Measurement Of Denitrification Research Articles

Denitrification Losses in Response to N Fertilizer Rates—Integrating High Temporal Resolution N2O, In Situ 15N2O and 15N2 Measurements and Fertilizer 15N Recoveries in Intensive Sugarcane Systems

AbstractDenitrification is a key process in the global nitrogen (N) cycle, causing both nitrous oxide (N2O) and dinitrogen (N2) emissions. However, estimates of seasonal denitrification losses (N2O + N2) are scarce, reflecting methodological difficulties in measuring soil‐borne N2 emissions against the high atmospheric N2 background and challenges regarding their spatio‐temporal upscaling. This study investigated N2O + N2 losses in response to N fertilizer rates (0, 100, 150, 200, and 250 kg N ha−1) on two intensively managed tropical sugarcane farms in Australia, by combining automated N2O monitoring, in situ N2 and N2O measurements using the 15N gas flux method and fertilizer 15N recoveries at harvest. Dynamic changes in the N2O/(N2O + N2) ratio (<0.01 to 0.768) were explained by fitting generalized additive mixed models (GAMMs) with soil factors to upscale high temporal‐resolution N2O data to daily N2 emissions over the season. Cumulative N2O + N2 losses ranged from 12 to 87 kg N ha−1, increasing non‐linearly with increasing N fertilizer rates. Emissions of N2O + N2 accounted for 31%–78% of fertilizer 15N losses and were dominated by environmentally benign N2 emissions. The contribution of denitrification to N fertilizer loss decreased with increasing N rates, suggesting increasing significance of other N loss pathways including leaching and runoff at higher N rates. This study delivers a blueprint approach to extrapolate denitrification measurements at both temporal and spatial scales, which can be applied in fertilized agroecosystems. Robust estimates of denitrification losses determined using this method will help to improve cropping system modeling approaches, advancing our understanding of the N cycle across scales.

Variation characteristics and influential factors of denitrification potential of typical ponds in the hilly region of the upper Lake Taihu Basin

水塘是连接上游集水坡地和下游水体的重要水文通道，也是氮素发生生物地球反应的重要场所.作为氮素高效去除的首要机制，水塘的反硝化脱氮潜力及其影响因素亟待揭示.本研究选择太湖上游丘陵区天目湖流域4类（茶园塘、村塘、养殖塘、林塘）共14个典型水塘为研究区，分别监测了其夏、秋季水质和沉积物变化特征，并利用膜进样质谱法（MIMS）直接测定水塘水体中溶存的反硝化产物N₂浓度.结果发现，14个水塘上覆水中N₂过饱和浓度在1.36~28.35 μmol/L之间，平均值为（8.23±6.04）μmol/L，夏季和秋季N₂过饱和浓度分别为（8.81±4.08）和（7.64±7.46）μmol/L，夏季高于秋季，不同类型水塘N₂过饱和浓度大小顺序为：茶园塘>村塘>养殖塘>林塘；结合水气交换通量模型估算了反硝化潜力，14个水塘的反硝化速率均值为（4.75±3.27） mmol/（m²·d），反硝化速率大小顺序为：茶园塘>村塘>养殖塘>林塘.相关分析结果表明，反硝化作用与水体中硝态氮浓度呈显著正相关，与溶解氧浓度呈显著负相关；反硝化作用同样与沉积物中硝态氮含量呈显著正相关，与容重呈负相关，说明水塘反硝化作用同时受上覆水中硝态氮、溶解氧以及沉积物容重等理化性质共同调节.进一步通过与传统的反硝化测算方法对比验证，发现采用膜进样质谱法并结合水气交换通量模型来估算水体反硝化潜力具有可行性，同时操作简便、所需样品少、测定速度快，适用于淹水环境反硝化作用的测定.;The ponds are important hydrological channels connecting the upstream hillslope and the downstream water bodies, and important hotspots for nitrogen processing. As the primary mechanism of efficient nitrogen removal, the potential of denitrification and nitrogen removal in ponds and its influential factors were rarely been documented. In this study, fourteen typical ponds of four types (tea garden pond, village pond, fish pond, and forested pond) in Lake Tianmu Basin in the hilly region of the upper reaches of Lake Taihu were selected as our study sites. The characteristics of water quality and sediment change in summer and autumn were monitored respectively, and the levels of dissolved denitrifying product-N₂ in the ponds were directly determined by membrane injection mass spectrometry (MIMS). The results showed that N₂ supersaturated concentration in the overlying water of 14 ponds ranged from 1.36 μmol/L to 28.35 μmol/L, with an average of (8.23±6.04) μmol/L. N₂ supersaturated concentration was (8.81±4.08) μmol/L in summer and (7.64±7.46) μmol/L in autumn, and the concentration of N₂ supersaturated in different ponds in summer is higher than that in autumn. The ΔN₂ was the highest in tea garden ponds, followed by village ponds and fish ponds, and the lowest in forested ponds. The mean value of denitrification rates of 14 ponds was (4.75±3.27) mmol/(m²·d), which was highest in tea garden ponds, followed by village ponds and fish ponds, and lowest in forested ponds. Correlation analysis showed that denitrification was significantly positively correlated with nitrate concentration, and significantly negatively correlated with dissolved oxygen. Denitrification was also significantly positively correlated with nitrate concentration in sediment, and negatively correlated with bulk density, indicating that denitrification in ponds was simultaneously regulated by substrate concentration, dissolved oxygen concentration, and sediment physical and chemical properties. Compared and validated with the traditional denitrification measurement method, the membrane injection mass spectrometry combined with the water-gas exchange flux model proves feasible to estimate the denitrification potential of water. It is convenient to operate, requires fewer samples, and has a fast determination speed, which is suitable for the determination of denitrification in a flooded environment.

