Diazotrophic Activity Research Articles

Bridging the Spatiotemporal Gap in Diazotroph Activity and Diversity With High-Resolution Measurements

The biological fixation of dinitrogen (N 2) by marine microbes called diazotrophs sustains ∼50% new production in the ocean, boosting CO 2 absorption by photoautotrophs and thus contributing to the mitigation of climate change. New environmental conditions sustaining N 2 fixation have been revealed in recent years, enabling more accurate forecasting of future nitrogen inputs and localized hot spots. However, at present the paucity and biased geographical coverage of N 2 fixation and diazotroph diversity measurements impede attempts to reconcile global nitrogen budgets with observed rates. Most studies have been conducted at disparate spatiotemporal scales, including: (i) discrete and short duration measurements in small seawater volumes isolated from the environment, and (ii) spatial extrapolations and global models of diazotrophy projected over decades to centuries. We argue that this knowledge gap lies at the fine scales: dynamic seawater structures < 200 km wide and < 2 months lifetime. However, the spatiotemporal resolution of conventional oceanographic cruises, with stations separated by tens to hundreds of kilometers, is too poor to resolve fine scale processes. Bridging this gap requires leveraging high spatiotemporal resolution measurements. Here we present and discuss the advantages and disadvantages of contemporary methods and equipment able to provide high-resolution measurements at sea. We also provide insights into high-resolution sampling approaches to be developed in the near future. Increasing the spatiotemporal resolution of diazotroph activity and diversity will provide more realistic quantifications of nitrogen fluxes in the dynamic ocean.

INFLUENCE OF RISOAGRINE SEED INOCULATION ON RHIZOAGRINE MICROFLORA AND WINTER WHEAT YIELDS IN WESTERN SIBERIA FOREST-STEPPE

The researchers conduct the field stationary experiment with the use of mineral fertilizers and straw in the grain and steam crop rotation with the withdrawal field of alfalfa and study the number of microflora in the rhizosphere of winter wheat of the new variety Priirtyshskaya after treatment of seeds with biospecimen of complex effect - risoagrine. The highest number of useful crop groups of microorganisms was observed in the variant with inoculation of crop seeds by risoagrine on the basis of applying mineral fertilizers, as well as in combination of inoculation techniques, application of mineral fertilizers and straw (N15P23 + straw + inoculation), respectively, 444 and 355 million UU/yr with 217 million UU/yr in the control group. In the variant with inoculation of winter wheat seeds by mineral fertilizers (N15P23 + inoculation), the number ofoligonitrophils and bacteria, mineralizing mineral phosphates, increased by 2.2 times, nitrifiers - by 60%, microorganisms that utilize organic nitrogen compounds on MPA - by 39, consuming mineral nitrogen on CAA - by 73% compared to the control group. The celluloseolytic soil activity under winter wheat sowing in variants N15P23 + inoculation and N15P23 + straw + inoculation increased to 66.5-67.0%, exceeded the control group by 1.7 times. The highest increase in the crop’s grains was observed in the combination of mineral, organic (straw) and bacterial (rizoagrine) fertilizers - 40.3% in comparison with the control group. Additional nitrogen removal by winter wheat crop due to the activity of associative diazotrophs varied from 6 to 16.5 kg/ha. Correlative relations of high (r=0.84-0.91) and average (r=0.62-0.72) degree of microorganisms in the rhizosphere were observed among the indicators of crop yield and number of microorganisms. The closest correlation took place between the value of winter wheat grain yield and the number of bacteria growing on MPA, including ammonifiers, and the yield and number of nitrifying bacteria.

Heterotrophic Nitrogen Fixation at the Hyper-Eutrophic Qishon River and Estuary System.

Planktonic heterotrophic diazotrophs (N2-fixers) are widely distributed in marine and freshwater systems, yet limited information is available on their activity, especially in environments with adverse conditions for diazotrophy (e.g., N-rich and oxygenated). Here, we followed the localization and activity of heterotrophic diazotrophs in the hyper-eutrophic N-rich Qishon River—an environment previously considered to be unfavorable for diazotrophy. Our results indicate high heterotrophic N2 fixation rates (up to 6.9 nmol N L–1 d–1), which were approximately three fold higher at an upstream location (freshwater) compared to an estuary (brackish) site. Further, active heterotrophic diazotrophs were capture associated with free-floating aggregates by a newly developed immunolocalization approach. These findings provide new insights on the activity of heterotrophic diazotrophs on aggregates in environments previously considered with adverse conditions for diazotrophy. Moreover, these new insights may be applicable to other aquatic regimes worldwide with similar N-rich/oxygenated conditions that should potentially inhibit N2 fixation.

