Azotosporobacter soli gen. nov., sp. nov., a novel nitrogen-fixing bacterium isolated from paddy soil.

A nitrogen-fixing strain designated SG130T was isolated from paddy soil in Fujian Province, China. Strain SG130T was Gram-staining-negative, rod-shaped, and strictly anaerobic. Strain SG130T showed the highest 16S rRNA gene sequence similarities with the type strains Dendrosporobacter quercicolus DSM 1736T (91.7%), Anaeroarcus burkinensis DSM 6283T (91.0%) and Anaerospora hongkongensis HKU 15T (90.9%). Furthermore, the phylogenetic and phylogenomic analysis also suggested strain SG130T clustered with members of the family Sporomusaceae and was distinguished from other genera within this family. Growth of strain SG130T was observed at 25-45 °C (optimum 30 °C), pH 6.0-9.5 (optimum 7.0) and 0-1% (w/v) NaCl (optimum 0.1%). The quinones were Q-8 and Q-9. The polar lipids were phosphatidylserine (PS), phosphatidylethanolamine (PE), glycolipid (GL), phospholipid (PL) and an unidentified lipid (UL). The major fatty acids (> 10%) were iso-C13:0 3OH (26.6%), iso-C17:1 (15.6%) and iso-C15:1 F (11.4%). The genomic DNA G + C content was 50.7%. The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between strain SG130T and the most closely related type strain D. quercicolus DSM 1736T (ANI 68.0% and dDDH 20.3%) were both below the cut-off level for species delineation. The average amino acid identity (AAI) between strain SG130T and the most closely related type strain D. quercicolus DSM 1736T was 63.2%, which was below the cut-off value for bacterial genus delineation (65%). Strain SG130T possessed core genes (nifHDK) involved in nitrogen fixation, and nitrogenase activity (106.38 µmol C2H4 g-1 protein h-1) was examined using the acetylene reduction assay. Based on the above results, strain SG130T is confirmed to represent a novel genus of the family Sporomusaceae, for which the name Azotosporobacter soli gen. nov., sp. nov. is proposed. The type strain is SG130T (= GDMCC 1.3312T = JCM 35641T).

Diversity of soil nitrogen-fixing bacteria in the rhizosphere and non-rhizophere soils of Ebinur Lake Wetland.

Ebinur Lake Wetland is an understudied desert wetland ecosystem, particularly regarding nitrogen cycling. This study aimed to ascertain the diversity and richness of nitrogen-fixing bacterial communities in the Ebinur Lake Wetland. The diversity of the nitrogen-fixing bacteria community of nifH genes from the rhizosphere and non-rhizosphere soils of four plants in different seasons were examined using Illumina HiSeq PE250 high-throughput sequencing technology. The correlation between soil environmental factors and diversity and richness of nitrogen-fixing bacteria was studied using the redundancy analysis (RDA). The results showed that the diversity of nitrogen-fixing bacteria in the rhizosphere soil of the constructive plants was higher than that in the non-rhizosphere soil; also, the diversity in July was higher than that in October and April. Geobacter, Pseudomonas and Bradyrhizobium were the dominant common bacteria in different samples of Ebinur Lake Wetland. The RDA showed that the total nitrogen, available potassium and available phosphoruswere significantly correlated with the diversity and richness of nitrogen-fixing bacteria. The diversity and community structure of nitrogen-fixing bacteria in soil samples also changed over time. The community structures of nitrogen-fixing bacteria in the rhizosphere and non-rhizosphere soils of the four plants were not the same during the same period. The correlation between soil environmental factors and the community structure and abundance of nitrogen-fixing bacteria can provide data basis and theoretical support for the degradation and restoration of Ebinur Lake Wetland.

Diversity and abundance of culturable nitrogen-fixing bacteria in the phyllosphere of maize.

