Nitrogen Fixation Genes Research Articles

Role of two RpoN in Bradyrhizobium sp. strain DOA9 in symbiosis and free-living growth.

RpoN is an alternative sigma factor (sigma 54) that recruits the core RNA polymerase to promoters of genes. In bacteria, RpoN has diverse physiological functions. In rhizobia, RpoN plays a key role in the transcription of nitrogen fixation (nif) genes. The Bradyrhizobium sp. DOA9 strain contains a chromosomal (c) and plasmid (p) encoded RpoN protein. We used single and double rpoN mutants and reporter strains to investigate the role of the two RpoN proteins under free-living and symbiotic conditions. We observed that the inactivation of rpoNc or rpoNp severely impacts the physiology of the bacteria under free-living conditions, such as the bacterial motility, carbon and nitrogen utilization profiles, exopolysaccharide (EPS) production, and biofilm formation. However, free-living nitrogen fixation appears to be under the primary control of RpoNc. Interestingly, drastic effects of rpoNc and rpoNp mutations were also observed during symbiosis with Aeschynomene americana. Indeed, inoculation with rpoNp, rpoNc, and double rpoN mutant strains resulted in decreases of 39, 64, and 82% in the number of nodules, respectively, as well as a reduction in nitrogen fixation efficiency and a loss of the bacterium's ability to survive intracellularly. Taken together, the results show that the chromosomal and plasmid encoded RpoN proteins in the DOA9 strain both play a pleiotropic role during free-living and symbiotic states.

Investigation at the molecular level for a nifgenes to bacteria klebsiella pneumoniea the use of polymerase chain reaction (PCR)

This research was aimed at detection on three nitrogen fixation genes nifK , nifD , nifH in 21 K. pneumonia pathogenic isolates which obtained from Prof. Jean-Marc in Leicester British university by using PCR technique and four specific primers P1, P2 , P5 , P6 and compare it with K. pneumoniae 342 standard strain. The results showed presence of two bands for each isolate with M.W 0.5 Kbp comparing with 1 Kbp of the marker.

Fundidesulfovibrio agrisoli sp. nov., A Nitrogen-Fixing Bacterium Isolated from Rice Field

A strictly anaerobic nitrogen-fixing strain, designated SG106T, was isolated from rice field. The 16S rRNA gene sequence analysis showed that strain SG106T was closely related to the type strain of Fundidesulfovibrio magnetotacticus (97.3%). In phylogenetic (based on 16S rRNA gene sequences) and phylogenomic (constructed using a concatenated alignment of 117 conserved bacterial single-copy genes with GTDB-Tk) trees, strain SG106T clustered with members of the genus Fundidesulfovibrio. Strain SG106T grew at 20-40°C and 0-0.4% (w/v) NaCl. Desulfoviridin was found in the strain SG106T. The genomic DNA G + C content of strain SG106T was 66.0%. The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between strain SG106T and the closely related F. magnetotacticus were 78.4% and 21.7%, respectively. Genome analysis showed that strain SG106T encodes genes for nitrogen fixation (nifHDK). Acetylene reduction experiments showed that the nitrogenase activity of strain SG106T could reach 224.7μmol C2H4 g-1 protein h-1. Based on the above results, strain SG106T represents a novel species of the genus Fundidesulfovibrio, for which the name Fundidesulfovibrio agrisoli sp. nov. is proposed. The type strain is SG106T (= GDMCC 1.3136T = JCM 35588T).

Diversity of common bean rhizobia in blackland of northeastern China and their symbiotic compatibility with two host varieties.

The common bean (Phaseolus vulgaris L.) is an important crop in the world that forms root nodules with diverse rhizobia. Aiming to learn the rhizobial communities associated with the common bean in the black soil of Northeast China, 79 rhizobia were isolated from root nodules of two host varieties (Cuican and Jiadouwang) grown in two sites of blackland and were characterized by comparative sequence analyses of 16S rRNA, recA, atpD, nodC, and nifH genes, and whole genome. As a result, Rhizobium indigoferae, R. anhuiense, and R. croatiense as minor groups and three dominant novel Rhizobium species were identified based on their average nucleotide identity and DNA-DNA hybridization values to the type strains of relative species. This community composition of rhizobia associated with the common bean in the tested black soils was unique. Despite their different species affiliations, all of them were identified into the symbiovar phaseoli according to the phylogenies of symbiotic genes, nodC and nifH. While the phylogenetic discrepancies found in nodC, nifH evidenced that the evolutions of nodulation (nod) and nitrogen fixation (nif ) genes were partially independent. In addition, only one dominant rhizobial species was shared by the two common bean varieties grown in the two soil samples, implying that both the plant variety and the soil characteristics affected the compatibility between rhizobia and their hosts. These findings further enlarged the spectrum of common bean-nodulating rhizobia and added more information about the interactions among the soil factors, rhizobial species, and host plants in the symbiosis.