Nutrient stoichiometry (N:P) controls nitrogen fixation and distribution of diazotrophs in a tropical eutrophic estuary

Nitrogen fixation and its ecological regulation are poorly understood in the tropical estuaries, which are highly influenced by anthropogenic disturbances. In this study, we investigated the role of nutrient stoichiometry in the diversity, abundance and activity of N2-fixing bacterial community and their seasonal variations in the water column of a tropical eutrophic estuary (Cochin estuary). The N2 fixation rates in the estuary ranged from 0.1 to 2.0 nmol N2 l−1 h−1, with higher activity during post-monsoon and lower during monsoon. The rates are appeared to be primarily controlled by dissolved inorganic nitrogen and phosphorous (N:P) ratio. Clone library analysis of nitrogenase (nifH) gene revealed that the major N2 fixing phylotypes belong to Cluster I and Cluster III diazotrophs. The overall findings of this study suggest that monsoon induced seasonal changes in nutrient stoichiometry control the distribution and activity of diazotrophs in a tropical estuary.

Evidence for phylogenetically and catabolically diverse active diazotrophs in deep-sea sediment

Diazotrophic microorganisms regulate marine productivity by alleviating nitrogen limitation. However, we know little about the identity and activity of diazotrophs in deep-sea sediments, a habitat covering nearly two-thirds of the planet. Here, we identify candidate diazotrophs from Pacific Ocean sediments collected at 2893 m water depth using 15N-DNA stable isotope probing and a novel pipeline for nifH sequence analysis. Together, these approaches detect an unexpectedly diverse assemblage of active diazotrophs, including members of the Acidobacteria, Firmicutes, Nitrospirae, Gammaproteobacteria, and Deltaproteobacteria. Deltaproteobacteria, predominately members of the Desulfobacterales and Desulfuromonadales, are the most abundant diazotrophs detected, and display the most microdiversity of associated nifH sequences. Some of the detected lineages, including those within the Acidobacteria, have not previously been shown to fix nitrogen. The diazotrophs appear catabolically diverse, with the potential for using oxygen, nitrogen, iron, sulfur, and carbon as terminal electron acceptors. Therefore, benthic diazotrophy may persist throughout a range of geochemical conditions and provide a stable source of fixed nitrogen over geologic timescales. Our results suggest that nitrogen-fixing communities in deep-sea sediments are phylogenetically and catabolically diverse, and open a new line of inquiry into the ecology and biogeochemical impacts of deep-sea microorganisms.

Amazon River influence on nitrogen fixation in the western tropical North Atlantic

We measured rates of N- and C-fixation with a direct tracer method in regions of the western tropical North Atlantic influenced by the Amazon River plume during the high flow period of 2010 (May–June 2010). We found distinct regional variations in N-fixation activity, with the lowest rates in the plume proper and the highest rates in the plume margins and in offshore waters. A comparison of our N- and C-fixation measurements showed that the relative contribution of N-fixation to total primary production increased from the plume core toward oceanic waters, and that most of the C-fixation in this system was supported by sources of nitrogen other than those derived from biological N-fixation, or diazotrophy. We complemented these rate experiments with measurements of the δ15N of suspended particles (δ15PN), which documented the important and often dominant role of diazotrophs in supplying nitrogen to particulate organic matter in the water column. These coupled measurements revealed that small phytoplankton contributed more new nitrogen to the particulate nitrogen pool than larger phytoplankton. We used a habitat classification method to assess the fac- tors that control diazotrophic activity and contribution to the suspended particle pool, both of which increased from the plume toward oceanic waters. Our findings provide an important constraint on the role of the Amazon plume in creating distinct niches and roles for diazotrophs in the nutrient and carbon budgets of the western tropical North Atlantic.

