Evidence For Nitrogen Fixation Research Articles

Functional Signatures of the Epiphytic Prokaryotic Microbiome of Agaves and Cacti.

Microbial symbionts account for survival, development, fitness and evolution of eukaryotic hosts. These microorganisms together with their host form a biological unit known as holobiont. Recent studies have revealed that the holobiont of agaves and cacti comprises a diverse and structured microbiome, which might be important for its adaptation to drylands. Here, we investigated the functional signatures of the prokaryotic communities of the soil and the episphere, that includes the rhizosphere and phyllosphere, associated with the cultivated Agave tequilana and the native and sympatric Agave salmiana, Opuntia robusta and Myrtillocactus geometrizans by mining shotgun metagenomic data. Consistent with previous phylogenetic profiling, we found that Proteobacteria, Actinobacteria and Firmicutes were the main represented phyla in the episphere of agaves and cacti, and that clustering of metagenomes correlated with the plant compartment. In native plants, genes related to aerobic anoxygenic phototrophy and photosynthesis were enriched in the phyllosphere and soil, while genes coding for biofilm formation and quorum sensing were enriched in both epiphytic communities. In the episphere of cultivated A. tequilana fewer genes were identified, but they belonged to similar pathways than those found in native plants. A. tequilana showed a depletion in several genes belonging to carbon metabolism, secondary metabolite biosynthesis and xenobiotic degradation suggesting that its lower microbial diversity might be linked to functional losses. However, this species also showed an enrichment in biofilm and quorum sensing in the epiphytic compartments, and evidence for nitrogen fixation in the rhizosphere. Aerobic anoxygenic phototrophic markers were represented by Rhizobiales (Methylobacterium) and Rhodospirillales (Belnapia) in the phyllosphere, while photosystem genes were widespread in Bacillales and Cyanobacteria. Nitrogen fixation and biofilm formation genes were mostly related to Proteobacteria. These analyses support the idea of niche differentiation in the rhizosphere and phyllosphere of agaves and cacti and shed light on the potential mechanisms by which epiphytic microbial communities survive and colonize plants of arid and semiarid ecosystems. This study establishes a guideline for testing the relevance of the identified functional traits on the microbial community and the plant fitness.

Basin scale variability of active diazotrophs and nitrogen fixation in the North Pacific, from the tropics to the subarctic Bering Sea

AbstractNitrogen‐fixing microorganisms (diazotrophs) provide biologically available nitrogen to plankton communities and thereby greatly influence the productivity in many marine regions. Various cyanobacterial groups have traditionally been considered the major oceanic diazotrophs, but later noncyanobacterial and presumably heterotrophic diazotrophs were also found to be widespread and potentially important in nitrogen fixation. However, the distribution and activity of different diazotroph groups is still poorly constrained for most oceanic ecosystems. Here we examined diazotroph community structure and activity along a 7500 km south‐north transect between the central equatorial Pacific and the Bering Sea. Nitrogen fixation contributed up to 84% of new production in the upper waters of the subtropical gyre, where the diazotroph community included the gammaproteobacterium γ‐24774A11 and highly active cyanobacterial phylotypes (>50% of total nifH transcript abundance). Nitrogen fixation was sometimes detectable down to 150 m depth and extended horizontally to the edge of the gyre at around 35°N. Nitrogen fixation was even detected far north on the Bering Sea shelf. In the Alaskan Coastal Waters on the Bering Sea shelf, low nitrate together with high dissolved iron concentrations seemed to foster diazotroph growth, including a prominent role of UCYN‐A2, which was abundant near the surface (1.2×105 nifH gene copies L−1). Our study provides evidence for nitrogen fixation in the Bering Sea and suggests a clear contrast in the composition of diazotrophs between the tropical/subtropical gyre and the separate waters in the cold northern regions of the North Pacific.

Growth stimulation of barley and biocontrol effect on plant pathogenic fungi by a Cellulosimicrobium sp. strain isolated from salt-affected rhizosphere soil in northwestern Algeria

