Abundance and Diversity of Soybean-Nodulating Rhizobia in Black Soil Are Impacted by Land Use and Crop Management

To investigate the effects of land use and crop management on soybean rhizobial communities, 280 nodule isolates were trapped from 7 fields with different land use and culture histories. Besides the known Bradyrhizobium japonicum, three novel genospecies were isolated from these fields. Grassland (GL) maintained a higher diversity of soybean bradyrhizobia than the other cultivation systems. Two genospecies (Bradyrhizobium spp. I and III) were distributed widely in all treatments, while Bradyrhizobium sp. II was found only in GL treatment. Cultivation with soybeans increased the rhizobial abundance and diversity, except for the soybean monoculture (S-S) treatment. In monoculture systems, soybeans favored Bradyrhizobium sp. I, while maize and wheat favored Bradyrhizobium sp. III. Fertilization decreased the rhizobial diversity indexes but did not change the species composition. The organic carbon (OC) and available phosphorus (AP) contents and pH were the main soil parameters positively correlated with the distribution of Bradyrhizobium spp. I and II and Bradyrhizobium japonicum and negatively correlated with Bradyrhizobium sp. III. These results revealed that different land uses and crop management could not only alter the diversity and abundance of soybean rhizobia, but also change interactions between rhizobia and legume or nonlegume plants, which offered novel information about the biogeography of rhizobia.     

Bradyrhizobium Populations Occur in Deep Soil under the Leguminous Tree Acacia albida

Soil cores were drilled under the leguminous tree Acacia albida growing in two different ecoclimatic zones of West Africa: the Sahelian area (100 to 500 mm of annual rainfall) and the Sudano-Guinean area (1,000 to 1,500 mm of annual rainfall). Soil samples were collected at different depths from the surface down to the water table level and analyzed for the presence of rhizobia able to nodulate A. albida. In both areas, population densities of rhizobia were substantially greater near the water table than near the surface. In the Sahelian area, rhizobia were present as deep as 34 m at a concentration of 1.3 × 10³/g of soil. In the Sudano-Guinean area, population densities at 0.5 to 4.5 m depth were higher than in the Sahelian area and, at several depths, comparable to that of temperate soils supporting legume crops (10⁴ rhizobia per g of soil). Surface and deep soil isolates from all four sites were found to be slow-growing rhizobia (Bradyrhizobium sp.). The proportion of effective isolates was almost the same within surface and deep soils.     

Modeling Symbiotic Performance of Introduced Rhizobia in the Field by Use of Indices of Indigenous Population Size and Nitrogen Status of the Soil

The ability to predict the symbiotic performance of rhizobia introduced into different environments would allow for a more judicious use of rhizobial inoculants. Data from eight standardized field inoculation trials were used to develop models that could be used to predict the success of rhizobial inoculation in diverse environments based on indices of the size of indigenous rhizobial populations and the availability of mineral N. Inoculation trials were conducted at five diverse sites on the island of Maui, Hawaii, with two to four legumes from among nine species, yielding 29 legume-site observations. The sizes of indigenous rhizobial populations were determined at planting. Soil N mineralization potential, total soil N, N accumulation and seed yield of nonnodulating soybean, and N derived from N₂ fixation in inoculated soybean served as indices of available soil N. Uninoculated, inoculated, and fertilizer N treatments evaluated the impact of indigenous rhizobial populations and soil N availability on inoculation response and crop yield potential. The ability of several mathematical models to describe the inverse relationship between numbers of indigenous rhizobia and legume inoculation responses was evaluated. Power, exponential, and hyperbolic functions yielded similar results; however, the hyperbolic equation provided the best fit of observed to estimated inoculation responses (r² = 0.59). The fact that 59% of the observed variation in inoculation responses could be accounted for by the relationship of inoculation responses to numbers of indigenous rhizobia illustrates the profound influence that the size of soil rhizobial populations has on the successful use of rhizobial inoculants. In the absence of indigenous rhizobia, the inoculation response was directly proportional to the availability of mineral N. Therefore, the hyperbolic response function was subsequently combined with several indices of soil N availability to generate models for predicting legume inoculation response. Among the models developed, those using either soil N mineralization potential or N derived from N₂ fixation in soybean to express the availability of mineral N were most useful in predicting the success of legume inoculation. Correlation coefficients between observed and estimated inoculation responses were r = 0.83 for the model incorporating soil N mineralization potential and r = 0.96 for the model incorporating N derived from N₂ fixation. Several equations collectively termed “soil N deficit factors” were also found to be useful in estimating inoculation responses. In general, models using postharvest indices of soil N were better estimators of observed inoculation responses than were those using laboratory measures of soil N availability. However, the latter, while providing less precise estimates, are more versatile because all input variables can be obtained through soil analysis prior to planting. These models should provide researchers, as well as regional planners, with a more precise predictive capability to determine the inoculation requirements of legumes grown in diverse environments.     