Fundamentally different global marine nitrogen cycling in response to severe ocean deoxygenation

The present-day marine nitrogen (N) cycle is strongly regulated by biology. Deficiencies in the availability of fixed and readily bioavailable nitrogen relative to phosphate (P) in the surface ocean are largely corrected by the activity of diazotrophs. This feedback system, termed the "nitrostat," is thought to have provided close regulation of fixed-N speciation and inventory relative to P since the Proterozoic. In contrast, during intervals of intense deoxygenation such as Cretaceous ocean anoxic event (OAE) 2, a few regional sedimentary δ15N records hint at the existence of a different mode of marine N cycling in which ammonium plays a major role in regulating export production. However, the global-scale dynamics during this time remain unknown. Here, using an Earth System model and taking the example of OAE 2, we provide insights into the global marine nitrogen cycle under severe ocean deoxygenation. Specifically, we find that the ocean can exhibit fundamental transitions in the species of nitrogen dominating the fixed-N inventory--from nitrate (NO3-) to ammonium (NH4+)--and that as this transition occurs, the inventory can partially collapse relative to P due to progressive spatial decoupling between the loci of NH4+ oxidation, NO3- reduction, and nitrogen fixation. This finding is relatively independent of the specific state of ocean circulation and is consistent with nitrogen isotope and redox proxy data. The substantive reduction in the ocean fixed-N inventory at an intermediate state of deoxygenation may represent a biogeochemical vulnerability with potential implications for past and future (warmer) oceans.

Divergent Capacity of Scleractinian and Soft Corals to Assimilate and Transfer Diazotrophically Derived Nitrogen to the Reef Environment.

Corals are associated with dinitrogen (N2)-fixing bacteria that potentially represent an additional nitrogen (N) source for the coral holobiont in oligotrophic reef environments. Nevertheless, the few studies investigating the assimilation of diazotrophically derived nitrogen (DDN) by tropical corals are limited to a single scleractinian species (i.e., Stylophora pistillata). The present study quantified DDN assimilation rates in four scleractinian and three soft coral species from the shallow waters of the oligotrophic Northern Red Sea using the 15N2 tracer technique. All scleractinian species significantly stimulated N2 fixation in the coral-surrounding seawater (and mucus) and assimilated DDN into their tissue. Interestingly, N2 fixation was not detected in the tissue and surrounding seawater of soft corals, despite the fact that soft corals were able to take up DDN from a culture of free-living diazotrophs. Soft coral mucus likely represents an unfavorable habitat for the colonization and activity of diazotrophs as it contains a low amount of particulate organic matter, with a relatively high N content, compared to the mucus of scleractinian corals. In addition, it is known to present antimicrobial properties. Overall, this study suggests that DDN assimilation into coral tissues depends on the presence of active diazotrophs in the coral’s mucus layer and/or surrounding seawater. Since N is often a limiting nutrient for primary productivity in oligotrophic reef waters, the divergent capacity of scleractinian and soft corals to promote N2 fixation may have implications for N availability and reef biogeochemistry in scleractinian versus soft coral-dominated reefs.

Effects of emergent aquatic plants on nitrogen transformation processes and related microorganisms in a constructed wetland in northern China

Wetland plants affect ammonia-oxidation rate (AOR) and nitrogen-fixation rate (NFR), resulting in changes in plant-available nitrogen and wetland nitrogen-cycling. This study aims to investigate the underlying mechanisms and influencing factors associated with ammonium oxidation and nitrogen fixation in response to wetland-emergent aquatic plants. A field experiment was conducted to study the response of activity, abundance, and community structure of ammonia-oxidizers and diazotrophs to three wetland-emergent aquatic plants in summer and autumn. The root exudates and soil physicochemical properties were determined due to their important role in rhizosphere interactions. AOR and NFR were significantly higher in rhizosphere than in bulk soils. The increases of AOR and NFR in rhizosphere were significantly positively affected by oxidation-reduction potential, additionally, NFR was also significantly affected by sugar-containing root exudates. Furthermore, NFR strongly correlated with diazotroph abundance, whereas AOR was linearly correlated with ammonia-oxidizing archaea (AOA) activity and mostly determined by a specific AOA taxon. Emergent plant rhizosphere exhibited strong positive effects on AOR and NFR by altering community structure and abundance of related microorganisms. Release of available oxygen and carbon in plant rhizospheres is vital for promoting ammonium oxidation and nitrogen fixation and might further mediate wetlands nitrogen-cycling.

Multiple factors drive the abundance and diversity of the diazotrophic community in typical farmland soils of China.

