Assessing the impact of non-urea ruminant urine nitrogen compounds on urine patch nitrous oxide emissions

Urea, the dominant form of N in ruminant urine, degrades in soil to produce N2O emissions. However, the fate of non‐urea urine N compounds (NUNCs) in soil and their contribution to urine patch N2O emissions remain unclear. This study evaluated five NUNCs: allantoin (10%), creatinine (3%), creatine (3%), uric acid (1%), and (hypo)xanthine (0.6%), where numbers in parentheses represent the average percentage of total urine N. The fates of NUNCs in a pasture soil were determined using 15N‐labeled NUNCs in a laboratory trial. Two NUNCs, hypoxanthine and creatine, were added to the soil with perennial ryegrass (Lolium perenne L.) present and sampled over time for soil inorganic N, N2O emissions, and plant N dynamics. The 15N enrichments of soil inorganic N and plant N were significantly increased within 24 h of NUNC application, indicating rapid microbial degradation and plant uptake of NUNCs in pasture soil. An autumn field trial was also conducted to evaluate the in situ impact of varying concentrations of NUNCs on urine patch N2O emissions. Increasing the proportion of urine N excreted as NUNCs did not alter the urine patch N2O emission factor, soil inorganic N concentrations, or plant N uptake. It is concluded that NUNCs rapidly degrade in pasture soil and that an increased ruminant excretion of urine N as NUNCs does not significantly alter the urine patch N2O emission factor

Global Research Alliance N2O chamber methodology guidelines: Introduction, with health and safety considerations

Non-steady state (NSS) chamber techniques have been used for decades to measure nitrous oxide (N2O) fluxes from agricultural soils. These techniques are widely used because they are relatively inexpensive, easy to adopt, versatile and adaptable to varying conditions. Much of our current understanding of the drivers of N2O emissions, efficacy of mitigation practices, as well as estimations of agricultural N2O emission inventories, are based on using NSS chambers. While easy to adopt, use of NSS chambers requires decisions regarding multiple methodological aspects including chamber materials and geometry, sample replication, timing and frequency, sample analysis, use of ancillary information, and data analysis and statistical methods, each of which may significantly impact the results. Variation in these methodological details can lead to challenges in comparing results between studies and assessment of reliability and uncertainty. Therefore, the New Zealand Government, in support of the objectives of the Livestock Research Group of the Global Research Alliance on Agricultural Greenhouse Gases (GRA), funded an international project to develop standardised guidelines on the use of NSS chambers. Building on these initial guidelines, an international team of scientists have since refined them based on the most up to date knowledge and method developments. This introductory paper summarizes a collection of papers, comprising this special section of the Journal of Environmental Quality, that represent the revised guidelines. Each article summarises existing knowledge and provides guidance and recommendations for key aspects, including design, deployment, sample collection, storage and analysis, automated chambers, flux calculations, statistical analysis, emission factor estimation and data reporting, modelling approaches, and guidelines for ‘backfilling’ missing measurements. In addition, health and safety (H&S) considerations are included in this introductory paper. Each paper defines minimum requirements; however, these are not meant to be highly prescriptive, but instead to provide researchers with guidance on best practice and factors that need to be considered

Protein Quantification in Label-Free LC-MS Experiments

The goal of many LC-MS proteomic investigations is to quantify and compare the abundance of proteins in complex biological mixtures. However, the output of an LC-MS experiment is not a list of proteins, but a list of quantified spectral features. To make protein-level conclusions, researchers typically apply ad hoc rules, or take an average of feature abundance to obtain a single protein-level quantity for each sample. We argue that these two approaches are inadequate. We discuss two statistical models, namely, fixed and mixed effects Analysis of Variance (ANOVA), which views individual features as replicate measurements of a protein’s abundance, and explicitly account for this redundancy. We demonstrate, using a spike-in and a clinical data set, that the proposed models improve the sensitivity and specificity of testing, improve the accuracy of patient-specific protein quantifications, and are more robust in the presence of missing data

Potential inhibition of urine patch nitrous oxide emissions by Plantago lanceolata and its metabolite aucubin

Plantain (Plantago lanceolata L.), a forage used in grazed pastures, contains active secondary metabolites that could potentially inhibit nitrification, a key step in nitrous oxide (N2O) production from grazing ruminant livestock urine patches. A field study was performed to determine the effects of aucubin, a secondary metabolite in plantain, on nitrification and soil N2O emissions under a ruminant urine patch. Soils were treated with bovine urine (700 kg ha−1) and either a plantain leaf extract (PLE), which contained all extractable compounds in plantain including aucubin, or an aucubin solution (AS). PLE and AS were applied at the same rate of aucubin (47 kg ha−1). N2O emissions were reduced by 50% and 70% in the PLE and AS treatments, respectively; however, there were no significant differences in soil inorganic nitrogen concentrations when urine was applied with PLE or AS