Gut microbiota contributes to lignocellulose deconstruction and nitrogen fixation of the larva of Apriona swainsoni.

Apriona swainsoni is a vital forest pest prevalent in China. The larvae of A. swainsoni live solely in the branches of trees and rely entirely on the xylem for nutrition. However, there is still a lack of in-depth research on the gut microbiota's use of almost nitrogen-free wood components to provide bio-organic macromolecular components needed for their growth. Thus, in this study, the metagenome, metaproteome, and metabolome of the A. swainsoni larvae in four gut segments (foregut; midgut; anterior hindgut; posterior hindgut) were analyzed by the multi-omics combined technology, to explore the metabolic utilization mechanism of the corresponding gut microbiota of A. swainsoni. Firstly, we found that the metagenome of different gut segments was not significantly different in general, but there were different combinations of dominant bacteria and genes in different gut segments, and the metaproteome and metabolome of four gut segments were significantly different in general. Secondly, the multi-omics results showed that there were significant gradient differences in the contents of cellulose and hemicellulose in different segments of A. swainsoni, and the expression of corresponding metabolic proteins was the highest in the midgut, suggesting the metabolic characteristics of these lignocellulose components in A. swainsoni gut segments. Finally, we found that the C/N ratio of woody food was significantly lower than that of frass, and metagenomic results showed that nitrogen fixation genes mainly existed in the foregut and two hindgut segments. The expression of the key nitrogen fixing gene nifH occurred in two hindgut parts, indicating the feature of nitrogen fixation of A. swainsoni. In conclusion, our results provide direct evidence that the larvae of A. swainsoni can adapt to the relatively harsh niche conditions through the highly organized gut microbiome in four gut segments, and may play a major role in their growth.

How to survive in the world's third poplar: Insights from the genome of the highest altitude woody plant, Hippophae tibetana (Elaeagnaceae).

Hippophae tibetana (Tibetan sea-buckthorn) is one of the highest distributed woody plants in the world (3,000-5,200 meters a.s.l.). It is characterized by adaptation to extreme environment and important economic values. Here, we combined PacBio Hifi platform and Hi-C technology to assemble a 1,452.75 Mb genome encoding 33,367 genes with a Contig N50 of 74.31 Mb, and inferred its sexual chromosome. Two Hippophae-specific whole-genome duplication events (18.7-21.2 million years ago, Ma; 28.6-32.4 Ma) and long terminal repeats retroelements (LTR-RTs) amplifications were detected. Comparing with related species at lower altitude, Ziziphus jujuba (<1, 700 meters a.s.l.), H. tibetana had some significantly rapid evolving genes involved in adaptation to high altitude habitats. However, comparing with Hippophae rhamnoides (<3, 700 meters a.s.l.), no rapid evolving genes were found except microtubule and microtubule-based process genes, H. tibetana has a larger genome, with extra 2, 503 genes (7.5%) and extra 680.46 Mb transposable elements (TEs) (46.84%). These results suggest that the changes in the copy number and regulatory pattern of genes play a more important role for H. tibetana adapting to more extreme and variable environments at higher altitude by more TEs and more genes increasing genome variability and expression plasticity. This suggestion was supported by two findings: nitrogen-fixing genes of H. tibetana having more copies, and intact TEs being significantly closer genes than fragmentary TEs. This study provided new insights into the evolution of alpine plants.

