Nitrate Isotope Research Articles

Isotopic signature of N2O produced during sulfur‐ and thiosulfate‐driven chemoautotrophic denitrification in freshwaters

AbstractChemoautotrophic denitrification plays an important role in nitrogen removal and the formation of a greenhouse gas (N2O) in aquatic environments. Natural stable isotopes support the tracing of nitrogen sources and the identification of biogeochemical processes in field research. However, the isotopic characteristics of N2O produced during chemoautotrophic denitrification have not been investigated so far. In this study, we analyzed isotopic signatures of nitrate and N2O in sulfur‐ and thiosulfate‐dependent denitrifying enrichments obtained from freshwater lakes. Chemoautotrophic denitrification exhibited a nitrate isotope pattern similar to heterotrophic denitrification: the 18ε/15ε‐nitrate followed a ratio close to 1. However, chemoautotrophic denitrification produced N2O with lower δ15N, δ18O, and higher site preference (SP = δ15Nα‐δ15Nβ) values, compared to heterotrophic denitrification. The SP value, approximately 5.1‰, was characteristic in detecting chemoautotrophic denitrification driven by different sulfur forms. δ18O varied with specific electron donors, around 20‰ and 40‰ during sulfur‐ and thiosulfate‐dependent denitrification, respectively. The unique N2O isotope characteristics were likely regulated by nitric oxide reductases of Burkholderiaceae populations during sulfur‐dependent denitrification and Sulfurovum during thiosulfate‐dependent denitrification. These findings improve our understanding of N2O production processes and have important implications for predicting N2O emissions at a greater spatial and temporal resolution.

Experimental Study on the Impact of Sorption-Desorption on Nitrate Isotopes

Nitrate pollution is a global environmental problem, and mean nitrate levels have risen by an estimated 36% in global waterways since 1990. Tracing nitrate sources is important for water quality management, and nitrate isotopes (δ15N-NO3− and δ18O-NO3−) are commonly used for this purpose because of the different isotopic compositions of different sources. However, the impact of nitrate sorption on matrix and desorption from matrix on N and O isotopic composition of nitrate in liquid phase has not been well clarified. To explore the mechanism for the changes in nitrate concentration and isotopes in liquid phase during sorption and desorption, this study took a shale sample (enriched in clay minerals and commonly exposed in the Earth), conducted a series of laboratory experiments for nitrate sorption and desorption, and studied the impact of sorption and desorption on nitrate N and O isotopic composition in liquid phase. The results showed that the shale sample exhibited a rapid sorption and desorption rate for nitrate in the surface water samples, with the nitrate concentration in the solution decreasing from 14.3 mg/L to 4.1 mg/L within 5 min. The sorption data fit the Langmuir model better than that of the Freundlich model. The maximum possible sorption (Qmax) for the shale sample was estimated to be 46 μg/g. Preliminary laboratory experiments showed that changes in δ15N-NO3− values were not obvious, and changes in δ18O-NO3− values in liquid phase were minor during sorption and desorption of the shale sample, suggesting that nitrogen isotopic fractionation can be neglected, and the sorption of nitrate by the shale sample has a very limited impact on the distribution of nitrate isotopes in liquid phase. However, the impact of nitrate desorption on the nitrate isotopes in liquid phase depends on the isotopic composition of exchangeable nitrate in the solid phase, which may be related to antecedent water–rock interactions. This study provides important information for elucidating the evolution mechanism of nitrate and its isotopic compositions following sorption-desorption, and is conducive to revealing the nitrogen cycle law in the environment.