AIMS: The present study aimed at gaining an insight into the abundance and genetic diversity of culturable N-fixing epiphyte bacteria on the phyllosphere of maize in arid and semi-arid regions of Iran. METHODS AND RESULTS: Leaf samples of the maize variety, 'single cross 704' (Zea mays L.) were collected from different locations in Iran. The community of culturable N-fixing epiphyte bacteria present was examined by 16S rRNA sequencing, BOXAIR-polymerase chain reaction (PCR) and restricted fragment length polymorphisms analysis of 16S rRNA gene (16S-RFLP). Approximately, 31·82% of the 242 isolates were identified as N-fixers by cultivation of bacteria in Rennie medium and detection of their nifH gene. The N-fixers were affiliated with four bacterial phyla: Firmicutes, Proteobacteria, Actinobacteria and Bacteroidetes. 16S rRNA sequencing detected 16 genera and 24 different species in the identified phyla. The most dominant genus was Bacillus and the species identified were B. pumilus, B. amyloliquefaciens, B. subtilis, B. paralicheniformis, B. licheniformis, B. niabensis and B. megaterium. In total, 22 RFLP groups were present among the isolates originally identified as N-fixing bacteria. BOXAIR-PCR showed that there was a low similarity level among the N-fixing bacteria isolates, and genetic differentiation of individual strains was relatively great. CONCLUSIONS: Our findings suggest that nitrogen-fixing epiphyte bacteria on the phyllosphere of maize may provide significant nitrogen input into arid and semi-arid ecosystem. SIGNIFICANCE AND IMPACT OF THE STUDY: This research implies that phyllosphere epiphyte diazotrophs have much to offer in sustainable agriculture and can be an alternative to chemical N-fertilizers for providing nitrogen to crops arid and semi-arid regions.

Plant-Growth-Promoting Rhizobacteria Emerging as an Effective Bioinoculant to Improve the Growth, Production, and Stress Tolerance of Vegetable Crops.

Vegetable cultivation is a promising economic activity, and vegetable consumption is important for human health due to the high nutritional content of vegetables. Vegetables are rich in vitamins, minerals, dietary fiber, and several phytochemical compounds. However, the production of vegetables is insufficient to meet the demand of the ever-increasing population. Plant-growth-promoting rhizobacteria (PGPR) facilitate the growth and production of vegetable crops by acquiring nutrients, producing phytohormones, and protecting them from various detrimental effects. In this review, we highlight well-developed and cutting-edge findings focusing on the role of a PGPR-based bioinoculant formulation in enhancing vegetable crop production. We also discuss the role of PGPR in promoting vegetable crop growth and resisting the adverse effects arising from various abiotic (drought, salinity, heat, heavy metals) and biotic (fungi, bacteria, nematodes, and insect pests) stresses.

Diversity of nitrogen-fixing rhizobacteria associated with sugarcane: a comprehensive study of plant-microbe interactions for growth enhancement in Saccharum spp.

BACKGROUND: Nitrogen is an essential element for sugarcane growth and development and is generally applied in the form of urea often much more than at recommended rates, causing serious soil degradation, particularly soil acidification, as well as groundwater and air pollution. In spite of the importance of nitrogen for plant growth, fewer reports are available to understand the application and biological role of N2 fixing bacteria to improve N2 nutrition in the sugarcane plant. RESULTS: In this study, a total of 350 different bacterial strains were isolated from rhizospheric soil samples of the sugarcane plants. Out of these, 22 isolates were selected based on plant growth promotion traits, biocontrol, and nitrogenase activity. The presence and activity of the nifH gene and the ability of nitrogen-fixation proved that all 22 selected strains have the ability to fix nitrogen. These strains were used to perform 16S rRNA and rpoB genes for their identification. The resulted amplicons were sequenced and phylogenetic analysis was constructed. Among the screened strains for nitrogen fixation, CY5 (Bacillus megaterium) and CA1 (Bacillus mycoides) were the most prominent. These two strains were examined for functional diversity using Biolog phenotyping, which confirmed the consumption of diverse carbon and nitrogen sources and tolerance to low pH and osmotic stress. The inoculated bacterial strains colonized the sugarcane rhizosphere successfully and were mostly located in root and leaf. The expression of the nifH gene in both sugarcane varieties (GT11 and GXB9) inoculated with CY5 and CA1 was confirmed. The gene expression studies showed enhanced expression of genes of various enzymes such as catalase, phenylalanine-ammonia-lyase, superoxide dismutase, chitinase and glucanase in bacterial-inoculated sugarcane plants. CONCLUSION: The results showed that a substantial number of Bacillus isolates have N-fixation and biocontrol property against two sugarcane pathogens Sporisorium scitamineum and Ceratocystis paradoxa. The increased activity of genes controlling free radical metabolism may at least in part accounts for the increased tolerance to pathogens. Nitrogen-fixation was confirmed in sugarcane inoculated with B. megaterium and B. mycoides strains using N-balance and 15N2 isotope dilution in different plant parts of sugarcane. This is the first report of Bacillus mycoides as a nitrogen-fixing rhizobacterium in sugarcane.