Effects of microplastics on nitrogen and phosphorus cycles and microbial communities in sediments

Sediments are the long-term sinks of microplastics (MPs) and nutrients in freshwater ecosystems. Therefore, understanding the effect of MPs on sediment nutrients is crucial. However, few studies have discussed the effects of MPs on nitrogen and phosphorus cycles in freshwater sediments. Herein, 0.5% (w/w) polyvinyl chloride (PVC), polylactic acid (PLA), and polypropylene (PP) MPs were added to freshwater sediments to evaluate their effects on microbial communities and nitrogen and phosphorus release. The potential biochemical functions of the bacterial communities in the sediments were predicted and assessed via 16S rRNA gene sequencing. The results showed that MPs significantly affected the microbial community composition and nutrient cycling in the sediments. PVC and PP MPs can promote microbial nitrification and nitrite oxidation, while PP can significantly promote alkaline phosphatase (ALP) activity and the abundance of the phosphorus-regulation (phoR) gene. PLA MPs had the potential to promote the abundance of microbial phosphorus transporter (ugpB), nitrogen fixation (nifD, nifH, and nifX), and denitrification (nirS, napA, and norB) genes and inhibit nitrification, resulting in massive accumulation and release of ammonia nitrogen. Although PLA MPs inhibited the activity of ALP and the abundance of the organophosphorus mineralization (phoD) gene, it could enhance dissimilatory iron and sulfite reduction, which may promote the release of sedimentary phosphorus. Our findings may help understand the mechanisms of nitrogen and phosphorus cycles and microbial communities driven by MPs in sediments and provide a basis for future assessments of the environmental behavior of MPs in freshwater ecosystems.

Whole-genome analysis revealed the growth-promoting mechanism of endophytic bacterial strain Q2H1 in potato plants.

Endophytes are non-pathogenic inhabitants of healthy plant tissues and have been found to promote plant growth and health. The endophytic bacterial strain Q2H1 was isolated from the roots of the potato and was identified to exhibit growth-promoting effects in potato plants. Whole-genome sequencing was performed to reveal the mechanism underlying its growth-promoting effect. The obtained sequencing data of approximately 5.65 MB encompassed 5,533 coding sequences. Of note, nine secondary metabolite gene clusters, including siderophore gene clusters, closely associated with plant growth promotion (PGP) were predicted by antiSMASH software. Comparative genomic analysis revealed that Q2H1 belongs to the genus Peribacillus. By gene function annotation, those genes related to plant growth-promoting activities, including indole-3-acetic acid (IAA) synthesis in tryptophan metabolism, siderophore biosynthetic activity, phosphate solubilization, nitrogen fixation, and related genes, were summarized. IAA (14.4 μg/ml) was presumptively produced by Q2H1 using the Salkowski colorimetric method. A total of five genes, namely, phoU, pstB, pstA1, pstC, and pstS, were annotated for phosphate solubilization, which is associated with the ability of the Q2H1 strain to solubilize phosphate under in vitro conditions. It is revealed that genes in the Q2H1 genome associated with nitrogen fixation belonged to three groups, namely, nitrogen fixation (nifU, sufU, salA, and nifS), nitrogen metabolism (nirA, nrtB, and nasA), and glutamate synthesis (glnA, gltB, gltD, and gudB), supported by evidence that Q2H1 grew on medium without nitrogen. We have also identified a siderophore gene cluster located on the chromosome of Q2H1, including seven genes (viz., rbsR, rhbf, rhbE, rhbD, rhbC, rhbA, ddc, and an unknown gene). In the in vitro assay, a prominent brown circle around the colony was produced on the chrome azurol S medium at 48 and 72 h post-inoculation, indicating that the siderophore gene cluster in Q2H1 harbored the ability to produce siderophores. In summary, these findings implied that identifying strain-specific genes for their metabolic pathways in bacterial endophytes may reveal a variety of significant functions of plant growth-promoting mechanisms.

Effects of Reductive Soil Disinfestation Combined with Liquid-Readily Decomposable Compounds and Solid Plant Residues on the Bacterial Community and Functional Composition.

