Rhizobial Communities Research Articles

Effects of Long-Term Fertilization Strategies on Soil Productivity and Soybean Rhizobial Diversity in a Chinese Mollisol

Rhizobial diversity is affected by interactions between soil features, fertilization strategy, and cropping system. However, interactions among the rhizobial community, chemical-organic manure fertilization, and plant production have not been well documented in Mollisols from long-term experiments. Aimed at maintaining and recovering the productivity of Chinese Mollisols, a long-term fertilization experiment had been carried out for 29 years under a wheat-maize-soybean rotation system, involving the application of recycled organic manure (ROM), chemical fertilizers (N, P, and/or K), or ROM plus N, P, and/or K. In the present study, the effects of different treatments were evaluated by determining soil physicochemical features, soybean production, and soybean rhizobial diversity. The results showed that application of ROM plus NPK maintained or increased soil fertility, which was accompanied by higher production and higher diversity of rhizobia, as compared with the other treatments. The negative association of Bradyrhizobium japonicum with N fertilizer, positive association of B. diazoefficiens with soil pH, and alleviation of N-inhibition on the diversity of Bradyrhizobium by the addition of ROM were recorded as new findings. Therefore, application of ROM or ROM plus NPK could be a feasible strategy for maintaining and recovering the fertility of Chinese Mollisols, whereas rhizobial diversity could be an indicator of soil fertility.

Is there sufficient Ensifer and Rhizobium species diversity in UK farmland soils to support red clover (Trifolium pratense), white clover (T. repens), lucerne (Medicago sativa) and black medic (M. lupulina)?

Rhizobia play important roles in agriculture owing to their ability to fix nitrogen through a symbiosis with legumes. The specificity of rhizobia-legume associations means that underused legume species may depend on seed inoculation with their rhizobial partners. For black medic (Medicago lupulina) and lucerne (Medicago sativa) little is known about the natural prevalence of their rhizobial partner Ensifer meliloti in UK soils, so that the need for inoculating them is unclear. We analysed the site-dependence of rhizobial seed inoculation effects on the subsequent ability of rhizobial communities to form symbioses with four legume species (Medicago lupulina, M. sativa, Trifolium repens and T. pratense). At ten organic farms across the UK, a species-diverse legume based mixture (LBM) which included these four species was grown. The LBM seed was inoculated with a mix of commercial inocula specific for clover and lucerne. At each site, soil from the LBM treatment was compared to the soil sampled prior to the sowing of the LBM (the control). From each site and each of the two treatments, a suspension of soils was applied to seedlings of the four legume species and grown in axenic conditions for six weeks. Root nodules were counted and their rhizobia isolated. PCR and sequencing of a fragment of the gyrB gene from rhizobial isolates allowed identification of strains. The number of nodules on each of the four legume species was significantly increased when inoculated with soil from the LBM treatment compared to the control. Both the proportion of plants forming nodules and the number of nodules formed varied significantly by site, with sites significantly affecting the Medicago species but not the Trifolium species. These differences in nodulation were broadly reflected in plant biomass where site and treatment interacted; at some sites there was a significant advantage from inoculation with the commercial inoculum but not at others. In particular, this study has demonstrated the commercial merit of inoculation of lucerne with compatible rhizobia.

Diversity and dynamics of rhizobial populations in acidic soils with aluminum and manganese toxicities in forest zones