The plant growth promoting effect of bacterial isolates from salt-affected agricultural rhizospheric soil from Bejaia, Algeria, on barley seedlings as well as biological control abilities of these isolates against phytopathogenic fungi were determined. Four isolates stimulated significantly germination and growth of barley seedlings in an axenic test system and in soil pots. Isolate S16 (Cellulosimicrobium sp.) stimulated the growth of barley seedlings by 185% (stem height of 13.0 ± 0.11 cm) over non-inoculated control seedlings (7.0 ± 0.12 cm). Cellulosimicrobium sp. S16 was found also superior in mycelial growth inhibition assays against the plant pathogenic fungi Botrytis cinerea, Fusarium oxysporum and Verticillium dahliae. Furthermore, several plant growth promoting traits (production of indole acetic acid, inorganic phosphate solubilization, siderophore production) and production of enzymes beneficial for soil fertility (protease, chitinase, amylase and urease) were identified. However, no evidence for nitrogen fixation was found by testing acetylene reduction and the presence of nif-genes. Based on comparative sequence analysis of almost full length 16S-rRNA coding gene fragments, Cellulosimicrobium sp. S16 exhibits the highest similarity of 99.7% to Cellulosimicrobium cellulans (accession number AY665978). Cellulosimicrobium sp. S16 could be a successful candidate for the application as a plant growth promoting inoculant.

Strain-level genomic variation in natural populations of Lebetimonas from an erupting deep-sea volcano

Chemolithoautotrophic Epsilonproteobacteria are ubiquitous in sulfidic, oxygen-poor habitats, including hydrothermal vents, marine oxygen minimum zones, marine sediments and sulfidic caves and have a significant role in cycling carbon, hydrogen, nitrogen and sulfur in these environments. The isolation of diverse strains of Epsilonproteobacteria and the sequencing of their genomes have revealed that this group has the metabolic potential to occupy a wide range of niches, particularly at dynamic deep-sea hydrothermal vents. We expand on this body of work by examining the population genomics of six strains of Lebetimonas, a vent-endemic, thermophilic, hydrogen-oxidizing Epsilonproteobacterium, from a single seamount in the Mariana Arc. Using Lebetimonas as a model for anaerobic, moderately thermophilic organisms in the warm, anoxic subseafloor environment, we show that genomic content is highly conserved and that recombination is limited between closely related strains. The Lebetimonas genomes are shaped by mobile genetic elements and gene loss as well as the acquisition of novel functional genes by horizontal gene transfer, which provide the potential for adaptation and microbial speciation in the deep sea. In addition, these Lebetimonas genomes contain two operons of nitrogenase genes with different evolutionary origins. Lebetimonas expressed nifH during growth with nitrogen gas as the sole nitrogen source, thus providing the first evidence of nitrogen fixation in any Epsilonproteobacteria from deep-sea hydrothermal vents. In this study, we provide a comparative overview of the genomic potential within the Nautiliaceae as well as among more distantly related hydrothermal vent Epsilonproteobacteria to broaden our understanding of microbial adaptation and diversity in the deep sea.

Bacillus simplex—A Little Known PGPB with Anti-Fungal Activity—Alters Pea Legume Root Architecture and Nodule Morphology When Coinoculated with Rhizobium leguminosarum bv. viciae

Two strains, 30N-5 and 30VD-1, identified as Bacillus simplex and B. subtilis, were isolated from the rhizospheres of two different plants, a Podocarpus and a palm, respectively, growing in the University of California, Los Angeles (UCLA) Mildred E. Mathias Botanical Garden. B. subtilis is a well-known plant-growth promoting bacterial species, but B. simplex is not. B. simplex 30N-5 was initially isolated on a nitrogen-free medium, but no evidence for nitrogen fixation was found. Nevertheless, pea plants inoculated with B. simplex showed a change in root architecture due to the emergence of more lateral roots. When Pisum sativum carrying a DR5::GUSA construct, an indicator for auxin response, was inoculated with either B. simplex 30N-5 or its symbiont Rhizobium leguminosarum bv. viciae 128C53, GUS expression in the roots was increased over the uninoculated controls. Moreover, when pea roots were coinoculated with either B. simplex 30N-5 or B. subtilis 30VD-1 and R. leguminosarum bv. viciae 128C53, the nodules were larger, clustered, and developed more highly branched vascular bundles. Besides producing siderophores and solubilizing phosphate, the two Bacillus spp., especially strain 30VD-1, exhibited anti-fungal activity towards Fusarium. Our data show that combining nodulating, nitrogen-fixing rhizobia with growth-promoting bacteria enhances plant development and strongly supports a coinoculation strategy to improve nitrogen fixation, increase biomass, and establish greater resistance to fungal disease.

