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In grazing systems, urine patches deposited by livestock are hotspots of nutrient cycling and the most important source of nitrous oxide (N 2 O) emissions. Studies of the effects of urine deposition, including, for example, the determination of country-specific N 2 O emission factors, require natural urine for use in experiments and face challenges obtaining urine of the same composition, but of differing concentrations. Yet, few studies have explored the importance of storage conditions and processing of ruminant urine for use in subsequent gaseous emission experiments. We conducted three experiments with sheep urine to determine optimal storage conditions and whether partial freeze-drying could be used to concentrate the urine, while maintaining the constituent profile and the subsequent urine-derived gaseous emission response once applied to soil. We concluded that filtering of urine prior to storage, and storage at - 20 °C best maintains the nitrogen-containing constituent profile of sheep urine samples. In addition, based on the 14 urine chemical components determined in this study, partial lyophilisation of sheep urine to a concentrate represents a suitable approach to maintain the constituent profile at a higher overall concentration and does not alter sheep urine-derived soil gaseous emissions.

Agricultural intensification and the subsequent decline of mixed farming systems has led to an increase in continuous cropping with only a few fallow or break years, undermining global soil health. Arable-ley rotations incorporating temporary pastures (leys) lasting 1-4 years may alleviate soil degradation by building soil fertility and improving soil structure. However, the majority of previous research on arable-ley rotations has utilized either grass or grassclover leys within ungrazed systems. Multispecies leys, containing a mix of grasses, legumes, and herbs, are rapidly gaining popularity due to their promotion in agri-environment schemes and potential to deliver greater ecosystem services than conventional grass or grass-clover leys. Livestock grazing in arable-ley rotations may increase the economic resilience of these systems, despite limited research of the effects of multispecies leys on ruminant health and greenhouse gas emissions. This review aims to evaluate previous research on multispecies leys, highlighting areas for future research and the potential benefits and disbenefits on soil quality and livestock productivity. The botanical composition of multispecies leys is crucial, as legumes, deep rooted perennial plants (e.g., Onobrychis viciifolia and Cichorium intybus) and herbs (e.g., Plantago lanceolata) can increase soil carbon, improve soil structure, reduce nitrogen fertilizer requirements, and promote the recovery of soil fauna (e.g., earthworms) in degraded arable soils while delivering additional environmental benefits (e.g., biological nitrification inhibition and enteric methane reduction). Multispecies leys have the potential to deliver biologically driven regenerative agriculture, but more long-term research is needed to underpin evidence-based policy and farmer guidance. [ABSTRACT FROM AUTHOR]

If a dry soil is wetted it can produce a large flush of nitrous oxide (N2O), which can be 10 times above the background rate, this phenomenon is called a hot moment. However, there is uncertainty in the literature regarding the mechanisms that cause hot moments. Therefore, this study aimed to induce two different hot moments on soil cores to investigate changes in nitrogen (N) cycling functional genes, and correlate these changes with subsequent N2O emissions, and transformations in soil chemistry. We induced a 'pre‐dry' treatment which wetted soils from 50% to 90% water‐filled pore space (WFPS) and 'pre‐wet' treatment which wetted soils from 70% to 90% WFPS to analyse changes in gas fluxes (N2O, nitric oxide [NO] and carbon dioxide [CO2]), soil chemistry (pH, dissolved organic carbon [DOC], total oxidised nitrogen [TON], ammonium [NH4+]) and microbiology (bacterial 16S, fungal ITS and nitrogen cycling functional genes associated with denitrification). The soil produced emissions of N2O that were significantly higher for the pre‐dry treatment compared with the pre‐wet treatment (p = <0.001; 9 times that of the pre‐wet). The pre‐dry treatments did not prime the soil any more than the pre‐wet treatments for CO2 and NO emissions, and the results showed that the soils TON and DOC concentrations were significantly higher in the pre‐wet conditions despite its lower N2O emissions. This suggests that the cause of the hot moment was not as dependant on the soil's substrate concentrations as previously thought. Moreover, the amount of denitrification genes in the pre‐dry soil was statistically similar to the pre‐wet (nirK, norB, nifH, nosZI, nxrA), apart from nirS which was marginally higher (p = 0.03). Contrary to other experiments, this study provides evidence that changes in soil substrates and functional gene abundance are not the cause of N2O hot moments. We therefore suggest that future studies should investigate changes in mRNA abundance related to N cycling genes. Highlights: Rewetting following drought caused a significant release of N2O.The pre‐dry soil produced higher N2O fluxes but contained less key substrates (TOC, NH4+ and DOC) than the pre‐wet soil.The pre‐dry and pre‐wet soil had a similar quantity of denitrification functional genes.Differences in soil chemistry and denitrification functional genes between the pre‐dry and pre‐wet soil cannot explain the differences in N2O emissions. [ABSTRACT FROM AUTHOR]