Paraburkholderia atlantica sp. nov. and Paraburkholderia franconis sp. nov., two new nitrogen-fixing nodulating species isolated from Atlantic forest soils in Brazil.

A polyphasic study was conducted with 11 strains trapped by Mimosa pudica and Phaseolus vulgaris grown in soils of the Brazilian Atlantic Forest. In the phylogenetic analysis of the 16S rRNA gene, one clade of strains (Psp1) showed higher similarity with Paraburkholderia piptadeniae STM7183T (99.6%), whereas the second (Psp6) was closely related to Paraburkholderia tuberum STM678T (99%). An MLSA (multilocus sequence analysis) with four (recA, gyrB, trpB and gltB) housekeeping genes placed both Psp1 and Psp6 strains in new clades, and BOX-PCR profiles indicated high intraspecific genetic diversity within each clade. Values of digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) of the whole genome sequences were of 56.9 and 94.4% between the Psp1 strain CNPSo 3157T and P. piptadeniae; and of 49.7% and 92.7% between the Psp6 strain CNPSo 3155T and P. tuberum, below the threshold for species delimitation. In the nodC analysis, Psp1 strains clustered together with P. piptadeniae, while Psp6 did not group with any symbiotic Paraburkholderia. Other phenotypic, genotypic and symbiotic properties were evaluated. The polyphasic analysis supports that the strains represent two novel species, for which the names Paraburkholderia franconis sp. nov. with type strain CNPSo 3157T (= ABIP 241, = LMG 31644) and Paraburkholderia atlantica sp. nov. with type strain CNPSo 3155T (= ABIP 236, = LMG 31643) are proposed.

Mining the roots of various species of the halophyte Suaeda for halotolerant nitrogen-fixing endophytic bacteria with the potential for promoting plant growth.

Saline area may tend to be a productive land; however, many of salt-affected soils have nitrogen limitation and depend on plant-associated diazotrophs as their source of 'new' nitrogen. Herein, a total of 316 salinity tolerant nitrogen-fixing endophytic bacteria were isolated from roots of the halophyte Suaeda sp. sampled from 22 different areas of Iran to prepare the collection of nitrogen-fixing bacterial endophytes and evaluate the plant growth-promoting effect of effective isolates on growth of the halophyte Suaeda maritima. All of the identified nitrogen-fixing endophytes were classified to Proteobacteria, Actinobacteria, Firmicutes, and Bacteroidetes phylum while we did not detect common nitrogen-fixing endophyte of glycophytes like Azospirillum. The genera Pseudomonas and Microbacterium were both encountered in high abundance in all samples, indicating that they might play an advanced role in the micro-ecosystem of the halophyte Suaeda. In addition, the results also showed that not only soil salinity can affect halophyte endophytic composition but also other factors such as geographical location, plant species, and other soil properties may be involved. Interestingly, only Zhihengliuella halotolerans and Brachybacterium sp. belonging to Actinobacteria could grow in semi-solid N-free (NFb) medium supplemented with 6% NaCl and highly enhanced growth of S. maritima in vitro. Overall, this study offers useful new resources for nitrogen-fixing endophytic bacteria which may be utilized to improve approaches for providing bio-fertilizer useful in saline-based agriculture.