Diversity of rhizobia and interactions among the host legumes and rhizobial genotypes in an agricultural-forestry ecosystem

To investigate the diversity of rhizobia and interactions among the host legumes and rhizobial genotypes in the same habitat, a total of 97 rhizobial strains isolated from nine legume species grown in an agricultural-forestry ecosystem were identified into seven genomic species and 12 symbiotic genotypes within the genera Bradyrhizobium, Mesorhizobium, Rhizobium and Sinorhizobium based upon analyses of genomic DNA regions and symbiotic genes. The results evidenced that the symbiotic genotypes of rhizobia were consistent with their hosts of origin; revealed that vertical transfer was the main mechanism in rhizobia to maintain the symbiotic genes but lateral transfer of symbiotic genes might have happened between the closely related rhizobial species; suggested the existence of co-distribution and co-evolution among the legume hosts and compatible rhizobia. All of these data demonstrated that the biogeography of rhizobia was a result of interactions among the host legumes, bacterial genomic backgrounds and environments.     

Nitrogen contributions from faba bean (Vicia faba L.) reliant on soil rhizobia or inoculation

Background and Aims

Understanding the impact of soil rhizobial populations and inoculant rhizobia in supplying sufficient nodulation is crucial to optimising N₂ fixation by legume crops. This study explored the impact of different rates of inoculant rhizobia and contrasting soil rhizobia on nodulation and N₂ fixation in faba bean (Vicia faba L.).

Methods

Faba beans were inoculated with one of seven rates of rhizobial inoculation, from no inoculant to 100 times the normal rate of inoculation, sown at two field sites, with or without soil rhizobia present, and their nodulation and N₂ fixation assessed.

Results

At the site without soil rhizobia, inoculation increased nodule number and increased N₂ fixation from 21 to 129 kg shoot N ha^?1, while N₂ fixation increased from 132 to 218 kg shoot N ha^?1 at the site with high background soil rhizobia. At the site without soil rhizobia, inoculation increased concentrations of shoot N from 14 to 24 mg g^?1, grain N from 32 to 45 mg g^?1, and grain yields by 1.0 Mg (metric tonne) ha^?1. Differences in nodulation influenced the contributions of fixed N to the system, which varied from the net removal of 20 kg N ha^?1 from the system in the absence of rhizobia, to a net maximum input of 199 kg N ha^?1 from legume shoot and root residues, after accounting for removal of N in grain harvest.

Conclusions

The impact of inoculation and soil rhizobia strongly influenced grain yield, grain N concentration and the potential contributions of legume cropping to soil N fertility. In soil with resident rhizobia, N₂ fixation was improved only with the highest inoculation rate.     

