Simulated Nitrogen Deposition Reduces CH4 Uptake and Increases N2O Emission from a Subtropical Plantation Forest Soil in Southern China

To date, few studies are conducted to quantify the effects of reduced ammonium (NH₄ ⁺) and oxidized nitrate (NO₃ ⁻) on soil CH₄ uptake and N₂O emission in the subtropical forests. In this study, NH₄Cl and NaNO₃ fertilizers were applied at three rates: 0, 40 and 120 kg N ha⁻¹ yr⁻¹. Soil CH₄ and N₂O fluxes were determined twice a week using the static chamber technique and gas chromatography. Soil temperature and moisture were simultaneously measured. Soil dissolved N concentration in 0–20 cm depth was measured weekly to examine the regulation to soil CH₄ and N₂O fluxes. Our results showed that one year of N addition did not affect soil temperature, soil moisture, soil total dissolved N (TDN) and NH₄ ⁺-N concentrations, but high levels of applied NH₄Cl and NaNO₃ fertilizers significantly increased soil NO₃ ⁻-N concentration by 124% and 157%, respectively. Nitrogen addition tended to inhibit soil CH₄ uptake, but significantly promoted soil N₂O emission by 403% to 762%. Furthermore, NH₄ ⁺-N fertilizer application had a stronger inhibition to soil CH₄ uptake and a stronger promotion to soil N₂O emission than NO₃ ⁻-N application. Also, both soil CH₄ and N₂O fluxes were driven by soil temperature and moisture, but soil inorganic N availability was a key integrator of soil CH₄ uptake and N₂O emission. These results suggest that the subtropical plantation soil sensitively responses to atmospheric N deposition, and inorganic N rather than organic N is the regulator to soil CH₄ uptake and N₂O emission.     

The global nitrogen cycle in the twenty-first century

Global nitrogen fixation contributes 413 Tg of reactive nitrogen (N_r) to terrestrial and marine ecosystems annually of which anthropogenic activities are responsible for half, 210 Tg N. The majority of the transformations of anthropogenic N_r are on land (240 Tg N yr⁻¹) within soils and vegetation where reduced N_r contributes most of the input through the use of fertilizer nitrogen in agriculture. Leakages from the use of fertilizer N_r contribute to nitrate (NO₃⁻) in drainage waters from agricultural land and emissions of trace N_r compounds to the atmosphere. Emissions, mainly of ammonia (NH₃) from land together with combustion related emissions of nitrogen oxides (NO_x), contribute 100 Tg N yr⁻¹ to the atmosphere, which are transported between countries and processed within the atmosphere, generating secondary pollutants, including ozone and other photochemical oxidants and aerosols, especially ammonium nitrate (NH₄NO₃) and ammonium sulfate (NH₄)₂SO₄. Leaching and riverine transport of NO₃ contribute 40–70 Tg N yr⁻¹ to coastal waters and the open ocean, which together with the 30 Tg input to oceans from atmospheric deposition combine with marine biological nitrogen fixation (140 Tg N yr⁻¹) to double the ocean processing of N_r. Some of the marine N_r is buried in sediments, the remainder being denitrified back to the atmosphere as N₂ or N₂O. The marine processing is of a similar magnitude to that in terrestrial soils and vegetation, but has a larger fraction of natural origin. The lifetime of N_r in the atmosphere, with the exception of N₂O, is only a few weeks, while in terrestrial ecosystems, with the exception of peatlands (where it can be 10²–10³ years), the lifetime is a few decades. In the ocean, the lifetime of N_r is less well known but seems to be longer than in terrestrial ecosystems and may represent an important long-term source of N₂O that will respond very slowly to control measures on the sources of N_r from which it is produced.     