Due to climate change, the frequency and duration of flood events are predicted to increase in many regions of the world. This is expected to cause large changes in soil functioning and to a progressive decline in soil quality such as reduced rates of nutrient cycling, enhanced greenhouse gas emissions and loss of soil biodiversity. There is a knowledge gap, however, on how temperate agricultural soils under different management practices (e.g. manure application) respond to prolonged river or coastal flooding. The main objective of this work was to determine the effects of a simulated prolonged flooding with saline and freshwater on soil N cycling, following application of a low C:N organic amendment (broiler litter) at two temperatures, representative of a winter and a spring flood event. Using laboratory mesocosms we simulated prolonged winter (6 °C) and spring (14 °C) flooding of soil amended with broiler litter. We also compared the effects of inundation with either river (freshwater) or coastal (saline) water. An agricultural grassland soil (Eutric Cambisol) was subjected to different combinations of treatments (flood with fresh or saline water, winter vs spring temperatures, with/without poultry manure). The impact of these treatments on soil solution N dynamics, greenhouse gas emissions (CO 2 , CH 4 , N 2 O) and microbial community structure (by PLFA analysis) were evaluated over an 11 week simulated flood event followed by an 8 week soil recovery period (without flood). Overall, potential losses of NH 4 + and cumulative GHG emissions were increased by flooding and the presence of manure. CH 4 emissions were found to dominate under freshwater flooding conditions and N 2 O under saline flooding. Significant releases of GHG occurred during both flooding and after floodwater removal. Temperature was less influential on regulating GHG under the different treatments. These releases in GHG were associated with disruption in N cycling and changes in soil microbial composition and these changes persisted after floodwater removal. Extreme flooding negatively impacts soil functioning, however, the magnitude of any changes remain critically dependent on flood duration and source of flood water, and management conditions. Further work is required at the field scale to understand the molecular basis of the responses observed in this study. [ABSTRACT FROM AUTHOR]

Nitrification inhibitors are a potential technology to mitigate N 2 O emissions from the urine patches of grazing animals. At present, there is limited information regarding the efficacy of the nitrification inhibitor 3,4-dimethylpyrazole phosphate (DMPP) in reducing N 2 O emissions from ruminant urine patches, as opposed to the well-studied nitrification inhibitor, dicyandiamide. In practical terms, urine patches would be deposited to soil at various times following the application of a nitrification inhibitor to soil. We hypothesised that the effectiveness of DMPP in reducing cumulative N 2 O emissions would decrease the longer the time since DMPP application. This study utilised an automated closed chamber technique, to monitor fluxes of N 2 O from sheep urine patches (725 kg N ha −1 ; 150 ml; 300 cm 2 ) deposited to a Eutric Cambisol, where DMPP was applied (1 kg ha −1 ) on the same day, 2 weeks before and 4 weeks before urine application. Fluxes were monitored continuously from 4 weeks before, to 9 weeks after, urine application. DMPP was found to be ineffective at reducing cumulative N 2 O emissions and 9-week urine-N 2 O emission factors when applied at the same time as the sheep urine, although a low number of replicates were used in this study. Some effect of DMPP in delaying the accumulation of soil NO 3 − was observed, with effects being greater the shorter the time since DMPP application. The temporal dynamics of N 2 O fluxes were also altered where DMPP was applied on the same day as the urine. Heterogeneity in soil conditions were deemed responsible for the large spatial variability of N 2 O emissions observed in this study. The use of Rhizon samplers were useful for detecting spatial variability within the soil solution directly beneath the urine patches (within the flux chambers), which may not have been detected within duplicate urine patches (outside the chambers), where soil sampling was conducted. Further work is required to determine the loading rate and duration of efficacy of DMPP to reduce emissions from urine patches under temperate summer conditions. [ABSTRACT FROM AUTHOR]