Soil acidity in the humid forest zones of Cameroon is one of the major constraints to agricultural productivity. This study was carried out to assess the rhizobial communities of two acidic soils; with aluminum toxicity (Nkoemvone) and manganese toxicity (Nkolbisson) for their potential to improve soil fertility in Cameroon. These two soils were used to inoculate to the host plants cowpea and siratro. At harvest, 120 rhizobacterial isolates were extracted from the nodules of these two hosts and subjected to morphological characterization. Twenty isolates per site were selected and analyzed for their 16S rDNA genetic profile following restrictions with endonucleases of PCR products and electrophoresis. The restriction patterns of the 16S rDNA of the 40 isolates showed 12 different profiles. Eight occurred in both types of soils, where as 4 were specific to the manganese-toxic-acidic soil. While the Al toxicity reduced the nodulation and growth of both plants, the Mn toxicity mostly affect the cowpea. This study ascertained the distribution of rhizobia based on soil characteristics. Further molecular analyses would allow the identification of the isolates recovered as well as their phylogenetical relationships.Int. J. Agril. Res. Innov. & Tech. 6 (2): 12-23, December, 2016

Cultivation may not dramatically alter rhizobial community diversity or structure associated with rooibos tea (Aspalathus linearis Burm.f.) in South Africa

Rooibos tea, Aspalathus linearis (Burm.f.), represents one of South Africa's most important indigenous crops, and monotypic plantations are rapidly replacing wild populations. Dwindling wild rooibos populations may drastically reduce possible, but hitherto overlooked, natural resources to bolster commercial productivity and long-term sustainability.Here, using next generation sequencing data, we seek to determine whether cultivation impacts on the diversity and community structure of mutualistic nitrogen fixing bacteria (rhizobia) associated with rooibos plants. We do this by generating operational taxonomic units (OTUs) from nodulation (nodC) and nitrogenase (nifH) DNA sequence data from rhizobia within root nodules of rooibos plants from cultivated and wild populations. For these rhizobial communities we found significant differences for various OTU diversity metrics due to geography (site), the interaction between site and status (cultivated vs. wild), but not for status. We also sequenced nodC soil microbiomes and found evidence for compositional differences in soil rhizobial communities due to geography, status (cultivated vs. wild), and the interaction between geography and status.Our data adds to existing evidence suggesting that tea cultivation in South Africa may have limited impacts on soil microbial community diversity and structure, and that such changes are mainly driven by larger geographic processes rather than human-mediated disturbances such as cultivation.

Ancient Heavy Metal Contamination in Soils as a Driver of Tolerant Anthyllis vulneraria Rhizobial Communities.

Phytostabilization of toxic mine spoils limits heavy metal dispersion and environmental pollution by establishing a sustainable plant cover. This eco-friendly method is facilitated by the use of selected and adapted cover crop legumes living in symbiosis with rhizobia that can stimulate plant growth naturally through biological nitrogen fixation. We studied microsymbiont partners of a metal-tolerant legume, Anthyllis vulneraria, which is tolerant to very highly metal-polluted soils in mining and nonmining sites. Site-specific rhizobial communities were linked to taxonomic composition and metal tolerance capacity. The rhizobial species Mesorhizobium metallidurans was dominant in all Zn-Pb mines but one. It was not detected in unpolluted sites where other distinct Mesorhizobium species occur. Given the different soil conditions at the respective mining sites, including their heavy-metal contamination, revegetation strategies based on rhizobia adapting to local conditions are more likely to succeed over the long term compared to strategies based on introducing less-well-adapted strains.

Presence of diverse rhizobial communities responsible for nodulation of common bean (Phaseolus vulgaris) in South African and Mozambican soils.

The diversity and phylogeny of root-nodule bacteria isolated from common bean grown in Mozambique and different provinces of South Africa was studied by restriction fragment length polymorphism (RFLP) and phylogenetic analysis. The combined restriction banding pattern of 16S rRNA and nifH profile-generated dendrogram grouped all test isolates into four major clusters with XXI restriction groups and three clusters with VIII restriction groups. Location-based clustering was observed with the 16S rRNA RFLP analysis. Phylogenetic analysis of 16S rRNA, glnII, gyrB and gltA sequences showed that common bean was nodulated specifically by Rhizobium etli in Mozambican soils, and by a diverse group of Rhizobium species in South African soils (e.g. R. etli, R. phaseoli, R. sophoriradicis, R. leucaenae and novel group of Rhizobium spp.). Isolates from the Eastern Cape region of South Africa were dominated by R. leucaenae Overall, the results suggested high nodulation promiscuity of common bean grown in Southern Africa. The nifH and nodC sequence analysis classified all the test isolates with R. etli group, except for isolates TUTPVSA117, TUTPVSA114 and TUTPVSA110 which delineated with R. tropici group. This finding was inconsistent with the phylogram of the housekeeping genes, and is probably an indication of horizontal gene transfer among the Rhizobium isolates tested.