Growth and Nitrogen Uptake Characteristics Reveal Outbreak Mechanism of the Opportunistic Macroalga Gracilaria tenuistipitata

Macroalgae has bloomed in the brackish lake of Shenzhen Bay, China continuously from 2010 to 2014. Gracilaria tenuistipitata was identified as the causative macroalgal species. The aim of this study was to explore the outbreak mechanism of G. tenuistipitata, by studying the effects of salinity and nitrogen sources on growth, and the different nitrogen sources uptake characteristic. Our experimental design was based on environmental conditions observed in the bloom areas, and these main factors were simulated in the laboratory. Results showed that salinity 12 to 20 ‰ was suitable for G. tenuistipitata growth. When the nitrogen sources'' (NH₄ ⁺, NO₃ ⁻) concentrations reached 40 µM or above, the growth rate of G. tenuistipitata was significantly higher. Algal biomass was higher (approximately 1.4 times) when cultured with NH₄ ⁺ than that with NO₃ ⁻ addition. Coincidentally, macroalgal bloom formed during times of moderate salinity (∼12 ‰) and high nitrogen conditions. The NH₄ ⁺ and NO₃ ⁻ uptake characteristic was studied to understand the potential mechanism of G. tenuistipitata bloom. NH₄ ⁺ uptake was best described by a linear, rate-unsaturated response, with the slope decreasing with time intervals. In contrast, NO₃ ⁻ uptake followed a rate-saturating mechanism best described by the Michaelis-Menten model, with kinetic parameters V_max = 37.2 µM g⁻¹ DM h⁻¹ and K_s = 61.5 µM. Further, based on the isotope ¹⁵N tracer method, we found that ¹⁵N from NH₄ ⁺ accumulated faster and reached an atom% twice than that of ¹⁵N from NO₃ ⁻, suggesting when both NH₄ ⁺ and NO₃ ⁻ were available, NH₄ ⁺ was assimilated more rapidly. The results of the present study indicate that in the estuarine environment, the combination of moderate salinity with high ammonium may stimulate bloom formation.     

Denitrification and Assimilatory Nitrate Reduction in Aquaspirillum magnetotacticum

Aquaspirillum magnetotacticum MS-1 grew microaerobically but not anaerobically with NO₃⁻ or NH₄⁺ as the sole nitrogen source. Nevertheless, cell yields varied directly with NO₃⁻ concentration under microaerobic conditions. Products of NO₃⁻ reduction included NH₄⁺, N₂O, NO, and N₂. NO₂⁻ and NH₂OH, each toxic to cells at 0.2 mM, were not detected as products of cells growing on NO₃⁻. NO₃⁻ reduction to NH₄⁺ was completely repressed by the addition of 2 mM NH₄⁺ to the growth medium, whereas NO₃⁻ reduction to N₂O or to N₂ was not. C₂H₂ completely inhibited N₂O reduction to N₂ by growing cells. These results indicate that A. magnetotacticum is a microaerophilic denitrifier that is versatile in its nitrogen metabolism, concomitantly reducing NO₃⁻ by assimilatory and dissimilatory means. This bacterium appears to be the first described denitrifier with an absolute requirement for O₂. The process of NO₃⁻ reduction appears well adapted for avoiding accumulation of several nitrogenous intermediates that are toxic to cells.     

Atmospheric Nitrogen Deposition at Two Sites in an Arid Environment of Central Asia

Arid areas play a significant role in the global nitrogen cycle. Dry and wet deposition of inorganic nitrogen (N) species were monitored at one urban (SDS) and one suburban (TFS) site at Urumqi in a semi-arid region of central Asia. Atmospheric concentrations of NH₃, NO₂, HNO₃, particulate ammonium and nitrate (pNH₄ ⁺ and pNO₃ ⁻) concentrations and NH₄-N and NO₃-N concentrations in precipitation showed large monthly variations and averaged 7.1, 26.6, 2.4, 6.6, 2.7 µg N m⁻³ and 1.3, 1.0 mg N L⁻¹ at both SDS and TFS. Nitrogen dry deposition fluxes were 40.7 and 36.0 kg N ha⁻¹ yr⁻¹ while wet deposition of N fluxes were 6.0 and 8.8 kg N ha⁻¹ yr⁻¹ at SDS and TFS, respectively. Total N deposition averaged 45.8 kg N ha⁻¹ yr⁻¹at both sites. Our results indicate that N dry deposition has been a major part of total N deposition (83.8% on average) in an arid region of central Asia. Such high N deposition implies heavy environmental pollution and an important nutrient resource in arid regions.     