Abstract The return of agricultural crop residues are vital to maintain or even enhance soil fertility. However, the influence of application rate of crop residues on denitrification and its related gaseous N emissions is not fully understood. We conducted a fully robotized continuous flow incubation experiment using a Helium/Oxygen atmosphere over 30 days to examine the effect of maize straw application rate on: i) the rate of denitrification, ii) denitrification product stoichiometry N 2 O/(N 2 O+N 2), and iii) the contribution of fungal denitrification to N 2 O fluxes. Five treatments were established using sieved, repacked sandy textured soil; i) non-amended control, ii) nitrate only, iii) low rate of straw + nitrate, iv) medium rate of straw + nitrate, and iv) high rate of straw + nitrate (n = 3). We simultaneously measured NO, N 2 O as well as direct N 2 emissions and used the N 2 O 15N site preference signatures of soil-emitted N 2 O to distinguish N 2 O production from fungal and bacterial denitrification. Uniquely, soil NO 3 − measurements were also made throughout the incubation. Emissions of N 2 O during the initial phase of the experiment (0–13 days) increased almost linearly with increasing rate of straw incorporation and with (almost) no N 2 production. However, the rate of straw amendment was negatively correlated with N 2 O, but positively correlated with N 2 fluxes later in the experimental period (13–30 days). Soil NO 3 − content, in all treatments, was identified as the main factor responsible for the shift from N 2 O production to N 2 O reduction. Straw amendment immediately lowered the proportion of N 2 O from bacterial denitrification, thus implying that more of the N 2 O emitted was derived from fungi (18 ± 0.7% in control and up to 40 ± 3.0% in high straw treatments during the first 13 days). However, after day 15 when soil NO 3 − content decreased to <40 mg NO 3 −-N kg−1 soil, the N 2 O 15N site preference values of the N 2 O produced in the medium straw rate treatment showed a sharp declining trend 15 days after onset of experiment thereby indicating a clear shift towards a more dominant bacterial source of N 2 O. Our study singularly highlights the complex interrelationship between soil NO 3 − kinetics, crop residue incorporation, fungal denitrification and N 2 O/(N 2 O + N 2) ratio. Overall we found that the effect of crop residue applications on soil N 2 O and N 2 emissions depends mainly on soil NO 3 − content, as NO 3 − was the primary regulator of the N 2 O/(N 2 O + N 2) product ratio of denitrification. Furthermore, the application of straw residue enhanced fungal denitrification, but only when the soil NO 3 − content was sufficient to supply enough electron acceptors to the denitrifiers. Highlights • Straw application rate does not directly affect N 2 O/(N 2 O+N 2) ratio. • N 2 O emission following straw application is controlled by soil NO 3 − content. • Straw addition increases fungal N 2 O. [ABSTRACT FROM AUTHOR]