Nitrous oxide and dinitrogen fluxes from grazed pasture soil after cattle urine deposition

Ruminant cattle grazing pasture result in urine deposition at high nitrogen (N) rates that can impact the environment including via gaseous emissions. Among those lost pathways, nitrogen gas emission, especially nitrous oxide (N₂O) and dinitrogen (N2), is the primary way where N can be lost to the atmosphere. There is limited information on the magnitude or fluxes of N2 losses from grazed-pasture systems after urine deposition due to the method limitation. We used the 15N flux method and high sampling frequency to explore N2 and N₂O fluxes over time after urine application at two rates (400 and 800 kg N ha-1) on a New Zealand grazed pasture soil. The higher N rate significantly increased daily N₂O fluxes but has no significant effect on daily N2 fluxes in our study compared with the lower rate. N2 is the predominant gaseous N form lost from the applied urinary-N which contributed 32.1 ± 4.1% and 14.4 ± 1.7% of the total deposited-N from 400 kg N ha-1 and 800 kg N ha-1 respectively, over 95 measurement days. Denitrification and codenitrification co-occurred in the pasture system, with denitrification being the predominant N2 production pathway, contributing 97.9 – 98.5 % of total N2 production. The N₂O/(N2+N₂O) product ratio was generally higher during periods of nitrification (the first month after urine application) but with no clear relationship to other measured variables. Contrary to our hypothesis, an elevated urine-N rate did not enhance N2 loss. This is speculated to be due to enhanced ammonia volatilisation and transfer of N as nitrate, to deeper soil layers. Soil relative gas diffusivity indicated that high N2 fluxes resulted from entrapped N2 diffusing from the draining soil

Potential for forage diet manipulation in New Zealand pasture ecosystems to mitigate ruminant urine derived N2O emissions: a review

Nitrous oxide (N2O) emissions from agricultural soils account for more than 10% of New Zealand’s greenhouse gas emissions. Livestock urine deposition drives N2O losses from these soils. It has been speculated that non-urea nitrogen compounds (UNCs) in ruminant urine could reduce or inhibit urine patch N2O emissions. However, we hypothesise that UNCs will have no effect on N2O emissions due to their potentially rapid degradation by plants and soil microbes. Our review suggests that plant secondary metabolites (PSMs) are more likely to perform a role in reducing N2O emissions since many PSMs have known antimicrobial properties. Aucubin, found in Plantago, and isothiocyanates, found in Brassica, have been shown to inhibit a key step in N2O production. Future studies should explore this promising research gap by evaluating forages for potential inhibitory PSMs, assessing whether PSMs are excreted in urine after consumption, and determine whether excretal PSM concentrations are sufficient to reduce N2O emissions

In situ nitrous oxide and dinitrogen fluxes from a grazed pasture soil following cow urine application at two nitrogen rates

Cattle grazing of pastures deposits urine onto the pasture soil at high nitrogen (N) rates that exceed the pasture's immediate N demands, increasing the risk of N loss. Nitrous oxide (N2O), a greenhouse gas, and dinitrogen (N2) are lost from the cattle urine patches. There is limited information on the in situ loss of N2 from grazed-pasture systems which is needed for understanding pasture soil N dynamics and balances. The 15N flux method was used to determine N2 and N2O fluxes over time following synthetic urine-15N application at either 400 or 800 kg N ha−1 to a grazed perennial pasture soil. Results showed that daily N2O fluxes were higher under 800 kg N ha−1 than under 400 kg N ha−1, but there was no significant difference in N2 fluxes. Cumulative N2O emissions from soil with 400 kg N ha−1 and 800 kg N ha−1 applied represented 0.16 ± 0.08% and 0.43 ± 0.08% of deposited N, respectively, while emitted N2 accounted for 32.1 ± 4.1% and 14.4 ± 1.7%, respectively, over 95 days after urine application. Codenitrification and denitrification co-occurred, with denitrification accounting for 97.9 to 98.5% of total N2 production. Recovery of urine-15N in pasture decreased with increasing N rate with 14.7 ± 0.5% and 9.9 ± 0.8% recovered at 400 and 800 kg N ha−1, respectively after 95 days. The N2O/(N2 + N2O) product ratio was generally higher during periods of nitrification of urine-N (the first month after urine application) but with no clear relationship to other measured variables. Contrary to our hypothesis, an elevated urine-N rate did not enhance N2 loss. This is speculated to be due to enhanced ammonia volatilisation and transfer of N as nitrate, to deeper soil layers. Soil relative gas diffusivity indicated that high N2 fluxes resulted from entrapped N2 diffusing from the draining soil