Nitrous oxide fluxes from long-term limed soils following P and glucose addition: Nonlinear response to liming rates and interaction from added P

Liming of acidic soils to regulate pH for crop growth may decrease emissions of nitrous oxide (N2O) due to direct effects of pH on the synthesis of N2O reductases by denitrifying bacteria. However, liming also changes general pH-dependent soil properties, including availability of phosphorus (P), with a feedback on N2O fluxes that remains largely unknown. Here we used a mesocosm approach to study the combined role of liming and P in regulating N2O fluxes from denitrification in an arable coarse sandy soil where N2O emissions under field condition coincided with rainfall events and irrigation, which facilitated anoxia. Soils from three long-term liming treatments (0, 4, and 12 Mg ha-1) with resulting pH(CaCl2) of 3.6, 4.7 and 6.3 were incubated at original bulk density first at 60% water filled pore space (WFPS) and successively at 75% WFPS with added nitrate, inorganic P (0 and 10 μg P g-1 soil) and glucose as labile carbon. N2O fluxes were measured during 28 days and were supplemented with measurements of CO2 fluxes, microbial biomass, potential denitrification, and acid phosphatase activity. The results showed a nonlinear response of N2O fluxes to liming rates, with highest fluxes at the intermediate liming level (4 Mg ha-1). Furthermore, inorganic P stimulated N2O fluxes only at the intermediate liming level. Assays of potential denitrification indicated that the N2O/(N2O + N2) product ratio decreased consistently with increasing liming rates, but total N2O fluxes responded nonlinearly likely due to combined effects on N2O/(N2O + N2) product ratios and total denitrification rates. The results suggest that liming and P addition interact on microbial properties and N2O emissions from acidic arable soils and may not follow linear trends. This makes it uncertain to predict and model the resulting net effect, which may depend on the actual pH range and P availability from the unlimed to the limed treatments.

Denitrification within the sediments and epiphyton of tropical macrophyte stands

ABSTRACT Excess nitrogen (N) is one of the most widespread and serious pollutants in the environment, but wetlands can reduce N loads, ameliorating its damaging effects downstream. Tropical wetlands are highly productive and experience high temperatures year-round, resulting in potentially high denitrification rates. However, few measurements of denitrification have been reported for tropical wetlands. In this study, we measured denitrification within stands of macrophytes at the edge of a tropical lake (900 m length, 4 m deep) in Australia. We compared denitrification rates among sediments under emergent grass (giant bulrush, Actinoscirpus grossus), sediments under floating waterlilies (Nymphae spp.), and sediments from the deeper sections of the lagoon with no macrophytes. We also measured the denitrification and primary productivity of the epiphyton on macrophytes and compared the rates with those from the sediment. Denitrification in the sediment was higher (Dt = 3.3–52 mg m−2 h−1) than denitrification of the epiphyton (Dt = 1.9–3.6 mg m−2 h−1) and was mostly coupled with nitrification (Dn). Denitrification was highest in sediments rich in organic carbon (32.3%) and N (1.4%), and during times of the year when nitrates (NOx −-N = NO2 −-N + NO3 −-N) concentrations were relatively high (>0.10 mg L−1). Denitrification was lowest in sediments with no macrophytes, which comprised most of the lake area. Denitrification rates of sediments under these macrophyte stands were among the highest values measured for natural wetlands and highlight the potential role of this process in ameliorating N pollution in tropical catchments.