Effects of pH,phosphate and ammonia on the rate of uptake of nitrate and ammonia by freshwater phytoplankton

The effect of phosphate (PO₄ ⁺³) and pH in regulating nitrate (NO₃) and ammonia (NH₃ ⁺) uptake by phytoplankton was investigated in two Oklahoma lakes using ¹⁵N tracers. Addition of PO₄ ⁺³ above ambient concentrations had a negligible effect on the rate of uptake of NO₃ ^– or NH₃ ⁺. Manipulation of pH of lake water had little effect on uptake of either NO₃ ^– or NH₃ ⁺. A correlation analysis suggested that NO₃ ^– is not used by phytoplankton when NH₃ ⁺ concentrations exceed about 210 µg NH₃ ⁺-N(1)^–1.     

Light-Limited Growth Rate Modulates Nitrate Inhibition of Dinitrogen Fixation in the Marine Unicellular Cyanobacterium Crocosphaera watsonii

Biological N₂ fixation is the dominant supply of new nitrogen (N) to the oceans, but is often inhibited in the presence of fixed N sources such as nitrate (NO₃ ⁻). Anthropogenic fixed N inputs to the ocean are increasing, but their effect on marine N₂ fixation is uncertain. Thus, global estimates of new oceanic N depend on a fundamental understanding of factors that modulate N source preferences by N₂-fixing cyanobacteria. We examined the unicellular diazotroph Crocosphaera watsonii (strain WH0003) to determine how the light-limited growth rate influences the inhibitory effects of fixed N on N₂ fixation. When growth (µ) was limited by low light (µ = 0.23 d⁻¹), short-term experiments indicated that 0.4 µM NH₄ ⁺ reduced N₂-fixation by ∼90% relative to controls without added NH₄ ⁺. In fast-growing, high-light-acclimated cultures (µ = 0.68 d⁻¹), 2.0 µM NH₄ ⁺ was needed to achieve the same effect. In long-term exposures to NO₃ ⁻, inhibition of N₂ fixation also varied with growth rate. In high-light-acclimated, fast-growing cultures, NO₃ ⁻ did not inhibit N₂-fixation rates in comparison with cultures growing on N₂ alone. Instead NO₃ ⁻ supported even faster growth, indicating that the cellular assimilation rate of N₂ alone (i.e. dinitrogen reduction) could not support the light-specific maximum growth rate of Crocosphaera. When growth was severely light-limited, NO₃ ⁻ did not support faster growth rates but instead inhibited N₂-fixation rates by 55% relative to controls. These data rest on the basic tenet that light energy is the driver of photoautotrophic growth while various nutrient substrates serve as supports. Our findings provide a novel conceptual framework to examine interactions between N source preferences and predict degrees of inhibition of N₂ fixation by fixed N sources based on the growth rate as controlled by light.     

Effects of Feedstock and Pyrolysis Temperature on Biochar Adsorption of Ammonium and Nitrate

Biochar produced by pyrolysis of biomass can be used to counter nitrogen (N) pollution. The present study investigated the effects of feedstock and temperature on characteristics of biochars and their adsorption ability for ammonium N (NH₄ ⁺-N) and nitrate N (NO₃ ⁻-N). Twelve biochars were produced from wheat-straw (W-BC), corn-straw (C-BC) and peanut-shell (P-BC) at pyrolysis temperatures of 400, 500, 600 and 700°C. Biochar physical and chemical properties were determined and the biochars were used for N sorption experiments. The results showed that biochar yield and contents of N, hydrogen and oxygen decreased as pyrolysis temperature increased from 400°C to 700°C, whereas contents of ash, pH and carbon increased with greater pyrolysis temperature. All biochars could sorb substantial amounts of NH₄ ⁺-N, and the sorption characteristics were well fitted to the Freundlich isotherm model. The ability of biochars to adsorb NH₄ ⁺-N followed: C-BC>P-BC>W-BC, and the adsorption amount decreased with higher pyrolysis temperature. The ability of C-BC to sorb NH₄ ⁺-N was the highest because it had the largest cation exchange capacity (CEC) among all biochars (e.g., C-BC400 with a CEC of 38.3 cmol kg⁻¹ adsorbed 2.3 mg NH₄ ⁺-N g⁻¹ in solutions with 50 mg NH₄ ⁺ L⁻¹). Compared with NH₄ ⁺-N, none of NO₃ ⁻-N was adsorbed to biochars at different NO₃ ⁻ concentrations. Instead, some NO₃ ⁻-N was even released from the biochar materials. We conclude that biochars can be used under conditions where NH₄ ⁺-N (or NH₃) pollution is a concern, but further research is needed in terms of applying biochars to reduce NO₃ ⁻-N pollution.     

