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Afforestation is an effective approach to rehabilitate degraded ecosystems, but often depletes deep soil moisture. Presently, it is not known how an afforestation-induced decrease in moisture affects soil microbial community and functionality, hindering our ability to understand the sustainability of the rehabilitated ecosystems. To address this issue, we examined the impacts of 20 years of afforestation on soil bacterial community, co-occurrence pattern, and functionalities along vertical profile (0-500 cm depth) in a semiarid region of China's Loess Plateau. We showed that the effects of afforestation with a deep-rooted legume tree on cropland were greater in deep than that of in top layers, resulting in decreased bacterial beta diversity, more responsive bacterial taxa and functional groups, increased homogeneous selection, and decreased network robustness in deep soils (120-500 cm). Organic carbon and nitrogen decomposition rates and multifunctionality also significantly decreased by afforestation, and microbial carbon limitation significantly increased in deep soils. Moreover, changes in microbial community and functionality in deep layer was largely related to changes in soil moisture. Such negative impacts on deep soils should be fully considered for assessing afforestation's eco-environment effects and for the sustainability of ecosystems because deep soils have important influence on forest ecosystems in semiarid and arid climates.

Predicting the distribution patterns of soil microbial communities requires consideration of more environmental drivers. The effects of soil micronutrients on composition of microbial communities are largely unknown despite micronutrients closely relating to soil fertility and plant communities. Here we used data from 228 agricultural fields to identify the importance of micronutrients (iron, zinc, copper and manganese) in shaping structure of soil microbial communities (bacteria, fungi and protist) along latitudinal gradient over 3400 km, across diverse edaphic conditions and climatic gradients. We found that micronutrients explained more variations in the structure of microbial communities than macronutrients in maize soils. Moreover, micronutrients, particularly iron and copper, explained a unique percentage of the variation in structure of microbial communities in maize soils even after controlling for climate, soil physicochemical properties and macronutrients, but these effects were stronger for fungi and protist than for bacteria. The ability of micronutrients to predict the structure of soil microbial communities declined greatly in paddy soils. Machine learning approach showed that the addition of micronutrients substantially increased the predictive power by 9–17% in predicting the structure of soil microbial communities with up to 69–78% accuracy. These results highlighted the considerable contributions of soil micronutrients to microbial community structure, and advocated that soil micronutrients should be considered when predicting the structure of microbial communities in a changing world.

It is becoming increasingly evident about interactions between plant hosts and their microbiotas, while the effects of plant communities on microbial communities in different geographic environments are poorly understood. Here, the differentiation of licorice plant ecotypes and the distribution of root-associated microbiotas were investigated across five sampling sites in northwest China. The interactions between the environment, plant and microbial communities, and their effects on licorice root secondary metabolites, were elucidated. The plant community was clearly differentiated into distinct ecotypes based on genotyping-by-sequencing and was primarily driven by geographic distance and soil available nitrogen. The bulk soil and root-associated microbiotas (rhizosphere soil and root endosphere) partially correlated with plant community, but all were significantly discriminated by plant clade. Moreover, these microbiotas were explained to different extents by distinct combinations of environment, geography and plant community. Similarly, three structural equation models showed that licorice root secondary metabolites were complicatedly modulated by multiple abiotic and biotic factors; and were mostly explained by these factors in the rhizosphere model. Collectively, the results provide novel insights into the role of environment-plant-microbiotas interactions in regulating root secondary metabolites. That should be accounted for when selecting appropriate licorice planting sites and management measures. This article is protected by copyright. All rights reserved.

Background Plant health and growth are negatively affected by pathogen invasion; however, plants can dynamically modulate their rhizosphere microbiome and adapt to such biotic stresses. Although plant-recruited protective microbes can be assembled into synthetic communities for application in the control of plant disease, rhizosphere microbial communities commonly contain some taxa at low abundance. The roles of low-abundance microbes in synthetic communities remain unclear; it is also unclear whether all the microbes enriched by plants can enhance host adaptation to the environment. Here, we assembled a synthetic community with a disease resistance function based on differential analysis of root-associated bacterial community composition. We further simplified the synthetic community and investigated the roles of low-abundance bacteria in the control of Astragalus mongholicus root rot disease by a simple synthetic community. Results Fusarium oxysporum infection reduced bacterial Shannon diversity and significantly affected the bacterial community composition in the rhizosphere and roots of Astragalus mongholicus. Under fungal pathogen challenge, Astragalus mongholicus recruited some beneficial bacteria such as Stenotrophomonas, Achromobacter, Pseudomonas, and Flavobacterium to the rhizosphere and roots. We constructed a disease-resistant bacterial community containing 10 high- and three low-abundance bacteria enriched in diseased roots. After the joint selection of plants and pathogens, the complex synthetic community was further simplified into a four-species community composed of three high-abundance bacteria (Stenotrophomonas sp., Rhizobium sp., Ochrobactrum sp.) and one low-abundance bacterium (Advenella sp.). Notably, a simple community containing these four strains and a thirteen-species community had similar effects on the control root rot disease. Furthermore, the simple community protected plants via a synergistic effect of highly abundant bacteria inhibiting fungal pathogen growth and less abundant bacteria activating plant-induced systemic resistance. Conclusions Our findings suggest that bacteria with low abundance play an important role in synthetic communities and that only a few bacterial taxa enriched in diseased roots are associated with disease resistance. Therefore, the construction and simplification of synthetic communities found in the present study could be a strategy employed by plants to adapt to environmental stress.

Outer membrane vesicles (OMVs) can function as nanoscale vectors that mediate bacterial interactions in microbial communities. How bacteria recognize and recruit OMVs inter-specifically remains largely unknown, thus limiting our understanding of the complex physiological and ecological roles of OMVs. Here, we report a ligand-receptor interaction-based OMV recruitment mechanism, consisting of a type VI secretion system (T6SS)-secreted lipopolysaccharide (LPS)-binding effector TeoL and the outer membrane receptors CubA and CstR. We demonstrated that Cupriavidus necator T6SS1 secretes TeoL to preferentially associate with OMVs in the extracellular milieu through interactions with LPS, one of the most abundant components of OMVs. TeoL associated with OMVs can further bind outer membrane receptors CubA and CstR, which tethers OMVs to the recipient cells and allows cargo to be delivered. The LPS-mediated mechanism enables bacterial cells to recruit OMVs derived from different species, and confers advantages to bacterial cells in iron acquisition, interbacterial competition, and horizontal gene transfer (HGT). Moreover, our findings provide multiple new perspectives on T6SS functionality in the context of bacterial competition and HGT, through the recruitment of OMVs.

"Fertile islands" are known as hotspots of soil nutrient spatial heterogeneity in dryland ecosystems. Although soil microorganisms play critical functional roles in nutrient capture and cycling within fertile islands, our understanding of polymicrobial community roles in regulating soil nutrient distribution in fertile islands remains limited. Herein, we aim to clarify the relationships between the complexity of soil microbial (bacterial, archaeal and fungal) communities and the nutrient distribution around fertile islands. Soil samples were collected along vertical profiles at varying depths in three patches under the canopy of Hedysarum scoparium (CENTRE), at the edge (EDGE) of the canopy and outside (OUTSIDE) the canopy in an area of flowing sand. All the three microbiota have showed spatial heterogeneity around the fertile islands. Among them, bacteria had the most significant heterogeneity, and bacterial community assembly was dominated by deterministic processes. Microbial interaction patterns also showed spatial heterogeneity among different patches. More interaction complexity within microbiota was found in the bacteria in the CENTRE patch and in the fungi in the OUTSIDE patch. In addition, the proportions of among-kingdom connections were reduced under the canopies. Bacteria had the highest connectivity and centrality in the polymicrobial networks and were the most important predictor of polymicrobial interaction complexity, which may have contributed to the distribution of soil nutrients. The random forest (RF) model provided evidence that bacterial beta-diversity and the polymicrobial network complexity index can be optimal predictors of the soil multinutrient cycling index. Our study highlighted the responses of bacteria and polymicrobial interactions to fertile islands and their importance in driving soil nutrient heterogeneity. This information will help in managing soil microorganisms to provide dryland ecosystem services.