Foraminiferal Ecology and Role in Nitrogen Benthic Cycle in the Hypoxic Southeastern Bering Sea

Southeastern Bering Sea is one of the highest surface productivity area in the open ocean due to strong upwelling along the Bering canyon. However, the benthic geochemistry and organisms living in the area have been largely overlooked. In August 2017, surface sediment was sampled from four stations along a transect at depths between 1536 and 103 meters in the Bering canyon with JAMSTEC R/V Mirai. Bottom-water hypoxia was recorded in the two deepest stations (1536 and 536 m). At these stations, the oxygen penetrated down to 5 mm in the sediment due to siltier and much organic-rich sediments in the deeper stations while oxygen penetration was about 20 mm at stations 103 and 197 meters deep with coarse-grained sediment stations. Foraminiferal number of species and abundances were higher in the Unimak pass depression station E2 (197 m). Abundance did not change significantly between stations, suggesting that foraminiferal densities are not affected by the hypoxic conditions but are rather controlled by organic matter and nutrients availability. At the upper bathyal and middle bathyal stations, living foraminiferal communities were in general dominated by Uvigerina peregrina, Nonionella pulchella, Elphidium batialis, Globobulimina pacifica, Reophax spp., and Bolivina spathulata while the shallower stations exhibited large densities of Uvigerina peregrina, Cibicidoides wuellerstorfi, Recurvoidella bradyi, Globocassidulina subglobosa, and Portatrochammina pacifica. More than 50% of the individuals have a potential to accumulate nitrate in their cell (from 3 to 648 mmol/L; which is from 100 to 4000 times larger than the highest concentration measured in pore water). Onboard denitrification measurements confirmed that B. spathulata, N. pulchella and G. pacifica could reduce nitrate through denitrification and foraminiferal denitrification could contribute over 6% to benthic nitrate reduction at the southeast Bering Sea. Although the foraminiferal contributions were smaller than those measured at other hypoxic areas, our study quantitatively revealed the significance of eukaryotic microbes on benthic nitrogen cycles at this area.

Simultaneous Measurements of Dinitrogen Fixation and Denitrification Associated With Coral Reef Substrates: Advantages and Limitations of a Combined Acetylene Assay

Nitrogen (N) cycling in coral reefs is of key importance for these oligotrophic ecosystems, but knowledge about its pathways is limited. While dinitrogen (N¬2) fixation is comparably well studied, the counteracting denitrification pathway is under-investigated, mainly because of expensive and relatively complex experimental techniques currently available. Here, we combined two established acetylene-based assays to one single setup to determine N2-fixation and denitrification performed by microbes associated with coral reef substrates/organisms simultaneously. Accumulating target gases (ethylene for N2-fixation, nitrous oxide for denitrification) were measured in gaseous headspace samples via gas chromatography. We measured N2-fixation and denitrification rates of two Red Sea coral reef substrates (filamentous turf algae, coral rubble), and demonstrated, for the first time, the co-occurrence of both N-cycling processes in both substrates. N2-fixation rates were up to eight times higher during light compared to dark, whereas denitrification rates during dark incubations were stimulated for turf algae and suppressed for coral rubble compared to light incubations. Our results highlight the importance of both substrates in fixing N, but their role in relieving N is potentially divergent. Absolute N2-fixation rates of the present study correspond with rates reported previously, even though likely underestimated due to an initial lag phase. Denitrification is also presumably underestimated due to incomplete nitrous oxide inhibition and/or substrate limitation. Besides these inherent limitations, we show that a relative comparison of N2-fixation and denitrification activity between functional groups is possible. Thus, our approach facilitates cost-efficient sample processing in studies interested in comparing relative rates of N2-fixation and denitrification.

The utility of a boundary line approach for simulating denitrification and nitrous oxide emissions from a Regosol under summer maize-winter wheat crop rotation in Southwest China

Despite the ever-increasing measurements of denitrification and nitrous oxide (N2O) emissions globally, boundary line analyses of these important processes have only been applied to a limited number of agroecosystems. The primary aims of this study were to determine the responses of denitrification and N2O emissions from a subtropical cropland to several soil parameters through boundary line analyses, and to evaluate the ability of site-specific boundary line models for simulating gaseous nitrogen (N) losses using available soil parameters. We performed 403 measurements of denitrification and N2O emissions from two fields cropped with a winter wheat-summer maize rotation in the Sichuan Basin, Southwest China, from October 2004 to September 2005. Boundary lines were fitted for denitrification and N2O emission data against soil temperature, water content, ammonium and/or nitrate substrates, and heterotrophic respiration rates. The boundary lines were largely in agreement with existing theoretical concepts. By incorporating the regulatory effects of soil temperature, water content, N substrates, and an unknown factor that could be related to ammonium content, the boundary line models reasonably simulated the seasonal variations in denitrification and N2O emissions that were collected from a field trial in the same region during the maize season in 2012. The models overestimated the denitrification and N2O emissions on the sampling dates by 8% and 22% on average, respectively. We conclude that boundary line models are useful tools for estimating gaseous N losses from a given cropland. However, the application of the models is limited by the availability of input data.

