Effects of Amorphous Silica on CO2 and N2O Emissions Mediated by Water-Filled Pore Space in Diverse Agricultural Soils

Effects of Amorphous Silica on CO2 and N2O Emissions Mediated by Water-Filled Pore Space in Diverse Agricultural Soils | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Effects of Amorphous Silica on CO 2 and N 2 O Emissions Mediated by Water-Filled Pore Space in Diverse Agricultural Soils Peter Onyisi Uhuegbue, Mathias Hoffmann, Matthias Lück, Kathrin Grahmann, and 2 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-7348495/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Silicon (Si) is abundant in the Earth’s crust; however, its amorphous form (ASi) is often depleted in agricultural soils. While ASi benefits plant nutrient uptake and growth, its effects on soil pore characteristics, such as water-filled pore space (WFPS), and regulating greenhouse gas (GHG) emissions remain poorly understood. We investigated the effect of ASi addition on soil bulk density, WFPS, and subsequent N 2 O and CO 2 emission dynamics in two soil types of differing texture: Luvisols (moderate silt and clay) and Arenosols (low silt and clay). In a first experiment, we assessed how varying ASi levels affected soil bulk density and WFPS. A second experiment investigated the impact of 1% ASi on N 2 O and CO 2 emissions. ASi addition altered soil bulk density, leading to a decrease in WFPS, especially at 10% ASi in Luvisols. In Arenosols, WFPS increased at 1% ASi before declining at higher rates. The 1% ASi addition increased CO 2 and N 2 O emissions in Luvisols but reduced both in Arenosols. These contrasting outcomes likely reflect a dual effect of ASi: in finer-textured Luvisols, ASi reduces bulk density and increases pore volume, which lowers WFPS under fixed water input, improves aeration, and enhances microbial respiration and nitrification, resulting in increased CO 2 and N 2 O emissions. In coarser-textured Arenosols, ASi may reduce macroporosity by clogging larger pores, resulting in higher WFPS and oxygen limitation, thereby decreasing emissions. Our findings suggest ASi has texture-dependent effects on soil properties and GHG emissions. These outcomes highlight the need for further field-based investigation under natural conditions. Amorphous silica Water-filled-pore space Greenhouse gas emissions bulk density Silicon Figures Figure 1 Figure 2 Figure 3 Figure 4 Introduction Silicon (Si), the second most abundant element in the earth's crust, occurs in many minerals [ 1 , 2 ]. The various silicon fractions present in soils consist of dissolved silicon (either free in the soil solution or adsorbed onto soil minerals), amorphous silica (ASi) such as biogenic forms (e.g., phytoliths, diatom shells, and sponge spicules), or minerogenic forms (e.g., silica nodules and silica in pedogenic oxides) as well as all the Si-containing minerals [ 2 ] However, the amorphous silica (ASi) concentration in agricultural soils is often depleted, typically falling below 1%, and in many cases approaching 0% [ 2 , 3 ]. The depletion of ASi in agricultural soils occurs because most crops accumulate Si, resulting in the removal of large amounts of Si from the field through crop harvest [ 2 , 4 ]. At the same time, unlike essential nutrients, Si is rarely replenished through fertilization, leading to a progressive decline in soil Si contents. Despite not being classified as an essential plant nutrient (Fertilizer and Service; [ 5 ]), Si has been shown to affect soil properties, improve nutrient uptake, and thus enhance plant resilience against biotic and abiotic stresses [ 6 ]. For instance, it has been reported that ASi changes soil physical properties, such as bulk density or porosity, as well as water retention, where the extent of change depends on the initial soil texture [ 7 ]. Moreover, studies found that ASi addition to soil improved N uptake in young rice, olive, maize, as well as wheat plants, leading to increased growth and yield [ 6 ]. Similarly, Hoffmann, et al. [ 8 ] showed that ASi enhanced plant nitrogen uptake, potentially limiting nitrogen availability for microbial processes like nitrification and denitrification, which in turn led to a roughly 30% reduction in N 2 O emissions. Additionally, ASi has been associated with higher CO 2 emissions [ 8 ], likely due to increased biomass production that enhances microbial and root respiration. This is thought to result from ASi boosting carbon input to the soil through root exudates and organic matter decomposition [ 8 ]. However, while previous research has mainly been focused on the role of ASi in plant nutrition and stress tolerance, its effect on soil greenhouse gas (GHG) emissions remains largely unexplored. ASi has a lower bulk density (0.056–0.230 g cm − 3 ) [ 9 ] than most mineral soils (1.3–1.8 g cm − 3 ) [ 10 ]. Its addition can thus reduce soil bulk density, thereby altering water-filled-pore space (WFPS). Changes in these properties can significantly impact GHG emissions by modifying microbial activity and gas diffusion in soils. For instance, a reduction in bulk density may improve soil aeration, potentially influencing N 2 O and CO 2 flux dynamics. Barbosa, et al. [ 7 ] found that ASi improved aggregate stability and modified pore size distribution, increasing meso- and micropore volume in coarse-textured soils, while no such effect was observed in finer-textured soils. While this change in pore structure enhances moisture retention, particularly in coarse-textured soils [ 11 ], it may also affect soil aeration, thereby lowering WFPS compared to soil without ASi under identical water input. As WFPS regulates the balance between aerobic and anaerobic conditions [ 12 , 13 ], changes in WFPS due to ASi can substantially impact N 2 O and CO 2 emissions. In well-aerated soils, CO 2 emissions typically peak around 60% WFPS, where microbial respiration is optimized, and decline at lower ( 60%) WFPS due to oxygen or water limitation [ 12 ]. For N 2 O, different microbial processes may dominate across varying WFPS. Nitrification has its optimum also around 60% WFPS but produces a relatively small amount of N 2 O. With higher WFPS (80–100%), a decrease in oxygen availability favors denitrification (dominant pathway for N 2 O production), resulting in higher N 2 O emissions [ 12 , 14 ]. However, above 80% WFPS, increased oxygen depletion might also result in complete denitrification, converting N 2 O to N 2 [ 14 , 15 ]. Hence, in combination with other factors such as carbon and nitrate availability, as well as microbial community composition, WFPS might affect the N 2 O and N 2 ratio [ 14 , 16 ]. However, in coarse-textured soils with low soil organic carbon (SOC), the typical WFPS-emission relationship and pattern observed in soils with sufficient substrate and microbial activity may be altered, as denitrification and nitrification dynamics may be affected due to limited or a lack of substrate availability (e.g., labile organic carbon and nitrate) and reduced microbial activity. Given the strong impact of WFPS, aeration, and nitrogen availability on microbial processes and GHG emissions, understanding the potential effect of ASi addition is critical, especially in the case of texture-diverse agricultural soils where responses might differ. However, the specific mechanisms by which ASi affects soil physical properties and microbial-driven soil GHG emissions are not yet well understood. While WFPS is widely recognized as a major control of soil GHG emissions [ 12 , 13 ], its effect is strongly shaped by the underlying soil pore structure. For example, Maillet, et al. [ 17 ] demonstrated that soils with lower meso- and macroporosity exhibited higher WFPS and greater N 2 O emission, underlying the critical role of pore connectivity and size distribution in regulating gas exchange and microbial processes. Hence, this study aims to investigate how ASi addition affects soil bulk density, WFPS, and the emission of key GHGs, particularly N 2 O and CO 2 , being most relevant in mineral, agricultural soils. We hypothesize that ASi addition changes soil bulk density and thus i) alters WFPS compared to soils without ASi addition and under equivalent water input and ii) subsequently changes CO 2 and N 2 O emissions. However, the magnitude and direction of these changes may depend on the soil’s texture. To test these hypotheses, two experiments (Experiment I and II) were performed using two different soil types varying in their silt and clay content: Luvisol (modest silt and clay content) and Arenosol (low silt and clay content) sampled at the patchCROP experiment near Tempelberg, Germany. In experiment I, soils were treated without (controls) and with different amounts (1%, 5% and 10%) of ASi to determine the effect of ASi on bulk density and WFPS. In experiment II, soils with 1% ASi and without ASi (control) were incubated to determine the impact of ASi on soil N 2 O and CO 2 emissions. By addressing the impact of ASi addition on bulk density, WFPS, and finally N 2 O and CO 2 emissions, we seek to provide new insights into the potential of ASi-based soil amendments as a measure to modify soil physical conditions and thereby contribute to the mitigation of agricultural N 2 O and CO 2 emissions, depending on soil texture and management. Materials and methods Sampling site For this study, we collected four soil samples within the patchCROP experiment (Fig. 1 ; [ 18 ] at Tempelberg, Germany (52°26'37"N 14°09'39" E). The experimental field covers a total area of 70 ha and features an average annual air temperature of 9.6°C and annual precipitation of 472 mm (ZALF climate station). Its soil mostly comprises high sandy glacial till derived from ground moraine material, with modest variations in silt and clay content. The four sampling locations within the experimental field were specifically chosen for their predominantly sandy texture, as the effects of ASi are most pronounced in such soils, particularly in enhancing soil moisture levels [ 19 ]. Collected soil samples represent variations in silt and clay content of the two dominant soil types present in the experimental field, namely Luvisol with modest silt and clay content and Arenosol with low silt and clay content (Table 1 ). Soil samples from each of the four sampling locations were taken as a mixed topsoil (0 to 30 cm) sample (n = 3) using an auger. After sampling, composite samples of each location were air dried, sieved through a 2 mm mesh, and homogenized for subsequent experiments and analyses. Soil physical and chemical characterization The water holding capacity (WHC) was determined using Luvisol I as a reference soil to represent all four soils. For this, 100 g of air-dried Luvisol I (in three replicates) was weighed into a steel core (250 cm 3 volume) with one of the ends of the core covered with a perforated material that allows water drainage. About 60 ml of water was added to the soil samples, and afterward, soil samples were allowed to drain for 24 hours at a temperature of 22°C. The WHC of the soil was then calculated by taking the difference in the weight of the soil before the addition of water and the final weight after allowing the water to drain for 24 hours. The soil texture analysis was performed using a hydrometer, following a modified Bouyoucos method [ 20 ]. Briefly, 50 g of dry soil (in triplicate) was dispersed in 100 mL of 25% sodium hexametaphosphate (technical Calgon, Bioquim, Montevideo, Uruguay) for 16 hours on a reciprocating shaker. The suspension was then mixed for five minutes using an electric mixer in a Bouyoucos blender cup. After transferring to a 2 L sedimentation cylinder, deionized water was added up to the 2 L mark, and the mixture was manually agitated. The first hydrometer reading, taken after 40 seconds, determined sand content, while the second reading, recorded after two hours, estimated clay content. The silt fraction was calculated as the difference between