Ammonium Removal by the Oxygen-Limited Autotrophic Nitrification-Denitrification System

The present lab-scale research reveals the potential of implementation of an oxygen-limited autotrophic nitrification-denitrification (OLAND) system with normal nitrifying sludge as the biocatalyst for the removal of nitrogen from nitrogen-rich wastewater in one step. In a sequential batch reactor, synthetic wastewater containing 1 g of NH₄⁺-N liter⁻¹ and minerals was treated. Oxygen supply to the reactor was double-controlled with a pH controller and a timer. At a volumetric loading rate (B_v) of 0.13 g of NH₄⁺-N liter⁻¹ day⁻¹, about 22% of the fed NH₄⁺-N was converted to NO₂⁻-N or NO₃⁻-N, 38% remained as NH₄⁺-N, and the other 40% was removed mainly as N₂. The specific removal rate of nitrogen was on the order of 50 mg of N liter⁻¹ day⁻¹, corresponding to 16 mg of N g of volatile suspended solids⁻¹ day⁻¹. The microorganisms which catalyzed the OLAND process are assumed to be normal nitrifiers dominated by ammonium oxidizers. The loss of nitrogen in the OLAND system is presumed to occur via the oxidation of NH₄⁺ to N₂ with NO₂⁻ as the electron acceptor. Hydroxylamine stimulated the removal of NH₄⁺ and NO₂⁻. Hydroxylamine oxidoreductase (HAO) or an HAO-related enzyme might be responsible for the loss of nitrogen.     

Short Term Studies of Nitrate Uptake into Barley Plants Using Ion-Specific Electrodes and ClO(3): II. Regulation of NO(3) Efflux by NH(4)

The influence of NH₄⁺, in the external medium, on fluxes of NO₃⁻ and K⁺ were investigated using barley (Hordeum vulgare cv Betzes) plants. NH₄⁺ was without effect on NO₃⁻ (³⁶ClO₃⁻) influx whereas inhibition of net uptake appeared to be a function of previous NO₃⁻ provision. Plants grown at 10 micromolar NO₃⁻ were sensitive to external NH₄⁺ when uptake was measured in 100 micromolar NO₃⁻. By contrast, NO₃⁻ uptake (from 100 micromolar NO₃⁻) by plants previously grown at this concentration was not reduced by NH₄⁺ treatment. Plants pretreated for 2 days with 5 millimolar NO₃⁻ showed net efflux of NO₃⁻ when roots were transferred to 100 micromolar NO₃⁻. This efflux was stimulated in the presence of NH₄⁺. NH₄⁺ also stimulated NO₃⁻ efflux from plants pretreated with relatively low nitrate concentrations. It is proposed that short term effects on net uptake of NO₃⁻ occur via effects upon efflux. By contrast to the situation for NO₃⁻, net K⁺ uptake and influx of ³⁶Rb⁺-labeled K⁺ was inhibited by NH₄⁺ regardless of the nutrient history of the plants. Inhibition of net K⁺ uptake reached its maximum value within 2 minutes of NH₄⁺ addition. It is concluded that the latter ion exerts a direct effect upon K⁺ influx.     

Dissimilatory Reduction of NO2− to NH4+ and N2O by a Soil Citrobacter sp.