Nitrogen Transport Pathways and Source Contributions in a Typical Agricultural Watershed Using Stable Isotopes and Hydrochemistry

The increasing global nitrogen input poses a significant threat to aquatic environments, particularly in agricultural watersheds, where intensive human activities and insufficient water protection infrastructure exacerbate the risk of nitrogen pollution. Accurate identification of nitrogen pollution sources and the associated transformation processes is essential for protecting watershed ecosystems. In this study, a combination of hydrochemical analysis, correlation and principal component analysis, and stable nitrate isotopes (δ15N-NO3− and δ18O-NO3−) were employed to trace nitrogen transport pathways and source contributions in both surface water and groundwater within a typical agricultural watershed. The results revealed the presence of nitrogen pollution, including total nitrogen (TN), ammonia nitrogen (NH3-N), and nitrate nitrogen (NO3−-N), with significant spatial and seasonal variations in both surface water and groundwater. The spatiotemporal evolution of hydrochemical indicators and nitrate isotope compositions highlighted multiple potential sources of nitrogen, including soil input, agricultural input, and manure and sewage input. The results from stable isotope analysis in an R (SIAR) model indicated that ammonium fertilizers (7.1~78.4%) and manure and sewage (2.6~69.7%) were the primary sources of nitrates in surface water, while manure and sewage were the main sources in groundwater (67.9~73.7%). This research demonstrated that nitrification, seasonal variations, and human activities significantly impact nitrogen migration and transformation in agricultural watersheds. However, the issue of groundwater severely polluted by manure and sewage has received insufficient attention. To effectively control nitrogen pollution in agricultural watersheds, it is necessary to improve septic tanks and sewage networks, as well as implement scientific fertilization practices.

High Frequency Monitoring and Nitrate Sourcing Reveals Baseflow and Stormflow Controls on Total Dissolved Nitrogen and Carbon Export Along a Rural‐Urban Gradient

AbstractEfforts to reduce nitrogen and carbon loading from developed watersheds typically target specific flows or sources, but across gradients in development intensity there is no consensus on the contribution of different flows to total loading or sources of nitrogen export. This information is vital to optimize management strategies leveraging source reductions, stormwater controls, and restorations. We investigate how solute loading and sources vary across flows and land‐use using high frequency monitoring and stable nitrate isotope analysis from five catchments with different sanitary infrastructure, along a gradient in development intensity. High frequency monitoring allowed estimation of annual loading and attribution to storm versus baseflows. Nitrate loads were 16 kg/km2/yr. from the forested catchment and ranged from 68 to 119 kg/km2/yr., across developed catchments, highest for the septic served site. Across developed catchments, baseflow contributions ranged from 40% of N loading to 75% from the septic served catchment, and the contribution from high stormflows increased with development intensity. Stormflows mobilized and mixed many surface and subsurface nitrate sources while baseflow nitrate was dominated by fewer sources which varied by catchment (soil, wastewater, or fertilizer). To help inform future sampling designs, we demonstrate that grab sampling and targeted storm sampling would likely fail to accurately predict annual loadings within the study period. The dominant baseflow loads and subsurface stormflows are not treated by surface water management practices primarily targeted to surface stormflows. Using a balance of green and gray infrastructure and stream/riparian restoration may target specific flow paths and improve management.

Infiltration of secondary treated wastewater into an oxic aquifer: Hydrochemical insights from a large-scale sand tank experiment

To mitigate groundwater level decline, managed aquifer recharge (MAR) with secondary treated wastewater (STWW) is increasingly considered and implemented. However, the effectiveness and potential risks of such systems need evaluation prior to implementation. In this study, we present a large-scale sand tank experiment to analyse processes related to the infiltration of real STWW through the vadose zone and subsequent mixing with oxic native groundwater. The varying composition of STWW from 15 infiltration cycles over six months of operation and the retention times were the main drivers of the observed processes, which were characterized by a wide range of analytical techniques such as in situ high-resolution oxidation–reduction potential (ORP) measurements, closed mass balances of solutes, characterization of dissolved organic carbon (DOC), stable nitrate isotopes analysis, as well as numerical flow and transport modelling. Depending on the composition and infiltration rates of the STWW, both nitrification and denitrification could be observed, even simultaneously at different locations in the tank. Furthermore, due to the variability of the real STWW we observed enhanced arsenic mobilisation during times of elevated phosphate concentrations of the infiltrating STWW. Additionally, uranium was mobilised in our experimental system via carbonate mineral dissolution caused by the infiltrating STWW which was undersaturated of calcite for all infiltration cycles. Overall, our results showed the importance of conducting studies with waters of complex matrix, such as real STWW, and considering mixing with groundwater to assess the full range of possible processes encountered at MAR field sites.

