobium ripae WR4T, Bradyrhizobium spp. (WR23/WR74/WR93/WR96), or Ensifer/Mesorhizobium (WR41/WR52). As many plant growth-promoting rhizobacteria (PGPR), nodule isolates produced siderophores and solubilized phosphate. Among them, only the Bradyrhizobium strains nodulated P. angolensis under controlled conditions in the laboratory. Isolates were further characterized by multilocus sequence analysis and were found to be distant from known Bradyrhizobium species. Among additional reference species tested for nodulation on P. angolensis, Bradyrhizobium vignae 7-2T and Bradyrhizobium namibiense 5-10T from the Kavango region of Namibia as well as Bradyrhizobium elkanii LMG6234T and Bradyrhizobium yuanmingense LMG21728T induced nitrogen-fixing nodules, while Bradyrhizobium diazoefficiens USDA110T and Bradyrhizobium tropiciagri SEMIA6148T did not. This suggests a broad microsymbiont range from Bradyrhizobium japonicum and B. elkanii lineages. Phylogenetic analysis of nodC genes indicated that nodulating bradyrhizobia did not belong to a specific symbiovar. Also, for I. rautanenii and Wiborgia, nodule isolates B. ripae WR4T or Mesorhizobium sp. WR52, respectively, were authenticated. Characterization of symbionts inducing effective root nodules in P. angolensis and other shrubs from Subsahara Africa (SSA) give insights in their symbiotic partners for the first time and might help in future to develop bioinoculants for young seedlings in nurseries, and for reforestation efforts in Southern Africa.

Isolation and Characterization of Native Rhizobium Strains Nodulating Some Legumes Species in South Brazzaville in Republic of Congo

Actually, in Republic of Congo, rhizobia have poorly phenotypically and biochemically characterized. This study aimed to characterize native rhizobia. Rhizobia strains were isolated using nodule roots collected on Milletia laurentii, Acacia spp., Albizia lebbeck, and Vigna unguiculata. The strains isolated were characterized microbiologically, biochemically, physiologically, and molecularly identified using 16S rRNA method. The results reported in this study are only for six strains of all 77 isolated: RhA1, RhAc4, RhAc15, RhAc13, RhW1, and RhV3. All native strains were positive to urease activity, negative to cellulase and pectinase activity except for one isolate that showed a positive cellulase activity. Moreover, isolates have grown at 12% of NaCl. On different effects of temperatures, isolates were able to grow up to 44°C and showed good growth at pH from 7 to 9 and the ability to use ten different carbon hydrates sources. The strains were identified as Rhizobium tropici, Rhizobium sp., Mesorhizobium sp. Bradyrhizobium yuanmingense and Bradyrhizobium elkanii. The phylogenetically analysis of the 16S rRNA genes, using a clustering method, allowed us to have a history that is both ancient and stable of four clades among genes with similar patterns. Expanding our awareness of the new legume-rhizobia will be a valuable resource for incorporating an alternative nitrogen fixation approach to consolidate the growth of legumes. These germs can be used in Congolese agriculture to improve yield of crops.

Phylogenetic diversity analysis reveals Bradyrhizobium yuanmingense and Ensifer aridi as major symbionts of mung bean (Vigna radiata L.) in Pakistan.

The present study was carried out to evaluate the diversity of rhizobia associated with nodules of mung bean in Pakistan, because this information is necessary for inoculum development. Based on sequence analysis of 16S rRNA gene of thirty-one bacteria, 11 were assigned to genus Bradyrhizobium, 17 to Ensifer, and 3 to Rhizobium. Phylogenetic analyses on the basis of 16S-23S ITS region, atpD, recA, nifH, and nodA of representative strains revealed that B. yuanmingense is the predominant species distributed throughout different mung bean-growing areas. Among the fast-growing rhizobia, Ensifer aridi was predominant in Faisalabad, Layyah, and Rawalpindi, while E. meliloti in Thal desert. Sequence variations and phylogeny of nifH and nodA genes suggested that these genes might have been co-evolved with the housekeeping genes and maintained by vertical gene transfer in rhizobia detected in the present study. Host infectivity assay revealed the successful nodulation of host by rhizobia related to genera Bradyrhizobium, Ensifer and Rhizobium. Among all, Bradyrhizobium and Ensifer spp. inoculation exhibited a significantly higher number of nodules (11-34 nodules plant-1) and nitrogenase activity (nodule ARA 60-110μmolg-1h-1). Contrary to the previous studies, our data reveal that B. yuanmingense and E. aridi are predominant species forming effective nodules in mung bean in Pakistan. Furthermore, to the best of our knowledge, this is the first report showing the effective symbiosis of E. aridi, E. meliloti, and Rhizobium pusense with mung bean. The diversity of rhizobia in different habitats revealed in the present study will contribute towards designing site-specific inocula for mung bean.