Although studies of biogeography in soil bacterial communities have attracted considerable attention, the generality of these patterns along with assembly processes and underlying drivers is poorly understood in the inner tissues of plants. Plant tissues provide unique ecological habitats for microorganisms, which play an essential role in plant performance. Here, we compared core bacterial communities among five soil-plant associated compartments of common bean across five sampling sites in China. Neutral and null modelling consistently suggested that stochastic processes dominated the core community assembly processes and escalated from the belowground compartments to the inner tissues of aerial plant parts. The multiple distance-decay relationships also varied and had flattened patterns in the stem endosphere, which were shaped by distinct environmental factors in each compartment. Coexistence patterns also varied in topological features, in addition with the sparsest networks in the stem endosphere resulted from the interaction with the stochastic processes. This study considerably expanded our understanding of various biogeographic patterns, assembly processes, and the underlying mechanisms of core bacterial communities between aerial and belowground compartments of common bean. That will provide a scientific basis for the reasonable regulation of core bacterial consortia to get better plant performance.

Aims Licorice (Glycyrrhiza uralensis Fisch.) is a crucial medicinal herb as it accumulates glycyrrhizin and liquiritin in roots. Licorice root-associated bacterial communities shaped by soil characteristics are supposed to regulate the accumulation of root secondary metabolites. Methods The soil characteristics, root secondary metabolites, and root-associated bacterial communities were analyzed in licorice plants of different ages to explore their temporal dynamics and interaction mechanisms. Results Temporal variation in soil characteristics and root secondary metabolites was distinct. The alpha-diversity of root-associated bacterial communities decreased with root proximity, and the community composition was clustered in the rhizosphere. Different taxa that were core-enriched from the dominant taxa in the bulk soil, rhizosphere soil, and root endosphere displayed varied time–decay relationships. Soil total potassium (TK) as a key factor regulated the temporal variation in some individual taxa in the bulk and rhizosphere soils; these taxa were associated with the adjustment of root secondary metabolites across different TK levels. Conclusions Licorice specifically selects root-associated core bacteria over the course of plant development, and TK is correlated with root secondary metabolites and individual core-enriched taxa in the bulk and rhizosphere soils, which may have implications for practical licorice cultivation. Electronic supplementary material The online version of this article (10.1007/s11104-020-04692-0) contains supplementary material, which is available to authorized users.

A better understanding of bacterial communities and metabolomic responses to pristine zinc oxide manufacture nanoparticles (ZnO MNPs) and its sulfidized product (s-ZnO MNPs), as well as their corresponding Zn ions in rhizocompartments, critical in the plant-microbe interactions, could contribute to the sustainable development of nano-enabled agriculture. In this study, soybean (Glycine max) were cultivated in soils amended with three Zn forms, namely ZnSO

As the main consumers of bacteria and fungi in farmed soils, protists remain poorly understood. The aim of this study was to explore protist community assembly and ecological roles in soybean fields. Here, we investigated differences in protist communities using high-throughput sequencing and their inferred potential interactions with bacteria and fungi between the bulk soil and rhizosphere compartments of three soybean cultivars collected from six ecological regions in China. Distinct protist community structures characterized the bulk soil and rhizosphere of soybean plants. A significantly higher relative abundance of phagotrophs was observed in the rhizosphere (25.1%) than in the bulk soil (11.3%). Spatial location (R2 = 0.37–0.51) explained more of the variation in protist community structures of soybean fields than either the compartment (R2 = 0.08–0.09) or cultivar type (R2 = 0.02–0.03). The rhizosphere protist network (76 nodes and 414 edges) was smaller and less complex than the bulk soil network (147 nodes and 880 edges), indicating a smaller potential of niche overlap and interactions in the rhizosphere due to the increased resources in the rhizosphere. Furthermore, more inferred potential predator-prey interactions occur in the rhizosphere. We conclude that protists have a crucial ecological role to play as an integral part of microbial co-occurrence networks in soybean fields.

Ammonia oxidizing archaea (AOA) and bacteria (AOB), nitrite-oxidizing bacteria (NOB), and comammox Nitrospira (CMX) play pivotal roles in global nitrogen-cycling network. Despite its importance, the driving forces for niche specialization of these nitrifiers, as well as their relative contributions to nitrification and crop yield have not been fully understood. Here, we investigated the niche specialization and environmental prevalence of nitrifying communities, and their importance for the nitrification rate and crop yield across a gradient of nitrogen inputs in a two-decade old field experiment. The results of

Phosphorus (P) is essential for crop growth as an indispensable nutrient; however, there has been growing concern over the low use efficiency of P used in current fertilizers. We synthesized and characterized a potential P fertilizer nanohydroxyapatite/biochar/sodium humate (nHAP/BC/HANa) composite. To study the impact of the composite on soil chemical properties and microbial community in sandy soils, we set up four treatments as follows: (1) biochar (BC), (2) nanohydroxyapatite (nHAP), (3) nHAP/BC/HANa composite, and (4) sodium humate (HANa) was added separately into soils amended with nHAP/BC (nHAP/BC + HANa) to compare its performance with that of the nHAP/BC/HANa composite. A key finding was that the nHAP/BC/HANa composite not only significantly increased the soil available P content and alkaline phosphatase activity but also the increased organic matter content compared to the control. Additionally, leaching losses of P in soils amended with the nHAP/BC/HANa composite were lower than those in soils amended with the nHAP/BC + HANa, which suggested that the nHAP/BC/HANa composite had great potential to decrease P loss in sandy soils. Moreover, bacterial communities were more sensitive than fungal communities to all treatments. The bacterial communities showed the most significant changes in the nHAP/BC/HANa treatments. Results from Mantel tests further indicated that the strongest correlation between bacterial communities and soil properties occurring in the nHAP/BC/HANa treatments. Random forest analysis was conducted to identify the dominant microbial taxa, such as Proteobacteria, Acidobacteria, and Gemmatimonadetes, for predicting changes in soil properties. There was an asymptotical transition in bacterial community assembly processes from stochastic to deterministic in the nHAP/BC/HANa treatments. In conclusion, we demonstrated that nHAP/BC/HANa composite had the remarkable contribution to soil P availability in sandy soils, and simultaneously promoted the bacterial functions potential for P cycling, which present valuable insights to the development of potential P fertilizer.