the sand and clay values. Measurements were performed at 20–22°C, with temperature corrections applied when necessary. Mineral nitrogen (N min ; sum of NH 4 + and NO 3 − ) was determined for both the initial soil samples and soils with and without 1% ASi after incubation (experiment II). For the extraction, 80 mL of 1 M potassium chloride (KCl) solution was added to 20 g of soil each in 100 mL extraction tubes. Afterward, the mixture was shaken for 60 minutes at ~ 20°C. After 30 minutes of shaking, it was filtered using a MACHEREY-NAGEL MN 619 1 / 4 G filter paper (Keeney and Nelson, 1982). Ammonium (NH 4 + ) analysis based on the Bertholet reaction [ 21 ] was determined by mixing 5 mL of the filtrate with Bertholet reagent and was allowed to react for 30 minutes, before measuring the absorbance at 660 nm. For nitrate determination, another 5 mL of the filtrate was treated with a cadmium reduction column to convert NO 3 − to NO 2 − , after which the sample was then mixed with Griess reagent and was allowed to react for 20 minutes before measuring at 540 nm. Both NH 4 + and NO 3 − were measured spectrophotometrically (Gallery Plus; ThermoFisher SCIENTIFIC GmbH). The total concentration of organic carbon (TOC) and total nitrogen (TN) of the original soil samples were analyzed using a TOC analyzer (TNM-L, Shimadzu, Japan). Experiment I: determination of the effect of ASi on bulk density and WFPS Figure 2 illustrates how ASi addition, through alteration of bulk density and thus WFPS, might affect N 2 O and CO 2 emissions as a result of the theoretical relationship between WFPS and microbial activity, adapted from Linn and Doran [ 12 ]. The addition of ASi (Fig. 2 a) may reduce bulk density (Fig. 2 b) and enhance water-holding capacity, thereby lowering WFPS compared to soil without ASi addition. This shift in WFPS, in turn, could modify N 2 O and CO 2 emissions (Fig. 1 c & d) depending on the amount of available water, potentially altering the balance between nitrification, denitrification, and microbial respiration. To determine the potential effect of ASi addition on soil bulk density and WFPS, 50 g of air-dried soil from each sampling location were mixed with ASi at application rates of 0% (control), 1%, 5%, and 10%. Each application rate was replicated three times (n = 3). Soil-ASi mixtures were transferred to a soil steel core with a volume of 250 cm 3 . The volume occupied by the soil-ASi mixture in each vessel was measured, and subsequently, bulk density was determined based on the mass of the air-dry soils. Although disturbed soil samples were used, care was taken to pack the soil to realistic field-like conditions. The resulting density for the control treatments falls within the range typically observed in similar soils under field conditions for undisturbed soil cores [ 22 ], supporting the suitability of the approach for examining treatment effects. To obtain a potential change in WFPS, 8.5 mL of water (i.e., 60% of WHC of Luvisol I—reference soil, which means different WFPS for the four soils) was added to each of the soil-ASi mixtures. Afterward, the WFPS was calculated using Eq. 1. %WFPS = \(\:\left(\frac{{\theta\:}_{v}}{TP}\right)\times\:100\) 1. where \(\:{\theta\:}_{v}\) is the volumetric water content (%), TP is the total pore space calculated as \(\:\left(I-\frac{{P}_{b}}{{P}_{P}}\right)\times\:100\) where \(\:{P}_{b}\) is the soil bulk density (g cm − 3 ) and \(\:{P}_{P}\) is the particle density (~ 2.65 g cm − 3 ). Experiment II: determination of the effect of ASi on CO 2 and N 2 O emissions To assess the effect of ASi addition on CO 2 and N 2 O emissions, we incubated 100 g of soil from each of the four sampling locations for 14 consecutive days, with and without 1% ASi addition. The 1% ASi treatment was chosen because agricultural soils are typically depleted by ~ 1% ASi compared to natural soils, and a difference of 1% ASi is also known to alter soil moisture levels [ 2 , 23 ]. Right before incubation, 17 mL of water (i.e., 60% of WHC) were added to each of the 100 g soil samples with and without ASi mixed in. This water addition was equivalent to 8.5 mL applied to 50 g of soil during the bulk density and WFPS assessment in Experiment I. While we use 100 g of soil in a fixed volume vessel (13 cm in diameter and height) in the incubation experiment (Experiment II), no direct adjustment was made for bulk density. However, the soil-ASi mixture was packed in the incubation vessels to approximate realistic field conditions, following the same procedure as in Experiment I. This ensured consistency in soil structure and porosity across treatments. Each treatment was replicated four times (n = 4). Soil CO 2 and N 2 O emissions were continuously measured using the soil incubation system described in detail by Rillig, et al. [ 24 ]. The system operates in a flow-through steady-state mode [ 25 ]. It consists of 16 airtight, cylindrical incubation vessels (13 cm in diameter and height), constructed from commercially available KG DN sewer pipes and accessories (Marley, Germany), and placed in a temperature-controlled (20°C) box. Ambient air from a pressure vessel flows continuously through the headspace of the incubation vessels at 32 mL min − 1 to a gas analyzer (Picarro G2508; PICARRO, INC., Santa Clara, USA). A control channel allows ambient air to bypass the soil samples and flow directly from the pressure vessel to the gas analyzer at the same rate. To prevent soil samples within the incubation vessels from drying out over incubation time, the incoming air is humidified to 100% relative humidity before entering the incubation vessels. For N 2 O and CO 2 concentration measurements (1 Hz), each channel is sequentially connected to the gas analyzer via a multiplexer and a dedicated circular channel for 7 minutes per cycle. Air circulates between the incubation vessel headspace and the CRDS analyzer at 250 mL min − 1 , driven by a low-leak diaphragm pump (A0702, Picarro, Santa Clara, CA, USA). The multiplexer directs air from each of the 17 measurement channels (16 incubation vessels and 1 control channel) into this circuit. Fluxes were calculated using the ideal gas law Eq. (2) based on measured CO 2 or N 2 O gas concentrations in each channel and the equivalent concentrations in the control channel [ 24 ] \(\:F=\frac{\varDelta\:C*v*\rho\:}{\varDelta\:t*A*R*\left(T+273.15\right)}\) 2. where F is the flux rate (µmol m − 2 s- 1 ), ρ is the atmospheric pressure (Pa), V is the air flow rate into the headspace and the channels (m 3 ), and Δc is the difference of gas concentrations [mol] between the outlet of a specific vessel and the control channel. A is the chamber basal area (m 2 ), R is the gas constant (8.314 m 3 Pa K − 1 mol − 1 ), Δt is the time over which the concentration change was observed, and T is the incubation temperature (K). A modified version of a modular R program script described by [ 26 ] was used to calculate the cumulative N 2 O and CO 2 fluxes over specified time intervals. Statistical analysis and data visualization Fluxes were calculated, and visualization was made using the R computational environment [ 27 ]. The effects of ASi treatments, specifically control (0%), 1%, 5%, and 10% ASi on bulk density and WFPS across the two different soil types were assessed using a one-way analysis of variance (ANOVA). Given the sample size (n = 3, experiment I), the ANOVA assumptions were thoroughly examined to guarantee the validity of the analysis, particularly assessing the normality of residuals using the Shapiro-Wilk test. A Q-Q plot was used to further validate the results of the normality tests. A Levene's test was applied to evaluate the homogeneity of variance between the treatment groups. These diagnostic tests confirmed that the ANOVA assumptions, and homogeneity of variance, were satisfied, justifying the use of the parametric ANOVA approach. Following the ANOVA, a Tukey's post hoc test ( p < 0.05) was used to identify specific differences between treatment groups (control (0%), 1%, 5%, and 10% ASi) for the bulk density and WFPS (experiment I). A permutation test ( p < 0.05) was performed to determine significant differences between measured N 2 O and CO 2 fluxes of the control and ASi-treated soils (experiment II). Results Physicochemical soil properties The studied soil varied in texture (Table 1 ), with Luvisols I and II showing a modest initial silt (22 and 25%) and lower sand content (74% for both) compared to Arenosol I and II, with a lower silt (12 and 11%) and higher sand content (86 and 88%). Adding 1%, 5%, and 10% ASi, respectively, increased silt and clay and reduced sand content in both Luvisol and Arenosol (Table 1 ). Total nitrogen (TN) content was similar for all studied soils of the four sampling locations, while the total organic carbon content (TOC) was slightly higher for the Luvisols (~ 1% – 1.2%) compared to the Arenosols (~ 0.7% – 0.9%). N min measured after incubation did not differ significantly with or without 1% ASi addition, nor between Luvisols and Arenosols of all four sampling locations (Table 1 ). Table 1 Summary of soil physicochemical properties and CO 2 and N 2 O fluxes under different ASi treatments at four sampling locations (Luvisol I and II, Arenosol I and II). Reported are total cumulative CO 2 and N 2 O fluxes (mean ± SD, n = 4), mineral nitrogen content (Nmin; mean ± SD, n = 4), bulk density (g cm − 3 ; mean ± SD, n = 3), and water-filled pore space (WFPS; mean ± SD, n = 3). Soil texture is given as sand, silt, and clay contents, and total nitrogen TN and total organic carbon content (TOC) are provided for each sampling location. TN and TOC refer to untreated soils only. For bulk density and WFPS, statistical comparisons were made between each ASi treatment and its respective control using a post hoc test, while a permutation test was applied for CO 2 and N 2 O to determine the difference between 1% ASi treatment and the respective control treatments. Treatments showing significant differences ( p -value > 0.05) are indicated using different superscript letters. Sampling location Treatment CO 2 flux N 2 O flux NH 4 -N KCl NO 3 -N KCl Bulk density WFPS Sand Silt Clay TOC TN mg CO 2 -C g − 1 µg N 2 O-N g − 1 mg 100 g − 1 mg 100 g − 1 g cm − 3 % % (n = 4) (n = 4) (n = 4) (n = 4) (n = 3) (n = 3) Luvisol I Control 33.6 a ± 12.8 3.1 a ± 1.3 0.2 ± 0.1 1.9 ± 0.8 1.6 a ± 0.0 66 a ± 2.6. 