Reductive soil disinfestation (RSD) incorporated with sole plant residues or liquid-readily decomposable compounds is an effective management strategy to improve soil health. However, the synthetic effects of RSD incorporated with liquid-readily decomposable compounds and solid plant residues on soil ecosystem services remain unclear. Field experiments were carried out to investigate the effects of untreated soil (CK), RSD incorporated with sawdust (SA), molasses (MO), and their combinations (SA + MO) on the bacterial community and functional composition. The results showed that RSD treatments significantly altered soil bacterial community structure compared to CK treatment. The bacterial community structure and composition in MO and SA + MO treatments were clustered compared to SA treatment. This was mainly attributed to the readily decomposable carbon sources in molasses having a stronger driving force to reshape the soil microbial community during the RSD process. Furthermore, the functional compositions, such as the disinfestation efficiency of F. oxysporum (96.4 - 99.1%), abundances of nitrogen functional genes, soil metabolic activity, and functional diversity, were significantly increased in all of the RSD treatments. The highest disinfestation efficiency and abundances of denitrification (nirS and nrfA) and nitrogen fixation (nifH) genes were observed in SA + MO treatment. Specifically, SA + MO treatment enriched more abundant beneficial genera, e.g., Oxobacter, Paenibacillus, Cohnella, Rummeliibacillus, and Streptomyces, which were significantly and positively linked to disinfestation efficiency, soil metabolic activity, and denitrification processes. Our results indicated that combining RSD practices with liquid-readily decomposable compounds and solid plant residues could effectively improve soil microbial community and functional composition.

The host niches of soybean rather than genetic modification or glyphosate application drive the assembly of root-associated microbial communities.

Plant roots significantly influence soil microbial diversity, and soil microorganisms play significant roles in both natural and agricultural ecosystems. Although the genetically modified (GM) crops with enhanced insect and herbicide resistance are thought to have unmatched yield and stress resistance advantages, thorough and in-depth case studies still need to be carried out in a real-world setting due to the potential effects of GM plants on soil microbial communities. In this study, three treatments were used: a recipient soybean variety Jack, a triple transgenic soybean line JD321, and the glyphosate-treated JD321 (JD321G). Three sampling stages (flowering, seed filling and maturing), as well as three host niches of soybean rhizosphere [intact roots (RT), rhizospheric soil (RS) and surrounding soil (SS)] were established. In comparison to Jack, the rhizospheric soil of JD321G had higher urease activity and lower nitrite reductase at the flowering stage. Different treatments and different sampling stages existed no significant effects on the compositions of microbial communities at different taxonomic levels. However, at the genus level, the relative abundance of three plant growth-promoting fungal genera (i.e. Mortierella, Chaetomium and Pseudombrophila) increased while endophytic bacteria Chryseobacterium and pathogenic bacteria Streptomyces decreased from the inside to the outside of the roots (i.e. RT → RS → SS). Moreover, two bacterial genera, Bradyrhizobium and Ensifer were more abundant in RT than in RS and SS, as well as three species, Agrobacterium radiobacter, Ensifer fredii and Ensifer meliloti, which are closely related to nitrogen-fixation. Furthermore, five clusters of orthologous groups (COGs) associated to nitrogen-fixation genes were higher in RT than in RS, whereas only one COG annotated as dinitrogenase iron-molybdenum cofactor biosynthesis protein was lower. Overall, the results imply that the rhizosphere host niches throughout the soil-plant continuum largely control the composition and function of the root-associated microbiome of triple transgenic soybean.

Characterization of diazotrophic root endophytes in Chinese silvergrass (Miscanthus sinensis)

BackgroundPhytoremediation is a potentially cost-effective way to remediate highly contaminated mine tailing sites. However, nutrient limitations, especially the deficiency of nitrogen (N), can hinder the growth of plants and impair the phytoremediation of mine tailings. Nevertheless, pioneer plants can successfully colonize mine tailings and exhibit potential for tailing phytoremediation. Diazotrophs, especially diazotrophic endophytes, can promote the growth of their host plants. This was tested in a mine-tailing habitat by a combination of field sampling, DNA-stable isotope probing (SIP) analysis, and pot experiments.ResultsBacteria belonging to the genera Herbaspirillum, Rhizobium, Devosia, Pseudomonas, Microbacterium, and Delftia are crucial endophytes for Chinese silvergrass (Miscanthus sinensis) grown in the tailing, the model pioneer plant selected in this study. Further, DNA-SIP using 15N2 identified Pseudomonas, Rhizobium, and Exiguobacterium as putative diazotrophic endophytes of M. sinensis. Metagenomic-binning suggested that these bacteria contained essential genes for nitrogen fixation and plant growth promotion. Finally, two diazotrophic endophytes Rhizobium sp. G-14 and Pseudomonas sp. Y-5 were isolated from M. sinensis. Inoculation of another pioneer plant in mine tailings, Bidens pilosa, with diazotrophic endophytes resulted in successful plant colonization, significantly increased nitrogen fixation activity, and promotion of plant growth.ConclusionsThis study indicated that diazotrophic endophytes have the potential to promote the growth of pioneer plant B. pilosa in mine tailings.EAsdjSo3ZCTEVznKh83dpJVideo