Dissimilatory reduction of NO₂⁻ to N₂O and NH₄⁺ by a soil Citrobacter sp. was studied in an attempt to elucidate the physiological and ecological significance of N₂O production by this mechanism. In batch cultures with defined media, NO₂⁻ reduction to NH₄⁺ was favored by high glucose and low NO₃⁻ concentrations. Nitrous oxide production was greatest at high glucose and intermediate NO₃⁻ concentrations. With succinate as the energy source, little or no NO₂⁻ was reduced to NH₄⁺ but N₂O was produced. Resting cell suspensions reduced NO₂⁻ simultaneously to N₂O and free extracellular NH₄⁺. Chloramphenicol prevented the induction of N₂O-producing activity. The K_m for NO₂⁻ reduction to N₂O was estimated to be 0.9 mM NO₂⁻, yet the apparent K_m for overall NO₂⁻ reduction was considerably lower, no greater than 0.04 mM NO₂⁻. Activities for N₂O and NH₄⁺ production increased markedly after depletion of NO₃⁻ from the media. Amendment with NO₃⁻ inhibited N₂O and NH₄⁺ production by molybdate-grown cells but not by tungstate-grown cells. Sulfite inhibited production of NH₄⁺ but not of N₂O. In a related experiment, three Escherichia coli mutants lacking NADH-dependent nitrite reductase produced N₂O at rates equal to the wild type. These observations suggest that N₂O is produced enzymatically but not by the same enzyme system responsible for dissimilatory reduction of NO₂⁻ to NH₄⁺.     

Growth and Nitrogen Uptake Kinetics in Cultured Prorocentrum donghaiense

We compared growth kinetics of Prorocentrum donghaiense cultures on different nitrogen (N) compounds including nitrate (NO₃ ⁻), ammonium (NH₄ ⁺), urea, glutamic acid (glu), dialanine (diala) and cyanate. P. donghaiense exhibited standard Monod-type growth kinetics over a range of N concentraions (0.5–500 μmol N L⁻¹ for NO₃ ⁻ and NH₄ ⁺, 0.5–50 μmol N L⁻¹ for urea, 0.5–100 μmol N L⁻¹ for glu and cyanate, and 0.5–200 μmol N L⁻¹ for diala) for all of the N compounds tested. Cultures grown on glu and urea had the highest maximum growth rates (μ_m, 1.51±0.06 d⁻¹ and 1.50±0.05 d⁻¹, respectively). However, cultures grown on cyanate, NO₃ ⁻, and NH₄ ⁺ had lower half saturation constants (K_μ, 0.28–0.51 μmol N L⁻¹). N uptake kinetics were measured in NO₃ ⁻-deplete and -replete batch cultures of P. donghaiense. In NO₃ ⁻-deplete batch cultures, P. donghaiense exhibited Michaelis-Menten type uptake kinetics for NO₃ ⁻, NH₄ ⁺, urea and algal amino acids; uptake was saturated at or below 50 μmol N L⁻¹. In NO₃ ⁻-replete batch cultures, NH₄ ⁺, urea, and algal amino acid uptake kinetics were similar to those measured in NO₃ ⁻-deplete batch cultures. Together, our results demonstrate that P. donghaiense can grow well on a variety of N sources, and exhibits similar uptake kinetics under both nutrient replete and deplete conditions. This may be an important factor facilitating their growth during bloom initiation and development in N-enriched estuaries where many algae compete for bioavailable N and the nutrient environment changes as a result of algal growth.     

Ammonium as a Driving Force of Plant Diversity and Ecosystem Functioning: Observations Based on 5 Years' Manipulation of N Dose and Form in a Mediterranean Ecosystem