Host species and environmental variation can influence rhizobial community composition

Summary Nitrogen‐fixing rhizobial mutualists display high levels of genetic and phenotypic diversity at multiple spatial scales. Identifying the ecological and environmental drivers of variation in the genetic composition and function of rhizobial communities is critical to understanding rhizobial impacts on community and ecosystem productivity. In this study, we examined how environmental factors and host species jointly affect rhizobial phenotypic diversity and community structure. Individual host plants of two native Australian legumes, Acacia salicina and A. stenophylla, were inoculated with a community of six rhizobial strains (belonging to three genera) previously characterized for symbiotic effectiveness. The plants were grown in a fully factorial experiment under high and low phosphorus and water, and high or no nitrogen. Half the plants were harvested after 19 weeks and the other half after 30 weeks. The relative abundance of the six rhizobial strains was characterized using amplicon sequencing of the rhizobial 16S rRNA gene from DNA extracted from the root nodules of plants after each harvest. Variation in the relative abundance of the six rhizobial strains was best explained by host species, although interactions between environment and host species also emerged as significant predictors of variation in rhizobial community composition. We detected strong host‐specific interactions for two strains, while two other strains were clearly generalists, being relatively abundant in both host species. For both host species, the two generalist strains were as abundant as the best specialist strain within the rhizobial community. Synthesis. Our study show host species identity as the key determinant of rhizobial community structure within the root nodules of Acacia plants. Strains that were efficient at promoting the growth of their host plant had high relative abundance in experimental communities. Furthermore, interaction among host species, rhizobial host‐range, and environmental variation determine the structure of rhizobial communities, although not in clearly predictable ways. Such interactions contributed to the maintenance (albeit at relatively low frequencies) of ineffective strains in nodule communities. These interacting effects could help to explain why rhizobial communities are functionally diverse, such that specialists, generalists and seemingly ineffective rhizobia, can be simultaneously maintained in rhizosphere communities.

Genetic diversity of indigenous soybean-nodulating rhizobia in response to locally-based long term fertilization in a Mollisol of Northeast China.

The influences of five different fertilizer treatments on diversity of rhizobia in soybean nodule were investigated in a long-term experiment with with four replicates: (1) control (without fertilization), (2) balanced NPK fertilizer (NPK), and (3-5) unbalanced chemical fertilizers without one of the major elements (NP, PK, and NK) in Mollisol in Northeast China. The highest soybean yield was observed in the NPK treatment. Total of 200 isolates were isolated and grouped into four Bradyrhizobium genospecies corresponding to B. japonicum, B. diazoefficiens, B. ottawaense and Bradyrhizobium sp. I, based upon the multilocus sequence analysis of 6 housekeeping genes. The Bradyrhizobium sp. I was extensively distributed throughout the study site and was recorded as the dominant soybean rhizobia (82.5-87.5%). Except the NK treatment, the other fertilizer treatments had no effect on rhizobial species composition. Compared with the CK treatment, all the fertilizer treatments decreased species richness, diversity and evenness. The soil organic carbon contents, available N content and pH were the key soil factors to rhizobial community structure. Results suggest that long-term fertilization can decrease rhizobial species diversity, while balanced fertilization with NPK is the most suitable fertilization regime if taking both soybean yields and rhizobial diversity into account.