Biological nitrogen fixation plays an important role in nitrogen cycling by transferring atmospheric N2 to plant-available N in the soil. However, the diazotrophic activity and distribution in different types of soils remain to be further explored. In this study, 152 upland soils were sampled to examine the diazotrophic abundance, nitrogenase activity, diversity and community composition by quantitative polymerase chain reaction, acetylene reduction assay and the MiSeq sequencing of nifH genes, respectively. The results showed that diazotrophic abundance and nitrogenase activity varied among the three soil types. The diazotrophic community was mainly dominated by Bradyrhizobium, Azospirillum, Myxobacter, Desulfovibrio and Methylobacterium. The symbiotic diazotroph Bradyrhizobium was widely distributed among soils, while the distribution of free-living diazotrophs showed large variation and was greatly affected by multiple factors. Crop type and soil properties directly affected the diazotrophic ɑ-diversity, while soil properties, climatic factors and spatial distance together influenced the diazotrophic community. Network structures were completely different among all three types of soils, with most complex interactions observed in the Red soil. These findings suggest that diazotrophs have various activities and distributions in the three soil types, which played different roles in nitrogen input in agricultural soil in China, being driven by multiple environmental factors.

High Rates of N2 Fixation in Temperate, Western North Atlantic Coastal Waters Expand the Realm of Marine Diazotrophy

AbstractDinitrogen (N2) fixation can alleviate N limitation of primary productivity by introducing fixed nitrogen (N) to the world's oceans. Although measurements of pelagic marine N2 fixation are predominantly from oligotrophic oceanic regions, where N limitation is thought to favor growth of diazotrophic microbes, here we report high rates of N2 fixation from seven cruises spanning four seasons in temperate, western North Atlantic coastal waters along the North American continental shelf between Cape Hatteras and Nova Scotia, an area representing 6.4% of the North Atlantic continental shelf area. Integrating average areal rates of N2 fixation during each season and for each domain in the study area, the estimated N input from N2 fixation to this temperate shelf system is 0.02 Tmol N/year, an amount equivalent to that previously estimated for the entire North Atlantic continental shelf. Unicellular group A cyanobacteria (UCYN‐A) were most often the dominant diazotrophic group expressing nifH, a gene encoding the nitrogenase enzyme, throughout the study area during all seasons. This expands the domain of these diazotrophs to include coastal waters where dissolved N concentrations are not always depleted. Further, the high rates of N2 fixation and diazotroph diversity along the western North Atlantic continental shelf underscore the need to reexamine the biogeography and the activity of diazotrophs along continental margins. Accounting for this substantial but previously overlooked source of new N to marine systems necessitates revisions to global marine N budgets.

Bleaching forces coral's heterotrophy on diazotrophs and Synechococcus.

Coral reefs are threatened by global warming, which disrupts the symbiosis between corals and their photosynthetic symbionts (Symbiodiniaceae), leading to mass coral bleaching. Planktonic diazotrophs or dinitrogen (N2)-fixing prokaryotes are abundant in coral lagoon waters and could be an alternative nutrient source for corals. Here we incubated untreated and bleached coral colonies of Stylophora pistillata with a 15N2-pre-labelled natural plankton assemblage containing diazotrophs. 15N2 assimilation rates in Symbiodiniaceae cells and tissues of bleached corals were 5- and 30-fold higher, respectively, than those measured in untreated corals, demonstrating that corals incorporate more nitrogen derived from planktonic diazotrophs under bleaching conditions. Bleached corals also preferentially fed on Synechococcus, nitrogen-rich picophytoplanktonic cells, instead of Prochlorococcus and picoeukaryotes, which have a lower cellular nitrogen content. By providing an alternative source of bioavailable nitrogen, both the incorporation of nitrogen derived from planktonic diazotrophs and the ingestion of Synechococcus may have profound consequences for coral bleaching recovery, especially for the many coral reef ecosystems characterized by high abundance and activity of planktonic diazotrophs.