Enhanced nitrogen (N) availability is one of the main drivers of biodiversity loss and degradation of ecosystem functions. However, in very nutrient-poor ecosystems, enhanced N input can, in the short-term, promote diversity. Mediterranean Basin ecosystems are nutrient-limited biodiversity hotspots, but no information is available on their medium- or long-term responses to enhanced N input. Since 2007, we have been manipulating the form and dose of available N in a Mediterranean Basin maquis in south-western Europe that has low ambient N deposition (<4 kg N ha⁻¹ yr⁻¹) and low soil N content (0.1%). N availability was modified by the addition of 40 kg N ha⁻¹ yr⁻¹ as a 1∶1 NH₄Cl to (NH₄)₂SO₄ mixture, and 40 and 80 kg N ha⁻¹ yr⁻¹ as NH₄NO₃. Over the following 5 years, the impacts on plant composition and diversity (richness and evenness) and some ecosystem characteristics (soil extractable N and organic matter, aboveground biomass and % of bare soil) were assessed. Plant species richness increased with enhanced N input and was more related to ammonium than to nitrate. Exposure to 40 kg NH₄ ⁺-N ha⁻¹ yr⁻¹ (alone and with nitrate) enhanced plant richness, but did not increase aboveground biomass; soil extractable N even increased under 80 kg NH₄NO₃-N ha⁻¹ yr⁻¹ and the % of bare soil increased under 40 kg NH₄ ⁺-N ha⁻¹ yr⁻¹. The treatment containing less ammonium, 40 kg NH₄NO₃-N ha⁻¹ yr⁻¹, did not enhance plant diversity but promoted aboveground biomass and reduced the % of bare soil. Data suggest that enhanced NH_y availability affects the structure of the maquis, which may promote soil erosion and N leakage, whereas enhanced NO_x availability leads to biomass accumulation which may increase the fire risk. These observations are relevant for land use management in biodiverse and fragmented ecosystems such as the maquis, especially in conservation areas.     

Measurement of net fluxes of ammonium and nitrate at the surface of barley roots using ion-selective microelectrodes

Neutral carrier-based liquid membrane ion-selective microelectrodes for NH₄⁺ and NO₃⁻ were developed and used to investigate inorganic nitrogen acquisition in two varieties of barley, Hordeum vulgare L. cv Olli and H. vulgare L. cv Prato, originating in cold and warm climates, respectively. In the present paper, the methods used in the fabrication of ammonium- and nitrate-selective microelectrodes are described, and their application in the study of inorganic nitrogen uptake is demonstrated. Net ionic fluxes of NH₄⁺ and NO₃⁻ were measured in the unstirred layer of solution immediately external to the root surface. The preference for the uptake of a particular ionic form was examined by measuring the net flux of the predominant form of inorganic nitrogen, with and without the alternative ion in solution. Net flux of NH₄⁺ into the cold-adapted variety remained unchanged when equimolar concentrations (200 micromolar) of NH₄⁺ and NO₃⁻ were present. Similarly, net flux of NO₃⁻ into the warm-adapted variety was not affected when NH₄⁺ was also present in solution. The high temporal and spatial resolution afforded by ammonium- and nitrate-selective microelectrodes permits a detailed examination of inorganic nitrogen acquisition and its component ionic interactions.     

Simultaneous Measurement of Denitrification and Nitrogen Fixation Using Isotope Pairing with Membrane Inlet Mass Spectrometry Analysis

A method for estimating denitrification and nitrogen fixation simultaneously in coastal sediments was developed. An isotope-pairing technique was applied to dissolved gas measurements with a membrane inlet mass spectrometer (MIMS). The relative fluxes of three N₂ gas species (²⁸N₂, ²⁹N₂, and ³⁰N₂) were monitored during incubation experiments after the addition of ¹⁵NO₃⁻. Formulas were developed to estimate the production (denitrification) and consumption (N₂ fixation) of N₂ gas from the fluxes of the different isotopic forms of N₂. Proportions of the three isotopic forms produced from ¹⁵NO₃⁻ and ¹⁴NO₃⁻ agreed with expectations in a sediment slurry incubation experiment designed to optimize conditions for denitrification. Nitrogen fixation rates from an algal mat measured with intact sediment cores ranged from 32 to 390 μg-atoms of N m⁻² h⁻¹. They were enhanced by light and organic matter enrichment. In this environment of high nitrogen fixation, low N₂ production rates due to denitrification could be separated from high N₂ consumption rates due to nitrogen fixation. Denitrification and nitrogen fixation rates were estimated in April 2000 on sediments from a Texas sea grass bed (Laguna Madre). Denitrification rates (average, 20 μg-atoms of N m⁻² h⁻¹) were lower than nitrogen fixation rates (average, 60 μg-atoms of N m⁻² h⁻¹). The developed method benefits from simple and accurate dissolved-gas measurement by the MIMS system. By adding the N₂ isotope capability, it was possible to do isotope-pairing experiments with the MIMS system.     