Bradyrhizobium inoculation alters indigenous AMF community assemblages and interacts positively with AMF inoculum to improve cowpea performance

Legumes are a major component in sustainable agricultural systems since they can fix atmospheric nitrogen and form effective associations with both nitrogen fixing bacteria (NFB) and arbuscular mycorrhizal fungi (AMF). However, the impact of NFB inoculation on the indigenous AMF community structure in legumes has been overlooked. In the current study, we investigated the influence of a Bradhyrhizobium isolate together with a commercial mycorrhizal inoculum on the performance of cowpea, along with its endophytic mycorrhizal community structure. A three-way factorial randomized design was established in sterilized and non-sterilized soils, including control plants (without inoculation), inoculated plants with a Bradyrhizobium (NFB) strain, a commercial mycorrhizal inoculum (AMF) and their combination (AMF+/NFB+). All inoculation treatments under sterile and non-sterile soil conditions resulted in a significant increase in the shoot biomass, which was positively correlated with AMF colonization, and the number of nodules found in cowpea roots. Mycorrhizal colonization and nodule formation in control plants grown in non-sterilized soils, suggested an active and functional local mycorrhizal and rhizobial community in the soil, which was further supported by the molecular fingerprints. Growth of control plants in sterile soil resulted in lower shoot biomass that was in line with the absence of the respective microbial assemblages. Interestingly, the local AMF community pattern of cowpea roots was drastically altered after the NFB application. Molecular fingerprinting, through cloning of PCR amplified sequences of 18sRNA revealed that cowpea plants were colonized by Rhizophagus, Funneliformis, Claroideoglomus and Paraglomus related species. The results support the notion that the introduced Bradyrhizobium strain interacted drastically with the indigenous AMF community, leading to a more diverse colonization pattern of cowpea roots. These findings enhance our understanding of how these microorganisms interact to affect plant performance and increase agricultural sustainability.

Identification of symbiotic nitrogen-fixing bacteria from three African leguminous trees in Gorongosa National Park

The symbiosis between leguminous plants and symbiotic nitrogen-fixing bacteria is a key component of terrestrial ecosystems. Woody legumes are well represented in tropical African forests but despite their ecological and socio-economic importance, they have been little studied for this symbiosis. In this study, we examined the identity and diversity of symbiotic-nitrogen fixing bacteria associated with Acacia xanthophloea, Faidherbia albida and Albizia versicolor in the Gorongosa National Park (GNP) in Mozambique. To the best of our knowledge, this is the first report on the identity of symbiotic-nitrogen fixing bacteria in this region. 166 isolates were obtained and subjected to molecular identification. BOX-A1R PCR was used to discriminate different bacterial isolates and PCR-sequencing of 16S rDNA, and two housekeeping genes, glnII and recA, was used to identify the obtained bacteria. The gene nifH was also analyzed to assess the symbiotic capacity of the obtained bacteria. All isolates from F. albida and Al. versicolor belonged to the Bradyrhizobium genus whereas isolates from Ac. xanthophloea clustered with Mesorhizobium, Rhizobium or Ensifer strains. Soil chemical analysis revealed significant differences between the soils occupied by the three studied species. Thus, we found a clear delimitation in the rhizobial communities and soils associated with Ac. xanthophloea, F. albida and Al. versicolor, and higher rhizobial diversity for Ac. xanthophloea than previously reported.

Biodiversity and biogeography of rhizobia associated with common bean (Phaseolus vulgaris L.) in Shaanxi Province