A large number of sheep graze extensively managed grasslands, including upland and hill areas. Excretal deposition of nitrogen (N) to upland soil is a potentially large source of the powerful greenhouse gas (GHG), nitrous oxide (N 2 O), however, few studies have assessed urine-patch N 2 O emissions from upland areas. Current default excretal N 2 O emission factors (EFs) are based on intensively managed lowland systems, with cattle excreta as the N source. We hypothesised that N 2 O emissions could differ substantially from those of lowland systems, due to differences in soil type, climate, topography, pasture composition and management factors along altitudinal and productivity gradients. We investigated N 2 O emission factors across two seasons (spring and autumn), for an extensive semi-improved, temperate grassland using IPCC-compliant and representative sheep urine patches (in terms of urine chemical composition, urine patch size and N loading rates). An automated GHG monitoring system provided high-frequency GHG data from sheep urine patches (756 and 1112 kg N ha −1 applied in spring and autumn, respectively), reference artificial sheep urine patches (1066 and 1004 kg N ha −1 applied in spring and autumn, respectively) and control treatments. In spring, urine patch N 2 O emission factors were −0.02 ± 0.04 (artificial sheep urine) and 0.03 ± 0.09% (real sheep urine) of the applied N; in autumn emission factors were 0.02 ± 0.03 (artificial sheep urine) and 0.08 ± 0.04% (real sheep urine) of the applied N. These values are much lower than default inventory values (1% of applied N) for excreta deposited by grazing livestock. There was a greater pasture foliar N content following urine application in spring as opposed to autumn, and a significantly longer residence time of extractable mineral N in autumn. Our findings demonstrate the importance of generating country-specific N 2 O EFs based on altitude/productivity gradients of livestock production, with implications for national inventories and the accuracy of sustainability metrics of lamb produced in the UK uplands. [ABSTRACT FROM AUTHOR]

Organic manure is widely applied in agricultural systems to improve soil nutrient cycling and other physicochemical properties. However, the biotic and abiotic mechanisms that drive C, N, P, and S cycling following manure application are not completely understood. In this study, soil samples were collected from long-term experimental plots that had been amended with farmyard manure or mineral fertilisers since 1964. Isotope labelling with 15N, 33P, and 35S; metagenomics; and high-throughput sequencing were used to reveal the relationships between C, N, P, and S dynamics and microbial community composition and functions depending on fertilisation. A clear niche differentiation was observed between bacteria and fungi under mineral and manure regimes. A network analysis showed that long-term manure application reduced the complexity and stability of soil microbial network. Furthermore, a variation partitioning analysis based on redundancy analysis indicated that microbial community variation was mainly driven by soil Cand N contents. Dissolved organic C was the most important factor regulating microbial community structure. Soil C and N contents explained 43.5% of bacterial and 37.9% of fungal community variations. In contrast, soil P and S contents explained 29.9% of bacterial and 20.3% of fungal community variations. Long-term manure application increased the abundance of most functional genes related to C, N, P, and S cycling. This led to increased C and N cycling rates under manure application, which provided sufficient substrates for microbial growth. Partial least squares path modelling indicated that soil physicochemical properties, especially dissolved organic carbon, directly influenced C and S cycling, whereas the N and P cycles were indirectly affected by the changes in microbial community composition. These results provide a new perspective on both direct and indirect effects of organic manure and inorganic fertilisers on the soil nutrient cycling processes mediated by soil microbial community. [Display omitted] • Long-term manure application reduced microbial complexity. • Dissolved organic C was the most important factor driving microbial succession. • Functional genes were enriched under manure application. [ABSTRACT FROM AUTHOR]

