Seasonal patterns of nitrogen fixation in termites

1. Termite nitrogenase activity was highest in autumn and spring (≈ 3 μg N₂ fixed termite fresh mass (g)^–1 day^–1) and lowest in winter and summer (≈ 0·8 μg N₂ fixed termite fresh mass (g)^–1 day^–1).
2. The nitrogenase activity of worker termites was significantly higher than all other castes (1·58 ± 0·27 μg N₂ fixed termite fresh mass (g)^–1 day^–1).
3. Worker termites constituted the largest proportion of all the castes throughout the study period (≈ 90%).
4. The localized input of fixed nitrogen by termites may reach 15·3 mg N log^–1 day^–1 and 5·6 g N log^–1 year^–1.     

Seasonal patterns of growth and nitrogen fixation in field-grown pea

The seasonal patterns of growth and symbiotic N₂ fixation under field conditions were studied by growth analysis and use of¹⁵N-labelled fertilizer in a determinate pea cultivar (Pisum sativum L.) grown for harvest at the dry seed stage. The patterns of fertilizer N-uptake were almost identical in pea and barley (the non-fixing reference crop), but more fertilizer-N was recovered in barley than in pea. The estimated rate of N₂ fixation in pea gradually increased during the pre-flowering and flowering growth stages and reached a maximum of 10 kg N fixed per ha per day nine to ten weeks after seedling emergence. This was the time of early pod-development (flat pod growth stage) and also the time for maximum crop growth rate and maximum green leaf area index. A steep drop in N₂ fixation rate occurred during the following week. This drop was simultaneous with lodging of the crop, pod-filling (round pod growth stage) and the initiation of mobilization of nitrogen from vegetative organs. The application of fertilizer-N inhibited the rate of N₂ fixation only during that period of growth, when the main part of fertilizer-N was taken up and shortly after. Total accumulation of fixed nitrogen was estimated to be 244, 238 and 213 kg N ha⁻¹ in pea supplied with nil, 25 or 50 kg NO ₃ ⁻ −N ha⁻¹, respectively. About one-fourth of total N₂ fixation was carried out during preflowering, one fourth during the two weeks of flowering and the remainder during post-flowering. About 55% of the amount of N present in pods at maturity was estimated to be derived from mobilization of N from vegetative organs. “Starter” N (25 or 50 kg NO ₃ ⁻ −N ha⁻¹) did not significantly influence either dry matter and nitrogen accumulation or the development of leaf area. Neither root length and root biomass determined 8 weeks after seedling emergence nor the yield of seed dry matter and nitrogen at maturity were influenced by fertilizer application.     

Microbial community shifts influence patterns in tropical forest nitrogen fixation

The role of biodiversity in ecosystem function receives substantial attention, yet despite the diversity and functional relevance of microorganisms, relationships between microbial community structure and ecosystem processes remain largely unknown. We used tropical rain forest fertilization plots to directly compare the relative abundance, composition and diversity of free-living nitrogen (N)-fixer communities to in situ leaf litter N fixation rates. N fixation rates varied greatly within the landscape, and ‘hotspots’ of high N fixation activity were observed in both control and phosphorus (P)-fertilized plots. Compared with zones of average activity, the N fixation ‘hotspots’ in unfertilized plots were characterized by marked differences in N-fixer community composition and had substantially higher overall diversity. P additions increased the efficiency of N-fixer communities, resulting in elevated rates of fixation per nifH gene. Furthermore, P fertilization increased N fixation rates and N-fixer abundance, eliminated a highly novel group of N-fixers, and increased N-fixer diversity. Yet the relationships between diversity and function were not simple, and coupling rate measurements to indicators of community structure revealed a biological dynamism not apparent from process measurements alone. Taken together, these data suggest that the rain forest litter layer maintains high N fixation rates and unique N-fixing organisms and that, as observed in plant community ecology, structural shifts in N-fixing communities may partially explain significant differences in system-scale N fixation rates.     