The biodiversity and biogeography of rhizobia associated with bean in Shaanxi Province were investigated. A total of 194 bacterial isolates from bean nodules collected from 13 sampling sites were characterized based on phylogenetic analyses of the 16S rRNA gene, the housekeeping genes recA, glnII and atpD, and the symbiotic genes nodC and nifH. Fifteen genospecies belonging to the genera Rhizobium, Agrobacterium, Ensifer, Bradyrhizobium and Ochrobactrum were defined among the isolates, with Rhizobium sp. II, Agrobacterium sp. II, E. fredii and R. phaseoli being the dominant groups. Four symbiotic gene lineages corresponding to Rhizobium sp. I, Rhizobium sp. II, R. phaseoli and B. liaoningense were detected in the nodC and nifH sequence analyses, indicating different origins for the symbiotic genes and their co-evolution with the chromosome of the bacteria. Moreover, the Ensifer isolates harbored symbiotic genes closely related to bean-nodulating Pararhizobium giardinii, indicating possible lateral gene transfer from Rhizobium to Ensifer. Correlation of rhizobial community composition with moisture, temperature, intercropping, soil features and nutrients were detected. All the results demonstrated a great diversity of bean rhizobia in Shaanxi that might be due to the adaptable evolution of the bean-nodulating rhizobia subjected to the diverse ecological conditions in the area.

Differential plant invasiveness is not always driven by host promiscuity with bacterial symbionts.

Identification of mechanisms that allow some species to outcompete others is a fundamental goal in ecology and invasive species management. One useful approach is to examine congeners varying in invasiveness in a comparative framework across native and invaded ranges. Acacia species have been widely introduced outside their native range of Australia, and a subset of these species have become invasive in multiple parts of the world. Within specific regions, the invasive status of these species varies. Our study examined whether a key mechanism in the life history of Acacia species, the legume-rhizobia symbiosis, influences acacia invasiveness on a regional scale. To assess the extent to which species varying in invasiveness correspondingly differ with regard to the diversity of rhizobia they associate with, we grew seven Acacia species ranging in invasiveness in California in multiple soils from both their native (Australia) and introduced (California) ranges. In particular, the aim was to determine whether more invasive species formed symbioses with a wider diversity of rhizobial strains (i.e. are more promiscuous hosts). We measured and compared plant performance, including aboveground biomass, survival, and nodulation response, as well as rhizobial community composition and richness. Host promiscuity did not differ among invasiveness categories. Acacia species that varied in invasiveness differed in aboveground biomass for only one soil and did not differ in survival or nodulation within individual soils. In addition, acacias did not differ in rhizobial richness among invasiveness categories. However, nodulation differed between regions and was generally higher in the native than introduced range. Our results suggest that all Acacia species introduced to California are promiscuous hosts and that host promiscuity per se does not explain the observed differences in invasiveness within this region. Our study also highlights the utility of assessing potential mechanisms of invasion in species' native and introduced ranges.

Partner diversity and identity impacts on plant productivity in Acacia–rhizobial interactions

Summary Genetic variation for functionally important traits is ubiquitous in communities of nitrogen‐fixing rhizobia, and while some studies have described significant effects of diversity on the functioning of plant‐associated microbial communities, we lack a systematic test of how rhizobial diversity influences plant productivity. The complexity of potential interactions among rhizobia and plants complicates the development of general predictions regarding causal relationships between rhizobial diversity and plant productivity. For example, while rhizobial complementarity may result in positive associations between symbiont diversity and plant productivity, antagonistic competition may reduce rhizobial community function. Using two widespread native Australian Acacia species (A. salicina, A. stenophylla) and experimental rhizobial communities derived from 16 bacterial genotypes naturally associated with these hosts, we examined how the provision of mutualistic benefit varies with rhizobial identity, diversity and phylogenetic relatedness. Analysis of plant performance in relation to rhizobial genotypic richness revealed that the presence of multiple rhizobial genotypes in the rhizosphere was associated with a general decrease in plant productivity compared to growth with single rhizobial genotypes. Importantly, these results appear to be robust in the face of variation in host identity and host diversity (i.e. one or two species mixtures). We also found that rhizobial genotypic identity and host species significantly influenced plant productivity in Acacia–rhizobia interactions, both in single‐ and multistrain inoculations. Synthesis. Together, our data show that multiple rhizobia interacting with a single host species creates opportunities for emergent or higher‐order effects that extend beyond those that could be simply predicted based upon outcomes of pairwise interactions and that increased mutualist diversity does not necessarily translate into positive effects on plant growth.