Elucidating the mechanisms underpinning the responses of abundant and rare microbial taxa to environmental disturbances is essential for understanding the biodiversity-stability relationship and maintaining microbial diversity. Here, we explored the response patterns of abundant and rare bacterial taxa to disturbances by invasive plant growth and oil contamination in agricultural soils across a large spatial scale (latitude gradient = 18.62°–46.51°). Our meta-analysis based on existing Illumina sequencing datasets showed that abundant taxa persisted under the disturbances whereas rare taxa were more easily affected, indicating the higher resilience or resistance of abundant taxa to disturbances. The responses of abundant taxa were associated with mean annual temperature at the sampling sites, while rare taxa instead showed stochastic responses. There were significantly negative linear regressions between bacterial α-diversity and community dissimilarities (disturbed vs. undisturbed soils), suggesting stronger resilience or resistance in those bacterial communities with higher α-diversity. This resilience or resistance was mainly associated with the α-diversity of abundant taxa. Our network analysis showed that the disturbances substantially decreased the strength of the connections, loosened the co-occurrence relationships, and reshaped the complex bacterial interactions. In the undisturbed soils, abundant taxa were located in central positions within the network more often than were rare taxa, while these trends were reversed in the disturbed soils. Our results suggest that abundant taxa play a dominant role in the stability and maintenance of agro-soil bacterial communities, while rare taxa could greatly influence local bacterial interactions under environmental disturbances.

Understanding the influences of microbial interactions and niche heterogeneities on microbial communities and functional traits is critical for determining its engineering and ecological significance. However, little is known about microbial community assemblage and functional gene expression throughout full-scale landfill leachate treatment plants. Here, we applied a combination of 16S rRNA and rDNA amplicon sequencing, shotgun metagenomic, and qPCR approaches to unveil the ecological associations between distinct communities, functional gene expression and nitrogen cycling processes. By comparing the rDNA and rRNA-derived communities, the rRNA/rDNA ratios suggested that 57.2% of rare taxa were active, and their abundance decreased as increasing of potential activities. In particular, rDNA- and rRNA-based communities exhibited divergent assemblage patterns, and stronger intra-associations among core taxa in the rRNA-based communities than in rDNA-based communities. Furthermore, results regarding both bacterial assemblage and functional traits indicated that the habitat filtering and niche differentiation (treatment units) exerted selection on microbial communities based on functional traits, particular for key ecological functions related to nitrogen cycling. Collectively, our findings provide insights into structure-function associations at the local level and shed light on ecological rules guiding rDNA- and rRNA-based community assembly in landfill leachate treatment systems.

Salinization is an important global environmental problem influencing sustainable development of terrestrial ecosystems. Salt-tolerant halophytes are often used as a promising approach to remedy the saline soils. Yet, how rhizosphere microbes' association and functions vary with halophytes in saline ecosystems remains unclear, restricting our ability to assess the role of halophytes in remedying saline ecosystems. Herein, we examined bacterial and fungal diversities, compositions, and co-occurrence networks in the rhizospheres of six halophytes and bulk soils in a semiarid inland saline ecosystem, and related these parameters to microbial functions. The microbiomes were more diverse and complex and microbial activity and residues were higher in rhizospheres than bulk soils. The connections of taxa in the rhizosphere microbial communities increased with fungi-fungi and bacteria-fungi connections and fungal diversity. The proportion of the fungi-related central connections were larger in rhizospheres (13-73%) than bulk soils (3%). Moreover, microbial activity and residues were significantly correlated with microbial composition and co-occurrence network complexity. These results indicated that enhanced association between fungi and bacteria increased microbial co-occurring network complexity in halophytes rhizosphere, which contributed to the higher microbial functions (microbial activities and residue) in this inland saline ecosystem.

Multiple heavy metals (HMs) commonly coexist in mining areas, which highlights the necessity to select multiple HM-resistant plant growth-promoting bacteria for improving phytoremediation efficiency. In this study, we isolated and characterized 82 endophytic bacteria from the root nodules of black locust (Robinia pseudoacacia) grown in a Pb-Zn mining area. There were 80 isolates showing resistance to four HMs, 0.01–18.0 mM/L for Cd, 0.2–40.0 mM/L for Zn, 0.3–2.2 mM/L for Pb, and 0.2–1.4 mM/L for Cu. Indole-3-acetic acid production, siderophore production, and 1-aminocyclopropane-1-carboxylate deaminase activity were detected in 43, 50, and 17 isolates, respectively. Two symbiotic isolates selected with the highest potential for HM resistance and PGP traits, designated Mesorhizobium loti HZ76 and Agrobacterium radiobacter HZ6, were evaluated for promotion of plant growth and metal uptake by R. pseudoacacia seedlings grown in pots containing different levels of Cd, Zn, Pb, or Cu. HZ76 significantly increased plant shoot biomass, while HZ6 did not, compared with non-inoculated controls. The results indicate that inoculation with HZ76 or HZ6 relieved HM stress in the plants, depending on the type and concentration of HM in the treatment. Mesorhizobium loti HZ76 may be a better candidate for application in phytoremediation than A. radiobacter HZ6. The microsymbiosis between HM-resistant rhizobia and R. pseudoacacia is an interesting mutualistic system for phytoremediation in mining areas contaminated with multiple HMs.

Astragalus membranaceus (Fisch.) Bge. var. mongholicus, which is used in traditional Chinese medicine, contains several bioactive ingredients. The root-associated microbial communities play a crucial role in the production of secondary metabolites in plants. However, the correlation of root-associated bacteria and fungi with the bioactive ingredients production in A. mongholicus has not been elucidated. This study aimed to examine the changes in soil properties, root bioactive ingredients, and microbial communities in different cultivation years. The root-associated bacterial and fungal composition was analyzed using high-throughput sequencing. The correlation between root-associated bacteria and fungi, soil properties, and six major bioactive ingredients were examined using multivariate correlation analysis. Results showed that soil properties and bioactive ingredients were distinct across different cultivation years. The composition of the rhizosphere microbiome was different from that of the root endosphere microbiome. The bacterial community structure was affected by the cultivation year and exhibited a time-decay pattern. Soil properties affected the fungal community composition. It was found that 18 root-associated bacterial operational taxonomic units (OTUs) and four fungal OTUs were positively and negatively correlated with bioactive ingredient content, respectively. The abundance of Stenotrophomonas in the rhizosphere was positively correlated with astragaloside content. Phyllobacterium and Inquilinus in the endosphere were positively correlated with the calycosin content. In summary, this study provided a new opportunity and theoretical reference for improving the production and quality of in A. mongholicus, which thus increase the pharmacological value of A. mongholicus.

Unraveling the succession of microbial communities is a core ecological research topic. Yet few studies have focused on how long-term secondary succession affects the functional profiles and ecological processes of abundant and rare microbial subcommunities. Here, we used amplicon sequencing and GeoChip analysis to explore the ecological functions of abundant and rare biospheres and their correlation with soil multinutrient cycling. Samples for this study were collected from a well-established secondary succession chronosequence that spans >30 years of dryland ecosystem development on the Loess Plateau of China. Although both abundant and rare subcommunities shifted with succession, the changing of beta-diversity of the microbial communities was primarily driven by species replacement of the rare biosphere. Phylogenetic changes of abundant and rare taxa were associated with their functional traits, which dominated the diversity-related selection along all succession ages. Neutral theory analysis indicated that the assemblage of abundant taxa over all successional ages was regulated by dispersal homogenizing and ecological drift. The null model revealed that homogeneous and variable selection were the dominant assembly processes for rare subcommunities compared with abundant species. pH and nitrogen content were the paramount drivers determining the assembly of microbial communities and functional genes, consistent with the importance of environmental filtering. Furthermore, the rare biosphere had a paramount role in the entire ecological network and was the major driver for most soil processes such as C, N, and S cycling. Nonetheless, a significant portion of soil P cycling was regulated by abundant taxa. Collectively, our study provides insight into the mechanisms underlying microbial community assembly and soil microbe-driven functional changes in biogeochemical processes during secondary succession.