Efficient fertiliser nitrogen (N) management is critical to global food production and ecosystem health. Considering sheep grazing systems as whole ecosystems, and quantifying key ecosystem services provided by the soil microbial community, including plant N supply and N pollution mitigation, is essential in assessments of N use efficiency (NUE). Using a systems approach, we disassembled a low-intensity sheep (>5 ewe ha−1) grazed grassland, dominated by Lolium perenne , into a series of interlinked 15N-tracer experiments in North Wales during a summer growing season to assess fertiliser-N partitioning. 15N was traced into soil microbial protein-N via compound-specific amino acid 15N-stable isotope probing, with subsequent integration to provide a whole-system perspective. Retention of feed-N into sheep was low (11 %), despite high grass 15N-fertiliser uptake (58 %). The majority of grazed-N re-entered the soil N-cycle as excreta (47 % of total 15N) during the peak growing season. Quantifying 15N-assimilation into soil microbial protein (0–15 cm) demonstrated the central role soil microbes occupy in capturing excess fertiliser (16 %) and urinary-N (8 %) of the total 15N-fertiliser applied, thereby reducing N losses and subsequently supporting plant N supply. This approach emphasises how future management of moderate intensity grazing systems should target sheep NUE, alongside the role of the soil microbial community to retain, and later recycle N, for plant supply, optimising essential ecosystem service provisioning. • Interlinked whole ecosystem 15N tracing study for holistic approach • Mass balance of N cycle through grazing system revealed importance of soil retention to reduce losses. • Microbial assimilation of fertiliser and urine is a major fate of applied 15N. [ABSTRACT FROM AUTHOR]

• Sheep urination data is important for quantifying downstream nitrogen losses. • Assessed frequency, volume and chemical composition of sheep urine events. • Greater N excretion rate for sheep on improved versus semi-improved pasture. • Site and seasonal differences in sheep urine chemical composition observed. • Data will be useful for modelling/experimental studies assessing urine-N losses. Ruminant urine patches are potential sites of reactive nitrogen (N) loss to the environment. Quantification of N losses from grazed grasslands requires measurement of the frequency of urine deposition, as well as its volume and chemical composition. However, studies to date are typically restricted to analyses of few replicate animals and urination events, especially for sheep. Here, we present data on urine frequency, volume, chemical composition (n = 193 events from n = 6 sheep) and metabolomic profile (n = 4–5 events from n = 4–5 sheep) from penned sheep. Differences in urine parameters and chemical composition data were compared seasonally and between two sites (improved and semi-improved pasture). Sheep urinated 8–11 times d−1, assuming time within pens represented a 24 h period. The mean urine event volume recorded was 289 ± 14 mL, from which we estimated a daily urine production value of 2.77 ± 0.15 L urine sheep−1 d−1. Daily urine N excretion and individual urine N concentrations were greater from sheep in improved pasture (26.7 ± 2.3 g N sheep−1 d−1; 7.0 ± 0.2 g N L−1) compared to those in semi-improved pasture (16.7 ± 1.2 g N sheep−1 d−1; 5.5 ± 0.4 g N L−1), but this did not equate to greater individual urine patch N loadings due to site differences in the urine-to-soil surface area influenced (17.5 L m-2 at the semi-improved site and 8.9 L m-2 at the improved site). Urine chemical composition varied seasonally and by site. Site- and season-specific urine should, therefore, be used in studies assessing N losses from urine patches. Based on the urine chemical composition data, we provide an updated artificial sheep urine 'recipe' which could be utilised to replicate natural sheep urine. The urine metabolomic profile clustered according to pasture quality, while clustering according to season was less evident. Our results provide important information for experimental and modelling studies assessing the scale and nature of N pollution arising from sheep-grazed pastures. [ABSTRACT FROM AUTHOR]