Study of nitrogen fixation in microbial communities of oil-contaminated marine sediment microcosms

Aerobic microbial degradation of pollutant oil (petroleum) in aquatic environments is often severely limited by the availability of combined nitrogen. We therefore studied whether the microbial community enriched in marine sediment microcosms with an added oil layer and exposure to light harboured nitrogenase activity. The acetylene reduction (AR) assay indeed indicated active nitrogenase; however, similar activity was observed in oil-free control microcosms. In both microcosms, the AR rate was significantly reduced upon a dark shift, indicating that enriched cyanobacteria were the dominant diazotrophs. Analysis of structural dinitrogenase reductase genes (nifH) amplified from both microcosms indeed revealed NifH sequences related mostly to those of heterocystous cyanobacteria. NifH sequences typically affiliating with those of heterotrophic bacteria were more frequently retrieved from the oil-containing sediment. Expression analyses showed that mainly nifH genes similar to those of heterocystous cyanobacteria were expressed in the light. Upon a dark shift, nifH genes related to those of non-heterocystous cyanobacteria were expressed. Expression of nifH assignable to heterotrophs was apparently not significant. It is concluded that cyanobacteria are the main contributors of fixed nitrogen to oil-contaminated and pristine sediments if nitrogen is a limiting factor and if light is available. Hence, also the oil-degrading heterotrophic community may thus receive a significant part of combined nitrogen from cyanobacteria, even though oil vice versa apparently does not stimulate an additional nitrogen fixation in the enriched community.     

Interactions between nitrogen fixation and oxegenic photosynthesis in a marine cyanobacterial mat

Abstract Cyanobacterial mats developed on fine sandy sediments of the upper littoral of the island of Mellum (North Sea). Freshly colonized sediment was dominated by the non-heterocystous, nitrogen-fixing cyanobacterium Oscillatoria limosa . Well established mats in which the cosmopolitan cyanobacterium Microcoleus chthonoplastes was the dominant organism also usually contained O. limosa as a minor component. This mat was about 1 mm thick and contained high biomass. Photosynthesis was maximal at about 150 μm depth and reached values of 280 μmol oxygen. 1⁻¹ · min⁻¹. On the other hand, in the dark, high respiratory activity turned the mat anaerobic within minutes. Freshly colonized sediment consisted of low cyanobacterial biomass loosely attached to the sand grains and present up to a depth of 2.5 mm. Respiratory activity was low and the sediment remained aerobic to a depth of 2 mm throughout the night. Nitrogen fixation (acetylene reduction) was measured during 24-h periods in both types of mats in order to elucidate interactions with oxygenic photosynthesis and oxygen concentration. Acetylene reduction in the mats showed very different diurnal patterns which depended on the type of mat investigated and the time of year. The results indicated that a temporary separation of oxygenic photosynthesis and nitrogen fixation occurred in the mat. Established mats fixed nitrogen predominantly during the transition from dark to light and vice versa, when oxygenic photosynthesis was reduced or absent. Freshly colonized sediment-fixed nitrogen throughout the night but often a stimulation was seen at dawn. The latter showed much higher specific activities than the established type. Also in spring, specific activities were much higher.     

Diversity and expression of nitrogen fixation genes in bacterial symbionts of marine sponges

Marine sponges contain complex assemblages of bacterial symbionts, the roles of which remain largely unknown. We identified diverse bacterial nifH genes within sponges and found that nifH genes are expressed in sponges. This is the first demonstration of the expression of any protein-coding bacterial gene within a sponge. Two sponges Ircinia strobilina and Mycale laxissima were collected from Key Largo, Florida and had delta(15)N values of c. 0-1 per thousand and 3-4 per thousand respectively. The potential for nitrogen fixation by symbionts was assessed by amplification of nifH genes. Diverse nifH genes affiliated with Proteobacteria and Cyanobacteria were detected, and expression of nifH genes affiliated with those from cyanobacteria was detected. The nifH genes from surrounding seawater were similar to those of Trichodesmium and clearly different from the cyanobacterial nifH genes detected in the two sponges. This study advances understanding of the role of bacterial symbionts in sponges and suggests that provision of fixed nitrogen is a means whereby symbionts benefit sponges in nutrient-limited reef environments. Nitrogen fixation by sponge symbionts is possibly an important source of new nitrogen to the reef environment that heretofore has been neglected and warrants further investigation.     