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

Under nitrogen-limiting conditions, symbiotic nodulation promotes the growth of legume plants via the fixation of atmospheric nitrogen to ammonia by rhizobia in root nodules. The rhizobial Nod factor (NF) and type III secretion system (T3SS) are two key signaling pathways for establishing the legume–rhizobium symbiosis. However, whether NF signaling is involved in the nodulation of Robinia pseudoacacia and Mesorhizobium amorphae CCNWGS0123, and its symbiotic differences compared with T3SS signaling remain unclear. Therefore, to elucidate the function of NF signaling in nodulation, we mutated nodC in M. amorphae CCNWGS0123, which aborted NF synthesis. Compared with the plants inoculated with the wild type strain, the plants inoculated with the NF-deficient strain exhibited shorter shoots with etiolated leaves. These phenotypic characteristics were similar to those of the plants inoculated with the T3SS-deficient strain, which served as a Nod− (non-effective nodulation) control. The plants inoculated with both the NF- and T3SS-deficient strains formed massive root hair swellings, but no normal infection threads were detected. Sections of the nodules showed that inoculation with the NF- and T3SS-deficient strains induced small, white bumps without any rhizobia inside. Analyzing the accumulation of 6 plant hormones and the expression of 10 plant genes indicated that the NF- and T3SS-deficient strains activated plant defense reactions while suppressing plant symbiotic signaling during the perception and nodulation processes. The requirement for NF signaling appeared to be conserved in two other leguminous trees that can establish symbiosis with M. amorphae CCNWGS0123. In contrast, the function of the T3SS might differ among species, even within the same subfamily (Faboideae). Overall, this work demonstrated that nodulation of R. pseudoacacia and M. amorphae CCNWGS0123 was both NF and T3SS dependent.

Nitrogen-transforming microorganisms play pivotal roles for the microbial nitrogen-cycling network in the anammox granular system. However, little is known about the effects of inorganic carbon (IC) stresses on the assemblage patterns and functional profiles of abundant and rare taxa. Herein, the community assemblage and functional traits of abundant and rare sub-communities were investigated. Results revealed that insufficient IC had adverse influences on the process performance, while anammox activity could be recovered by IC addition. Co-occurrence network analysis indicated that abundant and rare sub-communities present divergent co-occurrence patterns. Additionally, environmental filtering had different influences on the ecological adaptability of bacterial sub-communities. Furthermore, qPCR results illustrated that NH4+-N and NO2--N consumption were regulated by abundant sub-community, while their accumulation was mediated by rare sub-community. Collectively, these findings suggest that abundant and rare sub-communities present contrasting assemblage patterns and metabolic pathways, and functional profiles dominated selection of bacterial sub-communities under IC stresses.

Background Soil microbiomes play an important role in the services and functioning of terrestrial ecosystems. However, little is known of their vertical responses to restoration process and their contributions to soil nutrient cycling in the subsurface profiles. Here, we investigated the community assembly of soil bacteria, archaea, and fungi along vertical (i.e., soil depths of 0–300 cm) and horizontal (i.e., distance from trees of 30–90 cm) profiles in a chronosequence of reforestation sites that represent over 30 years of restoration. Results In the superficial layers (0–80 cm), bacterial and fungal diversity decreased, whereas archaeal diversity increased with increasing soil depth. As reforestation proceeded over time, the vertical spatial variation in bacterial communities decreased, while that in archaeal and fungal communities increased. Vertical distributions of the soil microbiomes were more related to the variation in soil properties, while their horizontal distributions may be driven by a gradient effect of roots extending from the tree. Bacterial and archaeal beta-diversity were strongly related to multi-nutrient cycling in the soil, respectively, playing major roles in deep and superficial layers. Conclusions Taken together, these results reveal a new perspective on the vertical and horizontal spatial variation in soil microbiomes at the fine scale of single trees. Distinct response patterns underpinned the contributions of soil bacteria, archaea, and fungi as a function of subsurface nutrient cycling during the reforestation of ex-arable land. Electronic supplementary material The online version of this article (10.1186/s40168-018-0526-0) contains supplementary material, which is available to authorized users.

Root-associated bacteria have profound effects on plant health and productivity, but their biogeographic patterns across large spatial scales remain poorly understood. Here, we used high-throughput sequencing to compare the bacterial distributions in the bulk soil, rhizosphere, and endosphere across 51 soybean fields in China. Environmental variables were more important than spatial variables, and edaphic variables were more important than climatic variables, for governing bacterial community turnover in each soil-root compartment. Both bacterial richness and community turnover were significantly correlated with different environmental and spatial variables among the three compartments. Their different spatial autocorrelation ranges for bacteria suggested distinct bacterial biogeographic patterns were present. The distributions of nearest taxon index (NTI) showed that deterministic processes dominated local bacterial communities, while its importance decreased from the bulk soil to the endosphere. These results provide new insights into the assembly of root-associated bacterial communities at a continental scale.

Heavy metals can cause serious contamination of soils, especially in mining regions. A detailed understanding of the effects of heavy metals on plants and root-associated microbial communities could help to improve phytoremediation systems. In this study, black locust (Robinia pseudoacacia) seedlings with or without rhizobial inoculation were planted in soils contaminated with different levels of heavy metals. Bacterial communities in rhizosphere and bulk soil samples were analyzed using 16S rRNA gene sequencing on the Illumina MiSeq platform and shotgun metagenome sequencing on the Illumina HiSeq platform. Soil bacterial communities varied significantly depending on the level of soil contamination, and planting also had some influence. Although inoculation of Mesorhizobium loti HZ76 (a natural microsymbiont of R. pseudoacacia) was a relatively minor factor, it did influence the soil bacterial community. Under the selective pressure, plant growth promotion-related biomarkers in the rhizosphere increased after inoculation compared with non-inoculated controls, especially those associated with Mesorhizobium, Variovorax, Streptomyces, and Rhodococcus genera. Genes encoding ATP-binding cassette transporters were up-regulated in the rhizosphere after inoculation compared with genes related to sulfur/nitrogen metabolism. These results provide insight into soil bacterial communities and their functions in the R. pseudoacacia rhizosphere in response to rhizobial inoculation and heavy metal contamination. This knowledge may prove useful for improving phytoremediation of metal-contaminated soils.

Revealing the biogeographies and ecologies of rare and abundant microorganisms is crucial to understand ecosystem diversity and function. In this study, we investigated the biogeographic assemblies and ecological diversity patterns of rare and abundant bacteria in long-term oil-contaminated soils at intervals of 46–360 km by performing high-throughput sequencing of 16S rRNA genes. The results clearly revealed distinct distribution patterns for rare and abundant bacteria in soil samples. Rare taxa were unevenly distributed; however, abundant taxa were ubiquitous across all samples. Both rare and abundant sub-communities showed significant distance-decay relationships, and their assemblies were driven by different factors. The rare sub-community primarily exhibited a spatially structured distribution (i.e., stochastic processes), while edaphic factors (i.e., deterministic processes) largely contributed to the structure of the abundant sub-community. A network analysis revealed closer relationships between abundant bacteria and their heightened influence on other co-occurrences in the community compared with rare species. In conclusion, rare microbial taxa may play potential roles in maintaining ecosystem diversity, although they do not appear to be central to microbial networks. Abundant microbes are vital for microbial co-occurrences in oil-contaminated soils, and high relative abundance and ubiquitous distribution suggest potential roles in the degradation of organic pollutants. This article is protected by copyright. All rights reserved.