Underestimation of denitrification rates from field application of the <sup>15</sup>N gas flux method and its correction by gas diffusion modelling

Abstract. Common methods for measuring soil denitrification in situ include monitoring the accumulation of 15N-labelled N2 and N2O evolved from 15N-labelled soil nitrate pool in closed chambers that are placed on the soil surface. Gas diffusion is considered to be the main transport process in the soil. Because accumulation of gases within the chamber decreases concentration gradients between soil and the chamber over time, the surface efflux of gases decreases as well, and gas production rates are underestimated if calculated from chamber concentrations without consideration of this mechanism. Moreover, concentration gradients to the non-labelled subsoil exist, inevitably causing downward diffusion of 15N-labelled denitrification products. A numerical 3-D model for simulating gas diffusion in soil was used in order to determine the significance of this source of error. Results show that subsoil diffusion of 15N-labelled N2 and N2O – and thus potential underestimation of denitrification derived from chamber fluxes – increases with chamber deployment time as well as with increasing soil gas diffusivity. Simulations based on the range of typical soil gas diffusivities of unsaturated soils showed that the fraction of N2 and N2O evolved from 15N-labelled NO3- that is not emitted at the soil surface during 1 h chamber closing is always significant, with values up to >50 % of total production. This is due to accumulation in the pore space of the 15N-labelled soil and diffusive flux to the unlabelled subsoil. Empirical coefficients to calculate denitrification from surface fluxes were derived by modelling multiple scenarios with varying soil water content. Modelling several theoretical experimental set-ups showed that the fraction of produced gases that are retained in soil can be lowered by lowering the depth of 15N labelling and/or increasing the length of the confining cylinder. Field experiments with arable silt loam soil for measuring denitrification with the 15N gas flux method were conducted to obtain direct evidence for the incomplete surface emission of gaseous denitrification products. We compared surface fluxes of 15N2 and 15N2O from 15N-labelled micro-plots confined by cylinders using the closed-chamber method with cylinders open or closed at the bottom, finding 37 % higher surface fluxes with the bottom closed. Modelling fluxes of this experiment confirmed this effect, however with a higher increase in surface flux of 89 %. From our model and experimental results we conclude that field surface fluxes of 15N-labelled N2 and N2O severely underestimate denitrification rates if calculated from chamber accumulation only. The extent of this underestimation increases with closure time. Underestimation also occurs during laboratory incubations in closed systems due to pore space accumulation of 15N-labelled N2 and N2O. Due to this bias in past denitrification measurements, denitrification in soils might be more relevant than assumed to date. Corrected denitrification rates can be obtained by estimating subsurface flux and storage with our model. The observed deviation between experimental and modelled subsurface flux revealed the need for refined model evaluation, which must include assessment of the spatial variability in diffusivity and production and the spatial dimension of the chamber.

Metagenomic analysis reveals distinct patterns of denitrification gene abundance across soil moisture, nitrate gradients.

This study coupled a landscape-scale metagenomic survey of denitrification gene abundance in soils with in situ denitrification measurements to show how environmental factors shape distinct denitrification communities that exhibit varying denitrification activity. Across a hydrologic gradient, the distribution of total denitrification genes (nap/nar + nirK/nirS + cNor/qNor + nosZ) inferred from metagenomic read abundance exhibited no consistent patterns. However, when genes were considered independently, nirS, cNor and nosZ read abundance was positively associated with areas of higher soil moisture, higher nitrate and higher annual denitrification rates, whereas nirK and qNor read abundance was negatively associated with these factors. These results suggest that environmental conditions, in particular soil moisture and nitrate, select for distinct denitrification communities that are characterized by differential abundance of genes encoding apparently functionally redundant proteins. In contrast, taxonomic analysis did not identify notable variability in denitrifying community composition across sites. While the capacity to denitrify was ubiquitous across sites, denitrification genes with higher energetic costs, such as nirS and cNor, appear to confer a selective advantage in microbial communities experiencing more frequent soil saturation and greater nitrate inputs. This study suggests metagenomics can help identify denitrification hotspots that could be protected or enhanced to treat non-point source nitrogen pollution.