Biological nitrogen fixation and nifH gene abundance in deadwood of 13 different tree species

Deadwood is an extremely nitrogen (N) poor plant litter whose decomposition may rely on external N sources. Biological N2 fixation (BNF) by free-living diazotrophs, encoded with the nifH gene, is a potential pathway of N acquisition in deadwood. Still, the control of this process by tree species specific traits is hardly known. Here, we examined (1) BNF rates and nifH gene abundances in deadwood of 13 tree species after 12 years of decomposition and (2) how BNF was related to nutrient concentrations and non-structural carbohydrates (NSC). Comparing our BNF rates with the literature revealed no difference for angiosperms but gymnosperms. Large variability prevented any significant difference in BNF and nifH gene abundance between tree species identities of deadwood. Still, higher BNF and nifH gene abundances occurred in the group of diffuse-porous angiosperms compared to ring-porous angiosperms and gymnosperms. Positive relationships between BNF rates and molybdenum, phosphor, sulfur, and N suggested co-limitation of BNF by several nutrients in deadwood. Relatively high NSC concentrations indicated good carbon availability for diazotrophs. Furthermore, structural equation modeling highlighted the importance of water content for BNF and nifH gene abundance, although the overall explanatory power was low. In conclusion, BNF is a ubiquitous microbial process in deadwood of native European tree species and the comparison with other studies suggests no down-regulation of BNF by high N deposition in Central Europe.

Selenium enhanced nitrogen accumulation in legumes in soil with rhizobia bacteria

The loss of nitrogen in soil directly leads to ecological pollution such as water eutrophication. Urgent need to develop green and efficient approaches to increase nitrogen absorption in plants benefits to improve soil nitrogen use efficiency. Selenium is one of the beneficial elements of plants and has been proven to promote efficient nitrogen uptake by plants. However, previous studies focused on the effects of selenium on plant physiological functions while ignoring the regulation of selenium on the soil nitrogen cycle. This study aims to investigate the regulatory effect of selenium on legume rhizosphere microecology and soil nitrogen cycle process, and the key to legume biomass and nitrogen accumulation, soil physicochemical factors, microbiome characteristics and soil nitrogen cycle process. The results showed that selenium application increased the biomass (increased by 7.26%∼21.85% compared with CK) and nitrogen accumulation of legumes (increased by 18.29% ∼ 54.09% compared with CK). By 16S rRNA sequencing of rhizosphere soil and roots, we found selenium increased nitrogen-fixing bacteria such as Rhizobiaceae and Frankia and nitrifying bacteria such as Proteobacteria, Firmicutes and Chloroflex in rhizosphere. Moreover, selenium up-regulated soil nitrogen fixation genes (nifH) and nitrification genes (Arch-amoA, nxrA), and down-regulated denitrification genes (narG, nirS). These changes lead to a significant decrease in soil NO3−/NH4+, which was helpful for the uptake and utilization of legumes. When soil inoculated with rhizobia, the nitrogen accumulation in legumes promoted by Se was much higher than soil non-inoculated with rhizobia bacteria. This study explored the feasibility and great potential of selenium to promote nitrogen accumulation in legumes from the perspective of rhizosphere microecology, proposes and proves the approach to reduce the application of large amounts of elements by introducing selenium, which is conducive to reducing the use of nitrogen fertilizer from the source and alleviating nitrogen pollution.

Endophytic Bosea spartocytisi sp. nov. Coexists with rhizobia in root nodules of Spartocytisus supranubius growing in soils of Teide National Park (Canary Islands)

Two rod-shaped Gram negative strains, SSUT16T and SSUT22, were isolated from root nodules of Spartocytisus supranubius in soils of the Teide National Park (Tenerife, Spain). The 16S rRNA gene sequences of these two novel strains classified them within genus Bosea with similarity values ranging from 97.65 % to 99.54 % with respect to the other species of this genus. The MLSA analysis from a concatenation of the two housekeeping- genes, recA and gyrB, showed that Bosea thiooxidans LMG 26210T and B. robiniae LMG 26381T are the two closest relative species with which they share similarity sequences values of 94.42 % and 94.27 %, respectively. The genome sequence analysis of strain SSUT16T showed average nucleotide identity percentages (ANIb) and digital DNA-DNA hybridization (dDDH) below 84 % and 33 %, respectively, with the type strains of all sequenced species of genus Bosea. These values are much lower than the currently accepted cut-off values for these two parameters to delineate bacterial species, confirming that the novel strains constitute a novel Bosea species. In addition, they are also distinguished from the other closest species in their fatty acid composition and in other phenotypic characteristics. Genome sequence analysis showed the absence of the common nodulation and nitrogen fixation genes in the novel strains. Therefore, based on the results of phylogenetic, genomic, chemotaxonomic and phenotypic characterization, we propose a new species named Bosea spartocytisi sp. nov., with type strain SSUT16T (=LMG 32510T = CECT 30526T = HAMBI 3759T).