74 25 2 0.9 0.1 ASi − 1% 49.3 b ± 2.6 4.9 a ± 0.8 0.1 ± 0.0 2.3 ± 0.2 1.5 a ± 0.0 63 a ± 2.8 71 26 4 ASi − 5% 1.4 b ± 0.0 52 b ± 2.5 68 26 5 ASi − 10% 0.5 c ± 0.0 11 c ± 0.8 67 27 6 Luvisol II Control 28.8 a ± 2.7 4.3 a ± 3.4 0.1 ± 0.0 4.9 ± 0.2 1.7 a ± 0.0 65 a ± 0.7 74 22 4 1.2 0.1 ASi − 1% 45.5 a ± 31.7 5.2 a ± 3.9 0.1 ± 0.0 4.6 ± 0.1 1.5 a ± 0.0 59 a ± 3.8 74 22 4 ASi − 5% 1.4 b ± 0.0 48 b ± 2.5 70 25 5 ASi − 10% 0.6 b ± 0.0 12 c ± 0.8 69 25 6 Arenosol I Control 68.1 a ± 16.4 3.9 a ± 1.5 0.1 ± 0.0 0.7 ± 0.6 1.5 a ± 0.0 68 a ± 3.5 86 12 2 0.7 0.1 ASi − 1% 37.4 b ± 8.5 2.2 b ± 0.2 0.3 ± 0.3 0.9 ± 0.8 1.6 a ± 0.0 71 a ± 3.0 81 16 3 ASi − 5% 1.6 a ± 0.6 66 a ± 4.6 79 19 5 ASi − 10% 0.6 b ± 0.7 12 b ± 1.6 74 19 6 Arenosol II Control 44.3 a ± 18.5 5.8 a ± 3.6 0.1 ± 0.0 3.0 ± 0.2 1.7 a ± 0.0 82 a ± 5.3 88 11 1 0.9 0.1 ASi − 1% 25.5 b ± 2.5 5.3 a ± 4.7 0.1 ± 0.0 3.1 ± 0.1 1.8 a ± 0.2 95 b ± 4.3 87 12 1 ASi − 5% 1.6 b ± 0.6 63 c ± 5.2 81 12 6 ASi − 10% 0.6 c ± 0.1 12 d ± 2.6 78 13 9 Effect of ASi addition on bulk density and WFPS Experiment I showed that the addition of 1%, 5%, and 10% ASi altered bulk density and WFPS in both Arensol and Luvisol. In Luvisol I and II, a slight but not significant decrease in bulk density due to 1% ASi addition was observed (Table 1 ), with a more pronounced, significant decrease ( p < 0.05) observed at both 5% and 10% ASi addition, respectively. In contrast, Arenosol I and II showed a slight but not significant increase in bulk density due to 1% ASi addition compared to the control treatment. However, similar to Luvisol I and II, bulk density decreased following 5% and 10% ASi addition treatments. This decrease was statistically significant for Arenosol I at both 5% and 10%, and for Arenosol II at 10% ASi addition (Table 1 ). These changes in bulk density were accompanied by an observed change in WFPS. In Luvisol I and II a slight but not significant decrease in WFPS by 4.5% and 8.9% through the addition of 1% ASi was observed when compared to soil samples without ASi addition (Table 1 ; Fig. 3 ). A more pronounced and significant ( p < 0.05) decrease in WFPS, however, was obtained when 5% and 10% ASi were added to Luvisol I and II, decreasing WFPS to 51% and 11.4% for Luvisol I, and 48.1% and 11.8% for Luvisol II, when compared to soil samples without ASi addition, respectively (Table 1 ). In contrast to that, the addition of 1% ASi increased WFPS in both Arenosols, though only Arenosol II showed a significant increase from 82.2–94.5%, whereas Arenosol I showed a non-significant increase from 67.5–70.5% (Table 1 ; Fig. 3 ). However, at 5% and 10% ASi, admixture, WFPS decreased in both Arenosols, similar to the studied Luvisols, even though that decrease was only significant for 10% ASi addition at Arenosol I and II (Table 1 ). Effect of ASi addition on N 2 O and CO 2 emissions We found clear differences in cumulative N 2 O and CO 2 emissions due to 1% ASi addition in both soil types during experiment II. These differences, however, varied with ASi addition in Arenosol, reducing and in Luvisol, increasing the emissions. Figure 4 a-b shows the mean N 2 O and CO 2 emissions for soils from the four sampling locations without and with the addition of 1% ASi. Figures 4 c–d illustrate the theoretical relationship between microbial activity (%) and water-filled pore space (WFPS, 0–100%) alongside the same measured cumulative N 2 O and CO 2 emissions for soils treated with and without 1% ASi. Figures 4 e–f depict the pore network characteristics for Arenosols and Luvisols with and without 1% ASi addition. In both soils, the representation for soils treated with 1% ASi shows a visibly reduced pore space compared to the corresponding controls, with fewer and smaller macro-and micropores evident in the ASi-treated samples. In Arenosols, the addition of ASi predominantly clogged the abundant meso and macropores, whereas in Luvisols, which naturally have fewer macropores, ASi not only clogged existing pores but also contributed to the formation of new ones (Fig. 4 e & f). In Arenosols, N 2 O emissions decreased at increased WFPS under 1% ASi treatment, though only Arenosol I showed a significant ( p < 0.05) reduction by 44.3%, while Arenosol II showed a non-significant reduction by 8% compared to Arenosol without ASi addition (Table 1 ; Fig. 4 a & c). In contrast to both Arenosols, Luvisol I and II at decreased WFPS showed a non-significant increase in N 2 O emissions with the addition of 1% ASi by 57% and 18%, respectively (Table 1 ; Fig. 4 a & c). Although the differences in N 2 O emissions between the ASi-treated soil and the control were not significant across all sampling locations, a trend was evident for the two soil types: decreasing N 2 O emissions for Arenosol and increasing N 2 O emissions for Luvisol due to the addition of 1% ASi. In case of CO 2 , we found a significant ( p < 0.05) decrease in CO 2 emissions due to 1% ASi addition (resulting in an increased WFPS) by 45% and 42% for Arenosol I and II, respectively (Table 1 ; Fig. 4 b & d). In contrast, Luvisol I and II showed an increase in CO 2 emissions by 46.8%, which was only significant ( p < 0.05) in the case of Luvisol I, following 1% ASi addition despite only a slight decrease in WFPS (Table 1 ; Fig. 4 b & d). Discussion This study investigated the effect of ASi on soil bulk density, WFPS, and subsequent N 2 O and CO 2 emissions in soils with different silt contents. Our findings show that ASi addition to soil modifies their bulk density and WFPS. Findings of experiment I support our hypothesis that ASi addition alters the soil’s WFPS through influencing the soil's bulk density. However, contrasting effects were found depending on the initial soil texture (Table 1 ; Fig. 3 ). We found that ASi reduced WFPS in the finer-textured Luvisol soils (with modest silt and clay content and more micropores). In contrast, in the coarser-textured Arenosol soils (with low silt and clay content and less micropores), ASi appeared to increase WFPS. Thus, the direction and magnitude of ASi-induced changes in WFPS are not uniform but depend on the soil’s initial texture and underlying pore structure. The reason for this is likely due to two contrasting effects through which ASi alters bulk density and thus, WFPS: i) when added to finer-texture Luvisol soils, ASi decreased the bulk density likely due to its low density and high internal particle porosity by clogging the few mesopores (with less ASi) while predominantly creating new mesopores. These may have reduced the packing efficiency and increasing the total pore volume of the soil (Fig. 4 f), this in turn may lower the percentage of the pores filled with water at fixed water input, thus lowering the WFPS, ii) when added to coarse-textured Arenosol soils, ASi may reduce macroporosity by clogging larger pores, thereby enhancing the bulk density and WFPS [ 7 ]. Depending on the amount of added ASi and which of these effects dominates at a particular soil texture, adding ASi can either increase or decrease bulk density, thus affecting WFPS. Texture analysis showed that ASi behaves like silt- and clay-sized particles, increasing the fine fraction while proportionally reducing sand content (Table 1 ). This shift towards finer texture likely reduced macroporosity in sandy Arenosol (Fig. 4 e), thereby increasing WFPS. In contrast, in the already finer-textured Luvisols, ASi slightly lowers WFPS, potentially due to changes in pore structure, such as improved pore volume (Fig. 4 f) that lowers WFPS under fixed water input. The resulting aeration shifts match the observed decrease (Arenosols) or increase (Luvisols) in CO 2 and N 2 O emissions observed during experiment II. We found that 1% ASi addition to Arenosols caused an increase in bulk density, likely due to ASi clogging meso- and macropores, thereby enhancing bulk density. In contrast, Luvisols showed a decrease in bulk density, which may be attributed to the low density and high internal porosity of ASi particles reducing packing efficiency and increasing total pore volume. Neither the increase nor the decrease in bulk density at 1% ASi treatment was significant (Table 1 ). However, at increased ASi levels (10%), both Arenosols and Luvisols showed a significant reduction in both bulk density and WFPS, suggesting that higher ASi levels may have increased the proportion of finer pores or enhanced internal porosity within soil aggregates, leading to a decrease in bulk density and WFPS. In our second hypothesis, we proposed that ASi-induced WFPS changes via bulk density alteration would affect N 2 O and CO 2 emission dynamics. In experiment II, contrasting N 2 O and CO 2 emissions responses in Arenosols and Luvisols following ASi treatment were found, confirming the second hypothesis. The variations can be attributed to differences in soil texture that influence how ASi alters pore structure and bulk density, subsequently modulating the WFPS and oxygen availability. For example, the incorporation of ASi in Arenosols resulted in an increase in WFPS, from 67.5–70.4% in Arenosol I and from 82.2–94.5% in Arenosol II (Fig. 4 c & d). This rise in WFPS may be attributed to a reduction in mesopore and macropore volume due to ASi addition. These changes likely limit oxygen availability and diffusion within the soil, thereby inhibiting microbial respiration, essential oxygen-dependent pathways for CO 2 production, supporting the decrease in CO 2 emissions in Arenosol (Fig. 4 b & d). These findings are in line with established relationships indicating that microbial activity and GHG emissions decrease at high WFPS in response to oxygen limitation [ 12 , 13 ]. We showed that N 2 O emissions were highest at elevated WFPS (~ 80%) compared to those at lower WFPS (~ 60), irrespective of treatment (Fig. 4 c). This aligns with expectations that denitrification is enhanced under high WFPS, a condition that typically generates more N 2 O than nitrification processes occurring at lower WFPS [ 28 ]. This could explain the overall higher N 2 O emissions at elevated WFPS in our study for both control and ASi-treated soil, particularly in Arenosol II (Fig. 4 a & c). However, with ASi treatment, further increases in WFPS beyond ~ 80% (up to ~ 95%) were associated with a slight reduction in N 2 O emissions compared to the control (Fig. 4 a & c), even though emissions at these high WFPS remained greater than at lower WFPS. This pattern suggests that while denitrification was a key contributor to the high N 2 O flux at elevated WFPS, the more extreme oxygen-limited condition induced by ASi at > 80% WFPS may have promoted complete denitrification, thereby shifting nitrogen transformations from N 2 O toward dinitrogen (N 2 ) production [ 12 , 14 ]. This shift in WFPS probably played a significant role in substantially reducing overall N 2 O emissions found in Arenosols over time. This suggests that beyond a certain threshold, an increase in WFPS could limit oxygen-dependent microbial processes, thereby reducing both CO 2 and N 2 O emissions. Maillet, et al. [ 17 ] reported that soils with lower meso-and macroporosity, associated with WFPS, showed higher N 2 O emissions. In our study, this pattern was not uniform: in Luvisol, at lower WFPS under ASi treatment coincided with higher N 2 O emission compared to the untreated control (Fig. 4 A & C), whereas in Arenosol, under higher WFPS, due to ASi likely clogging meso- and macropores, coincided with lower N 2 O emission. These differences highlight that although pore structure strongly affects emissions, the direction of response depends on the initial texture and how ASi alters the pore network or structure In contrast, the addition of ASi to Luvisol soils resulted in a decrease in WFPS, from 65.7–62.7% in Luvisol I and from 64.6–58.8% in Luvisol II (Fig. 4 a & c). This adjustment in WFPS moved the conditions closer to the optimal WFPS range (40–60%) for microbial nitrification and aerobic respiration [ 12 ]. However, N 2 O emissions were generally lower in Luvisols at these lower WFPS compared to Arenosols (Fig. 4 c), which, under higher WFPS, produced the highest N 2 O emission irrespective of the treatment. The reduction in WFPS found for Luvisols (Fig. 4 a & c) may have improved soil pore volume and enhanced aeration. These changes probably facilitated oxygen diffusion, which may have promoted aerobic microbial activity, thereby increasing N 2 O and CO 2 emissions. For Luvisols, no significant increase in N 2 O emissions was observed; the small sample size (n = 4) may have reduced statistical power; increasing the sample size could demonstrate differences, through increased sensitivity, however. While ASi may contribute to lower N 2 O emissions by altering bulk density and WFPS, improved plant nitrogen uptake [ 8 ] could further reduce available mineral N for microbial nitrification and denitrification, being a potential way to make agriculture more sustainable [ 3 ]. In conclusion, our study particularly highlights ASi's effect on soil bulk density and WFPS, which in turn affects the N 2 O and CO 2 emissions by shifting towards or from optimal conditions for microbial respiration, nitrification, and denitrification. Under our experimental conditions, the highest N 2 O emissions coincided with the highest WFPS (> 80%), indicating that denitrification contributed substantially to N 2 O production under high WFPS. It is still not clear, however, what the observed effect of ASi on WFPS and associated N 2 O and CO 2 emissions would mean under field conditions. ASi