Elucidating the driving forces behind the temporal dynamics of abundant and rare microbes is essential for understanding the assembly and succession of microbial communities. Here, we explored the successional trajectories and mechanisms of abundant and rare bacteria via soil-enrichment subcultures in response to various pollutants (phenanthrene, n-octadecane, and CdCl2) using time-series Illumina sequencing datasets. The results reveal different successional patterns of abundant and rare sub-communities in eighty pollutant-degrading consortia and two original soil samples. A temporal decrease in α-diversity and high turnover rate for β-diversity indicate that deterministic processes are the main drivers of the succession of the abundant sub-community; however, the high cumulative species richness indicates that stochastic processes drive the succession of the rare sub-community. A functional prediction showed that abundant bacteria contribute primary functions to the pollutant-degrading consortia, such as amino acid metabolism, cellular responses to stress, and hydrocarbon degradation. Meanwhile, rare bacteria contribute a substantial fraction of auxiliary functions, such as carbohydrate-active enzymes, fermentation, and homoacetogenesis, which indicates their roles as a source of functional diversity. Our study suggests that the temporal succession of microbes in polluted microcosms is mainly associated with abundant bacteria rather than the high proportion of rare taxa. The major forces (i.e., stochastic or deterministic processes) driving microbial succession could be dependent on the low- or high-abundance community members in temporal microcosms with pollutants.

Mesorhizobium alhagi, a legume-symbiont soil bacterium that forms nodules with the desert plant Alhagi sparsifolia, can produce large amounts of exopolysaccharide (EPS) using mannitol as carbon source. However, the role of EPS in M. alhagi CCNWXJ12-2T, an EPS-producing rhizobium with high salt resistance, remains uncharacterized. Here, we studied the role of EPS in M. alhagi CCNWXJ12-2T using EPS-deficient mutants constructed by transposon mutagenesis. The insertion sites of six EPS-deficient mutants were analyzed using single primer PCR, and two putative gene clusters were found to be involved in EPS synthesis. EPS was extracted and quantified, and EPS production in the EPS-deficient mutants was decreased by approximately 25 times compared with the wild-type strain. Phenotypic analysis revealed reduced salt resistance, antioxidant capacity, and cell motility of the mutants compared with the wild-type strain. In conclusion, our results indicate that EPS can influence cellular Na+ content and antioxidant enzyme activity, as well as play an important role in the stress adaption and cell motility of M. alhagi CCNWXJ12-2T.

A lack of knowledge of the microbial responses to environmental change at the species and functional levels hinders our ability to understand the intrinsic mechanisms underlying the maintenance of microbial ecosystems. Here, we present results from temporal microcosms that introduced inorganic and organic contaminants into agro-soils for 90 days, with three common legume plants. Temporal dynamics and assemblage of soil microbial communities and functions in response to contamination under the influence of growth of different plants were explored via sequencing of the 16S rRNA amplicon and by shotgun metagenomics. Soil microbial alpha diversity and structure at the taxonomic and functional levels exhibited resilience patterns. Functional profiles showed greater resilience than did taxonomic ones. Different legume plants imposed stronger selection on taxonomic profiles than on functional ones. Network and random forest analyses revealed that the functional potential of soil microbial communities was fostered by various taxonomic groups. Betaproteobacteria were important predictors of key functional traits such as amino acid metabolism, nucleic acid metabolism, and hydrocarbon degradation. Our study reveals the strong resilience of the soil microbiome to chemical contamination and sensitive responses of taxonomic rather than functional profiles to selection processes induced by different legume plants. This is pivotal to develop approaches and policies for the protection of soil microbial diversity and functions in agro-ecosystems with different response strategies from global environmental drivers, such as soil contamination and plant invasion. IMPORTANCE Exploring the microbial responses to environmental disturbances is a central issue in microbial ecology. Understanding the dynamic responses of soil microbial communities to chemical contamination and the microbe-soil-plant interactions is essential for forecasting the long-term changes in soil ecosystems. Nevertheless, few studies have applied multi-omics approaches to assess the microbial responses to soil contamination and the microbe-soil-plant interactions at the taxonomic and functional levels simultaneously. Our study reveals clear succession and resilience patterns of soil microbial diversity and structure in response to chemical contamination. Different legume plants exerted stronger selection processes on taxonomic than on functional profiles in contaminated soils, which could benefit plant growth and fitness as well as foster the potential abilities of hydrocarbon degradation and metal tolerance. These results provide new insight into the resilience and assemblage of soil microbiome in response to environmental disturbances in agro-ecosystems at the species and functional levels.

Zinc oxide nanoparticles (ZnO NPs) are attracting much interest due to their potential toxicity and ubiquity in consumer products. However, understanding of pristine and transformed ZnO NPs impact on soil microbial communities is still limited. Here, we explored changes in the microbial communities of soils treated with pristine and sulfidized ZnO NPs (s-ZnO NPs), and their corresponding Zn ions (ZnSO4) for 30 and 90 days exposures at 100 and 500 mg Zn kg−1. The similarity in bacterial community responses was observed between ZnO NPs and s-ZnO NPs, and these Zn treatments significantly affected the bacterial communities at 90 days, which exhibited distinct patterns compared to ZnSO4. The single-time tested DTPA and H2O extractable Zn ions could not fully explain the observed ZnO NPs and s-ZnO NPs impact on bacterial communities. The two most dominant phylum Nitrospirae and Actinobacteria, associated with the reduction of NH4+–N and dissolved organic carbon, demonstrated significant changes in soils exposed to ZnO NPs and s-ZnO NPs. This suggests the potential long-term impact of transformed ZnO NPs on soil carbon and nitrogen cycling. For fungal communities, we did not find the distinct response patterns of fungal communities between nanoparticulate and ionic Zn exposures.

Aiming to investigate the diversity and distribution of rhizobia associated with Ammopiptanthus, an endangered evergreen legume widely distributed in deserts, we characterized a total of 219 nodule isolates from nine sampling sites in Northwest China with different soil characteristics based upon restriction fragment length polymorphism (RFLP) analysis of 16S ribosomal RNA (rRNA) and symbiotic genes (nodC and nifH). Ten isolates representing different 16S rRNA-RFLP types were selected for further sequence analyses of 16S rRNA and four housekeeping genes. As results, nine genospecies belonging to the genera Ensifer, Neorhizobium, Agrobacterium, Pararhizobium, and Rhizobium could be defined among the isolates. The nodC and nifH phylogenies of 14 isolates representing different symbiotic-RFLP types revealed five lineages linked to Ensifer fredii, Ensifer meliloti, Rhizobium leguminosarum, Mesorhizobium amorphae, and Rhizobium gallicum, which demonstrated the various origins and lateral transfers of symbiotic genes between different genera and species. The rhizobial diversities of Ammopiptanthus mongolicus varied among regions, and the community compositions of rhizobia associated with A. mongolicus were significantly different in wild and cultured fields. Constrained correspondence analysis showed that the distribution of A. mongolicus rhizobia could be explained by available potassium content and that the assembly of symbiotic types was mainly affected by available phosphorus content and carbon-nitrogen ratio.