Developmental patterns related to nitrogen fixation in theNostoc-Gunnera magellanica Lam. symbiosis

Developmental patterns related to nitrogen fixation in the heterocystous cyanobacteriumNostoc harboured in distinct colonies along the stem ofGunnera magellanica Lam. plantlets were examined using successive plant sections. Pronounced morphological, physiological and biochemical alterations in the cyanobacterium were demonstrated. Close to the growing apex the cyanobacterial biomass, contained in smallGunnera cells, was low and consisted mostly of vegetative cells showing a high density of different storage structures except for cyanophycin granules. In contrast, both the total and specific nitrogenase activity and the relative nitrogenase protein level were at maximum within this part; while the frequency of heterocysts increased from zero to 30% within the same area. The nitrogenase protein was localized only in the heterocysts throughout the plant. Further down theGunnera stem there was a progressive increase in both the cyanobacterial biomass and the heterocyst frequency, which finally constituted about 60% of the cyanobacterial cell population. Throughout this part of the stem, cyanophycin granules were frequent in the vegetativeNostoc cells. At the base of the stem, degeneratedNostoc cells dominated and the nitrogenase activity was close to zero, although the nitrogenase protein remained. Degeneration of theNostoc cells and leaf shedding coincided. Both intact plants (approx. 20 mm in height) and plant stem sections (2 mm in length) showed substantial nitrogenase activity, although sectioning caused a 30% reduction in total nitrogenase activity.     

On the role of oxygen for nitrogen fixation in the marine cyanobacterium Trichodesmium sp

The marine, non-heterocystous, filamentous cyanobacterium Trichodesmium shows a distinct diurnal pattern of nitrogenase activity. In an attempt to reveal the factors that control this pattern, a series of measurements were carried out using online acetylene reduction assay. Light response curves of nitrogenase were recorded applying various concentrations of oxygen. The effect of oxygen depended on the irradiance applied. Above a photon irradiance of 16 mumol m(-2) s(-1) nitrogenase activity was highest under anoxic conditions. Below this irradiance the presence of oxygen was required to achieve highest nitrogenase activity and in the dark 5% oxygen was optimal. At any oxygen concentration a photon irradiance of 100 mumol m(-2) s(-1) was saturating. When Trichodesmium was incubated in the dark, nitrogenase activity gradually decreased and this decline was higher at higher levels of oxygen. The activity recovered when the cells were subsequently incubated in the light. This recovery depended on oxygenic photosynthesis because it did not occur in the presence of DCMU [3-(3,4-dichlorophenyl)-1,1-dimethylurea]. Recovery of nitrogenase activity in the light was faster at low oxygen concentrations. The results showed that under aerobic conditions nitrogenase activity was limited by the availability of reducing equivalents suggesting a competition for electrons between nitrogenase and respiration.     

Biological nitrogen fixation

And he gave it for his opinion, that whoever could make two ears of corn or two blades of grass to grow upon a spot of ground where only one grew before, would deserve better of mankind, and do more essential service to his country than the whole race of politicians put together. {Jonathan Swift, ‘Gulliver's Travels’, Voyage to Brobdingnag, Ch. 7.)     

Biological nitrogen fixation: rates,patterns and ecological controls in terrestrial ecosystems

New techniques have identified a wide range of organisms with the capacity to carry out biological nitrogen fixation (BNF)—greatly expanding our appreciation of the diversity and ubiquity of N fixers—but our understanding of the rates and controls of BNF at ecosystem and global scales has not advanced at the same pace. Nevertheless, determining rates and controls of BNF is crucial to placing anthropogenic changes to the N cycle in context, and to understanding, predicting and managing many aspects of global environmental change. Here, we estimate terrestrial BNF for a pre-industrial world by combining information on N fluxes with ¹⁵N relative abundance data for terrestrial ecosystems. Our estimate is that pre-industrial N fixation was 58 (range of 40–100) Tg N fixed yr⁻¹; adding conservative assumptions for geological N reduces our best estimate to 44 Tg N yr⁻¹. This approach yields substantially lower estimates than most recent calculations; it suggests that the magnitude of human alternation of the N cycle is substantially larger than has been assumed.     

Diurnal and seasonal patterns of nitrogen fixation in analnus hirsuta plantation of central Korea

We used an acetylene reduction assay to measure rates of nitrogen fixation on a 38-year-oldAlnus hirsuta plantation in central Korea. The diurnal pattern of acetylene reduction changed significantly during May, August, and October, typically varying by 3-fold throughout the course of the day. Maximum rates occurred at 3 p.m. in May and October, but at 6 p.m. in August. Increasing trends were evident during the early growing season, with sustained high rates from mid-May through late September; July had the highest rates, averaging 7.2 μmole g^-1 dry nodule h^-1. The average nodule biomass for this plantation was 220 kg ha ’. Rates of acetylene reduction were related to soil temperature, but not to soil moisture content. Combining these nodule biomass calculations with seasonal average acetylene reduction rates yielded an estimate of current annual nitrogen fixation of 60 kg N ha^-1 for the plantation. This rate of annual nitrogen addition was very large in relation to the yearly nitrogen requirements of coniferous and deciduous forests in central Korea.     