may decrease or increase the WFPS in the field depending on the initial soil texture, and this may either raise or lower N 2 O emissions. Although denitrification was evident under high WFPS in our study, its magnitude and relevance could vary under field conditions, where additional factors such as high organic matter availability may further impact this pathway. In addition, other ASi-induced processes, such as enhanced crop growth, improved N uptake, and interactions with variable water availability and precipitation, may dominate or counteract the mechanisms observed under controlled conditions. These interconnected processes highlight the complexity of ASi's role in real-world settings and the need for further field-based research, since optimizing the use of ASi as a soil amendment requires an understanding of the relationships among microbial activity, structural changes caused by ASi, and GHG emission dynamics. Declarations Acknowledgements We thank Oscar Monzon for help with analysis and Björn Gustav Wang and Felix Erbe for help during soil sampling and map visualization. Funding The overall project was funded by the Deutsche Forschungsgemeinschaft for funding (SCHA 1822/14-1 and KA 1737/21-1). Funding for the infrastructure of the patchCROP experiment was provided by the Leibniz Centre for Agricultural Landscape Research (ZALF) and the German Research Foundation under Germany’s Excellence Strategy (EXC-2070–390732324 – PhenoRob). KG acknowledges support from BMBF for the Junior Research Group SoilRob (Grant 031B1391). Ethics, Consent to Participate, and Consent to Publish declarations : Not applicable. Clinical trial number: Not applicable. Author contributions: P.U. was responsible for conceptualization, investigation, data curation, data analysis, visualization, and writing (original draft). M.H. contributed to experimental design, data validation, visualization, and writing (review and editing). M.L. contributed to experimental monitoring. K.G. contributed to visualization and writing (review and editing). K.K. contributed to writing (review and editing). J.S. contributed to experimental design and writing (review and editing). All authors reviewed the manuscript and approved the final version for submission. Competing Interests: All authors certify that they have no affiliations with or involvement in any organization or entity with any financial interest or non-financial interest in the subject matter or materials discussed in this manuscript. Dual Publication: The results/data/figures in this manuscript have not been published elsewhere, nor are they under consideration (from you or one of your Contributing Authors) by another publisher. Data Availability: I do not have any research data outside the submitted manuscript file. 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Schaller, J, Hines, J, Brackhage, C, Bäucker, E, and Gessner, M O, "Silica decouples fungal growth and litter decomposition without changing responses to climate warming and N enrichment," Ecology, vol. 95, no. 11, pp. 3181-3189, 2014, doi: 10.1890/13-2104.1. Maillet, E, Grossel, A, Cousin, I, Arbaret, L, Cottenot, L, and Lacoste, M, "What is the most relevant soil structure parameter to describe field-measured N2O emissions?," Geoderma, vol. 453, p. 117155, 2025. Grahmann, K, Reckling, M, Hernández-Ochoa, I, Donat, M, Bellingrath-Kimura, S, and Ewert, F, "Co-designing a landscape experiment to investigate diversified cropping systems," Agricultural Systems, vol. 217, p. 103950, 2024. Zarebanadkouki, M, Al Hamwi, W, Abdalla, M, Rahnemaie, R, and Schaller, J, "The effect of amorphous silica on soil-plant-water relations in soils with contrasting textures," Sci Rep, vol. 14, no. 1, p. 10277, May 4 2024, doi: 10.1038/s41598-024-60947-1. Beretta, A N et al. , "Soil texture analyses using a hydrometer: modification of the Bouyoucos method," Ciencia e investigación agraria, vol. 41, no. 2, pp. 25-26, 2014, doi: 10.4067/s0718-16202014000200013. Weatherburn, M, "Phenol-hypochlorite reaction for determination of ammonia," Analytical chemistry, vol. 39, no. 8, pp. 971-974, 1967. Engels, A M, Gaiser, T, Ewert, F, Grahmann, K, and Hernández-Ochoa, I, "Simulating soil moisture dynamics in a diversified cropping system under heterogeneous soil conditions," Agronomy, vol. 15, no. 2, p. 407, 2025. Schaller, J, Cramer, A, Carminati, A, and Zarebanadkouki, M, "Biogenic amorphous silica as main driver for plant available water in soils," Sci Rep, vol. 10, no. 1, p. 2424, Feb 12 2020, doi: 10.1038/s41598-020-59437-x. Rillig, M C, Hoffmann, M, Lehmann, A, Liang, Y, Lück, M, and Augustin, J, "Microplastic fibers affect dynamics and intensity of CO 2 and N 2 O fluxes from soil differently," Microplastics and Nanoplastics, vol. 1, pp. 1-11, 2021. Livingston, G, "Enclosure-based measurement of trace gas exchange: applications and sources of error," Methods in ecology: biogenic trace gas emissions from soil and water, pp. 14-17, 1994. Hoffmann, M et al. , "Automated modeling of ecosystem CO2 fluxes based on periodic closed chamber measurements: A standardized conceptual and practical approach," Agricultural and Forest Meteorology, vol. 200, pp. 30-45, 2015. R Core Team, R, "R: A language and environment for statistical computing," 2013. Davidson, E A, Keller, M, Erickson, H E, Verchot, L V, and Veldkamp, E, "Testing a conceptual model of soil emissions of nitrous and nitric oxides: using two functions based on soil nitrogen availability and soil water content, the hole-in-the-pipe model characterizes a large fraction of the observed variation of nitric oxide and nitrous oxide emissions from soils," Bioscience, vol. 50, no. 8, pp. 667-680, 2000. Additional Declarations No competing interests reported. Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. We do this by developing innovative software and high quality services for the global research community. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-7348495","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":513078793,"identity":"4208f39d-8d36-48d7-a82e-6d8dbadb1ec0","order_by":0,"name":"Peter Onyisi Uhuegbue","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAA+ElEQVRIiWNgGAWjYBCDBCBmA2IbBgZ2xgcMPHgVM6NoSQMKMBuQpOUwYS3m7P0HH1cwHM4zON5+7TFPxXl7/magIW8qcGux7DnMbHiG4XCxwZkz5cY8Z24zSxxmZmCccwa3FoMbyWySDQyHEzfcyEmT5m27zcZwmP8AM28bHi33H7P/BGu5/wao5d85HnmgLcy8//DZwszGCLGF/Zg0b8MBCQOwlgZ8fkk2lmwwSE+ceSaH3XDOsWQDQ6CWg3OO4dZizn7w4ceGCuvEvuPHnz14U2NnL3e8mRHIwOMwBMljABc9gFsDTAsYsD/Ap3AUjIJRMApGMAAAXTVQLemFY1kAAAAASUVORK5CYII=","orcid":"","institution":"Leibniz Center for Agricultural Landscape Research (ZALF)","correspondingAuthor":true,"prefix":"","firstName":"Peter","middleName":"Onyisi","lastName":"Uhuegbue","suffix":""},{"id":513078794,"identity":"c67c2e06-0744-4a7e-a449-a6a66f546f58","order_by":1,"name":"Mathias Hoffmann","email":"","orcid":"","institution":"Leibniz Center for Agricultural Landscape Research (ZALF)","correspondingAuthor":false,"prefix":"","firstName":"Mathias","middleName":"","lastName":"Hoffmann","suffix":""},{"id":513078795,"identity":"ed2b89fe-f0d3-4f03-87c1-1fc410a91df9","order_by":2,"name":"Matthias Lück","email":"","orcid":"","institution":"Leibniz Center for Agricultural Landscape Research (ZALF)","correspondingAuthor":false,"prefix":"","firstName":"Matthias","middleName":"","lastName":"Lück","suffix":""},{"id":513078796,"identity":"d71467d0-fad6-40af-bc48-29b0edaf4994","order_by":3,"name":"Kathrin Grahmann","email":"","orcid":"","institution":"Leibniz Centre for Agricultural Landscape Research (ZALF)","correspondingAuthor":false,"prefix":"","firstName":"Kathrin","middleName":"","lastName":"Grahmann","suffix":""},{"id":513078797,"identity":"8eadac9b-dcfb-441b-b355-8a3c9b09fc11","order_by":4,"name":"Karsten Kalbitz","email":"","orcid":"","institution":"Technische Universität Dresden Soil Resources and Land Use","correspondingAuthor":false,"prefix":"","firstName":"Karsten","middleName":"","lastName":"Kalbitz","suffix":""},{"id":513078798,"identity":"ae14c328-b811-4406-bf87-eb616284c395","order_by":5,"name":"Jörg Schaller","email":"","orcid":"","institution":"Leibniz Center for Agricultural Landscape Research (ZALF)","correspondingAuthor":false,"prefix":"","firstName":"Jörg","middleName":"","lastName":"Schaller","suffix":""}],"badges":[],"createdAt":"2025-08-11 16:53:20","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-7348495/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-7348495/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":91073260,"identity":"9273eafa-af59-4573-95b9-7ed11b691660","added_by":"auto","created_at":"2025-09-11 10:57:40","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":518038,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003eSoil sampling locations (red crosses) within the patchCROP experiment. The soil map depicts silt content (in %) in the top 30 cm soil depth based on proximal soil texture mapping. S-21 and S-73 represent the Luvisol I and II (modest silt content), while S-59 and S-119 represent the Arenosol I and II (low silt content) soil sampling locations.\u003c/em\u003e\u003c/p\u003e","description":"","filename":"floatimage1.png","url":"https://assets-eu.researchsquare.com/files/rs-7348495/v1/ec42d4bc91466df2f5847f65.png"},{"id":91070425,"identity":"ac981266-cc36-4e04-be69-a4b68fa3d08d","added_by":"auto","created_at":"2025-09-11 10:41:40","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":269089,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003ea) Scanning electron microscopy (SEM) image of amorphous silica (ASi); b) sample preparation (ASi addition) for Experiments 1 and II; c) theoretical impact adapted from Linn and Doran [12]of ASi on N₂O emissions; d) theoretical impact of ASi on CO₂emissions. Both c) and d) illustrate the proposed effect of ASi on GHG emission via its influence on soil bulk density and WFPS.\u003c/em\u003e\u003c/p\u003e","description":"","filename":"floatimage2.png","url":"https://assets-eu.researchsquare.com/files/rs-7348495/v1/d4441ed867aed0872f1f9135.png"},{"id":91070421,"identity":"f0f5b5d7-7a60-4401-a804-6ffa069112de","added_by":"auto","created_at":"2025-09-11 10:41:40","extension":"png","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":130781,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003eRelationship between silt content and water-filled pore space (WFPS) for the Arenosol and Luvisol of the four sampling locations with (1%; orange) and without ASi addition (Control; black). Data points represent mean WFPS per sampling location with error bars indicating \u003c/em\u003e± SD.\u003c/p\u003e","description":"","filename":"floatimage3.png","url":"https://assets-eu.researchsquare.com/files/rs-7348495/v1/d4fb450c1e89ff4d70911247.png"},{"id":91070424,"identity":"29927077-3fd7-4fb2-a1b9-7954f1f294a9","added_by":"auto","created_at":"2025-09-11 10:41:40","extension":"png","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":337274,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cem\u003eEffects of ASi on soil structure and consequent greenhouse gas fluxes in Arenosols and Luvisols. The bars represent (a) mean N₂O flux (µg N₂O–N g\u003c/em\u003e\u003csup\u003e\u003cem\u003e-1\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e soil), and (b) mean CO₂ flux (mg CO₂–C g\u003c/em\u003e\u003csup\u003e\u003cem\u003e-1\u003c/em\u003e\u003c/sup\u003e\u003cem\u003e soil) under control (C) and 1% ASi treatments for Arenosol I \u0026amp; II and Luvisol I \u0026amp; II. Dark purple bars = control, light purple bars = 1% ASi in (a); dark red bars = control, light red bars = 1% ASi in (b). Error bars indicate ± SE, and asterisks (**) denote significant differences (p \u0026lt; 0.05; n.s = not significant). (c) cumulative N₂O flux as a function of WFPS alongside relative microbial activity; data points follow the same color scheme as in (a), with dark purple for control and light purple for 1% ASi. (d) cumulative CO₂ flux as a function of WFPS alongside relative microbial activity (%); data points follow the same color scheme as in (b), with dark red for control and light red for 1% ASi. The conceptual pore network diagrams for Arenosols (e) and Luvisols (f): the yellow outlined zones indicate soils treated with 1% ASi, compared with untreated controls.