Infection and Invasion of Roots by Symbiotic, Nitrogen-Fixing Rhizobia during Nodulation of Temperate Legumes

Bacteria belonging to the genera Rhizobium, Mesorhizobium, Sinorhizobium, Bradyrhizobium, and Azorhizobium (collectively referred to as rhizobia) grow in the soil as free-living organisms but can also live as nitrogen-fixing symbionts inside root nodule cells of legume plants. The interactions between several rhizobial species and their host plants have become models for this type of nitrogen-fixing symbiosis. Temperate legumes such as alfalfa, pea, and vetch form indeterminate nodules that arise from root inner and middle cortical cells and grow out from the root via a persistent meristem. During the formation of functional indeterminate nodules, symbiotic bacteria must gain access to the interior of the host root. To get from the outside to the inside, rhizobia grow and divide in tubules called infection threads, which are composite structures derived from the two symbiotic partners. This review focuses on symbiotic infection and invasion during the formation of indeterminate nodules. It summarizes root hair growth, how root hair growth is influenced by rhizobial signaling molecules, infection of root hairs, infection thread extension down root hairs, infection thread growth into root tissue, and the plant and bacterial contributions necessary for infection thread formation and growth. The review also summarizes recent advances concerning the growth dynamics of rhizobial populations in infection threads.     

Denitrifying capacity of rhizobial strains of Argentine soils and herbicide sensitivity

Agrochemical application in soils is a matter of environmental concern, and among soil microorganisms, rhizobia and their action before different pesticides are interesting to study, due to their taxonomic and functional diversity. The objectives of the present work were to assess the capacity of rhizobial populations to use herbicides as source of nutrients, as well as their ability to reduce nitrates and / or denitrify. Eighty-one strains belonging to four populations of different genera of rhizobia (Rhizobium, Mesorhizobium, Ensifer and Bradyrhizobium) were assessed. The effect of glyphosate, 2,4-dichlorophenoxyacetic acid, and atrazine on growth of the strains, as well as the ability of the strains to act on herbicide transformation to reduce nitrate and denitrify, were evaluated. The genera studied showed different responses to pesticides. Bradyrhizobium had the greater capacity to utilize the herbicides and among the compounds evaluated, atrazine was the most used as a source of energy. To conclude, some Bradyrhizobium strains were able both to denitrify and to use the atrazine herbicide. The results obtained in this study increase expectations of the use of rhizobia as inoculants, causing changes at the agricultural and environmental level and allowing an appropriate management of agricultural soil fertilization, efficiency in nitrogen fixation and a faster biodegradation of pesticides in soil.     

Unexpectedly Diverse Mesorhizobium Strains and Rhizobium leguminosarum Nodulate Native Legume Genera of New Zealand, while Introduced Legume Weeds Are Nodulated by Bradyrhizobium Species

The New Zealand native legume flora are represented by four genera, Sophora, Carmichaelia, Clianthus, and Montigena. The adventive flora of New Zealand contains several legume species introduced in the 19th century and now established as serious invasive weeds. Until now, nothing has been reported on the identification of the associated rhizobia of native or introduced legumes in New Zealand. The success of the introduced species may be due, at least in part, to the nature of their rhizobial symbioses. This study set out to address this issue by identifying rhizobial strains isolated from species of the four native legume genera and from the introduced weeds: Acacia spp. (wattles), Cytisus scoparius (broom), and Ulex europaeus (gorse). The identities of the isolates and their relationship to known rhizobia were established by comparative analysis of 16S ribosomal DNA, atpD, glnII, and recA gene sequences. Maximum-likelihood analysis of the resultant data partitioned the bacteria into three genera. Most isolates from native legumes aligned with the genus Mesorhizobium, either as members of named species or as putative novel species. The widespread distribution of strains from individual native legume genera across Mesorhizobium spp. contrasts with previous reports implying that bacterial species are specific to limited numbers of legume genera. In addition, four isolates were identified as Rhizobium leguminosarum. In contrast, all sequences from isolates from introduced weeds aligned with Bradyrhizobium species but formed clusters distinct from existing named species. These results show that native legume genera and these introduced legume genera do not have the same rhizobial populations.     