Improvement of the 15 N gas flux method for in situ measurement of soil denitrification and its product stoichiometry.

Field measurement of denitrification in agricultural ecosystems using the 15 N gas flux method has been limited by poor sensitivity because current isotope ratio mass spectrometry is not precise enough to detect low 15 N2 fluxes in the presence of a high atmospheric N2 background. For laboratory studies, detection limits are improved by incubating soils in closed systems and under N2 -depleted atmospheres. We developed a new procedure to conduct the 15 N gas flux method suitable for field application using an artificially N2 -depleted atmosphere to improve the detection limit at the given precision of mass spectrometry. Laboratory experiments with and without 15 N-labelling and using different flushing strategies were conducted to develop a suitable field method. Subsequently, this method was tested in the field and results were compared with those obtained from the conventional 15 N gas flux method. Results of the two methods were in close agreement showing that the denitrification rates determined were not biased by the flushing procedure. Best sensitivity for N2 + N2 O fluxes was 10 ppb, which was 80-fold better than that of the reference method. Further improvement can be achieved by lowering the N2 background concentration below the values established in the present study. In view of this progress in sensitivity, the new method will be suitable to measure denitrification dynamics in the field beyond peak events.

Estimate of gas transfer velocity in the presence of emergent vegetation using argon as a tracer: Implications for whole-system denitrification measurements

Denitrification associated with emergent macrophytes is a pivotal process underlying the treatment performance of wetlands and slow-flow waterways. Laboratory scale experiments targeting N losses via denitrification in sediments colonized by emergent macrophytes require the use of mesocosms that are necessarily open to the atmosphere. Thus, the proper quantification of N2 effluxes relies on the accurate characterization of the air–water gas exchanges. In this study, we present a simple approach for direct measurements of the gas transfer velocity, in open-top mesocosms with Phragmites australis, by using argon as a tracer. Different conditions of water velocity (0, 1.5, 3, and 6 cm s−1) and temperature (8.5, 16, and 28 °C), were tested, along with, for the first time, the presence of emergent vegetation. The outcomes demonstrated that water velocity and temperature are not the only factors regulating aeration at the mesocosm scale. Indeed, the gas transfer velocity was systematically higher, in the range of 42–53%, in vegetated compared to unvegetated sediments. The increase of small-local turbulence patterns created within water parcels moving around plant stems translated into significant modifications of the reaeration process. The adopted approach may be used to improve the accuracy of denitrification measurements by N2 efflux-based methods in wetland and slow-flow waterway sediments colonized by emergent macrophytes. Moreover, the present outcomes may have multiple implications for whole-system metabolism estimations from which largely depend our understanding of biogeochemical dynamics in inland waters that have strong connections to worldwide issues, such as nitrate contamination and greenhouse gas emissions.

Predicting Nitrate Retention at the Groundwater‐Surface Water Interface in Sandplain Streams

AbstractGroundwater‐surface water ecotones present opportunities for nitrate retention because changes in organic carbon availability, redox potential, and nitrate demand often occur at these locations. Although there have been many measurements of nitrate retention and denitrification at the groundwater‐surface water interface, few investigators have quantitatively predicted where nitrogen transformation occurs in this zone. Our main objective was to describe and predict spatial variation in nitrate retention and removal in shallow groundwater among and within streams in the central sand plains of Wisconsin. We predicted that nitrate transformation rates would be positively related to nitrate and particulate organic carbon availability, a negative nonlinear function of dissolved oxygen availability, and would peak at intermediate rates of groundwater discharge. Five sites on four streams were chosen that spanned an order of magnitude in groundwater nitrate concentration. Nitrate retention and N2 production were quantified by determining how nitrate and N2 fluxes changed along nominal 55‐cm groundwater flow paths. Nitrogen transformation was widespread but highly variable within the study sites. Partial least squares regression models explained a much larger amount of variation in nitrate retention and N2 production at the local scale than at the regional scale. Dissolved oxygen availability and groundwater discharge were the most consistent predictors of nitrate retention at both scales. We conclude that nitrate retention at the groundwater‐surface water interface can be predicted but that more research is necessary to determine if nitrate transformation at this ecotone can be modeled at broader scales.