\u003c/em\u003e\u003c/p\u003e","description":"","filename":"floatimage4.png","url":"https://assets-eu.researchsquare.com/files/rs-7348495/v1/7c31de5b9c4256522f533043.png"},{"id":92695004,"identity":"d95b7568-d558-4be6-8752-1d893e93918d","added_by":"auto","created_at":"2025-10-03 06:39:19","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":2221121,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-7348495/v1/9b165983-b296-4286-becd-b31e411b230c.pdf"}],"financialInterests":"No competing interests reported.","formattedTitle":"\u003cp\u003eEffects of Amorphous Silica on CO\u003csub\u003e2\u003c/sub\u003e and N\u003csub\u003e2\u003c/sub\u003eO Emissions Mediated by Water-Filled Pore Space in Diverse Agricultural Soils\u003c/p\u003e","fulltext":[{"header":"Introduction","content":"\u003cp\u003eSilicon (Si), the second most abundant element in the earth's crust, occurs in many minerals [\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e, \u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]. The various silicon fractions present in soils consist of dissolved silicon (either free in the soil solution or adsorbed onto soil minerals), amorphous silica (ASi) such as biogenic forms (e.g., phytoliths, diatom shells, and sponge spicules), or minerogenic forms (e.g., silica nodules and silica in pedogenic oxides) as well as all the Si-containing minerals [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e]\u003c/p\u003e\u003cp\u003eHowever, the amorphous silica (ASi) concentration in agricultural soils is often depleted, typically falling below 1%, and in many cases approaching 0% [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e]. The depletion of ASi in agricultural soils occurs because most crops accumulate Si, resulting in the removal of large amounts of Si from the field through crop harvest [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e]. At the same time, unlike essential nutrients, Si is rarely replenished through fertilization, leading to a progressive decline in soil Si contents. Despite not being classified as an essential plant nutrient (Fertilizer and Service; [\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e]), Si has been shown to affect soil properties, improve nutrient uptake, and thus enhance plant resilience against biotic and abiotic stresses [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. For instance, it has been reported that ASi changes soil physical properties, such as bulk density or porosity, as well as water retention, where the extent of change depends on the initial soil texture [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Moreover, studies found that ASi addition to soil improved N uptake in young rice, olive, maize, as well as wheat plants, leading to increased growth and yield [\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e]. Similarly, Hoffmann, et al. [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e] showed that ASi enhanced plant nitrogen uptake, potentially limiting nitrogen availability for microbial processes like nitrification and denitrification, which in turn led to a roughly 30% reduction in N\u003csub\u003e2\u003c/sub\u003eO emissions. Additionally, ASi has been associated with higher CO\u003csub\u003e2\u003c/sub\u003e emissions [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e], likely due to increased biomass production that enhances microbial and root respiration. This is thought to result from ASi boosting carbon input to the soil through root exudates and organic matter decomposition [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e]. However, while previous research has mainly been focused on the role of ASi in plant nutrition and stress tolerance, its effect on soil greenhouse gas (GHG) emissions remains largely unexplored.\u003c/p\u003e\u003cp\u003eASi has a lower bulk density (0.056\u0026ndash;0.230 g cm\u003csup\u003e\u0026minus;\u0026thinsp;3\u003c/sup\u003e) [\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e] than most mineral soils (1.3\u0026ndash;1.8 g cm\u003csup\u003e\u0026minus;\u0026thinsp;3\u003c/sup\u003e) [\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e]. Its addition can thus reduce soil bulk density, thereby altering water-filled-pore space (WFPS). Changes in these properties can significantly impact GHG emissions by modifying microbial activity and gas diffusion in soils. For instance, a reduction in bulk density may improve soil aeration, potentially influencing N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e flux dynamics.\u003c/p\u003e\u003cp\u003eBarbosa, et al. [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e] found that ASi improved aggregate stability and modified pore size distribution, increasing meso- and micropore volume in coarse-textured soils, while no such effect was observed in finer-textured soils. While this change in pore structure enhances moisture retention, particularly in coarse-textured soils [\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e], it may also affect soil aeration, thereby lowering WFPS compared to soil without ASi under identical water input. As WFPS regulates the balance between aerobic and anaerobic conditions [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e], changes in WFPS due to ASi can substantially impact N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions. In well-aerated soils, CO\u003csub\u003e2\u003c/sub\u003e emissions typically peak around 60% WFPS, where microbial respiration is optimized, and decline at lower (\u0026lt;\u0026thinsp;60%) or higher (\u0026gt;\u0026thinsp;60%) WFPS due to oxygen or water limitation [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. For N\u003csub\u003e2\u003c/sub\u003eO, different microbial processes may dominate across varying WFPS. Nitrification has its optimum also around 60% WFPS but produces a relatively small amount of N\u003csub\u003e2\u003c/sub\u003eO. With higher WFPS (80\u0026ndash;100%), a decrease in oxygen availability favors denitrification (dominant pathway for N\u003csub\u003e2\u003c/sub\u003eO production), resulting in higher N\u003csub\u003e2\u003c/sub\u003eO emissions [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. However, above 80% WFPS, increased oxygen depletion might also result in complete denitrification, converting N\u003csub\u003e2\u003c/sub\u003eO to N\u003csub\u003e2\u003c/sub\u003e [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e, \u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e]. Hence, in combination with other factors such as carbon and nitrate availability, as well as microbial community composition, WFPS might affect the N\u003csub\u003e2\u003c/sub\u003eO and N\u003csub\u003e2\u003c/sub\u003e ratio [\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e, \u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e]. However, in coarse-textured soils with low soil organic carbon (SOC), the typical WFPS-emission relationship and pattern observed in soils with sufficient substrate and microbial activity may be altered, as denitrification and nitrification dynamics may be affected due to limited or a lack of substrate availability (e.g., labile organic carbon and nitrate) and reduced microbial activity.\u003c/p\u003e\u003cp\u003eGiven the strong impact of WFPS, aeration, and nitrogen availability on microbial processes and GHG emissions, understanding the potential effect of ASi addition is critical, especially in the case of texture-diverse agricultural soils where responses might differ. However, the specific mechanisms by which ASi affects soil physical properties and microbial-driven soil GHG emissions are not yet well understood. While WFPS is widely recognized as a major control of soil GHG emissions [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e], its effect is strongly shaped by the underlying soil pore structure. For example, Maillet, et al. [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e] demonstrated that soils with lower meso- and macroporosity exhibited higher WFPS and greater N\u003csub\u003e2\u003c/sub\u003eO emission, underlying the critical role of pore connectivity and size distribution in regulating gas exchange and microbial processes.\u003c/p\u003e\u003cp\u003eHence, this study aims to investigate how ASi addition affects soil bulk density, WFPS, and the emission of key GHGs, particularly N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e, being most relevant in mineral, agricultural soils. We hypothesize that ASi addition changes soil bulk density and thus i) alters WFPS compared to soils without ASi addition and under equivalent water input and ii) subsequently changes CO\u003csub\u003e2\u003c/sub\u003e and N\u003csub\u003e2\u003c/sub\u003eO emissions. However, the magnitude and direction of these changes may depend on the soil\u0026rsquo;s texture. To test these hypotheses, two experiments (Experiment I and II) were performed using two different soil types varying in their silt and clay content: Luvisol (modest silt and clay content) and Arenosol (low silt and clay content) sampled at the patchCROP experiment near Tempelberg, Germany. In experiment I, soils were treated without (controls) and with different amounts (1%, 5% and 10%) of ASi to determine the effect of ASi on bulk density and WFPS. In experiment II, soils with 1% ASi and without ASi (control) were incubated to determine the impact of ASi on soil N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions. By addressing the impact of ASi addition on bulk density, WFPS, and finally N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions, we seek to provide new insights into the potential of ASi-based soil amendments as a measure to modify soil physical conditions and thereby contribute to the mitigation of agricultural N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions, depending on soil texture and management.\u003c/p\u003e"},{"header":"Materials and methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e\u003ch2\u003eSampling site\u003c/h2\u003e\u003cp\u003eFor this study, we collected four soil samples within the patchCROP experiment (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e; [\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e] at Tempelberg, Germany (52\u0026deg;26'37\"N 14\u0026deg;09'39\" E). The experimental field covers a total area of 70 ha and features an average annual air temperature of 9.6\u0026deg;C and annual precipitation of 472 mm (ZALF climate station). Its soil mostly comprises high sandy glacial till derived from ground moraine material, with modest variations in silt and clay content.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eThe four sampling locations within the experimental field were specifically chosen for their predominantly sandy texture, as the effects of ASi are most pronounced in such soils, particularly in enhancing soil moisture levels [\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e]. Collected soil samples represent variations in silt and clay content of the two dominant soil types present in the experimental field, namely Luvisol with modest silt and clay content and Arenosol with low silt and clay content (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Soil samples from each of the four sampling locations were taken as a mixed topsoil (0 to 30 cm) sample (n\u0026thinsp;=\u0026thinsp;3) using an auger. After sampling, composite samples of each location were air dried, sieved through a 2 mm mesh, and homogenized for subsequent experiments and analyses.