A multidisciplinary approach using hydrogeochemistry, δ15NNO3 isotopes, land use, and statistical tools in evaluating nitrate pollution sources and biochemical processes in Costa Rican volcanic aquifers

Nitrate pollution threatens the Barva and Colima multi-aquifer system, the primary drinking water source in the Greater Metropolitan Area of Costa Rica. In addressing nitrate contamination dynamics, this study proposes an integrated approach by combining multivariate statistical analyses, hydrochemical parameters, sewage discharge, and regional land-use and land-cover patterns to assess the extent and degree of contamination, dominant biogeochemical processes, and refine the interpretation of nitrate sources previously derived solely from δ15NNO3 information. Over seven years (2015–2022), 714 groundwater samples from 43 sites were analyzed for nitrate and major ions, including two sampling campaigns for dissolved organic and inorganic carbon, nitrite, ammonium, FeTotal, MnTotal, and δ15NNO3 analyses. The findings presented elevated nitrate concentrations in urban and agricultural/urban areas, surpassing the Maximum Concentration Levels on several occasions, and oxidizing conditions favoring mineralization and nitrification processes in unconfined Barva and locally confined Upper Colima/Lower Colima aquifers. Similar nitrate contents and spatial patterns in agricultural and urban zones in the shallow Barva aquifer suggest comparable contributions from nitrogen fertilizers and urban wastewaters despite the gradual increase in urban land cover and the reduction of agricultural areas. Isotopic analyses and dissolved organic carbon (DOC) indicate a shift in nitrate sources from agricultural to urban areas in both Barva and Colima aquifers. Principal Component and Hierarchical Cluster Analyses link land use, nitrate sources, and water quality. Three distinct sample clusters aligned with forest/grassland, agricultural/urban, and urban land use, emphasizing the impact of anthropogenic activities on groundwater quality, even in the deeper Colima aquifers. The study challenges nitrate isotope mixing models, enhancing accuracy in identifying pollution sources and assessing the spatial extent of contamination by incorporating DOC and other hydrochemical parameters. Similar outcomes, with and without the use of nitrate isotopes, reinforce the usefulness of the integrated approach, providing a practical and cost-effective alternative.

Tracing groundwater nitrate sources in an intensive agricultural region integrated of a self-organizing map and end-member mixing model tool

The traceability of groundwater nitrate pollution is crucial for controlling and managing polluted groundwater. This study integrates hydrochemistry, nitrate isotope (δ15N-NO3− and δ18O-NO3−), and self-organizing map (SOM) and end-member mixing (EMMTE) models to identify the sources and quantify the contributions of nitrate pollution to groundwater in an intensive agricultural region in the Sha River Basin in southwestern Henan Province. The results indicate that the NO3−-N concentration in 74% (n = 39) of the groundwater samples exceeded the WHO standard of 10 mg/L. According to the results of EMMTE modeling, soil nitrogen (68.4%) was the main source of nitrate in Cluster-1, followed by manure and sewage (16.5%), chemical fertilizer (11.9%) and atmospheric deposition (3.3%). In Cluster-2, soil nitrogen (60.1%) was the main source of nitrate, with a significant increase in the contribution of manure and sewage (35.5%). The considerable contributions of soil nitrogen may be attributed to the high nitrogen fertilizer usage that accumulated in the soil in this traditional agricultural area. Moreover, it is apparent that most Cluster-2 sampling sites with high contributions of manure and sewage are located around residential land. Therefore, the arbitrary discharge and leaching of domestic sewage may be responsible for these results. Therefore, this study provides useful assistance for the continuous management and pollution control of groundwater in the Sha River Basin.

Mechanisms underlying episodic nitrate and phosphorus leaching from poorly drained agricultural soils.