Functionally diverse microbial communities show resilience in response to a record-breaking rain event

Estuaries provide many ecosystem services and host a majority of the world’s population. Here, the response of microbial communities after a record-breaking flood event in a highly urbanized estuary was followed. Hurricane Harvey (hereafter Harvey) was a category 4 hurricane that made landfall on the Texas coast in 2017 and lashed the Houston area with 1.4–1.7 × 1010 m3 of rainfall, disrupting the natural gradients of nutrients and salinity. Here, we utilized metagenomics to analyze how Harvey altered the microbial community of Galveston Bay over five weeks following the storm. We hypothesized that the community would shift from a marine dominated community to that of a terrestrial and freshwater origin. We found that following the storm there were changes in the distribution of species with specific metabolic capacities, such as Cyanobacteria, enriched in oxygenic photosynthesis and nitrogen fixation genes, as well as Verrucomicrobia and Betaproteobacteria, with high prevalence of the SOX complex and anoxygenic photosynthesis genes. On the other hand, dominant members of the community with more diverse metabolic capabilities showed less fluctuations in their distribution. Our results highlight how massive precipitation disturbances can alter microbial communities and how the coalescence of diverse microorganisms creates a resilient community able to maintain ecosystem services even when the system is in an altered state.

History of the Study of the Genus Thiothrix: From the First Enrichment Cultures to Pangenomic Analysis.

Representatives of the genus Thiothrix are filamentous, sulfur-oxidizing bacteria found in flowing waters with counter-oriented sulfide and oxygen gradients. They were first described at the end of the 19th century, but the first pure cultures of this species only became available 100 years later. An increase in the number of described Thiothrix species at the beginning of the 21st century shows that the classical phylogenetic marker, 16S rRNA gene, is not informative for species differentiation, which is possible based on genome analysis. Pangenome analysis of the genus Thiothrix showed that the core genome includes genes for dissimilatory sulfur metabolism and central metabolic pathways, namely the Krebs cycle, Embden–Meyerhof–Parnas pathway, glyoxylate cycle, Calvin–Benson–Bassham cycle, and genes for phosphorus metabolism and amination. The shell part of the pangenome includes genes for dissimilatory nitrogen metabolism and nitrogen fixation, for respiration with thiosulfate. The dispensable genome comprises genes predicted to encode mainly hypothetical proteins, transporters, transcription regulators, methyltransferases, transposases, and toxin–antitoxin systems.

Phylogenetically and catabolically diverse diazotrophs reside in deep-sea cold seep sediments

Microbially mediated nitrogen cycling in carbon-dominated cold seep environments remains poorly understood. So far anaerobic methanotrophic archaea (ANME-2) and their sulfate-reducing bacterial partners (SEEP-SRB1 clade) have been identified as diazotrophs in deep sea cold seep sediments. However, it is unclear whether other microbial groups can perform nitrogen fixation in such ecosystems. To fill this gap, we analyzed 61 metagenomes, 1428 metagenome-assembled genomes, and six metatranscriptomes derived from 11 globally distributed cold seeps. These sediments contain phylogenetically diverse nitrogenase genes corresponding to an expanded diversity of diazotrophic lineages. Diverse catabolic pathways were predicted to provide ATP for nitrogen fixation, suggesting diazotrophy in cold seeps is not necessarily associated with sulfate-dependent anaerobic oxidation of methane. Nitrogen fixation genes among various diazotrophic groups in cold seeps were inferred to be genetically mobile and subject to purifying selection. Our findings extend the capacity for diazotrophy to five candidate phyla (Altarchaeia, Omnitrophota, FCPU426, Caldatribacteriota and UBA6262), and suggest that cold seep diazotrophs might contribute substantially to the global nitrogen balance.