Genome sequence of Bradyrhizobium yuanmingense strain P10 130, a highly efficient nitrogen-fixing bacterium that could be used for Desmodium incanum inoculation.

Strain P10 130, an isolated Bradyrhizobium strain from Argentina which promotes the growth of the leguminous plant Desmodium incanum by different mechanisms was previously selected as the best candidate for D. incanum inoculation based on broader selective criteria. A close relationship between this strain and B. yuanmingense was determined by MALDI BioTyper identification and 16S rRNA gene phylogenetic analysis. This study aimed to analyse the genome sequence of B. yuanmingense P10 130 in order to deepen our knowledge regarding its plant growth-promoting traits and to establish its phylogenetic relationship with other species of Bradyrhizobium genus. The genome size of strain P10 130 was estimated to be 7.54Mb that consisted of 65 contigs. Genome Average Nucleotide Identity (ANI) analysis revealed that B. yuanmingense CCBAU 10071T was the closest strain to P10 130 with ca. 96% identity. Further analysis of the genome of B. yuanmingense P10 130 identified 20 nod/nol/NOE, 14 nif/18 fix, 5 nap/5 nor genes, which may be potentially involved in nodulation, nitrogen fixation, and denitrification process respectively. Genome sequence of B. yuanmingense P10 130 is a valuable source of information to continue the research of its potential industrial production as a biofertilizer of D. incanum.

Growth, Nodulation, and Yield Responses of Groundnut (Arachis hypogaea L.) as Influenced by Combined Application of Rhizobium Inoculant and Phosphorus in the Guinea Savanna Zone of Ghana

Groundnut yields obtained by farmers in northern Ghana are generally low due to low soil fertility resulting from continuous cropping coupled with low use of external inputs. There is therefore the need to use systems’ internal resources such as biological nitrogen fixation efficiently to enhance crop production. This on-station experiment investigated nodulation and pod yield responses of three groundnut varieties, namely Obolo, Oboshie and Samnut 22 to inoculation with rhizobium inoculants of exotic strains, namely Bradyrhizobium yuanmingense (BR 3267) and USDA 3456 in combination with 0 kg P ha−1, 15 kg P ha−1 and 30 kg P ha−1. Combined application of 30 kg P ha−1 and BR 3267 increased the nodule numbers in Obolo, Oboshie and Samnut 22 by 144%, 188% and 56%, respectively compared to their uninoculated counterparts. Inoculation with BR 3267 produced the highest pod yield in all the three varieties with yields increasing from 13 to 40% over that of the uninoculated treatments, with BR 3267-inoculated Samnut 22 giving the highest yield of 2013 kg ha−1. P fertilizer and rhizobium inoculant also had a significant interactive influence on the pod yield of groundnut. Combined application of 30 kg P ha−1 and rhizobium inoculation increased the groundnut yield by 64 to 68%. The study observed a positive interaction between the rhizobium strains and P fertilizer.

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 .

Phylogeny and distribution of Bradyrhizobium symbionts nodulating cowpea (Vigna unguiculata L. Walp) and their association with the physicochemical properties of acidic African soils