Efficient Nitrogen-Fixing Bacteria Isolated from Soybean Nodules in the Semi-arid Region of Northeast Brazil are Classified as Bradyrhizobium brasilense (Symbiovar Sojae).

Soybean (Glycine max L.) is an important legume that greatly benefits from inoculation with nitrogen-fixing bacteria. In a previous study, five efficient nitrogen-fixing bacterial strains, isolated from nodules of soybean inoculated with soil from semi-arid region, Northeast Brazil, were identified as a new group within the genus Bradyrhizobium. The taxonomic status of these strains was evaluated in this study. The phylogenetic analysis of the 16S rRNA gene showed the high similarity of the five strains to Bradyrhizobium brasilense UFLA03-321T (100%), B. pachyrhizi PAC48T (100%), B. ripae WR4T (100%), B. elkanii USDA 76T (99.91%), and B. macuxiense BR 10303T (99.91%). However, multilocus sequence analysis of the housekeeping genes atpD, dnaK, gyrB, recA, and rpoB, average nucleotide identity, and digital DNA-DNA hybridization analyses supported the classification of the group as B. brasilense. Some phenotypic characteristics allowed differentiating the five strains and the type strain of B. brasilense from the two neighboring species (B. pachyrhizi PAC48T and B. elkanii USDA 76T). The nodC and nifH genes' analyses showed that these strains belong to symbiovar sojae, together with B. elkanii (USDA 76T) and B. ferriligni (CCBAU 51502T). The present results support the classification of these five strains as Bradyrhizobium brasilense (symbiovar sojae).

Signalling in actinorhizal root nodule symbioses.

Plants able to establish a nitrogen-fixing root nodule symbiosis with the actinobacterium Frankia are called actinorhizal. These interactions lead to the formation of new root organs, called actinorhizal nodules, where the bacteria are hosted intracellularly and fix atmospheric nitrogen thus providing the plant with an almost unlimited source of nitrogen for its nutrition. Like other symbiotic interactions, actinorhizal nodulation involves elaborate signalling between both partners of the symbiosis, leading to specific recognition between the plant and its compatible microbial partner, its accommodation inside plant cells and the development of functional root nodules. Actinorhizal nodulation shares many features with rhizobial nodulation but our knowledge on the molecular mechanisms involved in actinorhizal nodulation remains very scarce. However recent technical achievements for several actinorhizal species are allowing major discoveries in this field. In this review, we provide an outline on signalling molecules involved at different stages of actinorhizal nodule formation and the corresponding signalling pathways and gene networks.

Elevated level of arsenic negatively influences nifH gene expression of isolated soil bacteria in culture condition as well as soil system.

Comprehensive studies on the effect of arsenic (As) on free-living diazotrophs that play a crucial role in soil fertility by nitrogen fixation are still scanty. Here, we isolated three free-living bacteria from rice field with potential nitrogen-fixing ability and investigated the impact of As on their nifH gene expression and extracellular polysaccharide (EPS) production in culture condition and soil system. 16S rRNA sequence analysis showed that the isolated bacteria were affiliated to ß-Proteobacteria, γ-Proteobacteria and Firmicutes. As(III) exposure to bacterial isolates followed by RT-qPCR analysis revealed that elevated levels of As reduced the expression of nifH gene in selective bacteria, both in culture medium and soil condition. We also noticed reduced production of EPS under higher concentration of As. All the three bacteria showed high tolerance to As(III), able to oxidize As and exhibited significant plant growth-promoting traits. This investigation indicated that an environment exposed with higher concentration of As might perturbed the activity of free-living diazotrophs in agricultural soil system.

Metagenomic Analysis of Some Potential Nitrogen-Fixing Bacteria in Arable Soils at Different Formation Processes.