Molecular Diversity of Rhizobia Occurring on Native Shrubby Legumes in Southeastern Australia

The structure of rhizobial communities nodulating native shrubby legumes in open eucalypt forest of southeastern Australia was investigated by a molecular approach. Twenty-one genomic species were characterized by small-subunit ribosomal DNA PCR-restriction fragment length polymorphism and phylogenetic analyses, among 745 rhizobial strains isolated from nodules sampled on 32 different legume host species at 12 sites. Among these rhizobial genomic species, 16 belonged to the Bradyrhizobium subgroup, 2 to the Rhizobium leguminosarum subgroup, and 3 to the Mesorhizobium subgroup. Only one genomic species corresponded to a known species (Rhizobium tropici). The distribution of the various genomic species was highly unbalanced among the 745 isolates, legume hosts, and sites. Bradyrhizobium species were by far the most abundant, and Rhizobium tropici dominated among the Rhizobium and Mesorhizobium isolates in the generally acid soils where nodules were collected. Although a statistically significant association occurred between the eight most common genomic species and the 32 hosts, there was sufficient overlap in distributions that no clear specificity between rhizobial genomic species and legume taxa was observed. However, for three legume species, some preference for particular genomic species was suggested. Similarly, no geographical partitioning was found.     

Influence of the Size of Indigenous Rhizobial Populations on Establishment and Symbiotic Performance of Introduced Rhizobia on Field-Grown Legumes

Indigenous rhizobia in soil present a competition barrier to the establishment of inoculant strains, possibly leading to inoculation failure. In this study, we used the natural diversity of rhizobial species and numbers in our fields to define, in quantitative terms, the relationship between indigenous rhizobial populations and inoculation response. Eight standardized inoculation trials were conducted at five well-characterized field sites on the island of Maui, Hawaii. Soil rhizobial populations ranged from 0 to over 3.5 × 10⁴ g of soil^-1 for the different legumes used. At each site, no less than four but as many as seven legume species were planted from among the following: soybean (Glycine max), lima bean (Phaseolus lunatus), cowpea (Vigna unguiculata), bush bean (Phaseolus vulgaris), peanut (Arachis hypogaea), Leucaena leucocephala, tinga pea (Lathyrus tingeatus), alfalfa (Medicago sativa), and clover (Trifolium repens). Each legume was (i) inoculated with an equal mixture of three effective strains of homologous rhizobia, (ii) fertilized at high rates with urea, or (iii) left uninoculated. For soybeans, a nonnodulating isoline was used in all trials as the rhizobia-negative control. Inoculation increased economic yield for 22 of the 29 (76%) legume species-site combinations. While the yield increase was greater than 100 kg ha^-1 in all cases, in only 11 (38%) of the species-site combinations was the increase statistically significant (P ≤ 0.05). On average, inoculation increased yield by 62%. Soybean (G. max) responded to inoculation most frequently, while cowpea (V. unguiculata) failed to respond in all trials. Inoculation responses in the other legumes were site dependent. The response to inoculation and the competitive success of inoculant rhizobia were inversely related to numbers of indigenous rhizobia. As few as 50 rhizobia g of soil^-1 eliminated inoculation response. When fewer than 10 indigenous rhizobia g of soil^-1 were present, economic yield was significantly increased 85% of the time. Yield was significantly increased in only 6% of the observations when numbers of indigenous rhizobia were greater than 10 cells g of soil^-1. A significant response to N application, significant increases in nodule parameters, and greater than 50% nodule occupancy by inoculant rhizobia did not necessarily coincide with significant inoculation responses. No less than a doubling of nodule mass and 66% nodule occupancy by inoculant rhizobia were required to significantly increase the yield of inoculated crops over that of uninoculated crops. However, lack of an inoculation response was common even when inoculum strains occupied the majority of nodules. In these trials, the symbiotic yield of crops was, on average, only 88% of the maximum yield potential, as defined by the fertilizer N treatment. The difference between the yield of N-fertilized crops and that of N₂-fixing crops indicates a potential for improving inoculation technology, the N₂ fixation capacity of rhizobial strains, and the efficiency of symbiosis. In this study, we show that the probability of enhancing yield with existing inoculation technology decreases dramatically with increasing numbers of indigenous rhizobia.     