Diazotroph activity in surface Narragansett Bay sediments in summer is stimulated by hypoxia and organic matter delivery

MEPS Marine Ecology Progress Series Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsTheme Sections MEPS 614:35-50 (2019) - DOI: https://doi.org/10.3354/meps12901 Diazotroph activity in surface Narragansett Bay sediments in summer is stimulated by hypoxia and organic matter delivery R. F. Spinette1,2, S. M. Brown1,3, A. L. Ehrlich1,4, G. Puggioni5, C. Deacutis6, B. D. Jenkins1,2,* 1Department of Cell and Molecular Biology, University of Rhode Island, Kingston, Rhode Island 02881, USA 2Graduate School of Oceanography, Narragansett, Rhode Island 02882, USA 3Sailors for the Sea, Newport, Rhode Island 02840, USA 4Sema4, Branford, Connecticut 06405, USA 5Department of Computer Science and Statistics, University of Rhode Island, Kingston, Rhode Island 02881, USA 6Rhode Island Department of Environmental Management, Providence, Rhode Island 02908, USA *Corresponding author: bjenkins@uri.edu ABSTRACT: Bacteria that carry out many processes of the nitrogen cycle inhabit estuarine sediments. Denitrification is known to be a dominant process causing estuarine sediments to behave as net nitrogen sinks. However, measurements of nitrogen fluxes in the sediments of Narragansett Bay, Rhode Island, USA, have at times revealed high rates of net nitrogen (N2) fixation. Whereas changes in primary production, in magnitude and phenology, within Narragansett Bay have been identified as possible causes for these changes in nitrogen cycling within the benthos, a factor that has not been examined thus far is seasonal hypoxia. Since anaerobic diazotrophs figure so prominently within the sediments of Narragansett Bay, we hypothesized that dissolved oxygen concentrations in the bottom waters affect their activity. In order to explore this relationship, we measured the activity of diazotrophs in the surface sediments of 3 study areas during the summers of 2013 and 2014 using the acetylene reduction assay. We explored the effects of several water quality parameters on nitrogenase activity including, among others, dissolved oxygen and chlorophyll concentrations. Our measurements of nitrogenase activity were generally low, ranging between 2 and 5 nmol ethylene g-1 d-1 but spiked to 16 nmol ethylene g-1 d-1 at an area experiencing severe hypoxia in July 2013. Our data suggest that diazotrophy in estuarine sediments is enhanced when the benthos experiences very low dissolved oxygen in conjunction with recent influxes of autochthonous organic matter. Experiments with sediment core incubations conducted in the laboratory support our hypothesis that low dissolved oxygen and organic matter additions promote N2 fixation. KEY WORDS: N2 fixation · Diazotrophy · Hypoxia · Estuary · Sediments Full text in pdf format Supplementary material PreviousNextCite this article as: Spinette RF, Brown SM, Ehrlich AL, Puggioni G, Deacutis C, Jenkins BD (2019) Diazotroph activity in surface Narragansett Bay sediments in summer is stimulated by hypoxia and organic matter delivery. Mar Ecol Prog Ser 614:35-50. https://doi.org/10.3354/meps12901 Export citation RSS - Facebook - Tweet - linkedIn Cited by Published in MEPS Vol. 614. Online publication date: April 04, 2019 Print ISSN: 0171-8630; Online ISSN: 1616-1599 Copyright © 2019 Inter-Research.

Evidence of high N&amp;lt;sub&amp;gt;2&amp;lt;/sub&amp;gt; fixation rates in the temperate northeast Atlantic

Abstract. Diazotrophic activity and primary production (PP) were investigated along two transects (Belgica BG2014/14 and GEOVIDE cruises) off the western Iberian Margin and the Bay of Biscay in May 2014. Substantial N2 fixation activity was observed at 8 of the 10 stations sampled, ranging overall from 81 to 384 µmol N m−2 d−1 (0.7 to 8.2 nmol N L−1 d−1), with two sites close to the Iberian Margin situated between 38.8 and 40.7∘ N yielding rates reaching up to 1355 and 1533 µmol N m−2 d−1. Primary production was relatively lower along the Iberian Margin, with rates ranging from 33 to 59 mmol C m−2 d−1, while it increased towards the northwest away from the peninsula, reaching as high as 135 mmol C m−2 d−1. In agreement with the area-averaged Chl a satellite data contemporaneous with our study period, our results revealed that post-bloom conditions prevailed at most sites, while at the northwesternmost station the bloom was still ongoing. When converted to carbon uptake using Redfield stoichiometry, N2 fixation could support 1 % to 3 % of daily PP in the euphotic layer at most sites, except at the two most active sites where this contribution to daily PP could reach up to 25 %. At the two sites where N2 fixation activity was the highest, the prymnesiophyte–symbiont Candidatus Atelocyanobacterium thalassa (UCYN-A) dominated the nifH sequence pool, while the remaining recovered sequences belonged to non-cyanobacterial phylotypes. At all the other sites, however, the recovered nifH sequences were exclusively assigned phylogenetically to non-cyanobacterial phylotypes. The intense N2 fixation activities recorded at the time of our study were likely promoted by the availability of phytoplankton-derived organic matter produced during the spring bloom, as evidenced by the significant surface particulate organic carbon concentrations. Also, the presence of excess phosphorus signature in surface waters seemed to contribute to sustaining N2 fixation, particularly at the sites with extreme activities. These results provide a mechanistic understanding of the unexpectedly high N2 fixation in productive waters of the temperate North Atlantic and highlight the importance of N2 fixation for future assessment of the global N inventory.