The main goal of the study was to determine the diversity of the potential nitrogen-fixing (PNF) bacteria inhabiting agricultural (A) soils versus wastelands serving as controls (C). The soils were classified into three groups based on the formation process: autogenic soils (Albic Luvisols, Brunic Arenosols, Haplic Phaeozem) formed on loess material, hydrogenic soils (Mollic Gleysols, Eutric Fluvisol, Eutric Histosol) formed under the effect of stagnant water and lithogenic soils (Rendzina Leptosols) formed on limestone. In order to determine the preferable conditions for PNF bacteria, the relationships between the soil chemical features and bacterial operational taxonomic units (OTUs) were tested. Additionally, the nitrogen content and fertilisation requirement of the lithogenic (LG), autogenic (AG) and hydrogenic (HG) soils were discussed. The composition of the bacterial communities was analysed with the next-generation sequencing (NGS) by the Ion Torrent™ technology. The sequences were clustered into OTU based on a 99 % similarity threshold. The arable soils tested were distinctly dominated by ß-Proteobacteria representatives of PNF bacteria belonging to the genus Burkholderia. Bacteria from the α-Proteobacteria class and Devosia genus were subdominants. A free-living Cyanobacteria population dominated in A rather than in C soils. We have found that both soil agricultural management and soil formation processes are the most conducive factors for PNF bacteria, as a majority of these microorganisms inhabit the AG group of soils, whilst the LG soils with the lowest abundance of PNF bacteria revealed the need for additional mineral fertilisation. Our studies have also indicated that there are close relationships between soil classification with respect to soil formation processes and PNF bacteria preference for occupation of soil niches.

Native rhizobia from Zn mining soil promote the growth of Leucaena leucocephala on contaminated soil.

Plants on contaminated mining soils often show a reduced growth due to nutrient depletion as well as trace elements (TEs) toxicity. Since those conditions threat plant's survival, plant growth-promoting rhizobacteria (PGPRs), such as rhizobia, might be of crucial importance for plant colonization on TE-contaminated soils. Native rhizobia from mining soils are promising candidates for bioaugmented phytoremediation of those soils as they are adapted to the specific conditions. In this work, rhizobia from Zn- and Cd-contaminated mining soils were in vitro screened for their PGP features [organic acids, indole-3-acetic acid (IAA), and siderophore (SID) production; 1-aminocyclopropane-1-carboxylate (ACC) deaminase activity; and Ca3(PO4)2 solubilization] and Zn and Cd tolerance. In addition, some type and reference rhizobia strains were included in the study as well. The in vitro screening indicated that rhizobia and other native genera have great potential for phytoremediation purposes, by exerting, besides biological N2 fixation, other plant growth-promoting traits. Leucaena leucocephala-Mesorhizobium sp. (UFLA 01-765) showed multielement tolerance and an efficient symbiosis on contaminated soil, decreasing the activities of antioxidative enzymes in shoots. This symbiosis is a promising combination for phytostabilization.

Nitrogen fixing bacterial diversity in a tropical estuarine sediments.

Microorganisms play a significant role in biogeochemical cycles, especially in the benthic and pelagic ecosystems. Role of environmental parameters in regulating the diversity, distribution and physiology of these microorganisms in tropical marine environment is not well understood. In this study, we have identified dinitrogen (N2) fixing bacterial communities in the sediments by constructing clone libraries of nitrogenase (nifH) gene from four different stations in the Cochin estuary, along the southeastern Arabian Sea. N2 fixing bacterial clones revealed that over 20 putative diazotrophs belong to alpha-, beta-, gamma-, delta- and epsilon- proteobacteria and firmicutes. Predominant genera among these were Bradyrhizobium sp. (α-proteobacteria), Dechloromonas sp. (ß-proteobacteria); Azotobactor sp., Teredinibacter sp., Methylobacter sp., Rheinheimera sp. and Marinobacterium sp. (γ-proteobacteria); Desulfobacter sp., Desulfobulbus sp. and Desulfovibrio sp. (δ -proteobacteria); Arcobacter sp. and Sulfurospirillum sp. (ε-proteobacteria). Nostoc sp. was solely identified among the cyanobacterial phylotype. Nitrogen fixing Sulfate reducing bacteria (SRBs) such as Desulfobulbus sp., Desulfovibrio sp., Desulfuromonas sp., Desulfosporosinus sp., Desulfobacter sp., were also observed in the study. Most of the bacterial nifH sequences revealed that the identities of N2 fixing bacteria were less than 95% similar to that available in the GenBank database, which suggested that the sequences were of novel N2 fixing microorganisms. Shannon-Weiner diversity index of nifH gene ranged from 2.95 to 3.61, indicating an inflated diversity of N2 fixing bacteria. Canonical correspondence analysis (CCA) implied positive correlation among nifH diversity, N2 fixation rate and other environmental variables.