Inoculation Response of Legumes in Relation to the Number and Effectiveness of Indigenous Rhizobium Populations

The response of legumes to inoculation with rhizobia can be affected by many factors. Little work has been undertaken to examine how indigenous populations or rhizobia affect this response. We conducted a series of inoculation trials in four Hawaiian soils with six legume species (Glycine max, Vigna unguiculata, Phaseolus lunatus, Leucaena leucocephala, Arachis hypogaea, and Phaseolus vulgaris) and characterized the native rhizobial populations for each species in terms of the number and effectiveness of the population for a particular host. Inoculated plants had, on average, 76% of the nodules formed by the inoculum strain, which effectively eliminated competition from native strains as a variable between soils. Rhizobia populations ranged from less than 6 × 10⁰/g of soil to 1 × 10⁴/g of soil. The concentration of nitrogen in shoots of inoculated plants was not higher than that in uninoculated controls when the most probable number MPN counts of rhizobia were at or above 2 × 10¹/g of soil unless the native population was completely ineffective. Tests of random isolates from nodules of uninoculated plants revealed that within most soil populations there was a wide range of effectiveness for N₂ fixation. All populations had isolates that were ineffective in fixing N₂. The inoculum strains generally did not fix more N₂ than the average isolate from the soil population in single-isolate tests. Even when the inoculum strain proved to be a better symbiont than the soil rhizobia, there was no response to inoculation. Enhanced N₂ fixation after inoculation was related to increased nodule dry weights. Although inoculation generally increased nodule number when there were less than 1 × 10² rhizobia per g of soil, there was no corresponding increase in nodule dry weight when native populations were effective. Most species compensated for reduced nodulation in soils with few rhizobia by increasing the size of nodules and therefore maintaining a nodule dry weight similar to that of inoculated plants with more nodules. Even when competition by native soil strains was overcome with a selected inoculum strain, it was not always possible to enhance N₂ fixation when soil populations were above a threshold number and had some effective strains.     

Distribution and biodiversity of rhizobia nodulating Chamaecrista mimosoides in the Shandong peninsula of china

Chamaecrista mimosoides is an annual herb legume widely distributed in tropical and subtropical Asia and Africa. It may have primitive and independently-evolved root nodule types but its rhizobia have not been systematically studied. Therefore, in order to learn the diversity and species affinity of its rhizobia, root nodules were sampled from C. mimosoides plants growing in seven geographical sites along the coast line of Shandong Peninsula, China. A total of 422 rhizobial isolates were obtained from nodules, and they were classified into 28 recA haplotypes. By using multilocus sequence analysis of the concatenated housekeeping genes dnaK, glnII, gyrB, recA and rpoB, the representative strains for these haplotypes were designated as eight defined and five candidate novel genospecies in the genus Bradyrhizobium. Bradyrhizobium elkanii and Bradyrhizobium ferriligni were predominant and universally distributed. The symbiotic genes nodC and nifH of the representative strains showed very similar topology in their phylogenetic trees indicating their co-evolution history. All the representative strains formed effective root nodules in nodulation tests. The correlation between genospecies and soil characteristics analyzed by CANOCO software indicated that available potassium (AK), organic carbon (OC) and available nitrogen (AN) in the soil samples were the main factors affecting the distribution of the symbionts involved in this current study. The study is the first systematic survey of Chamaecrista mimosoides-nodulating rhizobia, and it showed that Chamaecrista spp. were nodulated by bradyrhizobia in natural environments. In addition, the host spectrum of the corresponding rhizobial species was extended, and the study provided novel information on the biodiversity and biogeography of rhizobia.     