\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eSoil physical and chemical characterization\u003c/h3\u003e\n\u003cp\u003eThe water holding capacity (WHC) was determined using Luvisol I as a reference soil to represent all four soils. For this, 100 g of air-dried Luvisol I (in three replicates) was weighed into a steel core (250 cm\u003csup\u003e3\u003c/sup\u003e volume) with one of the ends of the core covered with a perforated material that allows water drainage. About 60 ml of water was added to the soil samples, and afterward, soil samples were allowed to drain for 24 hours at a temperature of 22\u0026deg;C. The WHC of the soil was then calculated by taking the difference in the weight of the soil before the addition of water and the final weight after allowing the water to drain for 24 hours.\u003c/p\u003e\u003cp\u003eThe soil texture analysis was performed using a hydrometer, following a modified Bouyoucos method [\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e]. Briefly, 50 g of dry soil (in triplicate) was dispersed in 100 mL of 25% sodium hexametaphosphate (technical Calgon, Bioquim, Montevideo, Uruguay) for 16 hours on a reciprocating shaker. The suspension was then mixed for five minutes using an electric mixer in a Bouyoucos blender cup. After transferring to a 2 L sedimentation cylinder, deionized water was added up to the 2 L mark, and the mixture was manually agitated. The first hydrometer reading, taken after 40 seconds, determined sand content, while the second reading, recorded after two hours, estimated clay content. The silt fraction was calculated as the difference between the sand and clay values. Measurements were performed at 20\u0026ndash;22\u0026deg;C, with temperature corrections applied when necessary.\u003c/p\u003e\u003cp\u003eMineral nitrogen (N\u003csub\u003emin\u003c/sub\u003e; sum of NH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e and NO\u003csub\u003e3\u003c/sub\u003e\u003csup\u003e\u0026minus;\u003c/sup\u003e) was determined for both the initial soil samples and soils with and without 1% ASi after incubation (experiment II). For the extraction, 80 mL of 1 M potassium chloride (KCl) solution was added to 20 g of soil each in 100 mL extraction tubes. Afterward, the mixture was shaken for 60 minutes at ~\u0026thinsp;20\u0026deg;C. After 30 minutes of shaking, it was filtered using a MACHEREY-NAGEL MN 619 \u003csup\u003e1\u003c/sup\u003e/\u003csub\u003e4\u003c/sub\u003e G filter paper (Keeney and Nelson, 1982). Ammonium (NH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e) analysis based on the Bertholet reaction [\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e] was determined by mixing 5 mL of the filtrate with Bertholet reagent and was allowed to react for 30 minutes, before measuring the absorbance at 660 nm. For nitrate determination, another 5 mL of the filtrate was treated with a cadmium reduction column to convert NO\u003csub\u003e3\u003c/sub\u003e\u003csup\u003e\u0026minus;\u003c/sup\u003e to NO\u003csub\u003e2\u003c/sub\u003e\u003csup\u003e\u0026minus;\u003c/sup\u003e, after which the sample was then mixed with Griess reagent and was allowed to react for 20 minutes before measuring at 540 nm. Both NH\u003csub\u003e4\u003c/sub\u003e\u003csup\u003e+\u003c/sup\u003e and NO\u003csub\u003e3\u003c/sub\u003e\u003csup\u003e\u0026minus;\u003c/sup\u003e were measured spectrophotometrically (Gallery Plus; ThermoFisher SCIENTIFIC GmbH). The total concentration of organic carbon (TOC) and total nitrogen (TN) of the original soil samples were analyzed using a TOC analyzer (TNM-L, Shimadzu, Japan).\u003c/p\u003e\n\u003ch3\u003eExperiment I: determination of the effect of ASi on bulk density and WFPS\u003c/h3\u003e\n\u003cp\u003eFigure \u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003e illustrates how ASi addition, through alteration of bulk density and thus WFPS, might affect N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions as a result of the theoretical relationship between WFPS and microbial activity, adapted from Linn and Doran [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. The addition of ASi (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003ea) may reduce bulk density (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eb) and enhance water-holding capacity, thereby lowering WFPS compared to soil without ASi addition. This shift in WFPS, in turn, could modify N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003ec \u0026amp; d) depending on the amount of available water, potentially altering the balance between nitrification, denitrification, and microbial respiration.\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eTo determine the potential effect of ASi addition on soil bulk density and WFPS, 50 g of air-dried soil from each sampling location were mixed with ASi at application rates of 0% (control), 1%, 5%, and 10%. Each application rate was replicated three times (n\u0026thinsp;=\u0026thinsp;3). Soil-ASi mixtures were transferred to a soil steel core with a volume of 250 cm\u003csup\u003e3\u003c/sup\u003e. The volume occupied by the soil-ASi mixture in each vessel was measured, and subsequently, bulk density was determined based on the mass of the air-dry soils. Although disturbed soil samples were used, care was taken to pack the soil to realistic field-like conditions. The resulting density for the control treatments falls within the range typically observed in similar soils under field conditions for undisturbed soil cores [\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e], supporting the suitability of the approach for examining treatment effects. To obtain a potential change in WFPS, 8.5 mL of water (i.e., 60% of WHC of Luvisol I\u0026mdash;reference soil, which means different WFPS for the four soils) was added to each of the soil-ASi mixtures. Afterward, the WFPS was calculated using Eq.\u0026nbsp;1.\u003cdiv class=\"BlockQuote\"\u003e\u003cp\u003e%WFPS = \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\left(\\frac{{\\theta\\:}_{v}}{TP}\\right)\\times\\:100\\)\u003c/span\u003e\u003c/span\u003e 1.\u003c/p\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003ewhere \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:{\\theta\\:}_{v}\\)\u003c/span\u003e\u003c/span\u003e is the volumetric water content (%), TP is the total pore space calculated as \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:\\left(I-\\frac{{P}_{b}}{{P}_{P}}\\right)\\times\\:100\\)\u003c/span\u003e\u003c/span\u003e where \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:{P}_{b}\\)\u003c/span\u003e\u003c/span\u003e is the soil bulk density (g cm\u003csup\u003e\u0026minus;\u0026thinsp;3\u003c/sup\u003e) and \u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:{P}_{P}\\)\u003c/span\u003e\u003c/span\u003e is the particle density (~\u0026thinsp;2.65 g cm\u003csup\u003e\u0026minus;\u0026thinsp;3\u003c/sup\u003e).\u003c/p\u003e\n\u003ch3\u003eExperiment II: determination of the effect of ASi on CO\u003csub\u003e2\u003c/sub\u003e and N\u003csub\u003e2\u003c/sub\u003eO emissions\u003c/h3\u003e\n\u003cp\u003eTo assess the effect of ASi addition on CO\u003csub\u003e2\u003c/sub\u003e and N\u003csub\u003e2\u003c/sub\u003eO emissions, we incubated 100 g of soil from each of the four sampling locations for 14 consecutive days, with and without 1% ASi addition. The 1% ASi treatment was chosen because agricultural soils are typically depleted by ~\u0026thinsp;1% ASi compared to natural soils, and a difference of 1% ASi is also known to alter soil moisture levels [\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e, \u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e]. Right before incubation, 17 mL of water (i.e., 60% of WHC) were added to each of the 100 g soil samples with and without ASi mixed in. This water addition was equivalent to 8.5 mL applied to 50 g of soil during the bulk density and WFPS assessment in Experiment I. While we use 100 g of soil in a fixed volume vessel (13 cm in diameter and height) in the incubation experiment (Experiment II), no direct adjustment was made for bulk density. However, the soil-ASi mixture was packed in the incubation vessels to approximate realistic field conditions, following the same procedure as in Experiment I. This ensured consistency in soil structure and porosity across treatments. Each treatment was replicated four times (n\u0026thinsp;=\u0026thinsp;4). Soil CO\u003csub\u003e2\u003c/sub\u003e and N\u003csub\u003e2\u003c/sub\u003eO emissions were continuously measured using the soil incubation system described in detail by Rillig, et al. [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e]. The system operates in a flow-through steady-state mode [\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e]. It consists of 16 airtight, cylindrical incubation vessels (13 cm in diameter and height), constructed from commercially available KG DN sewer pipes and accessories (Marley, Germany), and placed in a temperature-controlled (20\u0026deg;C) box. Ambient air from a pressure vessel flows continuously through the headspace of the incubation vessels at 32 mL min\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e to a gas analyzer (Picarro G2508; PICARRO, INC., Santa Clara, USA). A control channel allows ambient air to bypass the soil samples and flow directly from the pressure vessel to the gas analyzer at the same rate. To prevent soil samples within the incubation vessels from drying out over incubation time, the incoming air is humidified to 100% relative humidity before entering the incubation vessels. For N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e concentration measurements (1 Hz), each channel is sequentially connected to the gas analyzer via a multiplexer and a dedicated circular channel for 7 minutes per cycle. Air circulates between the incubation vessel headspace and the CRDS analyzer at 250 mL min\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e, driven by a low-leak diaphragm pump (A0702, Picarro, Santa Clara, CA, USA). The multiplexer directs air from each of the 17 measurement channels (16 incubation vessels and 1 control channel) into this circuit. Fluxes were calculated using the ideal gas law Eq.\u0026nbsp;(2) based on measured CO\u003csub\u003e2\u003c/sub\u003e or N\u003csub\u003e2\u003c/sub\u003eO gas concentrations in each channel and the equivalent concentrations in the control channel [\u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e]\u003cdiv class=\"BlockQuote\"\u003e\u003cp\u003e\u003cspan class=\"InlineEquation\"\u003e\u003cspan class=\"mathinline\"\u003e\\(\\:F=\\frac{\\varDelta\\:C*v*\\rho\\:}{\\varDelta\\:t*A*R*\\left(T+273.15\\right)}\\)\u003c/span\u003e\u003c/span\u003e 2.\u003c/p\u003e\u003c/div\u003e\u003c/p\u003e\u003cp\u003ewhere F is the flux rate (\u0026micro;mol m\u003csup\u003e\u0026minus;\u0026thinsp;2\u003c/sup\u003e s-\u003csup\u003e1\u003c/sup\u003e), ρ is the atmospheric pressure (Pa), V is the air flow rate into the headspace and the channels (m\u003csup\u003e3\u003c/sup\u003e), and Δc is the difference of gas concentrations [mol] between the outlet of a specific vessel and the control channel. A is the chamber basal area (m\u003csup\u003e2\u003c/sup\u003e), R is the gas constant (8.314 m\u003csup\u003e3\u003c/sup\u003e Pa K\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e mol\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e), Δt is the time over which the concentration change was observed, and T is the incubation temperature (K). A modified version of a modular R program script described by [\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e] was used to calculate the cumulative N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e fluxes over specified time intervals.