Phylogenomics Uncovers Evolutionary Trajectory of Nitrogen Fixation in Cyanobacteria.

Biological nitrogen fixation (BNF) by cyanobacteria is of significant importance for the Earth’s biogeochemical nitrogen cycle but is restricted to a few genera that do not form monophyletic group. To explore the evolutionary trajectory of BNF and investigate the driving forces of its evolution, we analyze 650 cyanobacterial genomes and compile the database of diazotrophic cyanobacteria based on the presence of nitrogen fixation gene clusters (NFGCs). We report that 266 of 650 examined genomes are NFGC-carrying members, and these potentially diazotrophic cyanobacteria are unevenly distributed across the phylogeny of Cyanobacteria, that multiple independent losses shaped the scattered distribution. Among the diazotrophic cyanobacteria, two types of NFGC exist, with one being ancestral and abundant, which have descended from diazotrophic ancestors, and the other being anaerobe-like and sparse, possibly being acquired from anaerobic microbes through horizontal gene transfer. Interestingly, we illustrate that the origin of BNF in Cyanobacteria coincide with two major evolutionary events. One is the origin of multicellularity of cyanobacteria, and the other is concurrent genetic innovations with massive gene gains and expansions, implicating their key roles in triggering the evolutionary transition from nondiazotrophic to diazotrophic cyanobacteria. Additionally, we reveal that genes involved in accelerating respiratory electron transport (coxABC), anoxygenic photosynthetic electron transport (sqr), as well as anaerobic metabolisms (pfor, hemN, nrdG, adhE) are enriched in diazotrophic cyanobacteria, representing adaptive genetic signatures that underpin the diazotrophic lifestyle. Collectively, our study suggests that multicellularity, together with concurrent genetic adaptations contribute to the evolution of diazotrophic cyanobacteria.

Synergistic metabolism of carbon and nitrogen: Cyanate drives nitrogen cycle to conserve nitrogen in composting system

In this study, HCO3– was used as a co-substrate for cyanate metabolism to investigate its effect on nitrogen cycle in composting. The results showed that the carbamate content in experimental group (T) with HCO3– added was higher than that in control group (CP) during cooling period. Actinobacteria and Proteobacteria were the dominant phyla for cyanate metabolism, and the process was mediated by cyanase gene (cynS). The cynS abundance was 16.6% higher in T than CP. In cooling period, the nitrification gene hao in T was 8.125% higher than CP. Denitrification genes narG, narH, nirK, norB, and nosZ were 25.64%, 35.33%, 45.93%, 36.62%, and 36.12% less than CP, respectively. The nitrogen fixation gene nifH in T was consistently higher than CP in the late composting period. Conclusively, cyanate metabolism drove the nitrogen cycle by promoting nitrification, nitrogen fixation, and inhibiting denitrification, which improved nitrogen retention and compost quality.

Isolation, genomic characterization, and mushroom growth-promoting effect of the first fungus-derived Rhizobium.

Polyporus umbellatus is a well-known edible and medicinal mushroom, and some bacteria isolated from mushroom sclerotia may have beneficial effects on their host. These mushroom growth-promoting bacteria (MGPBs) are of great significance in the mushroom production. In this work, we aimed to isolate and identify MGPBs from P. umbellatus sclerotia. Using the agar plate dilution method, strain CACMS001 was isolated from P. umbellatus sclerotia. The genome of CACMS001 was sequenced using PacBio platform, and the phylogenomic analysis indicated that CACMS001 could not be assigned to known Rhizobium species. In co-culture experiments, CACMS001 increased the mycelial growth of P. umbellatus and Armillaria gallica and increased xylanase activity in A. gallica. Comparative genomic analysis showed that CACMS001 lost almost all nitrogen fixation genes but specially acquired one redox cofactor cluster with pqqE, pqqD, pqqC, and pqqB involved in the synthesis of pyrroloquinoline quinone, a peptide-derived redox participating in phosphate solubilization activity. Strain CACMS001 has the capacity to solubilize phosphate using Pikovskaya medium, and phnA and phoU involved in this process in CACMS001 were revealed by quantitative real-time PCR. CACMS001 is a new potential Rhizobium species and is the first identified MGPB belonging to Rhizobium. This novel bacterium would play a vital part in P. umbellatus, A. gallica, and other mushroom cultivation.