Phylogenetically diverse group of native bacterial symbionts isolated from root nodules of groundnut (Arachis hypogaea L.) in South Africa

Groundnut is an economically important N_?2-fixing legume that can contribute about 100–190 kg N ha^?1 to cropping systems. In this study, groundnut-nodulating native rhizobia in South African soils were isolated from root nodules. Genetic analysis of isolates was done using restriction fragment length polymorphism (RFLP)-PCR of the intergenic spacer (IGS) region of 16S-23S rDNA. A total of 26 IGS types were detected with band sizes ranging from 471 to 1415 bp. The rhizobial isolates were grouped into five main clusters with Jaccard's similarity coefficient of 0.00–1.00, and 35 restriction types in a UPGMA dendrogram. Partial sequence analysis of the 16S rDNA, IGS of 16S rDNA-23S rDNA, atpD, gyrB, gltA, glnII and symbiotic nifH and nodC genes obtained for representative isolates of each RFLP-cluster showed that these native groundnut-nodulating rhizobia were phylogenetically diverse, thus confirming the extent of promiscuity of this legume. Concatenated gene sequence analysis showed that most isolates did not align with known type strains, and may represent new species from South Africa. This underscored the high genetic variability associated with groundnut Rhizobium and Bradyrhizobium in South African soils, and the possible presence of a reservoir of novel groundnut-nodulating Bradyrhizobium and Rhizobium in the country.     

<Emphasis Type="Italic">Bradyrhizobium</Emphasis> spp. and <Emphasis Type="Italic">Sinorhizobium fredii</Emphasis> are predominant in root nodules of <Emphasis Type="Italic">Vigna angularis</Emphasis>, a native legume crop in the subtropical region of China

Adzuki bean (Vigna angularis) is an important legume crop native to China, but its rhizobia have not been well characterized. In the present study, a total of 60 rhizobial strains isolated from eight provinces of China were analyzed with amplified 16S rRNA gene RFLP, IGS-RFLP, and sequencing analyses of 16S rRNA, atpD, recA, and nodC genes. These strains were identified as genomic species within Rhizobium, Sinorhizobium, Mesorhizobium, Bradyrhizobium, and Ochrobactrum. The most abundant groups were Bradyrhizobium species and Sinorhizobium fredii. Diverse nodC genes were found in these strains, which were mainly co-evolved with the housekeeping genes, but a possible lateral transfer of nodC from Sinorhizobium to Rhizobium was found. Analyses of the genomic and symbiotic gene backgrounds showed that adzuki bean shared the same rhizobial gene pool with soybean (legume native to China) and the exotic Vigna species. All of these data demonstrated that nodule formation is the interaction of rhizobia, host plants, and environment characters. Electronic Supplementary Material  Supplementary material is available for this article at and is accessible for authorized users.     

Genetic Diversity and Evolution of Bradyrhizobium Populations Nodulating Erythrophleum fordii,an Evergreen Tree Indigenous to the Southern Subtropical Region of China

The nodulation of Erythrophleum fordii has been recorded recently, but its microsymbionts have never been studied. To investigate the diversity and biogeography of rhizobia associated with this leguminous evergreen tree, root nodules were collected from the southern subtropical region of China. A total of 166 bacterial isolates were obtained from the nodules and characterized. In a PCR-based restriction fragment length polymorphism (RFLP) analysis of ribosomal intergenic sequences, the isolates were classified into 22 types within the genus Bradyrhizobium. Sequence analysis of 16S rRNA, ribosomal intergenic spacer (IGS), and the housekeeping genes recA and glnII classified the isolates into four groups: the Bradyrhizobium elkanii and Bradyrhizobium pachyrhizi groups, comprising the dominant symbionts, Bradyrhizobium yuanmingense, and an unclassified group comprising the minor symbionts. The nodC and nifH phylogenetic trees defined five or six lineages among the isolates, which was largely consistent with the definition of genomic species. The phylogenetic results and evolutionary analysis demonstrated that mutation and vertical transmission of genes were the principal processes for the divergent evolution of Bradyrhizobium species associated with E. fordii, while lateral transfer and recombination of housekeeping and symbiotic genes were rare. The distribution of the dominant rhizobial populations was affected by soil pH and effective phosphorus. This is the first report to characterize E. fordii rhizobia.     