Sub-Parts-Per-Billion Nitrate Method: Use of an N2O-Producing Denitrifier to Convert NO3− or 15NO3− to N2O

A more sensitive analytical method for NO₃⁻ was developed based on the conversion of NO₃⁻ to N₂O by a denitrifier that could not reduce N₂O further. The improved detectability resulted from the high sensitivity of the ⁶³Ni electron capture gas chromatographic detector for N₂O and the purification of the nitrogen afforded by the transformation of the N to a gaseous product with a low atmospheric background. The selected denitrifier quantitatively converted NO₃⁻ to N₂O within 10 min. The optimum measurement range was from 0.5 to 50 ppb (50 μg/liter) of NO₃⁻ N, and the detection limit was 0.2 ppb of N. The values measured by the denitrifier method compared well with those measured by the high-pressure liquid chromatographic UV method above 2 ppb of N, which is the detection limit of the latter method. It should be possible to analyze all types of samples for nitrate, except those with inhibiting substances, by this method. To illustrate the use of the denitrifier method, NO₃⁻ concentrations of <2 ppb of NO₃⁻ N were measured in distilled and deionized purified water samples and in anaerobic lake water samples, but were not detected at the surface of the sediment. The denitrifier method was also used to measure the atom% of ¹⁵N in NO₃⁻. This method avoids the incomplete reduction and contamination of the NO₃⁻ -N by the NH₄⁺ and N₂ pools which can occur by the conventional method of ¹⁵NO₃⁻ analysis. N₂O-producing denitrifier strains were also used to measure the apparent K_m values for NO₃⁻ use by these organisms. Analysis of N₂O production by use of a progress curve yielded K_m values of 1.7 and 1.8 μM NO₃⁻ for the two denitrifier strains studied.     

In Vivo Blue-Light Activation of Chlamydomonas reinhardii Nitrate Reductase

Chlamydomonas reinhardii cells, growing photoautotrophically under air, excreted to the culture medium much higher amounts of NO₂⁻ and NH₄⁺ under blue than under red light. Under similar conditions, but with NO₂⁻ as the only nitrogen source, the cells consumed NO₂⁻ and excreted NH₄⁺ at similar rates under blue and red light. In the presence of NO₃⁻ and air with 2% CO₂ (v/v), no excretion of NO₂⁻ and NH₄⁺ occurred and, moreover, if the bubbling air of the cells that were currently excreting NO₂⁻ and NH₄⁺ was enriched with 2% CO₂ (v/v), the previously excreted reduced nitrogen ions were rapidly reassimilated. The levels of total nitrate reductase and active nitrate reductase increased several times in the blue-light-irradiated cells growing on NO₃⁻ under air. When tungstate replaced molybdate in the medium (conditions that do not allow the formation of functional nitrate reductase), blue light activated most of the preformed inactive enzyme of the cells. Furthermore, nitrate reductase extracted from the cells in its inactive form was readily activated in vitro by blue light. It appears that under high irradiance (90 w m⁻²) and low CO₂ tensions, cells growing on NO₃⁻ or NO₂⁻ may not have sufficient carbon skeletons to incorporate all the photogenerated NH₄⁺. Because these cells should have high levels of reducing power, they might use NO₃⁻ or, in its absence, NO₂⁻ as terminal electron acceptors. The excretion of the products of NO₂⁻ and NH₄⁺ to the medium may provide a mechanism to control reductant level in the cells. Blue light is suggested as an important regulatory factor of this photorespiratory consumption of NO₃⁻ and possibly of the whole nitrogen metabolism in green algae.     