Rhizobia and legume plants are famous mutualistic symbiosis partners who provide nitrogen nutrition to the natural environment. Rhizobial type III secretion systems (T3SSs) deliver effectors that manipulate the metabolism of eukaryotic host cells. Mesorhizobium amorphae CCNWGS0123 (GS0123) contains two T3SS gene clusters, T3SS-I and T3SS-II. T3SS-I contains all the basal components for an integrated T3SS, and the expression of T3SS-I genes is up-regulated in the presence of flavonoids. In contrast, T3SS-II lacks the primary extracellular elements of T3SSs, and the expression of T3SS-II genes is down-regulated in the presence of flavonoids. Inoculation tests on Robinia pseudoacacia displayed considerable differences in gene expression patterns and levels among roots inoculated with GS0123 and T3SS-deficient mutant (GS0123ΔrhcN1 (GS0123ΔT1), GS0123ΔrhcN2 (GS0123ΔT2) and GS0123ΔrhcN1ΔrhcN2 (GS0123ΔS)). Compared with the GS0123-inoculated plants, GS0123ΔT1-inoculated roots formed very few infection threads and effective nodules, while GS0123ΔT2-inoculated roots formed a little fewer infection threads and effective nodules with increased numbers of bacteroids enclosed in one symbiosome. Moreover, almost no infection threads or effective nodules were observed in GS0123ΔS-inoculated roots. In addition to evaluations of plant immunity signals, we observed that the coexistence of T3SS-I and T3SS-II promoted infection by suppressing host defense response in the reactive oxygen species defense response pathway. Future studies should focus on identifying rhizobial T3SS effectors and their host target proteins.

A Gram-positive, facultative anaerobic, rod-shaped, and endospore-forming strain, designated 53-2(T) was isolated from the root nodule of Oxytropis ochrocephala Bunge growing on Qilian mountain, China. The strain can grow at pH 7.0-8.0, 10-50 °C and tolerate up to 11% NaCl. Optimal growth occurred at pH 7.2 and 37 °C. The result of BLASTn search based on 16S rRNA gene sequence revealed that strain 53-2(T) , being closest related to Bacillus acidicola 105-2(T) , possessed remote similarity (less than 95.64%) to the species within genus Bacillus. The DNA G + C content was 37.8%. Chemotaxonomic data (major quinone is MK-7; major polar lipids are diphosphatidylglycerol, phosphatidylglycerol, unknown phospholipid, and aminoglycophospholipid; fatty acids are anteiso-C15: 0 , iso-C15:0 and anteiso-C17: 0 ) supported the affiliation of the isolate to the genus Bacillus. On the basis of physiological, phylogenetic, and biochemical properties, strain 53-2(T) represents a novel species within genus Bacillus, for which the name Bacillus radicibacter is proposed. The type strain is 53-2(T) (=DSM27302(T) =ACCC06115(T) =CCNWQLS5(T) ).

Rhizobia typically produce a lower level of ACC deaminase compared with free-living plant growth-promoting bacteria. While the endogenous rhizobial ACC deaminase is important in legume nodulation, it is not sufficient to protect host plants against environmental stresses. The main goal of this study was to assess the effects of a genetically engineered Sinorhizobium meliloti strain overproducing ACC deaminase, and its symbiotic performance in Medicago lupulina under copper stress conditions. The engineered strain was transformed with an exogenous acdS gene by triparental conjugation. A plant growth assay was conducted to assess its plant growth promotion ability under copper stress conditions. The expressions of antioxidant genes in these plants were analyzed using quantitative real-time PCR. Plants nodulated with the engineered strain showed a greater dry weight, a decreased ethylene level in roots, a higher total copper uptake but a lower level of copper translocation to aerial parts, as compared with the plants nodulated with the wild-type strain under copper stress conditions. These results were positively correlated with higher expression of antioxidant genes in the roots of these plants exposed to severe copper stress. The engineered strain could improve plant growth as well as copper tolerance of M. lupulina, and enhance the antioxidant defense system.

To better understand the diversity of metal resistance genetic determinant from microbes that survived at metal tailings in northwest of China, a highly elevated level of heavy metal containing region, genomic analyses was conducted using genome sequence of three native metal-resistant plant growth promoting bacteria (PGPB). It shows that: Mesorhizobium amorphae CCNWGS0123 contains metal transporters from P-type ATPase, CDF (Cation Diffusion Facilitator), HupE/UreJ and CHR (chromate ion transporter) family involved in copper, zinc, nickel as well as chromate resistance and homeostasis. Meanwhile, the putative CopA/CueO system is expected to mediate copper resistance in Sinorhizobium meliloti CCNWSX0020 while ZntA transporter, assisted with putative CzcD, determines zinc tolerance in Agrobacterium tumefaciens CCNWGS0286. The greenhouse experiment provides the consistent evidence of the plant growth promoting effects of these microbes on their hosts by nitrogen fixation and/or indoleacetic acid (IAA) secretion, indicating a potential in-site phytoremediation usage in the mining tailing regions of China.

Although the rhizomicrobiome has been extensively studied, little is known about the interactions between soil properties and the assemblage of plant growth-promoting microbes in the rhizosphere. Herein, we analysed the composition and structure of rhizomicrobiomes associated with soybean and alfalfa plants growing in different soil types using deep Illumina 16S rRNA sequencing. Soil pH, P and K significantly affected the composition of the soybean rhizomicrobiome, whereas soil pH and N had a significant effect on the alfalfa rhizomicrobiome. Plant biomass was influenced by plant species, the composition of the rhizomicrobiome, soil pH, N, P and plant growth stage. The beta diversity of the rhizomicrobiome was the second most influential factor on plant growth (biomass). Rhizomicrobes associated with plant biomass were identified and divided into four groups: (1) positively associated with soybean biomass; (2) negatively associated with soybean biomass; (3) positively associated with alfalfa biomass; and (4) negatively associated with alfalfa biomass. Genera assemblages among the four groups differentially responded to soil properties; Group 1 and Group 2 were significantly correlated with soil pH and P, whereas Group 3 and Group 4 were significantly correlated with soil N, K and C. The influence of soil properties on the relative abundance of plant biomass-associated rhizomicrobes differed between soybean and alfalfa. The results suggest the rhizomicrobiome has a pronounced influence on plant growth, and the rhizomicrobiome assemblage and plant growth-associated microbes are differentially structured by soil properties and leguminous plant species.

Understanding the impacts of the ecological patterns and niche-based selection on microbial community assembly and nitrogen-cycling network is crucial for achieving energy-neutral wastewater treatment. However, little is known about the niche differentiation and microbial nitrogen-cycling traits of keystone taxa in flocs and granules in anammox-based systems. Herein, the aspects of community assemblage patterns, metabolic functions and nitrogen transformation pathways were explored. The findings discovered that the treatment performance and bacterial community assembly were regulated by core taxa and flocs and granules communities harbored core taxa based on their functional traits. Both niche differentiation and environmental filtering have profound influences on functional bacteria. Furthermore, a combined analysis showed that nitrogen removal in flocs and granules was regulated by different nitrogen transformation pathways. These results suggest that core taxa are the key drivers for the microbial nitrogen-cycling network and improve the understanding of cross-feeding and metabolic pathways between anammox and nitrogen-cycling-related microorganisms.