In the N2-fixing symbiosis, the choice of a symbiotic partner is largely influenced by the host plant, the rhizobial symbiont, as well as soil factors. Understanding the soil environment conducive for the survival and multiplication of root-nodule bacteria is critical for microbial ecology. In this study, we collected cowpea-nodules from acidic soils in Ghana and South Africa, and nodule DNA isolates were characterized using 16S–23S rRNA-RFLP, phylogenetic analysis of housekeeping and symbiotic genes, and bradyrhizobial community structure through canonical correspondence analysis (CCA). The CCA ordination plot results showed that arrow of soil pH was overlapping on CCA2 axis and was the most important to the ordination. The test nodule DNA isolates from Ghana were positively influenced by soil Zn, Na and K while nodule DNA isolates from South Africa were influenced by P. The amplified 16S–23S rRNA region yielded single polymorphic bands of varying lengths (573–1298bp) that were grouped into 28 ITS types. The constructed ITS-dendrogram placed all the nodule DNA isolates in five major clusters at low cut-off of approx. 0.1 Jaccard’s similarity coefficient. The phylogenetic analysis of 16S rRNA and housekeeping genes (glnII, gyrB, and atpD) formed distinct Bradyrhizobium groups in the phylogenetic trees. It revealed the presence of highly diverse bradyrhizobia (i.e. Bradyrhizobium vignae, Bradyrhizobium elkanii, Bradyrhizobium iriomotense, Bradyrhizobium pachyrhizi, and Bradyrhizobium yuanmingense) together with novel/unidentified bradyrhizobia in the acidic soils from Ghana and South Africa. Discrepancies noted in the phylogenies of some nodule DNA isolates could be attributed to horizontal gene transfer or recombination.

Sinergismo entre Azotobacter chroococcum y Bradyrhizobium yuanmingense en el crecimiento de Lactuca sativa “lechuga”

Las rizobacterias promotoras de crecimiento vegetal estan siendo consideradas como una alternativa para el desarrollo agricola. Se evaluo el efecto del sinergismo entre las rizobacterias, nativas de la region norte del Peru, como Azotobacter chroococcum y Bradyrhizobium yuanmingense en el crecimiento de Lactuca sativa “lechuga”. Se realizo una suspension de cada bacteria, a una concentracion de 109 UFC/mL y con tres tratamientos a semillas germinadas de L. sativa. En un tratamiento se inoculo 5 mL de A. chroococcum, al otro B. yuanmingense y al ultimo, una mezcla de suspension bacteriana, 2,5 mL de A. chroococcum y 2,5 mL de B. yuanmingense. Ademas, se considero un control (sin bacterias). Esta experiencia fue realizada con cuatro repeticiones. Se recolecto informacion a partir de las variables agronomicas: numero de hojas, longitud del tallo y raiz, peso seco de la parte aerea y de la raiz, peso seco total de la planta e indice de efectividad de la inoculacion (IEI) despues de 40 dias. Se observo un incremento significativo en cada una de las variables procesadas en condiciones de laboratorio, por lo que la sinergia entre las bacterias nativas utilizadas beneficia el crecimiento de L. sativa.

Distribution and correlation between phylogeny and functional traits of cowpea (Vigna unguiculata L. Walp.)-nodulating microsymbionts from Ghana and South Africa

Cowpea (Vigna unguiculata L. Walp.) is indigenous to Africa, and highly valued for its N2-fixing trait and the nutritional attributes of its grain and leaves. The species’ ability to establish effective symbiosis with diverse rhizobial populations gives it survival and growth advantage in N-limited environments. To explore the functional diversity and phylogenetic positions of rhizobia nodulating cowpea in Africa, nodules were collected from various cowpea varieties grown in soils from the Guinea savanna and Sudano-sahelian agroecologies of Northern Ghana, and from the lowveld and middleveld areas of Mpumalanga Province in South Africa. Box-PCR profiling and multilocus sequence analysis revealed the presence of diverse microsymbionts responsible for cowpea nodulation across the study sites. BOX-PCR amplifications yielded variable band sizes, ranging from 618 bp to 5354 bp, which placed the isolates in six major clusters (Cluster A–F). Phylogenetic analysis based on 16S rRNA, atpD, glnII, gyrB, rpoB, nifH and nodC genes revealed the presence of diverse Bradyrhizobium sp. closely related to Bradyrhizobium daqingense, Bradyrhizobium subterraneum, Bradyrhizobium yuanmingense, Bradyrhizobium embrapense, Bradyrhizobium pachyrhizi, Bradyrhizobium elkanii and novel Bradyrhizobium species in the soils studied, a finding that could be attributed to the unique edapho-climatic conditions of the contrasting environments. The test isolates exhibited distinct symbiotic efficiencies, and also induced variable (p ≤ 0.001) photosynthetic rates, leaf transpiration, total chlorophyll and shoot biomass accumulation on cowpea (their homologous host). Canonical correspondence analysis showed that the distribution of these microsymbionts was influenced by the concentrations of macro- and micronutrients in soils. The pairwise genetic distances derived from phylogenies and nodule functioning showed significant (p < 0.05) correlation, which suggests that local environmental factors played a major role in the cowpea-Bradyrhizobium symbiosis.