\u003c/p\u003e\n\u003ch3\u003eStatistical analysis and data visualization\u003c/h3\u003e\n\u003cp\u003eFluxes were calculated, and visualization was made using the R computational environment [\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e]. The effects of ASi treatments, specifically control (0%), 1%, 5%, and 10% ASi on bulk density and WFPS across the two different soil types were assessed using a one-way analysis of variance (ANOVA). Given the sample size (n\u0026thinsp;=\u0026thinsp;3, experiment I), the ANOVA assumptions were thoroughly examined to guarantee the validity of the analysis, particularly assessing the normality of residuals using the Shapiro-Wilk test. A Q-Q plot was used to further validate the results of the normality tests. A Levene's test was applied to evaluate the homogeneity of variance between the treatment groups. These diagnostic tests confirmed that the ANOVA assumptions, and homogeneity of variance, were satisfied, justifying the use of the parametric ANOVA approach. Following the ANOVA, a Tukey's post hoc test (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05) was used to identify specific differences between treatment groups (control (0%), 1%, 5%, and 10% ASi) for the bulk density and WFPS (experiment I). A permutation test (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05) was performed to determine significant differences between measured N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e fluxes of the control and ASi-treated soils (experiment II).\u003c/p\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec9\" class=\"Section2\"\u003e\u003ch2\u003ePhysicochemical soil properties\u003c/h2\u003e\u003cp\u003eThe studied soil varied in texture (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e), with Luvisols I and II showing a modest initial silt (22 and 25%) and lower sand content (74% for both) compared to Arenosol I and II, with a lower silt (12 and 11%) and higher sand content (86 and 88%). Adding 1%, 5%, and 10% ASi, respectively, increased silt and clay and reduced sand content in both Luvisol and Arenosol (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). Total nitrogen (TN) content was similar for all studied soils of the four sampling locations, while the total organic carbon content (TOC) was slightly higher for the Luvisols (~\u0026thinsp;1% \u0026ndash; 1.2%) compared to the Arenosols (~\u0026thinsp;0.7% \u0026ndash; 0.9%). N\u003csub\u003emin\u003c/sub\u003e measured after incubation did not differ significantly with or without 1% ASi addition, nor between Luvisols and Arenosols of all four sampling locations (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e\u003cp\u003e\u003cdiv class=\"gridtable\"\u003e\u003ctable float=\"Yes\" id=\"Tab1\" border=\"1\"\u003e\u003ccaption language=\"En\"\u003e\u003cdiv class=\"CaptionNumber\"\u003eTable 1\u003c/div\u003e\u003cdiv class=\"CaptionContent\"\u003e\u003cp\u003eSummary of soil physicochemical properties and CO\u003csub\u003e2\u003c/sub\u003e and N\u003csub\u003e2\u003c/sub\u003eO fluxes under different ASi treatments at four sampling locations (Luvisol I and II, Arenosol I and II). Reported are total cumulative CO\u003csub\u003e2\u003c/sub\u003e and N\u003csub\u003e2\u003c/sub\u003eO fluxes (mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD, n\u0026thinsp;=\u0026thinsp;4), mineral nitrogen content (Nmin; mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD, n\u0026thinsp;=\u0026thinsp;4), bulk density (g cm\u003csup\u003e\u0026minus;\u0026thinsp;3\u003c/sup\u003e; mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD, n\u0026thinsp;=\u0026thinsp;3), and water-filled pore space (WFPS; mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD, n\u0026thinsp;=\u0026thinsp;3). Soil texture is given as sand, silt, and clay contents, and total nitrogen TN and total organic carbon content (TOC) are provided for each sampling location. TN and TOC refer to untreated soils only. For bulk density and WFPS, statistical comparisons were made between each ASi treatment and its respective control using a post hoc test, while a permutation test was applied for CO\u003csub\u003e2\u003c/sub\u003e and N\u003csub\u003e2\u003c/sub\u003eO to determine the difference between 1% ASi treatment and the respective control treatments. Treatments showing significant differences (\u003cem\u003ep\u003c/em\u003e-value\u0026thinsp;\u0026gt;\u0026thinsp;0.05) are indicated using different superscript letters.\u003c/p\u003e\u003c/div\u003e\u003c/caption\u003e\u003ccolgroup cols=\"13\"\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c1\" colnum=\"1\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c2\" colnum=\"2\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c3\" colnum=\"3\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c4\" colnum=\"4\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c5\" colnum=\"5\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c6\" colnum=\"6\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c7\" colnum=\"7\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c8\" colnum=\"8\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c9\" colnum=\"9\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c10\" colnum=\"10\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c11\" colnum=\"11\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c12\" colnum=\"12\"\u003e\u003c/div\u003e\u003cdiv align=\"left\" class=\"colspec\" colname=\"c13\" colnum=\"13\"\u003e\u003c/div\u003e\u003cthead\u003e\u003ctr\u003e\u003cth align=\"left\" colname=\"c1\"\u003e\u003cp\u003eSampling location\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c2\"\u003e\u003cp\u003eTreatment\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c3\"\u003e\u003cp\u003eCO\u003csub\u003e2\u003c/sub\u003e flux\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c4\"\u003e\u003cp\u003eN\u003csub\u003e2\u003c/sub\u003eO flux\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c5\"\u003e\u003cp\u003eNH\u003csub\u003e4\u003c/sub\u003e-N\u003csub\u003eKCl\u003c/sub\u003e\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c6\"\u003e\u003cp\u003eNO\u003csub\u003e3\u003c/sub\u003e-N\u003csub\u003eKCl\u003c/sub\u003e\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c7\"\u003e\u003cp\u003eBulk density\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c8\"\u003e\u003cp\u003eWFPS\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c9\"\u003e\u003cp\u003eSand\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c10\"\u003e\u003cp\u003eSilt\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c11\"\u003e\u003cp\u003eClay\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c12\"\u003e\u003cp\u003eTOC\u003c/p\u003e\u003c/th\u003e\u003cth align=\"left\" colname=\"c13\"\u003e\u003cp\u003eTN\u003c/p\u003e\u003c/th\u003e\u003c/tr\u003e\u003c/thead\u003e\u003ctbody\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003emg CO\u003csub\u003e2\u003c/sub\u003e-C g\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e\u0026micro;g N\u003csub\u003e2\u003c/sub\u003eO-N g\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003emg 100 g\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003emg 100 g\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003eg cm\u003csup\u003e\u0026minus;\u0026thinsp;3\u003c/sup\u003e\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colspan=\"5\" nameend=\"c13\" namest=\"c9\"\u003e\u003cp\u003e%\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e(n\u0026thinsp;=\u0026thinsp;4)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e(n\u0026thinsp;=\u0026thinsp;4)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e(n\u0026thinsp;=\u0026thinsp;4)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e(n\u0026thinsp;=\u0026thinsp;4)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e(n\u0026thinsp;=\u0026thinsp;3)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e(n\u0026thinsp;=\u0026thinsp;3)\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colspan=\"5\" nameend=\"c13\" namest=\"c9\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eLuvisol I\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eControl\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e33.6\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;12.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e3.1\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;1.3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e0.2\u0026thinsp;\u0026plusmn;\u0026thinsp;0.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e1.9\u0026thinsp;\u0026plusmn;\u0026thinsp;0.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e1.6\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;0.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e66\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;2.6.\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e74\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e25\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c12\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003e0.9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c13\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003e0.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eASi \u0026minus;\u0026thinsp;1%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e49.3\u003csup\u003eb\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;2.6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e4.9\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;0.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e0.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e2.3\u0026thinsp;\u0026plusmn;\u0026thinsp;0.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e1.5\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;0.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e63\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;2.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e71\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e26\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e4\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eASi \u0026minus;\u0026thinsp;5%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e1.4\u003csup\u003eb\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;0.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e52\u003csup\u003eb\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;2.5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e68\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e26\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c13\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eASi \u0026minus;\u0026thinsp;10%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.5\u003csup\u003ec\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;0.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e11\u003csup\u003ec\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;0.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e67\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e27\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c13\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003eLuvisol II\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eControl\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e28.8\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;2.7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e4.3\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;3.4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e0.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e4.9\u0026thinsp;\u0026plusmn;\u0026thinsp;0.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e1.7\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;0.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e65\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;0.7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e74\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e22\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e4\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c12\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003e1.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c13\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003e0.