The main aim of this work was to study molecular characterization of a DNA fragment conferring resistance to Cu(II) in Sinorhizobium meliloti CCNWSX0020. The strain CCNWSX0020, resistant to 1.4 mmol l(-1) Cu(II) in tryptone-yeast extract medium was isolated from Medicago lupulina growing in mine tailings of Fengxian County, China. The availability of the complete genome sequence of S. meliloti CCNWSX0020 provides an opportunity for investigating genes that play significant roles in Cu(II) resistance. A copper resistance gene, with a length of 1,445 bp, encoding 481 amino acids, designated omp, was identified by cDNA-amplified fragment length polymorphism from S. meliloti CCNWSX0020. The expression of omp gene strongly increased in the presence of Cu(II). The omp-defective mutants display sensitivities to Cu(II) compared with their wild types. The Cu(II)-sensitive phenotype of the mutant was complemented by a 1.5-kb DNA fragment containing omp gene. BLAST analysis revealed that this gene encoded a hypothetical outer membrane protein with 75 % similarity to outer membrane efflux protein in Rhizobium leguminosarum bv. viciae 3841. These studies suggested that the omp product was involved in the Cu(II) tolerance of S. meliloti CCNWSX0020.

Elucidating the mechanisms underlying microbial succession is a major goal of microbial ecology research. Given the increasing human pressure on the environment and natural resources, responses to the repeated introduction of organic and inorganic pollutants are of particular interest. To investigate the temporal dynamics of microbial communities in response to pollutants, we analyzed the microbial community structure in batch microcosms that were inoculated with soil bacteria following exposure to individual or combined pollutants (phenanthrene, n-octadecane, phenanthrene + n-octadecane, and phenanthrene + n-octadecane + CdCl2). Subculturing was performed at 10-day intervals, followed by high-throughput sequencing of 16S rRNA genes. The dynamics of microbial communities in response to different pollutants alone and in combination displayed similar patterns during enrichment. Specifically, the repression and induction of microbial taxa were dominant, and the fluctuation was not significant. The rate of appearance for new taxa and the temporal turnover within microbial communities were higher than the rates reported in other studies of microbial communities in air, water, and soil samples. In addition, conditionally rare taxa that were specific to the treatments exhibited higher betweenness centrality values in the co-occurrence network, indicating a strong influence on other interactions in the community. These results suggest that the repeated introduction of pollutants could accelerate microbial succession in microcosms, resulting in the rapid re-equilibration of microbial communities. This article is protected by copyright. All rights reserved.

The microbiomes of rhizocompartments (nodule endophytes, root endophytes, rhizosphere and root zone) in soybean and alfalfa were analyzed using high-throughput sequencing to investigate the interactions among legume species, microorganisms and soil types. A clear hierarchical filtration of microbiota by plants was observed in the four rhizocompartments—the nodule endosphere, root endosphere, rhizosphere and root zone—as demonstrated by significant variations in the composition of the microbial community in the different compartments. The rhizosphere and root zone microbial communities were largely influenced by soil type, and the nodule and root endophytes were primarily determined by plant species. Diverse microbes inhabited the root nodule endosphere, and the corresponding dominant symbiotic rhizobia belonged to Ensifer for alfalfa and Ensifer-Bradyrhizobium for soybean. The non-symbiotic nodule endophytes were mainly Proteobacteria, Actinobacteria, Firmicutes and Bacteroidetes. The variation in root microbial communities was also affected by the plant growth stage. In summary, this study demonstrated that the enrichment process of nodule endophytes follows a hierarchical filtration and that the bacterial communities in nodule endophytes vary according to the plant species. This article is protected by copyright. All rights reserved.

Mine drainage leads to serious contamination of soil. To assess the effects of mine drainage on microbial communities in riparian soils, we used an Illumina MiSeq platform to explore the soil microbial composition and diversity along a settling pond used for mine drainage treatment. Non-metric multidimensional scaling analysis showed that the microbial communities differed significantly among the four sampling zones (influent, upstream, downstream and effluent), but not seasonally. Constrained analysis of principal coordinates indicated heavy metals (zinc, lead and copper), total sulphur, pH and available potassium significantly influenced the microbial community compositions. Heavy metals were the key determinants separating the influent zone from the other three zones. Lower diversity indices were observed in the influent zone. However, more potential indicator species, related to sulphur and organic matter metabolism were found there, such as the sulphur-oxidizing genera Acidiferrobacter, Thermithiobacillus, Limnobacter, Thioprofundum and Thiovirga, and the sulphur-reducing genera Desulfotomaculum and Desulfobulbus; the organic matter degrading genera, Porphyrobacter and Paucimonas, were also identified. The results indicated that more microorganisms related to sulphur- and carbon-cycles may exist in soils heavily contaminated by mine drainage.

A total of 159 endophytic bacteria were isolated from surface-sterilized root nodules of wild perennial Glycyrrhiza legumes growing on 40 sites in central and northwestern China. Amplified fragment length polymorphism (AFLP) genomic fingerprinting and sequencing of partial 16S rRNA genes revealed that the collection mainly consisted of Mesorhizobium, Rhizobium, Sinorhizobium, Agrobacterium and Paenibacillus species. Based on symbiotic properties with the legume hosts Glycyrrhiza uralensis and Glycyrrhiza glabra, we divided the nodulating species into true and sporadic symbionts. Five distinct Mesorhizobium groups represented true symbionts of the host plants, the majority of strains inducing N2-fixing nodules. Sporadic symbionts consisted of either species with infrequent occurrence (Rhizobium galegae, Rhizobium leguminosarum) or species with weak (Sinorhizobium meliloti, Rhizobium gallicum) or no N2 fixation ability (Rhizobium giardinii, Rhizobium cellulosilyticum, Phyllobacterium sp.). Multivariate analyses revealed that the host plant species and geographic location explained only a small part (14.4%) of the total variation in bacterial AFLP patterns, with the host plant explaining slightly more (9.9%) than geography (6.9%). However, strains isolated from G. glabra were clearly separated from those from G. uralensis, and strains obtained from central China were well separated from those originating from Xinjiang in the northwest, indicating both host preference and regional endemism.

A total of 115 endophytic bacteria were isolated from root nodules of the wild legume Sphaerophysa salsula grown in two ecological regions of Loess Plateau in China. The genetic diversity and phylogeny of the strains were revealed by restriction fragment length polymorphism and sequencing of 16S rRNA gene and enterobacterial repetitive intergenic consensus-PCR. Their symbiotic capacity was checked by nodulation tests and analysis of nifH gene sequence. This is the first systematic study on endophytic bacteria associated with S. salsula root nodules. Fifty of the strains found were symbiotic bacteria belonging to eight putative species in the genera Mesorhizobium, Rhizobium and Sinorhizobium, harboring similar nifH genes; Mesorhizobium gobiense was the main group and 65 strains were nonsymbiotic bacteria related to 17 species in the genera Paracoccus, Sphingomonas, Inquilinus, Pseudomonas, Serratia, Mycobacterium, Nocardia, Streptomyces, Paenibacillus, Brevibacillus, Staphylococcus, Lysinibacillus and Bacillus, which were universally coexistent with symbiotic bacteria in the nodules. Differing from other similar studies, the present study is the first time that symbiotic and nonsymbiotic bacteria have been simultaneously isolated from the same root nodules, offering the possibility to accurately reveal the correlation between these two kinds of bacteria. These results provide valuable information about the interactions among the symbiotic bacteria, nonsymbiotic bacteria and their habitats.