Perturbation-free measurement of in situ di-nitrogen emissions from denitrification in nitrate-rich aquatic ecosystems

Increased production of reactive nitrogen (Nr) from atmospheric di-nitrogen (N2) has greatly contributed to increased food production. However, enriching the biosphere with Nr has also caused a series of negative effects on global ecosystems, especially aquatic ecosystems. The main pathway converting Nr back into the atmospheric N2 pool is the last step in the denitrification process. Despite several attempts, there is still a need for perturbation-free methods for measuring in situ N2 fluxes from denitrification in aquatic ecosystems at the field scale. Such a method is needed to comprehensively quantify the N2 fluxes from aquatic ecosystems. Here we observed linear relationships between the δ15N-N2O signatures and the logarithmically transformed N2O/(N2+N2O) emission ratios. Through independent measurements, we verified that the perturbation-free N2 flux from denitrification in nitrate-rich aquatic ecosystems can be inferred from these linear relationships. Our method allowed the determination of field-scale in situ N2 fluxes from nitrate-rich aquatic ecosystems both with and without overlaying water. The perturbation-free in situ N2 fluxes observed by the new method were almost one order of magnitude higher than those by the sediment core method. The ability of aquatic ecosystems to remove Nr may previously have been severely underestimated

Impact of nitrogen compounds on fungal and bacterial contributions to codenitrification in a pasture soil

Ruminant urine patches on grazed grassland are a significant source of agricultural nitrous oxide (N2o) emissions. Of the many biotic and abiotic N2O production mechanisms initiated following urine-urea deposition, codenitrification resulting in the formation of hybrid N2O, is one of the least understood. Codenitrification forms hybrid N2O via biotic N-nitrosation, co-metabolising organic and inorganic N compounds (N substrates) to produce N2O. The objective of this study was to assess the relative significance of different N substrates on codenitrification and to determine the contributions of fungi and bacteria to codenitrification. 15N-labelled ammonium, hydroxylamine (NH2OH) and two amino acids (phenylalanine or glycine) were applied, separately, to sieved soil mesocosms eight days after a simulated urine event, in the absence or presence of bacterial and fungal inhibitors. Soil chemical variables and n2O fluxes were monitored and the codenitrified N2O fluxes determined. Fungal inhibition decreased N2O fluxes by ca. 40% for both amino acid treatments, while bacterial inhibition only decreased the N2O flux of the glycine treatment, by 14%. Hydroxylamine (NH2OH) generated the highest N2O fluxes which declined with either fungal or bacterial inhibition alone, while combined inhibition resulted in a 60% decrease in the N2O flux. All the N substrates examined participated to some extent in codenitrification. Trends for codenitrification under the NH2OH substrate treatment followed those of total N2O fluxes (85.7% of total N2O flux). Codenitrification fluxes under non-NH2OH substrate treatments (0.7–1.2% of total N2O flux) were two orders of magnitude lower, and significant decreases in these treatments only occurred with fungal inhibition in the amino acid substrate treatments. These results demonstrate that in situ studies are required to better understand the dynamics of codenitrification substrates in grazed pasture soils and the associated role that fungi have with respect to codenitrificatio

Response to nitrogen addition reveals metabolic and ecological strategies of soil bacteria

The nitrogen (N) cycle represents one of the most well-studied systems, yet the taxonomic diversity of the organisms that contribute to it is mostly unknown, or linked to poorly characterized microbial groups. While new information has allowed functional groups to be refined, they still rely on a priori knowledge of enzymes involved and the assumption of functional conservation, with little connection to the role the transformations, plays for specific organisms. Here, we use soil microcosms to test the impact of N deposition on prokaryotic communities. By combining chemical, genomic and transcriptomic analysis, we are able to identify and link changes in community structure to specific organisms catalysing given chemical reactions. Urea deposition led to a decrease in prokaryotic richness, and a shift in community composition. This was driven by replacement of stable native populations, which utilize energy from N-linked redox reactions for physiological maintenance, with fast responding populations that use this energy for growth. This model can be used to predict response to N disturbances and allows us to identify putative life strategies of different functional and taxonomic groups, thus providing insights into how they persist in ecosystems by niche differentiation