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 β-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.

Dam construction alters function and community composition of diazotrophs in riparian soils across an environmental gradient

Dam construction significantly influences riparian ecological functions and environmental conditions in the reservoir region. Despite the importance of diazotrophic mediation of nitrogen cycling in riparian ecosystems, knowledge concerning the impacts of dam construction on diazotrophic function and community composition remains very limited. Here, riparian soils were collected from control sites and dam-affected sites located in six cascade dam-formed reservoirs on the Lancang River, China. Diazotrophic activity was assessed by measuring nitrogen fixation rate, while bacterial abundance and community composition were determined with the nifH gene, using quantitative real-time polymerase chain reaction and MiSeq sequencing, respectively. The results indicate that the activity, abundance, and ɑ-diversity of diazotrophs in the affected sites were significantly higher than those in the control sites. The diazotrophic community composition was differed between the control and affected sites, with the latter showing significantly lower community dissimilarity than the control. Both geographic distance and environmental heterogeneity varied in the diazotrophic community, while environmental heterogeneity was the primary factor controlling diazotrophic assemblage in riparian soils, especially at the affected sites. Moreover, co-occurrence network analyses revealed a much higher possible network connectivity at the affected sites than at the control sites. This study suggests distinct activity, abundance, distribution pattern, and network structure of the diazotrophic populations in riparian soils between dam-affected and control sites; thus, providing a novel insight into the potential ecological effects of dam construction on nitrogen cycling.

Free-living diazotrophs drive castor bean nitrogen input in tropical semiarid soils

ABSTRACT: There are few studies of microbial diversity in castor bean soils in tropical semiarid environmental. Castor bean products have been widely used around the world justifying the commercial importance of ricinoculture in Brazil’ northeastern semiarid. There is no fertilization or maintenance of litter on the soils from the present study, so we hypothesized that the free-living diazotrophs microorganisms drive the nitrogen (N) input into these soils. We evaluated the communities’ structure and diversity of diazotrophs in tropical semiarid soils with i-Castor bean 50 years of cultivation intercropping with maize (CB-50); ii-Only Castor bean 10 years of cultivation (CB-10) and iii-soil under Caatinga vegetation (CAA). Nitrogenase enzyme activity was performed by the acetylene reduction assay (ARA) and the community profile of nifH gene was separated by denaturing gradient gel polyacrylamide electrophoresis (DGGE). Diazotrophs diversity was estimated by the Shannon (H’) and Simpson (D) indexes. Based on the maintenance of soil N stocks and N-microbial biomass over time, the activity of free-living diazotrophs was determinant in the N-input into these soils. The nifH gene was present and its profiles grouped the CB-50 with CAA treatments, so it reinforced the N-fixing by diazotrophs, which presented a greater diversity in the cultivated soils, even after the land use change for the castor bean implementation.

Diazotrophic community and associated dinitrogen fixation within the temperate coral Oculina patagonica.