Anaerobic methane oxidation and aerobic methane production in an east African great lake (Lake Kivu)

We investigated CH4 oxidation in the water column of Lake Kivu, a deep meromictic tropical lake with CH4-rich anoxic deep waters. Depth profiles of dissolved gases (CH4 and N2O) and a diversity of potential electron acceptors for anaerobic CH4 oxidation (NO3−, SO42−, Fe and Mn oxides) were determined during six field campaigns between June 2011 and August 2014. Denitrification measurements based on stable isotope labelling experiments were performed twice. In addition, we quantified aerobic and anaerobic CH4 oxidation, NO3− and SO42− consumption rates, with and without the presence of an inhibitor of SO42−-reducing bacteria activity. Aerobic CH4 production was also measured in parallel incubations with the addition of an inhibitor of aerobic CH4 oxidation. The maximum aerobic and anaerobic CH4 oxidation rates were estimated to be 27 ± 2 and 16 ± 8 μmol/L/d, respectively. We observed a difference in the relative importance of aerobic and anaerobic CH4 oxidation during the rainy and the dry season, with a greater role for aerobic oxidation during the dry season. Lower anaerobic CH4 oxidation rates were measured in presence of molybdate in half of the measurements, suggesting the occurrence of linkage between SO42− reduction and anaerobic CH4 oxidation. NO3− consumption and dissolved Mn production rates were never high enough to sustain the measured anaerobic CH4 oxidation, reinforcing the idea of a coupling between SO42− reduction and CH4 oxidation in the anoxic waters of Lake Kivu. Finally, significant rates (up to 0.37 μmol/L/d) of pelagic CH4 production were also measured in oxygenated waters.

Comprehensive effects of a sedge plant on CH4 and N2O emissions in an estuarine marsh

Although there have been numerous studies focusing on plants' roles in methane (CH4) emissions, the influencing mechanism of wetland plants on nitrous oxide (N2O) emissions has rarely been studied. Here, we test whether wetland plants also play an important role in N2O emissions. Gas fluxes were determined using the in situ static flux chamber technique. We also carried out pore-water extractions, sedge removal experiments and tests of N2O transportation. The brackish marsh acted as a net source of both CH4 and N2O. However, sedge plants played the opposite role in CH4 and N2O emissions. The removal of the sedges led to reduced CH4 emissions and increased accumulation of CH4 inside the sediment. Apart from being a conduit for CH4 transport, the sedges made a greater contribution to CH4 oxidation than CH4 production. The sedges exerted inhibitory effects on the release of N2O. The N2O was barely detectable inside the sediment in both vegetated and vegetation-removed plots. The denitrification measurements and nitrogen addition (the addition rates were equal to 0.028, 0.056 and 0.112 g m−2) experiments suggest that denitrification associated with N2O production occurred mainly in the surface sediment layer. The vascular sedge could transport atmospheric N2O downward into the rhizosphere. The rhizospheric sediment, together with the vascular sedge, became an effective sink of atmospheric N2O.

Open-channel measurement of denitrification in a large lowland river

Denitrification is considered to be the most important pathway removing nitrogen from terrestrial and freshwater ecosystems. However, field studies that quantify this process under in situ conditions are sparse, especially in large rivers. Here, we measured N2, the end product of denitrification, directly in the water column of a large 8th order lowland river (Elbe, Germany) using N2/Ar ratios measured by Membrane Inlet Mass Spectrometry (MIMS). Denitrification was calculated according to the open-channel two-station approach based on Lagrangian sampling along a 580 km long, mostly free flowing river section. Gas exchange was computed by several empirical equations to bound uncertainty in air–water exchange and the resulting fluxes were used to estimate ranges in N2-production. In summer 2011 and spring 2012, we found slight but distinct N2 super saturations in the river water averaging 2.8 and 3.5 µM, respectively. Denitrification rates averaged 18 and 13 mg N m− 2 h− 1 for summer 2011 and spring 2012, respectively. On an annual cycle this corresponds to a nitrogen removal of 10,000 t N year− 1 that is 10% of the total N inputs along the studied river section. These results show that large rivers can remove large amounts of nitrogen during downstream transport and demonstrate that the open-channel N2 method provides a valuable tool to study in situ denitrification not only in small, but also in large rivers.