Bacterial Diversity and Community Structure in Two Bornean Nepenthes Species with Differences in Nitrogen Acquisition Strategies.

Carnivorous plants of the genus Nepenthes have been studied for over a century, but surprisingly little is known about associations with microorganisms. The two species Nepenthes rafflesiana and Nepenthes hemsleyana differ in their pitcher-mediated nutrient sources, sequestering nitrogen from arthropod prey and arthropods as well as bat faeces, respectively. We expected bacterial communities living in the pitchers to resemble this diet difference. Samples were taken from different parts of the pitchers (leaf, peristome, inside, outside, digestive fluid) of both species. Bacterial communities were determined using culture-independent high-throughput amplicon sequencing. Bacterial richness and community structure were similar in leaves, peristomes, inside and outside walls of both plant species. Regarding digestive fluids, bacterial richness was higher in N. hemsleyana than in N. rafflesiana. Additionally, digestive fluid communities were highly variable in structure, with strain-specific differences in community composition between replicates. Acidophilic taxa were mostly of low abundance, except the genus Acidocella, which strikingly reached extremely high levels in two N. rafflesiana fluids. In N. hemsleyana fluid, some taxa classified as vertebrate gut symbionts as well as saprophytes were enriched compared to N. rafflesiana, with saprophytes constituting potential competitors for nutrients. The high variation in community structure might be caused by a number of biotic and abiotic factors. Nitrogen-fixing bacteria were present in both study species, which might provide essential nutrients to the plant at times of low prey capture and/or rare encounters with bats.

The bacterial communities of Alaskan mosses and their contributions to N2-fixation.

BACKGROUND: Mosses in high-latitude ecosystems harbor diverse bacterial taxa, including N2-fixers which are key contributors to nitrogen dynamics in these systems. Yet the relative importance of moss host species, and environmental factors, in structuring these microbial communities and their N2-fixing potential remains unclear. We studied 26 boreal and tundra moss species across 24 sites in Alaska, USA, from 61 to 69° N. We used cultivation-independent approaches to characterize the variation in moss-associated bacterial communities as a function of host species identity and site characteristics. We also measured N2-fixation rates via 15N2 isotopic enrichment and identified potential N2-fixing bacteria using available literature and genomic information. RESULTS: Host species identity and host evolutionary history were both highly predictive of moss microbiome composition, highlighting strong phylogenetic coherence in these microbial communities. Although less important, light availability and temperature also influenced composition of the moss microbiome. Finally, we identified putative N2-fixing bacteria specific to some moss hosts, including potential N2-fixing bacteria outside well-studied cyanobacterial clades. CONCLUSIONS: The strong effect of host identity on moss-associated bacterial communities demonstrates mosses' utility for understanding plant-microbe interactions in non-leguminous systems. Our work also highlights the likely importance of novel bacterial taxa to N2-fixation in high-latitude ecosystems. Video Abstract.

The Flora Compositions of Nitrogen-Fixing Bacteria and the Differential Expression of nifH Gene in Pennisetum giganteum z.x.lin Roots.