Poorly drained depressions within tile-drained croplands can have disproportionate environmental and agronomic impacts, but mechanisms controlling nutrient leaching remain poorly understood. We monitored nitrate and soluble reactive phosphorus (SRP) leaching using zero-tension soil lysimeters across a depression to upland gradient over 2 years in a corn-soybean (Zea mays L.-Glycine max [L.] Merr.) field in Iowa. We also measured stable isotopes (δ15N and δ18O) of nitrate to examine its sources and transformations. SRP concentrations peaked during winter and early spring after phosphorus (P) fertilization (mean=3mg P L-1), with highest values in the depression, and SRP was relatively stable thereafter (mean=0.3mg P L-1) irrespective of periods of high soil moisture that led to widespread iron (Fe) reduction across the field. During a near-average precipitation year, nitrate stable isotopes indicated direct leaching of fertilizer nitrate within days of application, followed by nitrification of fertilizer ammonium and several weeks of denitrification in depressional soils. Nevertheless, nitrate concentrations remained high (mean=28mg N L-1) in the depression despite strong isotopic evidence for denitrification (>48% N removal). During a wet year, nitrate concentrations were lower in the depression than upland and nitrate isotopes were highly variable, consistent with nearly complete nitrate removal by denitrification in the depression and significant denitrification in upland soils. We conclude that poorly drained depressional soils can potentially decrease nitrate leaching via denitrification under sustained wet conditions, but they inconsistently denitrify and are vulnerable to high nitrate and SRP losses when soils are not saturated, especially following fertilization.

Deciphering and quantifying nitrate sources and processes in the central Yellow Sea using dual isotopes of nitrate

Anthropogenic activities pose significant challenges to the accumulation of coastal nitrogen (N). Accurate identification of nitrate (NO3−) sources is thus essential for mitigating excessive N in many marginal seas. We investigated the dual isotopes of NO3− in the central Yellow Sea to elucidate the sources and cycling processes of NO3−. The results revealed significant spatial variability in NO3− concentrations among the Yellow Sea Surface Water (YSSW), Changjiang Diluted Water (CDW), Yellow Sea Cold Water Mass (YSCWM), and Taiwan Warm Current Water (TWCW). Stratification played a crucial role in restricting vertical nutrient transport, leading to distinct nutrient sources and concentrations in different water masses. The dual NO3− isotopic signature indicated that atmospheric deposition was the primary source of surface NO3−, contributing approximately 30 % to the NO3− in the YSSW. In the NO3−-rich CDW, the heavier δ15N-NO3− and δ18O-NO3− suggested incomplete NO3− assimilation. Organic matter mineralization and water stratification played crucial roles in the accumulation of nutrients within the YSCWM and TWCW. Notably, regenerated NO3− accounted for approximately half of the NO3− stored in the YSCWM. A synthesis of NO3− dual isotope data across the coastal China seas revealed significant spatial and seasonal variations in the N source. The study emphasized the dynamics of coastal NO3− supply, which are shaped by the complex interconnections among marine, terrestrial, and atmospheric processes. Our approach is a feasible method for exploring the origins of N amidst the escalating pressures of anthropogenic nutrient pollution in coastal waters.

Hemispherical scale mechanisms of nitrate formation in global marine aerosols

Nitrogen oxides (NOx) in the atmosphere directly affect air quality; however, their oxidation products (e.g., nitrate) are essential nutrients in terrestrial and marine ecosystems. To date, the mechanism and rate of nitrate formation in the global-scale marine boundary layer remains uncertain. Herein, aerosol nitrate isotopes covering the global ocean were analysed and observations indicated that nitrate formation was dominated by the proportion between the hydroxyl radical (daytime) and ozone (nighttime) pathways in the Northern Hemisphere (NH), whereas it changed to the BrO (Antarctic iceberg emission) pathway in the Southern Hemisphere (SH). These differences in the pathways suggested that the NOx removal (nitrate formation) efficiency was higher in the NH, which could be responsible for the much higher nitrate concentrations in the NH than in the SH (over twice). This study can assist in formulating effective policies to mitigate global NOx pollution and improve our understanding of the impact of increasing NOx concentrations on global ocean productivity.