Emerging patterns of epigenomic variation

Fuelled by new sequencing technologies, epigenome mapping projects are revealing epigenomic variation at all levels of biological complexity, from species to cells. Comparisons of methylation profiles among species reveal evolutionary conservation of gene body methylation patterns, pointing to the fundamental role of epigenomes in gene regulation. At the human population level, epigenomic changes provide footprints of the effects of genomic variants within the vast nonprotein-coding fraction of the genome, and comparisons of the epigenomes of parents and their offspring point to quantitative epigenomic parent-of-origin effects confounding classical Mendelian genetics. At the organismal level, comparisons of epigenomes from diverse cell types provide insights into cellular differentiation. Finally, comparisons of epigenomes from monozygotic twins help dissect genetic and environmental influences on human phenotypes and longitudinal comparisons reveal aging-associated epigenomic drift. The development of new bioinformatic frameworks for comparative epigenome analysis is putting epigenome maps within the reach of researchers across a wide spectrum of biological disciplines.     

Problems and promises of assaying the genetic potential for nitrogen fixation in the marine environment

Nitrogen fixation in the sea has attracted the attention of ecologists for decades. Much is known about the habitats in which it occurs and some of the factors that limit N₂ fixation activity in different environments, but we still know little about the organisms that fix nitrogen, and what limits the growth and distribution of these organisms in marine environments. Molecular biology technological developments have provided tools for detecting and characterizing N₂-fixing organisms in the environment. These techniques hold great promise for unraveling the mysteries and paradoxes of N₂ fixation in the sea. In this review, we address the theoretical basis for the use of a molecular approach to N₂ fixation, highlight the strengths and weaknesses of the approach, and provide case studies that demonstrate the potential contribution of molecular biology approaches to studies of N₂ fixation in the sea. Correspondence to: J.P. Zehr.     

Germination, field establishment patterns and nitrogen fixation of indigenous legumes on nutrient-depleted soils

Integrating N₂-fixing indigenous legumes in smallholder farming systems has potential to alleviate some of the major soil fertility constraints associated with lack of nitrogen (N) inputs in many parts of Sub-SaharanAfrica. Studies were conducted under low (450–650 mm yr^?1) and high (>800 mm yr^?1) rainfall areas in Zimbabwe to investigate the establishment and nitrogen fixation patterns of fifteen indigenous legume species. The legume seeds were broadcast in mixtures at 120 seeds m^?2 species^?1 during 2004/05 and 2005/06 rainfall seasons.Eriosema ellipticum, Crotalaria ochroleuca andC. pallida had emergence rates above 15% compared with <10% forTephrosia radicans andIndigofera astragalina. Seed hardness accounted for >50% germination failure, while low viability explained 10–30%.Crotalaria ochroleuca andC. pallida attained a maximum biomass of 5–9 t ha^?1 (dry weight) over six months, while species that reached peak biomass over three months (e.g.C. cylindrostachys andC. glauca) gave lowest yields of ≈0.5 t ha^?1. Biennials,Neonotonia wightii, E. ellipticum and Tephrosia radicans, exhibited slow growth rates and only attained their maximum biomass of ≈2 t ha^?1 in the second season. The legumes derived 60–99% of their N from the atmosphere, fixing 5–120 kg N ha^?1 under low rainfall and 78–267 kg N ha^?1 under high rainfall. These findings suggest that the legumes could contribute in restoring productivity of soils continuously cultivated with little or no nutrient inputs in most of Zimbabwe and similar agro-ecologies in SubSaharan Africa.     

Microwave fixation of marine invertebrates

Marine invertebrates may be rapidly fixed for histological examination using microwave irradiation generated by household microwave ovens. Ten-second irradiation of whole intact clams gave tissue fixation equal or superior to standard procedures using formaldehyde solutions and eliminated the need for that hazardous chemical. We suggest that invertebrates can be fixed while relaxed in sea-water baths, without having to remove or open the shell, and that invertebrates in bottom sediment cores also may be fixed in situ without being disturbed.