The flora compositions of nitrogen-fixing bacteria in roots of Pennisetum giganteum z.x.lin at different growth stages and the expression and copy number of nitrogen-fixing gene nifH were studied by Illumina Miseq second-generation sequencing technology and qRT-PCR. The results showed that there were more than 40,000~50,000 effective sequences in 5 samples from the roots of P. giganteum, with Proteobacteria and Cyanobacteria as the dominant nitrogen-fixing bacteria based on the OTU species annotations for each sample and Bradyrhizobium as the core bacterial genera. The relative expression and quantitative change of nifH gene in roots of P. giganteum at different growth stages were consistent with the changes in the flora compositions of nitrogen-fixing microbia. Both revealed a changing trend with an initial increase and a sequential decrease, as well as changing order as jointing stage>maturation stage>tillering stage>seedling stage>dying stage. The relative expression and copy number of nifH gene were different in different growth stages, and the difference among groups basically reached a significant level (p < 0.05). The relative expression and copy number of nifH gene at the jointing stage were the highest, and the 2-△△CT value was 4.43 folds higher than that at the seedling stage, with a copy number of 1.32 × 107/g. While at the dying stage, it was the lowest, and the 2-△△CT value was 0.67 folds, with a copy number of 0.31 × 107/g.

Denitrifiers, nitrogen-fixing bacteria and N2O soil gas flux in high Arctic ice-wedge polygon cryosols.

Climate warming and subsequent permafrost thaw may result in organic carbon and nutrient stores being metabolized by microbial communities, resulting in a positive feedback loop of greenhouse gas (GHG) soil emissions. As the third most important GHG, understanding nitrous oxide (N2O) flux in Arctic mineral ice-wedge polygon cryosols and its relationship to the active microbial community is potentially a key parameter for understanding future GHG emissions and climatic warming potential. In the present study, metatranscriptomic analyses of active layer Arctic cryosols, at a representative ice-wedge polygon site, identified active nitrogen-fixing and denitrifying bacteria that included members of Rhizobiaceae, Nostocaceae, Cyanothecaceae, Rhodobacteraceae, Burkholderiaceae, Chloroflexaceae, Azotobacteraceae and Ectothiorhodospiraceae. Unique microbial assemblages with higher proportion of Rhodobacteriales and Rhocyclales were identified by targeted functional gene sequencing at locations with higher (P = 0.053) N2O emissions in the wetter trough soils compared with the dryer polygon interior soils. This coincided with a higher relative abundance of the denitrification nirS gene and higher nitrate/nitrite concentrations in trough soils. The elevated N2O flux observed from wetter trough soils compared with drier polygon interior soils is concerning from a climate warming perspective, since the Arctic is predicted to become warmer and wetter.

Unlocking a high bacterial diversity in the coralloid root microbiome from the cycad genus Dioon.

Cycads are among the few plants that have developed specialized roots to host nitrogen-fixing bacteria. We describe the bacterial diversity of the coralloid roots from seven Dioon species and their surrounding rhizosphere and soil. Using 16S rRNA gene amplicon sequencing, we found that all coralloid roots are inhabited by a broad diversity of bacterial groups, including cyanobacteria and Rhizobiales among the most abundant groups. The diversity and composition of the endophytes are similar in the six Mexican species of Dioon that we evaluated, suggesting a recent divergence of Dioon populations and/or similar plant-driven restrictions in maintaining the coralloid root microbiome. Botanical garden samples and natural populations have a similar taxonomic composition, although the beta diversity differed between these populations. The rhizosphere surrounding the coralloid root serves as a reservoir and source of mostly diazotroph and plant growth-promoting groups that colonize the coralloid endosphere. In the case of cyanobacteria, the endosphere is enriched with Nostoc spp and Calothrix spp that are closely related to previously reported symbiont genera in cycads and other early divergent plants. The data reported here provide an in-depth taxonomic characterization of the bacterial community associated with coralloid root microbiome. The functional aspects of the endophytes, their biological interactions, and their evolutionary history are the next research step in this recently discovered diversity within the cycad coralloid root microbiome.