Spectral and molecular insights into the characteristics of dissolved organic matter in nitrate-contaminated groundwater

The characteristics of dissolved organic matter (DOM) serve as indicators of nitrate pollution in groundwater. However, the specific DOM components associated with nitrate in groundwater systems remain unclear. In this study, dual isotopes of nitrate, three-dimensional Excitation emission matrices (EEMs) and Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) were utilized to uncover the sources of nitrate and their associations with DOM characteristics. The predominant nitrate in the targeted aquifer was derived from soil organic nitrogen (mean 46.0%) and manure &sewage (mean 34.3%). The DOM in nitrate-contaminated groundwater (nitrate-nitrogen >20 mg/L) exhibited evident exogenous characteristics, with a bioavailable content 2.58 times greater than that of uncontaminated groundwater. Regarding the molecular characteristics, DOM molecules characterized by CHO + 3N, featuring lower molecular weights and H/C ratios, indicated potential for mineralization, while CHONS formulas indicated the exogenous features, providing the potential for accurate traceability. These findings provided insights at the molecular level into the characterization of DOM in nitrate-contaminated groundwater and offer scientific guidance for decision-making regarding the remediation of groundwater nitrate pollution.

Improved determination of nitrate isotopes in irrigated agricultural areas and Bayesian mixing model quantification of nitrate sources and fate

Technological advancements in isotope techniques have been widely utilized in environmental pollution research, particularly for the monitoring of nitrate pollution in water. The utilization of pretreatment with an aqueous nitric acid solution offers numerous advantages in analyzing isotopic composition; however, there is still room for improvement regarding conversion efficiency, reduced reaction time, and cost control. In light of this, this study proposes an enhanced chemical conversion method that involves mitigating the interference caused by the nitrogen azide reaction during the pretreatment process, optimizing the purge gas to remove interfering factors, and improving the inlet device for measuring nitrous oxide. The improved method was applied to determine nitrate isotopes in the Ulansuhai Basin, Inner Mongolia and its reliability was verified through analytical results obtained from bacterial denitrification methods. The mean δ15N-NO3- and δ18O-NO3- values determined using the chemical transformation method were 4.85‰ and 23.44‰, respectively. Compared with the previous method, the improved method enhanced the maneuverability of precise control, the pretreatment time was reduced from 2 to 3 hours to 1.5 hours, and the experimental cost was scaled down to half of the original one. In addition, the contribution of each source was calculated by combining nitrate isotopes with a Bayesian stable isotope mixing model. It was found that manure sewage accounted for (50.1±22.6) % while soil nitrogen sources contributed (25.1±20.4) % to nitrate levels in Ulansuhai Lake's water. The chemical conversion methods successfully improved within this study can also be applied to other watersheds or regions. These research findings provide a valuable theoretical foundation for managing nitrogen sources within watersheds.

Spatio-Temporal Variations in Nitrate Sources and Transformations in the Midstream of the Yellow River Determined Based on Nitrate Isotopes and Hydrochemical Compositions

Nitrate pollution is a major environmental problem threatening rivers, and nitrogen and oxygen isotopes have proved to be an effective means of analyzing the sources and transformations of nitrate in rivers. However, a low monitoring frequency cannot accurately reflect the changes in nitrate. In this study, the sources and transformations of nitrate in the middle reaches of the Yellow River and its tributaries during the dry season and the wet season were analyzed based on water quality parameters and nitrate isotopes. Stable isotope analysis conducted using the R (SIAR) model was used to estimate the proportions of different nitrate sources. The results showed that the main nitrate sources in the main stream were soil nitrogen (40.95–45.83%) and domestic sewage and manure (30.93–32.60%), respectively, with little variation between the dry season and wet season because of the large flow of the Yellow River. During the dry season, the nitrate sources of the two tributaries were mainly domestic sewage and manure (45.23–47.40%), followed by soil nitrogen (31.35–34.00%). However, the primary nitrate source of T2 (Qin River) became soil nitrogen (40.05%) during the wet season, a phenomenon that was mainly caused by the significant increase in river discharge and in soil erosion in the basin. During the wet season, the concentrations of total nitrogen (TN) and nitrate (NO3−) significantly decreased in the main stream and tributaries, and nitrification and denitrification processes occurred in both the main stream and tributaries of the Yellow River. In addition, the T2 tributary (Qin River) was also significantly affected by mixed dilution. High-frequency sampling can reflect the isotopic information of nitrate in the river more comprehensively, which helps us to understand the conversion process of nitrate more accurately.