Invasive plants decrease microbial capacity to nitrify and denitrify compared to native California grassland communities

Exotic plant invasions are a major driver of global environmental change that can significantly alter the availability of limiting nutrients such as nitrogen (N). Beginning with European colonization of California, native grasslands were replaced almost entirely by annual exotic grasses, many of which are now so ubiquitous that they are considered part of the regional flora (“naturalized”). A new wave of invasive plants, such as Aegilops triuncialis (Barb goatgrass) and Elymus caput-medusae (Medusahead), continue to spread throughout the state today. To determine whether these new-wave invasive plants alter soil N dynamics, we measured inorganic N pools, nitrification and denitrification potentials, and possible mediating factors such as microbial biomass and soil pH in experimental grasslands comprised of A. triuncialis and E. caput-medusae. We compared these measurements with those from experimental grasslands containing: (1) native annuals and perennials and (2) naturalized exotic annuals. We found that A. triuncialis and E. caput-medusae significantly reduced ion-exchange resin estimates of nitrate (NO3 −) availability as well as nitrification and denitrification potentials compared to native communities. Active microbial biomass was also lower in invaded soils. In contrast, potential measurements of nitrification and denitrification were similar between invaded and naturalized communities. These results suggest that invasion by A. triuncialis and E. caput-medusae may significantly alter the capacity for soil microbial communities to nitrify or denitrify, and by extension alter soil N availability and rates of N transformations during invasion of remnant native-dominated sites.

Application of the <sup>15</sup>N gas-flux method for measuring in situ N<sub>2</sub> and N<sub>2</sub>O fluxes due to denitrification in natural and semi-natural terrestrial ecosystems and comparison with the acetylene inhibition technique

Abstract. Soil denitrification is considered the most un-constrained process in the global N cycle due to uncertain in situ N2 flux measurements, particularly in natural and semi-natural terrestrial ecosystems. 15N tracer approaches can provide in situ measurements of both N2 and N2O simultaneously, but their use has been limited to fertilized agro-ecosystems due to the need for large 15N additions in order to detect 15N2 production against the high atmospheric N2. For 15N–N2 analyses, we have used an “in-house” laboratory designed and manufactured N2 preparation instrument which can be interfaced to any commercial continuous flow isotope ratio mass spectrometer (CF-IRMS). The N2 prep unit has gas purification steps and a copper-based reduction furnace, and allows the analysis of small gas injection volumes (4 µL) for 15N–N2 analysis. For the analysis of N2O, an automated Tracegas Preconcentrator (Isoprime Ltd) coupled to an IRMS was used to measure the 15N–N2O (4 mL gas injection volume). Consequently, the coefficient of variation for the determination of isotope ratios for N2 in air and in standard N2O (0.5 ppm) was better than 0.5 %. The 15N gas-flux method was adapted for application in natural and semi-natural land use types (peatlands, forests, and grasslands) by lowering the 15N tracer application rate to 0.04–0.5 kg 15N ha−1. The minimum detectable flux rates were 4 µg N m−2 h−1 and 0.2 ng N m−2 h−1 for the N2 and N2O fluxes respectively. Total denitrification rates measured by the acetylene inhibition technique in the same land use types correlated (r = 0.58) with the denitrification rates measured under the 15N gas-flux method, but were underestimated by a factor of 4, and this was partially attributed to the incomplete inhibition of N2O reduction to N2, under a relatively high soil moisture content, and/or the catalytic NO decomposition in the presence of acetylene. Even though relatively robust for in situ denitrification measurements, methodological uncertainties still exist in the estimation of N2 and N2O fluxes with the 15N gas-flux method due to issues related to non-homogenous distribution of the added tracer and subsoil gas diffusion using open-bottom chambers, particularly during longer incubation duration. Despite these uncertainties, the 15N gas-flux method constitutes a more reliable field technique for large-scale quantification of N2 and N2O fluxes in natural terrestrial ecosystems, thus significantly improving our ability to constrain ecosystem N budgets.

Modelling flow and inorganic nitrogen dynamics on the Hampshire Avon: Linking upstream processes to downstream water quality

Managing diffuse pollution in catchments is a major issue for environmental managers planning to meet water quality standards and comply with the EU Water Framework Directive. A major source of diffuse pollution is from nitrogen, with high nitrate concentrations affecting water supplies and in-stream ecology. A dynamic, process based model of flow, nitrate and ammonium (INCA-N) has been applied to the Hampshire Avon as part of the NERC Macronutrient Cycles Programme to link upstream and downstream measurements of water chemistry. The model has been calibrated and validated against Environment Agency discharge and solute chemistry data, as well as a data set collected from a river site immediately upstream of the estuary tidal limit. Upstream measurements of denitrification at six sites have been used to evaluate nitrate removal rates in vegetated and non-vegetated conditions. Results show that sediments underlying vegetation were associated with significantly higher rates of nitrate removal than un-vegetated sediments (with an average increase of 245%). These data have been used to scale up rates of nitrate loss to the whole catchment scale and have been implemented via the model. The effects of streambed geology and macrophyte cover on catchment-scale nitrogen dynamics are explored and nutrient fluxes entering the estuary are evaluated. The model is used to test a strategy for nitrogen reduction assessed using a nitrate vulnerable zone (NVZ) methodology. It suggests that nitrate and ammonium concentrations could be reduced by 10% in 10years and much lower nitrogen level can be achieved but only over a long time period.