Genetic Diversity of <Emphasis Type="Italic">Acacia seyal</Emphasis> Del. Rhizobial Populations Indigenous to Senegalese Soils in Relation to Salinity and pH of the Sampling Sites

The occurrence and the distribution of rhizobial populations naturally associated to Acacia seyal Del. were characterized in 42 soils from Senegal. The diversity of rhizobial genotypes, as characterized by polymerase chain reaction restriction fragment length polymorphism (RFLP) analysis of 16S–23S rDNA, performed on DNA extracted from 138 nodules resulted in 15 clusters. Results indicated the widespread occurrence of compatible rhizobia associated to A. seyal in various ecogeographic areas. However, the clustering of rhizobial populations based on intergenic spacer (IGS) RFLP profiles did not reflect their geographic origin. Four genera were discriminated on the basis of 16S rRNA gene sequences of the strains representative for the IGS-RFLP profiles. The majority of rhizobia associated to A. seyal were affiliated to Mesorhizobium and Sinorhizobium 64 and 29%, respectively, of the different IGS-RFLP profiles. Our results demonstrate the coexistence inside the nodule of plant-pathogenic non-N₂-fixing Agrobacterium and Burkholderia strains, which induced the formation of ineffective nodules, with symbiotic rhizobia. Nodulation was recorded in saline soils and/or at low pH values or in alkaline soils, suggesting adaptability of natural rhizobial populations to major ecological environmental stress and their ability to establish symbiotic associations within these soil environments. These results contribute to the progressing research efforts to uncover the biodiversity of rhizobia and to improve nitrogen fixation in agroforestry systems in sub-Saharan Africa.     

Eco-geographical diversity of cowpea bradyrhizobia in Senegal is marked by dominance of two genetic types

The genetic diversity of native cowpea rhizobia originating from 60 sites across four eco-geographic zones in Senegal was studied. More than 300 cowpea nodules were analyzed by PCR-RFLP of the 16S–23S rDNA InterGenic Spacer region (IGS). Alignments of IGS sequences indicated that all genotypes were grouping within the Bradyrhizobium genus. The geographical distribution showed that apart from five IGS types, the others were specifically found in only one region. The diversity was significantly higher in the Senegal River valley zone, which presents lower mean annual rainfalls and slightly alkaline soils. Interestingly, two IGS types dominated the Senegalese rhizobial collection, one IGS type (VI) was found on more than half of the nodules collected in the northern Senegal River valley while another IGS type (I) was recovered from the great majority of nodules in the three other regions sampled. Two representative strains from each of these two dominant types were isolated and further analyzed. Multi Locus Sequence Analyses using 6 housekeeping genes indicate that they belong to a new Bradyrhizobium species closely related to B. yuanmingense. Phylogenetic analyses of 2 symbiotic genes nodC and nifH show that they are clustered with B. arachidis. Physiological tests on these strains have shown that under laboratory conditions, the growth of the IGS type VI strains was slightly less affected by a higher osmotic strength in the medium and to alkaline pH, which corroborates the soil physico-chemical parameters.     

Indigenous bradyrhizobia populations in tropical farm-lands

Using the plant-infection technique, and soybeans as test plants, counts of root nodule bacteria in seven farm-lands in Southern Nigeria were estimated. All sites including those without a recent legume cover history contained bradyrhizobia. Abundance, however correlated with the presence of a legume. Populations varied from 0.95 to 3.63 log₁₀ rhizobia per g soil.
In addition to the presence of a legume, soil moisture and pH appeared to influence the rhizobial distribution in these soils. Most of the nodules produced on the test plant were ineffective, indicating that the rhizobial presence alone does not guarantee adequate N₂ fixation in a farming set-up.     