Anaerobic Ammonium Oxidation Measured in Sediments along the Thames Estuary, United Kingdom

Until recently, denitrification was thought to be the only significant pathway for N₂ formation and, in turn, the removal of nitrogen in aquatic sediments. The discovery of anaerobic ammonium oxidation in the laboratory suggested that alternative metabolisms might be present in the environment. By using a combination of ¹⁵N-labeled NH₄⁺, NO₃⁻, and NO₂⁻ (and ¹⁴N analogues), production of ²⁹N₂ and ³⁰N₂ was measured in anaerobic sediment slurries from six sites along the Thames estuary. The production of ²⁹N₂ in the presence of ¹⁵NH₄⁺ and either ¹⁴NO₃⁻ or ¹⁴NO₂⁻ confirmed the presence of anaerobic ammonium oxidation, with the stoichiometry of the reaction indicating that the oxidation was coupled to the reduction of NO₂⁻. Anaerobic ammonium oxidation proceeded at equal rates via either the direct reduction of NO₂⁻ or indirect reduction, following the initial reduction of NO₃⁻. Whether NO₂⁻ was directly present at 800 μM or it accumulated at 3 to 20 μM (from the reduction of NO₃⁻), the rate of ²⁹N₂ formation was not affected, which suggested that anaerobic ammonium oxidation was saturated at low concentrations of NO₂⁻. We observed a shift in the significance of anaerobic ammonium oxidation to N₂ formation relative to denitrification, from 8% near the head of the estuary to less than 1% at the coast. The relative importance of anaerobic ammonium oxidation was positively correlated (P < 0.05) with sediment organic content. This report of anaerobic ammonium oxidation in organically enriched estuarine sediments, though in contrast to a recent report on continental shelf sediments, confirms the presence of this novel metabolism in another aquatic sediment system.     

Nitrate-ammonium synergism in rice. A subcellular flux analysis

Many reports have shown that plant growth and yield is superior on mixtures of NO₃⁻ and NH₄⁺ compared with provision of either N source alone. Despite its clear practical importance, the nature of this N-source synergism at the cellular level is poorly understood. In the present study we have used the technique of compartmental analysis by efflux and the radiotracer ¹³N to measure cellular turnover kinetics, patterns of flux partitioning, and cytosolic pool sizes of both NO₃⁻ and NH₄⁺ in seedling roots of rice (Oryza sativa L. cv IR72), supplied simultaneously with the two N sources. We show that plasma membrane fluxes for NH₄⁺, cytosolic NH₄⁺ accumulation, and NH₄⁺ metabolism are enhanced by the presence of NO₃⁻, whereas NO₃⁻ fluxes, accumulation, and metabolism are strongly repressed by NH₄⁺. However, net N acquisition and N translocation to the shoot with dual N-source provision are substantially larger than when NO₃⁻ or NH₄⁺ is provided alone at identical N concentrations.     

The influence of ammonium and chloride on potassium and nitrate absorption by barley roots depends on time of exposure and cultivar

Net uptakes of K⁺ and NO₃⁻ were monitored simultaneously and continuously for two barley (Hordeum vulgare) cultivars, Prato and Olli. The cultivars had similar rates of net K⁺ and NO₃⁻ uptake in the absence of NH₄⁺ or Cl⁻. Long-term exposure (over 6 hours) to media which contained equimolar mixtures of NH₄⁺, K⁺, Cl⁻, or NO₃⁻ affected the cultivars very differently: (a) the presence of NH₄⁺ as NH₄Cl stimulated net NO₃⁻ uptake in Prato barley but inhibited net NO₃⁻ uptake in Olli barley; (b) Cl⁻ inhibited net NO₃⁻ uptake in Prato but had little effect in Olli; and (c) NH₄⁺ as (NH₄)₂SO₄ inhibited net K⁺ uptake in Prato but had little effect in Olli. Moreover, the immediate response to the addition of an ion often varied significantly from the long-term response; for example, the addition of Cl⁻ initially inhibited net K⁺ uptake in Olli barley but, after a 4 hour exposure, it was stimulatory. For both cultivars, net NH₄⁺ and Cl⁻ uptake did not change significantly with time after these ions were added to the nutrient medium. These data indicate that, even within one species, there is a high degree of genotypic variation in the control of nutrient absorption.