Denitrification in a subtropical, semi-arid North American savanna: field measurements and intact soil core incubations

Information on denitrification (particularly N2) losses from dry ecosystems is limited despite their large area. Here, we present the first direct denitrification measurements for a northern hemisphere savanna, a Prosopis-dominated grassland/grove matrix in south Texas. We used the gas-flow intact soil core method to quantify N2, N2O and CO2 losses and compared these with field measurements of N2O, NOy, NH3 and CO2. Under field-realistic soil moisture and O2 conditions (average 17.5–20 % O2, minimum 15 %) incubated soils produced no measurable N2 flux (detection limit 52.2 µg N m−2 h−1). Only in a subset of grove soils were fluxes of 70–75 µg N m−2 h−1 recorded after 102 h of incubation at 5–10 % O2 following wetting of very dry soils. Making the assumption that potential N2 production falls just below the detection limit (likely an overestimate given the conditions needed to generate measurable fluxes), N2 flux rates would fall on the low end of that recorded for a tropical Australian savanna (45–110 µg N m−2 h−1) under comparable abiotic conditions. Assuming maximum possible production rates, N2 could comprise <32–76 % of total soil N gas flux following soil wetting in summer. Lack of flux response to soil wetting in winter suggests that cold-season N2 fluxes are negligible. N2O fluxes for core incubations were significantly higher than for field chambers; thus it is likely that incubations may overestimate N2O flux by reducing soil column consumption. Overall, results indicate that soil N2 fluxes are less dominant in this savanna than in other ecosystems investigated.

Predicting the denitrification capacity of sandy aquifers from in situ measurements using push–pull &amp;lt;sup&amp;gt;15&amp;lt;/sup&amp;gt;N tracer tests

Abstract. Knowledge about the spatial variability of in situ denitrification rates (Dr(in situ)) and their relation to the denitrification capacity in nitrate-contaminated aquifers is crucial to predict the development of groundwater quality. Therefore, 28 push–pull 15N tracer tests for the measurement of in situ denitrification rates were conducted in two sandy Pleistocene aquifers in northern Germany. The 15N analysis of denitrification-derived 15N-labelled N2 and N2O dissolved in water samples collected during the push–pull 15N tracer tests was performed using isotope ratio mass spectrometry (IRMS) in the lab and additionally for some tracer tests online in the field with a quadrupole membrane inlet mass spectrometer (MIMS) in order to test the feasibility of on-site real-time 15N analysis. Aquifer material from the same locations and depths as the push–pull injection points was incubated, and the initial and cumulative denitrification after 1 year of incubation (Dcum(365)) as well as the stock of reduced compounds (SRC) was compared with in situ measurements of denitrification. This was done to derive transfer functions suitable to predict Dcum(365) and SRC from Dr(in situ). Dr(in situ) ranged from 0 to 51.5 μg N kg−1 d−1. Denitrification rates derived from on-site isotope analysis using MIMS satisfactorily coincided with laboratory analysis by conventional IRMS, thus proving the feasibility of in situ analysis. Dr(in situ) was significantly higher in the sulfidic zone of both aquifers compared to the zone of non-sulfidic aquifer material. Overall, regressions between the Dcum(365) and SRC of the tested aquifer material with Dr(in situ) exhibited only a modest linear correlation for the full data set. However, the predictability of Dcum(365) and SRC from Dr(in situ) data clearly increased for aquifer samples from the zone of NO3−-bearing groundwater. In the NO3−-free aquifer zone, a lag phase of denitrification after NO3− injections was observed, which confounded the relationship between reactive compounds and in situ denitrification activity. This finding was attributed to adaptation processes in the microbial community after NO3− injections. It was also demonstrated that the microbial community in the NO3−-free zone just below the NO3−-bearing zone can be adapted to denitrification by NO3− injections into wells for an extended period. In situ denitrification rates were 30 to 65 times higher after pre-conditioning with NO3−. Results from this study suggest that such pre-conditioning is crucial for the measurement of Dr(in situ) in deeper aquifer material from the NO3−-free groundwater zone and thus for the prediction of Dcum(365) and SRC from Dr(in situ).