Evidence of nitrogen fixation in lodgepole pine inoculated with diazotrophicPaenibacillus polymyxa

Diazotrophic bacteria previously isolated from internal tissues of naturally regenerating lodgepole pine ( Pinus contorta var. latifolia (Dougl.) Engelm.) seedlings were tested for their ability to colonize and fix nitrogen (N) in pine germinants in two experiments. Surface sterilized pine seed was sown in glass tubes containing an autoclaved sand – montmorillonite clay mixture that contained a N-limited nutrient solution labeled with15N as 0.35 mmol·L–1Ca(15NO3)2(5%15N label). Pine seed was inoculated with one of three of the following bacterial strains: Paenibacillus polymyxa P2b-2R, P. polymyxa P18b-2R, or Dyadobacter fermentans P19a-2R, and seedlings grew for either 27 or 35 weeks. At the end of each plant growth period, P. polymyxa strain P2b-2R was detected in the pine rhizosphere but not inside plant tissues. Pine foliar N concentrations were not affected by bacterial inoculation but significant foliar15N dilution was observed in seedlings treated with strain P2b-2R (30% and 66%, P < 0.05, in the first and second experiments, respectively). This strain also reduced seedling biomass in both experiments but effects were significant only in the second experiment (36%, P < 0.05). Notwithstanding the negative effect of bacterial inoculation on seedling growth, pine seedlings inoculated with strain P2b-2R derived 30% and 66%, respectively, of their foliar N from bacterial N fixation in two seedling growth experiments. These results demonstrate the possibility that some endophytic diazotrophs facilitate pine seedling growth in N-poor soils.

New evidence for nitrogen fixation within the Italian white truffle Tuber magnatum

Diversity of nitrogen-fixing bacteria and the nitrogen-fixation activity was investigated in Tuber magnatum, the most well-known prized species of Italian white truffle. Degenerate PCR primers were applied to amplify the nitrogenase gene nifH from T. magnatum ascomata at different stages of maturation. Putative amino acid sequences revealed mainly the presence of Alphaproteobacteria belonging to Bradyrhizobium spp. and expression of nifH genes from Bradyrhizobia was detected. The nitrogenase activity evaluated by acetylene reduction assay was 0.5–7.5 μmol C 2H 4 h −1 g −1, comparable with early nodules of legumes associated with specific nitrogen-fixing bacteria. This is the first demonstration of nitrogenase expression gene and activity within truffle.

Active Nanostructures at Interfaces for Photocatalytic Reactors and Low-power Consumption Sensors

AbstractActive nanostructures which provide unique transformations are being introduced to phase matched porous silicon (PS) nano/micropores to form a platform for low power consumption highly selective sensors and microreactors. TiO2-xNx photocatalysts have been formed in seconds at room temperature at the nanoscale via the direct nitration of anatase TiO2 nanocolloids. Tunability throughout the visible depends upon the degree of agglomeration and the ability to seed these nanoparticles with metal ions. Co metal ion seeding leads to the efficient room temperature phase transformation, of anatase to rutile TiO2, where normally much higher temperatures are required. Seeding of a properly nitridated TiO2 nanocolloid with transition metal ions (Co, Ni) allows for the enhancement of the infrared spectra of the TiO2-xNx nitridated titania surface in excess of 10-fold, providing a means to analyze for minor contaminants and intermediates. Evidence for nitrogen fixation is found in Fe treated systems. The TiO2-xNx systems act as visible light absorbing photocatalyts. These photocatalysts and additional nanostructured metal oxides can be placed on the surface of PS-based sensor and microreactor configurations to greatly improve the interface response.

Nitrate anomaly in the upper nutricline in the northern South China Sea ‐ Evidence for nitrogen fixation

Up to 2 μM of nitrate anomaly, N*, were found in the upper nutricline at the South East Asia Time‐series Study (SEATS) site in the northern South China Sea (SCS). These concentrations were among the higher values reported in the Pacific and indicate the significant contribution of the remineralization of nitrogen‐rich organic matter formed by nitrogen fixation to the nutrient dynamics of the area. The concentrations were systematically higher, by up to 2.5 μM, in the Fall through the early Spring, during the northeast monsoon, than in the Summer, suggesting that the impact of nitrogen fixation was higher during the former time period. This pattern is in phase with that of the atmospheric deposition of Asian dust to the northern SCS. The coherence is consistent with a coupling between nitrogen fixation and the availability of atmospherically derived iron.