Salt tolerance of rhizobial populations from contrasting environmental conditions: understanding the implications of climate change.

It is predicted that global climate change may alter environmental parameters such as rainfall distribution which in turn may alter the salinity of soils with unpredictable effects upon soil microbial populations. In the present work the tolerance to salinity of rhizobia, isolated from locations with contrasting climatic conditions, and the potential of strains to fix nitrogen symbiotically under saline conditions were investigated. Since plasmids may encode key genes related to growth and survival under environmental stress conditions, which will reflect on protein synthesis, both the plasmid and protein profiles were analyzed. A multivariate statistical approach related salt tolerance to the origin of the isolates, identifying rainfall and water availability as a possible factor explaining the differences in salt tolerance displayed by rhizobia isolates. The classification analysis allowed the subdivision of isolates in terms of salt tolerance into extremely sensitive (≤0.15%), sensitive (0.15-0.6%), moderately tolerant (0.9-1.5%), tolerant (2.1-3.6%) and extremely tolerant (≥5.4%). Taken all together it was shown that plasmids are involved in salt tolerance and that the impact of salinity on the protein profile and nitrogen fixation varied according to the salt tolerance of the strains, evidencing the susceptibility of rhizobial communities to changes in rainfall regimes.

Rhizobia Indigenous to the Okavango Region in Sub-Saharan Africa: Diversity, Adaptations, and Host Specificity

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.

Diversity and distribution of rhizobia nodulated with Phaseolus vulgaris in two ecoregions of China

To reveal the diversity and geographic distribution of bean (Phaseolus vulgaris L.) rhizobia in China, a total of 226 strains of nodule bacteria were isolated from bean plants grown in fields of nine sites in northern (ecoregion I) and southern (ecoregion II) regions of China, with soil pH ranged from 8.06 to 4.97. Based upon the phylogenies of housekeeping and symbiotic genes, 96.9% of the isolates were Rhizobium corresponding to four defined species and four unnamed genospecies, while the remaining were five Bradyrhizobium genospecies. Rhizobium leguminosarum, Rhizobium etli and Rhizobium sp. II or IV were dominant in different ecoregions, with varied relative abundances. Therefore, diverse and distinctive bean rhizobial communities exist in different ecoregions of China, demonstrating that the introduction of bean plants has driven the evolution of rhizobia to fit the necessity to nodulate the host plant under the local conditions, mainly the soil pH and the nutrient availability. The predominance of R. etli and the high similarities in the symbiotic genes (nodC and nifH) of all the Rhizobium genospecies with R. etli evidenced that symbiotic genes were transferred from R. etli, possibly introduced to China accompanying the seeds, to the related indigenous rhizobia. These findings enlarged the diversity of bean rhizobia, evidenced the biogeographic patterns of these rhizobia, and demonstrated the possible evolution and emerging of novel rhizobia under the combined selection of host plant and soil conditions.

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.

Phenotypic and molecular fingerprinting of fast growing rhizobia of field-grown pigeonpea from the eastern edge of the Brazilian Pantanal

The aim of this study was to evaluate the diversity of rhizobial isolates obtained from root nodules of pigeonpea plants grown at the eastern edge of the Brazilian Pantanal. The bacterial isolates were isolated from root nodules from field-growing pigeonpea grown in two rural settlements of the Aquidauana municipality. The bacterial isolates were characterized phenotypically by means of cultural characterization, intrinsic antibiotic resistance (IAR), salt and high incubation temperature tolerance, and amylolytic and cellulolytic activities. The molecular characterization of the bacterial isolates was carried out using amplified ribosomal DNA restriction analysis (ARDRA) and Box-polymerase chain reaction (PCR) techniques. In addition, the symbiotic performance of selected rhizobial isolates was evaluated in a greenhouse experiment using sterile substrate. The phenotypic characterization revealed that the bacterial strains obtained from pigeonpea root nodules presented characteristics that are uncommon among rhizobial isolates, indicating the presence of new species nodulating the pigeonpea plants in the Brazilian Pantanal. The molecular fingerprinting of these bacterial isolates also showed a highly diverse collection, with both techniques revealing less than 25% similarity among bacterial isolates. The evaluation of symbiotic performance also indicated the presence of microorganisms with high potential to increase the growth and nitrogen content at the shoots of pigeonpea plants. The results obtained in this study indicate the presence of a highly diversified rhizobial community nodulating the pigeonpea at the eastern edge of the Brazilian Pantanal.

