The rhizobial community indigenous to the Okavango region has not yet been characterized. The isolation of indigenous rhizobia can provide a basis for the formulation of a rhizobial inoculant. Moreover, their identification and characterization contribute to the general understanding of species distribution and ecology. Isolates were obtained from nodules of local varieties of the pulses cowpea, Bambara groundnut, peanut, hyacinth bean, and common bean. Ninety-one of them were identified by BOX repetitive element PCR (BOX-PCR) and sequence analyses of the 16S-23S rRNA internally transcribed spacer (ITS) and the recA, glnII, rpoB, and nifH genes. A striking geographical distribution was observed. Bradyrhizobium pachyrhizi dominated at sampling sites in Angola which were characterized by acid soils and a semihumid climate. Isolates from the semiarid sampling sites in Namibia were more diverse, with most of them being related to Bradyrhizobium yuanmingense and Bradyrhizobium daqingense. Host plant specificity was observed only for hyacinth bean, which was nodulated by rhizobia presumably representing yet-undescribed species. Furthermore, the isolates were characterized with respect to their adaptation to high temperatures, drought, and local host plants. The adaptation experiments revealed that the Namibian isolates shared an exceptionally high temperature tolerance, but none of the isolates showed considerable adaptation to drought. Moreover, the isolates' performance on different local hosts showed variable results, with most Namibian isolates inducing better nodulation on peanut and hyacinth bean than the Angolan strains. The local predominance of distinct genotypes implies that indigenous strains may exhibit a better performance in inoculant formulations.T he catchment area of the Okavango River extends across parts of the three sub-Saharan African countries of Angola, Namibia, and Botswana and is mainly characterized by smallholder and subsistence farming. Since cultivation techniques remain underdeveloped, e.g., in terms of irrigation and the application of agrochemicals or rhizobial inoculants, local farmers are confronted with low yields and decreasing soil fertilities (1). Pulses like cowpea (Vigna unguiculata), Bambara groundnut (Vigna subterranea), peanut (Arachis hypogaea), hyacinth bean (Lablab purpureus), and common bean (Phaseolus vulgaris) are cultivated and occasionally intercropped, but natural nodulation rates are often low (2), presumably due to a seasonal reduction of the rhizobial population caused by drought and heat. Thus, nitrogen input via symbiotic N 2 fixation might be very limited.Field-grown cowpea plants in Ghana have been reported to fix more than 200 kg N/ha (89% of plant N) (3). In contrast, values of only 4 to 29 kg N-fixed/ha (15 to 56% of plant N) were observed in semiarid southwestern Zimbabwe (4). In such areas, the application of a rhizobial inoculant may thus hold the potential to increase plant nutrition and soil fertility. Many effective rhizobial strains have been identif...

Marama bean (Tylosema esculentum) is an indigenous non-nodulating legume to the arid agro-ecological parts of Southern Africa. It is a staple food for the Khoisan and Bantu people from these areas. It is intriguing how it is able to synthesize the high-protein content in the seeds since its natural habitat is nitrogen deficient. The aim of the study was to determine the presence of seed transmittable bacterial endophytes that may have growth promoting effects, which may be particularly important for the harsh conditions. Marama bean seeds were surface sterilized and gnotobiotically grown to 2 weeks old seedlings. From surface-sterilized shoots and roots, 123 distinct bacterial isolates were cultured using three media, and identified by BOX-PCR fingerprinting and sequence analyses of the 16S rRNA and nifH genes. Phylogenetic analyses of 73 putative endophytes assigned them to bacterial species from 14 genera including Proteobacteria (Rhizobium, Massilia, Kosakonia, Pseudorhodoferax, Caulobacter, Pantoea, Sphingomonas, Burkholderia, Methylobacterium), Firmicutes (Bacillus), Actinobacteria (Curtobacterium, Microbacterium) and Bacteroidetes (Mucilaginibacter, Chitinophaga). Screening for plant growth-promoting activities revealed that the isolates showed production of IAA, ACC deaminase, siderophores, endoglucanase, protease, AHLs and capacities to solubilize phosphate and fix nitrogen. This is the first report that marama bean seeds may harbor endophytes that can be cultivated from seedlings; in this community of bacteria, physiological characteristics that are potentially plant growth promoting are widespread.

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Twenty one strains of symbiotic bacteria from root nodules of local races of cowpea (Vigna unguiculata), Bambara groundnut (Vigna subterranea) and peanuts (Arachis hypogaea) grown on subsistence farmers' fields in the Kavango region of Namibia, were previously characterized as a novel group within the genus Bradyrhizobium. To verify their taxonomic position, the strains were further analysed using a polyphasic approach. 16S rRNA gene sequences were most similar to Bradyrhizobium manausense BR 3351 T , with Bradyrhizobium ganzhouense RITF806 T being the most closely related type strain in the phylogenetic analysis, and Bradyrhizobium yuanmingense CCBAU 10071 T in the ITS sequence analysis. Phylogenetic analysis of concatenated glnII-recA-rpoB-dnaK placed the strains in a highly supported lineage distinct from species of the genus Bradyrhizobium with validly published names; they were most closely related to Bradyrhizobium subterraneum 58 2-1 T . The status of the species was validated by results of DNA-DNA hybridization. The combination of phenotypic characteristics from several tests, including carbon source utilization and antibiotic resistance, could be used to differentiate representative strains of species of the genus Bradyrhizobium with validly published names. Novel strain 7-2 T induced effective nodules on Vigna subterranea, Vigna unguiculata, Arachis hypogaea and on Lablab purpureus. The DNA G+C content of strain 7-2 T was 65.4 mol% (Tm). Based on the data presented, we conclude that these strains represent a novel species for which the name Bradyrhizobium vignae sp. nov. is proposed, with strain 7-2 T [LMG 28791 T , DSMZ 100297 T , NTCCM0018 T (Windhoek)] as the type strain.In the Kavango region of Namibia, agriculture is largely dominated by smallholder farms, without irrigation or agrochemical inputs. Local farmers are confronted with low yields and decreasing soil fertility (Pröpper et al., 2010). The intercropping of grain legumes is common practice with local cereals interspersed in an irregular pattern (Grönemeyer et al., 2013). Cowpea (Vigna unguiculata (L.) Walp., local name in Kavango: makunde) is the main grain legume grown by farmers, but also Bambara groundnut [Vigna subterranea (L.) Verdc., local name: nongomene ] and peanuts (Arachis hypogaea L.) are planted, albeit to a lesser extent (Grönemeyer et al., 2013). Inoculants adapted to the crops and the harsh environmental conditions (long dry seasons and high temperatures) may help to increase yields for local smallholders.In a previous study (Grönemeyer et al., 2014), populations of symbiotic bacteria from nodules of pulses collected in the Kavango region of Namibia and in the Angolan province Bié were sampled. Pure cultures of Bradyrhizobium were isolated and characterized. On the basis of multilocus sequence analysis (MLSA) of concatemers of three protein-coding genes (glnII-recA-rpoB) and intergenic spacer (ITS) sequences, several novel lineages within the genus Bradyrhizobium were suggested by phylogenetic analyses (Gröne-meyer et al....

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.