Evidence of nitrogen fixation by non-heterocystous cyanobacteria in the Baltic Sea and re-calculation of a budget of nitrogen fixation

Nitrogen fixation rates and related plankton parameters were determined at 8 sta- tions in the Baltic proper and Mecklenburg Bay in 1997 and 1998. Nitrogen fixation was mea- sured with 15 N-tracer method in unenriched samples. Measurable nitrogen fixation rates were found from July to October, with highest rates in August when heterocystous cyanobacteria formed blooms. Nitrogen fixation rates measured during a moderate bloom in 1998 were related to biomass of heterocystous and coccoid (non-heterocystous) cyanobacteria and primary produc- tion. The size fraction <10 µm contributed significantly to total nitrogen fixation both during day and night. It is discussed whether this may be due to small, non-heterocystous cyanobacteria, which were abundant in summer and autumn. They may separate photosynthesis and nitrogen fixation temporally (day and night) and may be especially responsible for the high nitrogen fix- ation rates observed in the dark. As the fraction of pico- and nanoplankton was not considered in earlier studies, a new budget of nitrogen fixation in the Baltic proper has been estimated. On average, daily nitrogen fixation rates of 2.5 mmol N m -2 d -1 (in July/August 1997/1998) and mean annual nitrogen fixation of 125 mmol N m -2 yr -1 were estimated for the Baltic proper. The high variability is discussed. For summer 1998, a budget of the nitrogen cycle at the main station in the Gotland Sea is given.

The nitrogen isotopic composition of nitrate in the Kuroshio Water northeast of Taiwan: evidence for nitrogen fixation as a source of isotopically light nitrate

The Kuroshio Water, which provides nutrients to the shelf water of the East China Sea through upwelling at the shelf break northeast of Taiwan, is abundant with nitrogen fixing cyanobacteria. Accordingly, the δ15N values of nitrate at three stations in the sea northeast of Taiwan were analyzed to estimate the flux of biologically fixed nitrogen in the Kuroshio Water by characterization of the isotopic signature of the upwelled nitrate. Prior to sample analysis, we verified using standard KNO3 solution the validity of the sample preparation procedure, which has been modified by reducing the amount of Devarda's alloy to lower the reagent blank. However, the concentrations of nitrate in seawaters estimated from the amount of extracted nitrogen were consistently higher than those measured colorimetrically, suggesting interference from dissolved organic nitrogen of which the δ15N value was estimated to be −3.0 ± 0.2%., Correction was made on each sample based on the estimated δ15N for DON. The δ15N values of waters from 500 to 780 m in the sea northeast of Taiwan fell in the range of + 5.5 to + 6.1%c, which was consistent with those observed in deep waters elsewhere. The δ15N values decreased in the overlying Kuroshio Water by 1–3%. in general. In order to achieve isotope balance in the water column, the input of isotopically light nitrogen, presumably from nitrogen fixation, is required. The nitrogen flux from this source was estimated to be 0.5 mg m−2 day−1 N which is only 8% of the estimated new nitrogen to the Kuroshio surface water. On the other hand, this nitrogen may account for 20–40% of the nitrate in the subsurface water (100–400 m) as a result of accumulation and recycling. The nitrate in the source upwelling water had a δ15N value of +4.7 ± 0.6%., implying 20 ± 9% of it originating from nitrogen fixation.

Discovery and evidence of nitrogen fixation by thermophilic heterotrophs in hot springs

A previously unknown phenomenon, nitrogenase activity at high temperatures by nonphotosynthetic prokaryotes in thermal springs, is reported. This activity was revealed by dark-bottle incubations during cyanobacterial nitrogenase studies in hot springs of Yellowstone National Park. Benthic mat cores from alkaline thermal streams reduced acetylene in the dark from 30° to 60°C. the bacterial activity was distinguished from that of sympatric cyanobacteria by its sensitivity to oxygen (light bottles), heterotrophic response to organic amendments, and distinctive spatial distribution. Field tests for other biotic or abiotic explanations of the activity were negative. Anaerobic enrichment cultures supported the field evidence. Fourteen morphological types of presumptive diazotrophs, including sulfate-reducing and spore-forming rods. have been observed in these cultures.

The physiological significance of the leaf nodules of psychotria

In 6-month growth experiments it was found that leaf-nodulatedPsychotria mucronata seedlings grown in N-poor soil showed a restricted growth and developed severe nitrogen-deficiency symptoms in the leaves. Plants in the same soil supplied with NO3-N showed healthy growth and dark green leaves. Detached Psychotria leaves bearing leaf nodules exposed to an atmosphere containing N15-labelled nitrogen gas or acetylene gas gave no evidence of nitrogen fixation, either in the light or in the dark or in both in succession. Therefore nitrogen fixation is probably not associated with the leaf nodules.

Possibility of Fixation of Atmospheric Nitrogen by the Rice Plant

A RECENT communication1 by Chakraborty and Sen Gupta suggested that rice plants might be capable of fixing atmospheric nitrogen. We have made an attempt to confirm their result using the variety ‘Lead 35’ (a pure-line selection of material derived from British Guiana). No evidence of nitrogen-fixation has been obtained.