Research progress on nitrate isotope coupled multi-tracer tracing groundwater nitrate pollution

Nitrate pollution in groundwater has become a global concern. One of the most important issues in controlling the nitrate pollution of groundwater is to identify the pollution source quickly and accurately. In this review, we firstly summarized the isotopic background values of potential sources of nitrate pollution in groundwater in 17 provinces (cities, autonomous regions) and 29 study areas in China, which could provide the fundamental database for subsequent research. Secondly, we reviewed the research progress of nitrate isotopes combined with multiple tracers for tracing nitrate in groundwater, and discussed their applicable conditions, advantages, and disadvantages. We found that halides and microorganisms combined with nitrate isotopes could accurately trace the pollution sources of domestic sewage, excrement and agricultural activities. The combination of Δ17O and nitrate isotopes could effectively distinguish the source of atmospheric deposition of nitrate in groundwater. The combination of groundwater age and nitrate isotopes could further determine the time scale of nitrate pollution. In addition, we summarized the application cases and compared the characteristics of mass balance mixing model, IsoSource model, Bayesian isotope mixing model, and EMMTE model for quantitative identification of nitrate pollution in groundwater. For the complexity and concealment of groundwater pollution sources, the coupling of nitrate isotopes with other chemical and biological tracing methods, as well as the application of nitrate isotope quantitative models, are effective tools for reliably identifying groundwater nitrate sources and transformation processes.

Vadose zone flushing of fertilizer tracked by isotopes of water and nitrate

AbstractA substantial fraction of nitrogen (N) fertilizer applied in agricultural systems is not incorporated into crops and moves below the rooting zone as nitrate (NO3−). Understanding mechanisms for soil N retention below the rooting zone and leaching to groundwater is essential for our ability to track the fate of added N. We used dual stable isotopes of nitrate (δ15N–NO3− and δ18O–NO3−) and water (δ18O–H2O and δ2H–H2O) to understand the mechanisms driving nitrate leaching at three depths (0.8, 1.5, and 3.0 m) of an irrigated corn field sampled every 2 weeks from 2016 to 2020 in the southern Willamette Valley, Oregon, USA. Distinct periods of high nitrate concentrations with lower δ15N–NO3− values indicated that a portion of that nitrate was from recent fertilizer applications. We used a mixing model to quantify nitrate fluxes associated with recently added fertilizer N versus older, legacy soil N during these “fertilizer signal periods.” Nitrate leached below 3.0 m in these periods made up a larger proportion of the total N leached at that depth (∼52%) versus the two shallower depths (∼13%–16%), indicating preferential movement of recently applied fertilizer N through the deep soil into groundwater. Further, N associated with recent fertilizer additions leached more easily when compared to remobilized legacy N. A high volume of fall and winter precipitation may push residual fertilizer N to depth, potentially posing a larger threat to groundwater than legacy N. Optimizing fertilizer N additions could minimize fertilizer losses and reduce nitrate leaching to groundwater.

Increase in Agricultural-Derived NHx and Decrease in Coal Combustion-Derived NOx Result in Atmospheric Particulate N-NH4+ Surpassing N-NO3- in the South China Sea.