The aim of this work is to describe the diversity of potentially symbiotic bacteria associated with the invasive introduced legume Robinia pseudoacacia in China. Thirty-three isolates from 33 separate trees and nodules were characterized using restriction length fragment polymorphism and sequencing of 16S rRNA, nodA, nodC and nifH genes. Their 16S rRNA gene patterns and sequences placed them in three clades: 85% of isolates were related to the Mesorhizobium mediterraneum/temperatum group, whereas the remaining were similar either to Mesorhizobium amorphae or to Sinorhizobium meliloti. However, despite their diverse taxonomic positions, the nodA, nodC and nifH genes' phylogenies indicated that these R. pseudoacacia symbionts share similar symbiosis genes, implying gene transfers and a degree of host specificity. Comparison of R. pseudoacacia symbiotic diversity in native and other invaded areas suggests that most Chinese symbionts may not have arrived with the seed but were local bacteria that acquired specific symbiotic genes from native American rhizobia.

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.

Many legumes have been selected as model plants to degrade organic contaminants with their special associated rhizosphere microbes in soil. However, the function of root nodules during microbe-assisted phytoremediation is not clear. A pot study was conducted to examine phenanthrene (PHE) utilizing bacteria associated with root nodules and the effects of cowpea root nodules on phytoremediation in two different types of soils (freshly contaminated soil and aged contaminated soil). Cowpea nodules in freshly-contaminated soil showed less damage in comparison to the aged-contaminated soil, both morphologically and ultra-structurally by scanning electron microscopy. The study of polycyclic aromatic hydrocarbon (PAH) attenuation conducted by high performance liquid chromatography revealed that more PAH was eliminated from liquid culture around nodulated roots than nodule-free roots. PAH sublimation and denaturation gradient gel electrophoresis were applied to analyze the capability and diversity of PAH degrading bacteria from the following four parts of rhizo-microzone: bulk soil, root surface, nodule surface and nodule inside. The results indicated that the surface and inside of cowpea root nodules were colonized with bacterial consortia that utilized PHE. Our results demonstrated that root nodules not only fixed nitrogen, but also enriched PAH-utilizing microorganisms both inside and outside of the nodules. Legume nodules may have biotechnological values for PAH degradation.

A gram-negative bacterium designated BN30(T), which is motile with a polar flagellum, non-endospores forming, oxidase- and catalase-positive, was isolated from soybean root nodule. The organism is facultative anaerobic and surface-wrinkled rod. It can grow at 10-40 °C, pH 6-8 and 6 % (w/v) NaCl. BLASTn search based on 16S rRNA gene sequence revealed that the strain is closely related to Diaphorobacter aerolatus 8604S-37(T), Alicycliphilus denitrificans K601(T), Simplicispira limi EMB325(T), Diaphorobacter nitroreducens NA10B(T) and Diaphorobacter oryzae RF3(T), which all belonged to the family Comamonadaceae in class Betaproteobacteria. Phylogenetic analysis showed that the strain formed a firm clade with three Diaphorobacter species, being closest to Diaphorobacter aerolatus 8604S-37(T) with similarity of 98.64 %. DNA-DNA relatedness values between strain BN30(T) and five reference strains ranged from 11.5 to 35.9 %. All the results of phylogenetic analysis, chemotaxonomical data (predominant fatty acids are C16:0, sum feature 3, sum feature 8 and C17:0 cyclo; major polar lipids are diphosphatidylglycerol, phosphatidylethanolamine and phosphatidylglycerol; major quinone is Q-8; G+C of total DNA is 65.2 %), physiological and phenotypic results supported that BN30(T) represented a novel species within the genus Diaphorobacter. The name Diaphorobacter ruginosibacter was proposed, and the type strain is BN30(T) (=ACCC06116(T) = DSM 27467(T)).

Biodegradation of polycyclic aromatic hydrocarbons (PAHs) is normally limited by their low solubility and poor bioavailability. Prior research suggests that biosurfactants are synthesized as intermediates during the production of mucilage at the root tip. To date the effects of mucilage on PAH degradation and microbial community response have not been directly examined. To address this question, our research compared 3 cowpea breeding lines (Vigna unguiculata) that differed in mucilage production for their effects on phenanthrene (PHE) degradation in soil. The High Performance Liquid Chromatography results indicated that the highest PHE degradation rate was achieved in soils planted with mucilage producing cowpea line C1, inoculated with Bradyrhizobium, leading to 91.6% PHE disappearance in 5 weeks. In root printing tests, strings treated with mucilage and bacteria produced larger clearing zones than those produced on mucilage treated strings with no bacteria or bacteria inoculated strings. Experiments with 14C-PHE and purified mucilage in soil slurry confirmed that the root mucilage significantly enhanced PHE mineralization (82.7%), which is 12% more than the control treatment without mucilage. The profiles of the PHE degraders generated by Denaturing gradient gel electrophoresis suggested that cowpea C1, producing a high amount of root mucilage, selectively enriched the PHE degrading bacteria population in rhizosphere. These findings indicate that root mucilage may play a significant role in enhancing PHE degradation and suggests that differences in mucilage production may be an important criterion for selection of the best plant species for use in phytoremediation of PAH contaminated soils.

Legumes are important for nitrogen cycling in the environment and agriculture due to the ability of nitrogen fixation by rhizobia. In this review, we introduce an important and potential role of legume-rhizobia symbiosis in aiding phytoremediation of some metal contaminated soils as various legumes have been found to be the dominant plant species in metal contaminated areas. Resistant rhizobia used for phytoremediation could act on metals directly by chelation, precipitation, transformation, biosorption and accumulation. Moreover, the plant growth promoting (PGP) traits of rhizobia including nitrogen fixation, phosphorus solubilization, phytohormone synthesis, siderophore release, and production of ACC deaminase and the volatile compounds of acetoin and 2, 3-butanediol may facilitate legume growth while lessening metal toxicity. The benefits of using legumes inoculated with naturally resistant rhizobia or recombinant rhizobia with enhanced resistance, as well as co-inoculation with other plant growth promoting bacteria (PGPB) are discussed. However, the legume-rhizobia symbiosis appears to be sensitive to metals, and the effect of metal toxicity on the interaction between legumes and rhizobia is not clear. Therefore, to obtain the maximum benefits from legumes assisted by rhizobia for phytoremediation of metals, it is critical to have a good understanding of interactions between PGP traits, the symbiotic plant-rhizobia relationship and metals.

The plant-growth-promoting bacterium Agrobacterium tumefaciens CCNWGS0286, isolated from the nodules of Robinia pseudoacacia growing in zinc-lead mine tailings, both displayed high metal resistance and enhanced the growth of Robinia plants in a metal-contaminated environment. Our goal was to determine whether bacterial metal resistance or the capacity to produce phytohormones had a larger impact on the growth of host plants under zinc stress. Eight zinc-sensitive mutants and one zinc-sensitive mutant with reduced indole-3-acetic acid (IAA) production were obtained by transposon mutagenesis. Analysis of the genome sequence and of transcription via reverse transcriptase PCR (RT-PCR) combined with transposon gene disruptions revealed that ZntA-4200 and the transcriptional regulator ZntR1 played important roles in the zinc homeostasis of A. tumefaciens CCNWGS0286. In addition, interruption of a putative oligoketide cyclase/lipid transport protein reduced IAA synthesis and also showed reduced zinc and cadmium resistance but had no influence on copper resistance. In greenhouse studies, R. pseudoacacia inoculated with A. tumefaciens CCNWGS0286 displayed a significant increase in biomass production over that without inoculation, even in a zinc-contaminated environment. Interestingly, the differences in plant biomass improvement among A. tumefaciens CCNWGS0286, A. tumefaciens C58, and zinc-sensitive mutants 12-2 ( zntA ::Tn 5 ) and 15-6 (low IAA production) revealed that phytohormones, rather than genes encoding zinc resistance determinants, were the dominant factor in enhancing plant growth in contaminated soil.