Rhizobial counts in peat inoculants vary amongst legume inoculant groups at manufacture and with storage: implications for quality standards

Background and aims

Inoculation of legumes at sowing with rhizobia has arguably been one of the most cost-effective practices in modern agriculture. Critical aspects of inoculant quality are rhizobial counts at manufacture/registration and shelf (product) life.

Methods

In order to re-evaluate the Australian standards for peat-based inoculants, we assessed numbers of rhizobia (rhizobial counts) and presence of contaminants in 1,234 individual packets of peat–based inoculants from 13 different inoculant groups that were either freshly manufactured or had been stored at 4 °C for up to 38 months to determine (a) rates of decline of rhizobial populations, and (b) effects of presence of contaminants on rhizobial populations. We also assessed effects of inoculant age on survival of the rhizobia during and immediately after inoculation of polyethylene beads.

Results

Rhizobial populations in the peat inoculants at manufacture and decline rates varied substantially amongst the 13 inoculant groups. The most stable were Sinorhizobium, Bradyrhizobium and Mesorhizobium with Rhizobium, particularly R. leguminosarum bv. trifolii the least stable. The presence of contaminants at the 10^?6 level of dilution, i.e. >log 6.7 g^?1 peat, reduced rhizobial numbers in the stored inoculants by an average of 37 %. Survival on beads following inoculation improved 2–3 fold with increasing age of inoculant.

Conclusions

We concluded that the Australian standards for peat-based rhizobial inoculants should be reassessed to account for the large differences amongst the groups in counts at manufacture and survival rates during storage. Key recommendations are to increase expiry counts from log 8.0 to log 8.7 rhizobia g^?1 peat and to have four levels of inoculant shelf life ranging from 12 months to 3 years.     

Lima bean nodulates efficiently with <Emphasis Type="Italic">Bradyrhizobium</Emphasis> strains isolated from diverse legume species

Lima bean (Phaseolus lunatus L.) is an important legume species that establishes symbiosis with rhizobia, mainly of the Bradyrhizobium genus. The aim of this study was to evaluate the efficiency of rhizobia of the genus Bradyrhizobium in symbiosis with lima bean, in both Leonard jars and in pots with a Latossolo Amarelo distrófico (Oxisol). In the experiment in Leonard jars, 17 strains isolated from nodules of the three legume subfamilies, Papilionoideae (Vigna unguiculata, Pterocarpus sp., Macroptilium atropurpureum, Swartzia sp., and Glycine max), Mimosoideae (Inga sp.), and Caesalpinioideae (Campsiandra surinamensis) and two uninoculated controls, one with a low concentration (5.25 mg L^?1) and another with a high concentration (52.5 mg L^?1) of mineral nitrogen (N) were evaluated. The six strains that exhibited the highest efficiency in Leonard jars, isolated from nodules of Vigna unguiculata (UFLA 03–144, UFLA 03–84, and UFLA 03–150), Campsiandra surinamensis (INPA 104A), Inga sp. (INPA 54B), and Swartzia sp. (INPA 86A), were compared to two uninoculated controls, one without and another with 300 mg N dm^?3 (NH₄NO₃) applied to pots with samples of an Oxisol in the presence and absence of liming. In this experiment, liming did not affect nodulation and plant growth; the INPA 54B and INPA 86A strains stood out in terms of shoot dry matter production and provided increases of approximately 48% in shoot N accumulation compared to the native rhizobia populations. Our study is the first to indicate Bradyrhizobium strains isolated from the three legume subfamilies are able to promote lima bean growth via biological nitrogen fixation in soil conditions.