[Effects of Transgenic Maize with cry1Ab and Epsps Genes C0030.3.5 on the Abundance and Community Structure of Soil Nitrogen-fixing Bacteria].

In order to evaluate the potential risk of planting transgenic corn on soil nitrogen-fixing microorganisms, in 2015, rhizosphere and non-rhizosphere soil samples were collected at the jointing stage, tassel stage, milky stage, and ripening stage, and the effects of transgenic maize with the cry1Ab and epsps genes on the abundance and diversity of soil nitrogen-fixing bacteria were studied by real-time quantitative PCR and T-RFLP. The results showed that the copy number of the diazotrophic nifH gene in the rhizosphere and non-rhizosphere soil of transgenic maize with the cry1Ab and epsps genes (C0030.3.5) and its parental maize (DBN318) showed a trend where it first increased and then decrease with the growth stages, ranging between 2.99×107 and 7.02×107 copies·g-1. The abundance of the diazotrophic nifH gene in the rhizosphere soil and non-rhizosphere soil gene showed no significant difference between TM and PM in the same growth stage (P>0.05). The correlation analysis showed that the abundance of the diazotrophic nifH gene was positively correlated with the content of organic matter, but negatively correlated with water content. T-RFLP analysis yielded 14 T-RFs of different lengths, and 43-bp and 155-bp fragments were the dominant population. The community composition of nitrogen-fixing bacteria was the same as that of TM and PM in the rhizosphere soil and non-rhizosphere soil, and there was no significant difference between the TM and PM populations in the same growth period (P>0.05). The Shannon index and Evenness index of the diazotrophic nifH gene showed a trend where they first increased and then decreased with the growth period, and there was no significant difference in the Shannon index and Evenness index in the same growth stage between the rhizosphere and non-rhizosphere soil samples. Principal component analysis(PCA) indicated that the composition of nitrogen-fixing bacteria was not different between TM and PM. Redundancy analysis (RDA) showed that soil ammonium, nitrogen, and pH were significantly correlated with composition of nitrogen-fixing bacteria.

Phylogenetic Diversity of Nitrogenase Reductase Genes and Possible Nitrogen-Fixing Bacteria in Thermophilic Chemosynthetic Microbial Communities in Nakabusa Hot Springs.

Chemosynthetic microbial communities develop and form dense cell aggregates in slightly alkaline sulfidic hot springs in the temperature range of 70-86°C at Nakabusa, Japan. Nitrogenase activity has recently been detected in the microbial communities collected. To identify possible members capable of nitrogen fixation, we examined the diversities of 16S rRNA and nitrogenase reductase (NifH) gene sequences in four types of chemosynthetic communities with visually different colors and thicknesses. The results of a 16S rRNA gene analysis indicated that all four microbial communities had similar bacterial constituents; the phylum Aquificae was the dominant member, followed in abundance by Thermodesulfobacteria, Firmicutes, and Thermotogae. Most of the NifH sequences were related to sequences reported in hydrothermal vents and terrestrial hot springs. The results of a phylogenetic analysis of NifH sequences revealed diversity in this gene among the communities collected, distributed within 7 phylogenetic groups. NifH sequences affiliated with Aquificae (Hydrogenobacter/Thermocrinis) and Firmicutes (Caldicellulosiruptor) were abundant. At least two different energy metabolic pathways appeared to be related to nitrogen fixation in the communities analyzed; aerobic sulfur/hydrogen-oxidizing bacteria in Aquificae and fermentative bacteria in Firmicutes. The metabolic characteristics of these two dominant phyla differed from those previously inferred from nitrogenase activity assays on chemosynthetic communities, which were associated with hydrogen-dependent autotrophic sulfate reduction. These assays may correspond to the observed NifH sequences that are distantly related to the known species of Thermodesulfovibrio sp. (Nitrospirae) detected in the present study. The activities of nitrogen-fixing organisms in communities may depend on redox states as well as the availability of electron donors, acceptors, and carbon sources.