The atmospheric deposition of anthropogenic active nitrogen significantly influences marine primary productivity and contributes to eutrophication. The form of nitrogen deposition has been evolving annually, alongside changes in human activities. A disparity arises between observation results and simulation conclusions due to the limited field observation and research in the ocean. To address this gap, our study undertook three field cruises in the South China Sea in 2021, the largest marginal sea of China. The objective was to investigate the latest atmospheric particulate inorganic nitrogen deposition pattern and changes in nitrogen sources, employing nitrogen-stable isotopes of nitrate (δ15N-NO3-) and ammonia (δ15N-NH4+) linked to a mixing model. The findings reveal that the N-NH4+ deposition generally surpasses N-NO3- deposition, attributed to a decline in the level of NOx emission from coal combustion and an upswing in the level of NHx emission from agricultural sources. The disparity in deposition between N-NH4+ and N-NO3- intensifies from the coast to the offshore, establishing N-NH4+ as the primary contributor to oceanic nitrogen deposition, particularly in ocean background regions. Fertilizer (33 ± 21%) and livestock (20 ± 6%) emerge as the primary sources of N-NH4+. While coal combustion continues to be a significant contributor to marine atmospheric N-NO3-, its proportion has diminished to 22 (Northern Coast)-35% (background area) due to effective NOx emission controls by the countries surrounding the South China Sea, especially the Chinese Government. As coal combustion's contribution dwindles, the significance of vessel and marine biogenic emissions grows. The daytime higher atmospheric N-NO3- concentration and lower δ15N-NO3- compared with nighttime further underscore the substantial role of marine biogenic emissions.

Changes in isotope fractionation during nitrate assimilation by marine eukaryotic and prokaryotic algae under different pH and CO2 conditions

AbstractThe impact of environmental factors on nitrogen (N) and oxygen (O) isotope effects during algal nitrate assimilation causes uncertainty in the field application of sedimentary N isotope records and nitrate isotopes to understand the marine nitrogen cycle. Ocean acidification is predicted to change nitrogen cycling including nitrate assimilation, but how N and O isotope effects during algal nitrate assimilation vary in response to changes in seawater pH and partial pressure CO2 (pCO2) remains unknown. We measured N and O isotope effects during nitrate assimilation and physiological states of the marine diatom Thalassiosira weissflogii and Synechococcus under different pH (8.1 or 7.8) and pCO2 (400 or 800 μatm) conditions. Low pH and/or high pCO2 equally decreased N and O isotope effects during nitrate assimilation by diatoms possibly due to reducing cellular nitrate efflux/uptake ratio and decreased isotope effects for nitrate uptake, whereas they did not affect those by Synechococcus with low intracellular nitrate concentration and limited nitrate efflux. Our results provide compelling experimental evidence showing different changes in N and O isotope effects during nitrate assimilation by marine eukaryotic and prokaryotic phytoplankton at low pH and/or high pCO2. These findings suggest new insight into environmental controls on variability in the isotope effect during algal nitrate assimilation, and have implications for improving a predictive understanding of N and O isotope tools in acidified oceans.

Sources and transformation of nitrate in an agricultural riverine reservoir watershed of the Sichuan Basin

Elucidating the origins and transformations of nitrate in groundwater and surface water within agricultural regions is imperative for effectively managing watershed contamination and environmental improvement. This study conducted a comprehensive geochemical investigation using multiple stable isotopes (δD-H2O and δ18O–H2O, δ15N–NO3- and δ18O–NO3-, and δ15N–NH4+) and hydrogeochemical characteristics to explore the potential sources and transformation of nitrate in groundwater and reservoir water within an agricultural riverine reservoir watershed. The results indicated that nitrate pollution in groundwater exhibited a higher severity compared to the reservoir water within this watershed, thereby posing potential risks to the quality of the reservoir water through hydraulic connection. The integration of δ15N–NH4+ and dual-isotopes of nitrate, combined with hydrogeochemical analyses, revealed that the presence of NO3− in groundwater and reservoir water primarily originated from manure/sewage sources. Moreover, intricate biogeochemical processes containing denitrification, nitrification, and DNRA were identified within this watershed. The findings of the Bayesian stable isotope mixing model (SIAR) further corroborated the conclusion that manure/sewage was the primary source of nitrate. It revealed that manure/sewage accounted for an average of 46 % of groundwater NO3− concentration, followed by atmospheric precipitation (25 %), soil nitrogen (18 %), and chemical fertilizer (11 %). There were no distinct differences in nitrate source contributions between groundwater and reservoir water. For reservoir water, manure or sewage (45 % on average) > atmospheric precipitation (28 %) > soil nitrogen (16 %) > chemical fertilizer (11 %). This study demonstrates the effectiveness of incorporating δ15N–NH4+ with dual isotopes of nitrate and hydrogeochemical analyses, serving as a reliable methodology that yields valuable insights into the origins and transformations of NO3− in an agricultural reservoir watershed, with potential applicability to other regions.