Dinitrogen (N2 ) fixing bacteria (diazotrophs) are an important source of new nitrogen in oligotrophic environments and represent stable members of the microbiome in tropical corals, while information on corals from temperate oligotrophic regions is lacking. Therefore, this study provides new insights into the diversity and activity of diazotrophs associated with the temperate coral Oculina patagonica from the Mediterranean Sea by combining metabarcoding sequencing of amplicons of both the 16S rRNA and nifH genes and 15 N2 stable isotope tracer analysis to assess diazotroph-derived nitrogen (DDN) assimilation by the coral. Results show that the diazotrophic community of O. patagonica is dominated by autotrophic bacteria (i.e. Cyanobacteria and Chlorobia). The majority of DDN was assimilated into the tissue and skeletal matrix, and DDN assimilation significantly increased in bleached corals. Thus, diazotrophs may constitute an additional nitrogen source for the coral host, when nutrient exchange with Symbiodinium is disrupted (e.g. bleaching) and external food supply is limited (e.g. oligotrophic summer season). Furthermore, we hypothesize that DDN can facilitate the fast proliferation of endolithic algae, which provide an alternative carbon source for bleached O. patagonica. Overall, O. patagonica could serve as a good model for investigating the importance of diazotrophs in coral recovery from bleaching.

Community and abundance of heterotrophic diazotrophs in the northern South China Sea: Revealing the potential importance of a new alphaproteobacterium in N2 fixation

The present study quantified nifH gene abundances of heterotrophic diazotrophs by qPCR in the northern South China Sea (SCS), a region where primary productivity and new production supported by N2 fixation are highest in the winter and unicellular N2-fixers contribute more to nitrogen budget than filamentous N2-fixers. Vertical distributions and seasonal abundances of four prevailing Proteobacteria, two novel α-Proteobacteria (named Alpha 1 and Alpha 2), and two γ-Proteobacteria (γ-24774A11 and Gamma 4) were investigated. The Proteobacteria accounted for 77% of the overall nifH clone library. Both Alpha 1 and Alpha 2 were distributed between 0 and 1000 m depths, while γ-Proteobacteria were concentrated mainly in the upper 200 m. Water-column abundances of Alpha 1, with a maximum of 109 copies m−2, were 1–2 orders of magnitude greater than Alpha 2 or γ-Proteobacteria. Alpha 1 and γ-24774A11 were most abundant in the winter and summer, respectively. Together with UCYN-A and UCYN-B, they were the most important N2-fixers in the northern SCS. Our results suggested that diazotrophic activity of heterotrophic Proteobacteria should be considered an integrated part of nitrogen cycle in oceanic systems in which unicellular diazotrophs prevail over filamentous diazotrophs in N2 fixation.

Response of symbiotic and asymbiotic nitrogen-fixing microorganisms to nitrogen fertilizer application

Biological nitrogen fixation (BNF) plays an important role in nitrogen cycling by transferring atmospheric dinitrogen to the soil. BNF is performed by symbiotic and asymbiotic nitrogen-fixing microorganisms. However, the abundance, activity, and community structure of diazotrophs under different nitrogen fertilizer application rates and how root exudates influence diazotrophs remain unclear. 15N-N2 and 13C-CO2 labeling, DNA-based stable isotope probing (SIP), and molecular biological techniques were used in this study. The abundance, activity, and structure of symbiotic and asymbiotic diazotrophs under different nitrogen fertilizer applications in paddy soil were investigated. We found that the nitrogen fixation capacity in milk vetch (Astragalus sinicus L.) and nifH gene abundance in the root nodules were significantly higher in the low-nitrogen treatment than in the control (zero) and high-nitrogen treatments. Nitrogen-fixing bacteria were abundant in the soils with a high biodiversity. Soil nifH gene sequences were dominated by α-, β-, and δ-proteobacteria, as well as by Cyanobacteria. The relative abundance of α-proteobacteria was lower, and the relative abundance of Cyanobacteria was higher under high nitrogen. Incubation of soil with 13CO2 and subsequent DNA-SIP analysis demonstrated that OTU65 from α-proteobacteria was relatively more abundant in heavy fractions of the 13C-labeled soils than that in the unlabeled soils, indicating that α-proteobacteria may prefer rhizodeposition carbon to other organic carbon. However, OTU24 and OTU73 from δ-proteobacteria had relatively high abundances in light fractions both in the 13C-labeled and unlabeled samples, indicating that δ-proteobacteria may prefer other soil organic carbon to rhizodeposition carbon. The results suggested that soil N availability and rhizodeposition strongly modified the microbial communities of nitrogen-fixing bacteria. Moderate nitrogen application increased the symbiotic biological N fixing activity in the Astragalus sinicus L. rhizosphere. The BNF activity in the legume-rhizobia system is regulated by the exchange of organic C and N nutrient between the host plant and N-fixing bacteria.