Co-introduction of exotic rhizobia to the rhizosphere of the invasive legume Acacia saligna, an intercontinental study

Invasive woody legumes have profound impacts in the nitrogen content and cycling of invaded ecosystems due to the ability to enter into symbiosis with nitrogen-fixing bacteria. In spite of the relevance of this symbiosis, the identity and origin of the symbionts involved in invasion are not well understood. We conducted a study to assess the diversity of symbiotic root-nodulating bacteria associated with the invasive Acacia saligna, in newly colonized areas in Portugal and Australia. BOX-PCR was used to discriminate the isolated bacteria and 16S rRNA and nifD genes were sequenced to identify the different isolates and their geographic origin. Bradyrhizobium and Mesorhizobium nodulated A. saligna in Australia while only Bradyrhizobium spp. were found in Portugal. The dominant strains nodulating A. saligna were related to Bradyrhizobium liaoningense and Bradyrhizobium canariense. Co-occurring Acacia longifolia and A. saligna in Australia harbor different rhizobial communities. As an example, we found Mesorhizobium sp. and Phyllobacterium trifolii in A. saligna and A. longifolia respectively, being this the first report for this association. The analysis of the phylogeographic marker nifD clustered most of the sequences obtained in this study with sequences of Australian origin, indicating that exotic bradyrhizobia might have been co-introduced with A. saligna in Portugal. This result highlights the risks of introducing exotic inoculants that might facilitate the invasion of new areas and alter native soil bacterial communities, hindering the recovery of ecosystems.

Mutualisms are not constraining cross‐continental invasion success of Acacia species within Australia

AbstractAimStudying plant–soil interactions of introduced species in different parts of their global range could assist in managing biological invasions by elucidating the level of host specificity of key mutualists. We assessed the role of the soil microbial community (with an emphasis on symbiotic nitrogen‐fixing bacteria, collectively termed rhizobia) in determining cross‐continental invasion success of five woody legume species.LocationAustralia.MethodsFor each species, we compared growth of plants in soils from their native and non‐native ranges using a glasshouse study, a soil dilution method (most probable number) and T‐RFLP to assess rhizobial abundance and community composition, respectively.ResultsAcacia longifolia was the only species that had significantly larger above‐ground biomass when grown in soils from its non‐native range. Rhizobial abundance was equally high across species and ranges, indicating plants are unlikely to be limited by soil rhizobial abundance in non‐native ranges. Acacia cyclops, A. saligna and Paraserianthes lophantha formed associations with different rhizobial communities in non‐native vs. native range soils. Acacia longifolia and A. melanoxylon associated with similar rhizobial communities in their native and non‐native ranges, suggesting that rhizobia may have been accidentally introduced into their novel range with seeds or seedlings.Main conclusionsInvasive success of these five legume species is not constrained by the abundance of rhizobia in novel ranges for established legume populations, at least within Australia. Although differences in rhizobial community composition were evident between the native and non‐native ranges for three of the five species, these were not associated with differences in plant growth. Increased above‐ground biomass of A. longifolia when grown in soil from its non‐native range suggests that invasive success of this species may be associated with differences in the non‐rhizobial components of soil microbial communities in the novel range. This information could assist in management practises by facilitating a more instructive and effective screening for invasiveness.