Diversity of Bradyrhizobia in Subsahara Africa: A Rich Resource.

Making use of biological nitrogen fixation (BNF) with pulses and green manure legumes can help to alleviate nitrogen deficiencies and increase soil fertility, problems faced particularly in smallholder agriculture in Subsahara Africa (SSA). The isolation of indigenous rhizobia provides a basis for the formulation of rhizobial inoculants. Moreover, their identification and characterization contribute to the general understanding of species distribution and ecology. Here we discuss global species discovery of Bradyrhizobium spp. Although recently the number of validly published Bradyrhizobium species is rapidly increasing, their diversity in SSA is not well-represented. We summarize the recent knowledge on species diversity in the Bradyrhizobium yuanmingense lineage to which most SSA isolates belong, and their biogeographic distribution and adaptations. Most indigenous rhizobia appear to differ from species found on other continents. We stress that an as yet hidden diversity may be a rich resource for inoculant development in future. As some species are exceptionally temperature tolerant, they may be potential biofertilizer candidates for global warming scenarios.

Is there a need for Bradyrhizobium yuanmingense and B. japonicum reinoculation in subsequent cropping seasons under smallholder farmers’ conditions?

Reliable information on the persistence of rhizobium in soil in the absence of host between growing periods is important in deciding whether inoculation on the same plot in subsequent seasons is necessary. This study determined the survival of introduced rhizobium strains and predominant factors that influence the declining rates of their populations. Bradyrhizobium yuanmingense (BR 3267) and B. japonicum (USDA 110) were manually incorporated into soils at four different locations (Kpalga, Tanina, Tunayilli and Busa) in northern Ghana at 2.5 × 108 (log10 8.4) and 2.5 × 107 (log10 7.4) cells g−1 peat, respectively, per 6 m2. The populations of surviving cells were estimated at 0, 21, 42, 81, 142 and 296 days using the Most Probable Number (MPN) count technique. Several decline functions were applied to the data with hyperbolic regression function emerging as the option that provides the best fit for B. yuanmingense strain BR 3267 and B. japonicum strain USDA 110 at all locations. There was no significant difference in the declining rates between the different locations; however, there were differences in the declining rates for the sampling times. At 296 days, the numbers of surviving cells of B. yuanmingense strain BR 3267 and B. japonicum strain USDA 110 were log10 1.9 and log10 1.7, respectively. Native rhizobium population and soil moisture were the predominant factors that affected the survival of the introduced strains. It is evident from the studies that these strains can survive in sufficient numbers at least within a year; therefore, re-inoculation may not be necessary for a following season especially when using B. yuanmingense strain BR 3267.

Bacteria related to Bradyrhizobium yuanmingense from Ghana are effective groundnut micro-symbionts

The identification of locally-adapted rhizobia for effective inoculation of grain legumes in Africa’s semiarid regions is strategic for developing and optimizing cheap nitrogen fixation technologies for smallholder farmers. This study was aimed at selecting and characterising effective native rhizobia, from Ghanaian soils for groundnut (Arachis hypogaea L.) inoculation. From surface-disinfected root nodules of cowpea and groundnut plants grown on farmers’ fields, 150 bacterial isolates were obtained, 30 of which were eventually found to nodulate groundnut plants. After testing the symbiotic potential of these isolates on groundnut on sterilized substrate, seven of them, designated as KNUST 1001–1007, were evaluated in an open field pot experiment using 15N-labelled soil. Although 15N dilution analyses did not indicate differences among treatments in the proportion of nitrogen (N) derived from the atmosphere (%Ndfa), all seven strains increased total N derived from N2 fixation by inoculated groundnut plants as compared to the non-inoculated control. Inoculation with KNUST 1002 led to total N accumulation as high as that of the groundnut reference strain 32H1. Genetic characterisation of the isolates by sequence analysis of 16S rRNA gene, 16S – 23S rRNA intergenic transcribed spacer (ITS) region and nodC gene revealed that isolates KNUST 1003 and 1007 were related to Rhizobium tropici, a common bean symbiont. The other five isolates, including KNUST 1002 belonged to the Bradyrhizobium genus, being closely related to Bradyrhizobium yuanmingense. Therefore, this study revealed novel native Ghanaian rhizobia with potential for the development of groundnut inoculants.