Riverine nitrate source identification combining δ15N/δ18O-NO3− with Δ17O-NO3− and a nitrification 15N-enrichment factor in a drinking water source region

Dual nitrate isotopes (δ15N/δ18O-NO3−) are an effective tool for tracing nitrate sources in freshwater systems worldwide. However, the initial δ15N/δ18O values of different nitrate sources might be altered by isotopic fractionation during nitrification, thereby limiting the efficiency of source apportionment results. This study integrated hydrochemical parameters, site-specific isotopic compositions of potential nitrate sources, multiple stable isotopes (δD/δ18O-H2O, δ15N/δ18O-NO3− and Δ17O-NO3−), soil incubation experiments assessing the nitrification 15N-enrichment factor (εN), and a Bayesian mixing model (MixSIAR) to reduce/eliminate the influence of 15N/18O-fractionations on nitrate source apportionment. Surface water samples from a typical drinking water source region were collected quarterly (June 2021 to March 2022). Nitrate concentrations ranged from 0.35 to 3.06 mg/L (mean = 0.78 ± 0.46 mg/L), constituting ∼70 % of total nitrogen. A MixSIAR model was developed based on δ15N/δ18O-NO3− values of surface waters and the incorporation of a nitrification εN (−6.9 ± 1.8 ‰). Model source apportionment followed: manure/sewage (46.2 ± 10.7 %) > soil organic nitrogen (32.3 ± 18.5 %) > nitrogen fertilizer (19.7 ± 13.1 %) > atmospheric deposition (1.8 ± 1.6 %). An additional MixSIAR model coupling δ15N/δ18O-NO3− with Δ17O-NO3− and εN was constructed to estimate the potential nitrate source contributions for the June 2021 water samples. Results revealed similar nitrate source contributions (manure/sewage = 43.4 ± 14.1 %, soil organic nitrogen = 29.3 ± 19.4 %, nitrogen fertilizer = 19.8 ± 13.8 %, atmospheric deposition = 7.5 ± 1.6 %) to the original MixSIAR model based on εN and δ15N/δ18O-NO3−. Finally, an uncertainty analysis indicated the MixSIAR model coupling δ15N/δ18O-NO3− with Δ17O-NO3− and εN performed better as it generated lower uncertainties with uncertainty index (UI90) of 0.435 compared with the MixSIAR model based on δ15N/δ18O-NO3− (UI90 = 0.522) and the MixSIAR model based on δ15N/δ18O-NO3− and εN (UI90 = 0.442).

Substantial contribution of tree canopy nitrifiers to nitrogen fluxes in European forests

Human activities have greatly increased the reactive nitrogen in the biosphere, thus profoundly altering global nitrogen cycling. The large increase in nitrogen deposition over the past few decades has led to eutrophication in natural ecosystems, with negative effects on forest health and biodiversity. Recent studies, however, have reported oligotrophication in forest ecosystems, constraining their capacity as carbon sinks. Here we demonstrate the widespread biological transformation of atmospheric reactive nitrogen in the canopies of European forests by combining nitrogen deposition quantification with measurements of the stable isotopes in nitrate and molecular analyses across ten forests through August–October 2016. We estimate that up to 80% of the nitrate reaching the soil via throughfall was derived from canopy nitrification, equivalent to a flux of up to 5.76 kg N ha−1 yr−1. We also document the presence of autotrophic nitrifiers on foliar surfaces throughout European forests. Canopy nitrification thus consumes deposited ammonium and increases nitrate inputs to the soil. The results of this study highlight widespread canopy nitrification in European forests and its important contribution to forest nitrogen cycling.