Land application of farmyard manure (FYM) is a widespread agronomic practice used to enhance soil fertility, but its long-term effects on soil microbial carbon (C) and nitrogen (N) cycling have not been investigated in detail. Topsoils (0–23 cm) and subsoils (23–38 cm) were collected from a field trial on a sandy-textured soil where FYM had been applied at high (50–25 t ha−1 yr−1, 28 yr) and low rates (10 t ha−1 yr−1, 16 yr), and compared to soil treated only with synthetic NPK fertilisers. The turnover rate of key components of soil organic matter (SOM; proteins, peptides, amino acids, cellulose, and glucose) were evaluated by 14C labelling and measuring cellobiohydrolase, β-glucosidase, β-1,4-N-acetylglucosaminidase, L-leucine aminopeptidase, protease, and deaminase activities, whereas gross NH 4 + and NO 3 − production and consumption were determined by 15N-isotope pool dilution. Microbial communities were determined using phospholipid fatty acid (PLFA) profiling. Our results indicate that long-term FYM addition significantly enhanced the accumulation of soil C and N, soil organic N (SON) turnover, exoenzyme activity, and gross NO 3 − production and assimilation. Rates of protein, peptide, and amino acid processing rate were 169–248, 87–147, and 85–305 mg N kg DWsoil −1 d−1, respectively, gross NH 4 + and NO 3 − production and consumption were 1.8–5.8 mg N kg DWsoil −1 d−1, and the highest rates were shown under the high FYM treatment in topsoil and subsoil. The half-life of cellulose and glucose decomposition under the high FYM treatment were 16.4% and 31.0% lower than them in the synthetic NPK fertiliser treatment, respectively, indicating higher rates of C cycling under high manure application as also evidenced by the higher rate of CO 2 production. This was ascribed to an increase in microbial biomass rather than a change in microbial community structure. Based on the high pool sizes and high turnover rate, this suggests that peptides may represent one of the dominant forms of N taken up by soil microorganisms. We conclude that long-term FYM application builds SOM reserves and induces faster rates of nutrient cycling by boosting microbial biomass rather than by changing its community composition. • The influence of manure application on soil C/N cycling was evaluated. • 14C labeling and 15N-pool dilution techniques were used. • Manure application significantly affects C/N cycling in soil by boosting microbial biomass. • Peptides may be important for terrestrial cycling and supply of N. [ABSTRACT FROM AUTHOR]

Extensively grazed grasslands are understudied in terms of their contribution to greenhouse gas (GHG) emissions from livestock production. Mountains, moorlands and heath occupy 18% of the UK land area, however, in situ studies providing high frequency N 2 O emissions from sheep urine deposited to such areas are lacking. Organic soils typical of these regions may provide substrates for denitrification-related N 2 O emissions, however, acidic and anoxic conditions may inhibit nitrification (and associated emissions from nitrification and denitrification). We hypothesised urine N 2 O-N emission factors (EFs) would be lower than the UK country-specific and IPCC default value for urine, which is based on lowland measurements. Using automated GHG sampling chambers, N 2 O emissions were determined from real sheep urine (930 kg N ha−1) and artificial urine (920 kg N ha−1) applied in summer, and from an artificial urine treatment (1120 kg N ha−1) and a combined NO 3 − and glucose treatment (106 kg N ha−1; 213 kg C ha−1) in autumn. The latter treatment provided an assessment of the soils capacity for denitrification under non-substrate limiting conditions. The artificial urine-N 2 O EF was 0.01 ± 0.00% of the N applied in summer and 0.00 ± 0.00% of the N applied in autumn. The N 2 O EF for real sheep urine applied in summer was 0.01 ± 0.02%. A higher flux was observed in only one replicate of the real urine treatment, relating to one chamber where an increase in soil solution NO 3 − was observed. No lag phase in N 2 O emission was evident following application of the NO 3 − and glucose treatment, which emitted 0.69 ± 0.15% of the N applied. This indicates nitrification rates are the bottle-neck for N 2 O emissions in upland organic soils. We calculated the potential impact of using hill-grazing specific urine N 2 O EFs on the UK inventory of N 2 O emissions from sheep excreta, and found a reduction of ca. 43% in comparison to the use of a country-specific excretal EF. Unlabelled Image • Mountains, moorlands and heath occupy 18% of the UK land area. • These areas are understudied in terms of N 2 O emissions from livestock urine. • N 2 O-N EFs for sheep urine were determined across two seasonal grazing periods. • Low rates of nitrification of urine-N resulted in extremely low N 2 O-N EFs (<0.01%). • National inventory sheep excretal N 2 O-N estimates reduced by 43% using new EFs. [ABSTRACT FROM AUTHOR]