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eASi \u0026minus;\u0026thinsp;1%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e45.5\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;31.7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e5.2\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;3.9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u003cp\u003e0.1\u0026thinsp;\u0026plusmn;\u0026thinsp;0.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u003cp\u003e4.6\u0026thinsp;\u0026plusmn;\u0026thinsp;0.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e1.5\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;0.0\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e59\u003csup\u003ea\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;3.8\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e74\u003c/p\u003e\u003c/td\u003e\u003ctd 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align=\"left\" colname=\"c10\"\u003e\u003cp\u003e12\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c12\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003e0.7\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c13\" morerows=\"1\" rowspan=\"2\"\u003e\u003cp\u003e0.1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eASi \u0026minus;\u0026thinsp;1%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u003cp\u003e37.4\u003csup\u003eb\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;8.5\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u003cp\u003e2.2\u003csup\u003eb\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;0.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" 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colname=\"c8\"\u003e\u003cp\u003e95\u003csup\u003eb\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;4.3\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e87\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e12\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e1\u003c/p\u003e\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eASi \u0026minus;\u0026thinsp;5%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e1.6\u003csup\u003eb\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;0.6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e63\u003csup\u003ec\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;5.2\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e81\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e12\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c13\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003ctr\u003e\u003ctd align=\"left\" colname=\"c1\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c2\"\u003e\u003cp\u003eASi \u0026minus;\u0026thinsp;10%\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c3\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c4\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c5\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c6\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c7\"\u003e\u003cp\u003e0.6\u003csup\u003ec\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;0.1\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c8\"\u003e\u003cp\u003e12\u003csup\u003ed\u003c/sup\u003e\u0026thinsp;\u0026plusmn;\u0026thinsp;2.6\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c9\"\u003e\u003cp\u003e78\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c10\"\u003e\u003cp\u003e13\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c11\"\u003e\u003cp\u003e9\u003c/p\u003e\u003c/td\u003e\u003ctd align=\"left\" colname=\"c12\"\u003e\u0026nbsp;\u003c/td\u003e\u003ctd align=\"left\" colname=\"c13\"\u003e\u0026nbsp;\u003c/td\u003e\u003c/tr\u003e\u003c/tbody\u003e\u003c/colgroup\u003e\u003c/table\u003e\u003c/div\u003e\u003c/p\u003e\u003c/div\u003e\n\u003ch3\u003eEffect of ASi addition on bulk density and WFPS\u003c/h3\u003e\n\u003cp\u003eExperiment I showed that the addition of 1%, 5%, and 10% ASi altered bulk density and WFPS in both Arensol and Luvisol. In Luvisol I and II, a slight but not significant decrease in bulk density due to 1% ASi addition was observed (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e), with a more pronounced, significant decrease (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05) observed at both 5% and 10% ASi addition, respectively. In contrast, Arenosol I and II showed a slight but not significant increase in bulk density due to 1% ASi addition compared to the control treatment. However, similar to Luvisol I and II, bulk density decreased following 5% and 10% ASi addition treatments. This decrease was statistically significant for Arenosol I at both 5% and 10%, and for Arenosol II at 10% ASi addition (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). These changes in bulk density were accompanied by an observed change in WFPS. In Luvisol I and II a slight but not significant decrease in WFPS by 4.5% and 8.9% through the addition of 1% ASi was observed when compared to soil samples without ASi addition (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e; Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). A more pronounced and significant (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05) decrease in WFPS, however, was obtained when 5% and 10% ASi were added to Luvisol I and II, decreasing WFPS to 51% and 11.4% for Luvisol I, and 48.1% and 11.8% for Luvisol II, when compared to soil samples without ASi addition, respectively (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eIn contrast to that, the addition of 1% ASi increased WFPS in both Arenosols, though only Arenosol II showed a significant increase from 82.2\u0026ndash;94.5%, whereas Arenosol I showed a non-significant increase from 67.5\u0026ndash;70.5% (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e; Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). However, at 5% and 10% ASi, admixture, WFPS decreased in both Arenosols, similar to the studied Luvisols, even though that decrease was only significant for 10% ASi addition at Arenosol I and II (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cdiv id=\"Sec11\" class=\"Section2\"\u003e\u003ch2\u003eEffect of ASi addition on N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions\u003c/h2\u003e\u003cp\u003eWe found clear differences in cumulative N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions due to 1% ASi addition in both soil types during experiment II. These differences, however, varied with ASi addition in Arenosol, reducing and in Luvisol, increasing the emissions. Figure\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea-b shows the mean N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions for soils from the four sampling locations without and with the addition of 1% ASi. Figures\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ec\u0026ndash;d illustrate the theoretical relationship between microbial activity (%) and water-filled pore space (WFPS, 0\u0026ndash;100%) alongside the same measured cumulative N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions for soils treated with and without 1% ASi. Figures\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ee\u0026ndash;f depict the pore network characteristics for Arenosols and Luvisols with and without 1% ASi addition. In both soils, the representation for soils treated with 1% ASi shows a visibly reduced pore space compared to the corresponding controls, with fewer and smaller macro-and micropores evident in the ASi-treated samples. In Arenosols, the addition of ASi predominantly clogged the abundant meso and macropores, whereas in Luvisols, which naturally have fewer macropores, ASi not only clogged existing pores but also contributed to the formation of new ones (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ee \u0026amp; f).\u003c/p\u003e\u003cp\u003e\u003c/p\u003e\u003cp\u003eIn Arenosols, N\u003csub\u003e2\u003c/sub\u003eO emissions decreased at increased WFPS under 1% ASi treatment, though only Arenosol I showed a significant (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05) reduction by 44.3%, while Arenosol II showed a non-significant reduction by 8% compared to Arenosol without ASi addition (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e; Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea \u0026amp; c). In contrast to both Arenosols, Luvisol I and II at decreased WFPS showed a non-significant increase in N\u003csub\u003e2\u003c/sub\u003eO emissions with the addition of 1% ASi by 57% and 18%, respectively (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e; Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea \u0026amp; c). Although the differences in N\u003csub\u003e2\u003c/sub\u003eO emissions between the ASi-treated soil and the control were not significant across all sampling locations, a trend was evident for the two soil types: decreasing N\u003csub\u003e2\u003c/sub\u003eO emissions for Arenosol and increasing N\u003csub\u003e2\u003c/sub\u003eO emissions for Luvisol due to the addition of 1% ASi.\u003c/p\u003e\u003cp\u003eIn case of CO\u003csub\u003e2\u003c/sub\u003e, we found a significant (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05) decrease in CO\u003csub\u003e2\u003c/sub\u003e emissions due to 1% ASi addition (resulting in an increased WFPS) by 45% and 42% for Arenosol I and II, respectively (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e; Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eb \u0026amp; d). In contrast, Luvisol I and II showed an increase in CO\u003csub\u003e2\u003c/sub\u003e emissions by 46.8%, which was only significant (\u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05) in the case of Luvisol I, following 1% ASi addition despite only a slight decrease in WFPS (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e; Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eb \u0026amp; d).\u003c/p\u003e\u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eThis study investigated the effect of ASi on soil bulk density, WFPS, and subsequent N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions in soils with different silt contents. Our findings show that ASi addition to soil modifies their bulk density and WFPS.\u003c/p\u003e\u003cp\u003eFindings of experiment I support our hypothesis that ASi addition alters the soil\u0026rsquo;s WFPS through influencing the soil's bulk density. However, contrasting effects were found depending on the initial soil texture (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e; Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003e). We found that ASi reduced WFPS in the finer-textured Luvisol soils (with modest silt and clay content and more micropores). In contrast, in the coarser-textured Arenosol soils (with low silt and clay content and less micropores), ASi appeared to increase WFPS. Thus, the direction and magnitude of ASi-induced changes in WFPS are not uniform but depend on the soil\u0026rsquo;s initial texture and underlying pore structure. The reason for this is likely due to two contrasting effects through which ASi alters bulk density and thus, WFPS: i) when added to finer-texture Luvisol soils, ASi decreased the bulk density likely due to its low density and high internal particle porosity by clogging the few mesopores (with less ASi) while predominantly creating new mesopores. These may have reduced the packing efficiency and increasing the total pore volume of the soil (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ef), this in turn may lower the percentage of the pores filled with water at fixed water input, thus lowering the WFPS, ii) when added to coarse-textured Arenosol soils, ASi may reduce macroporosity by clogging larger pores, thereby enhancing the bulk density and WFPS [\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e]. Depending on the amount of added ASi and which of these effects dominates at a particular soil texture, adding ASi can either increase or decrease bulk density, thus affecting WFPS. Texture analysis showed that ASi behaves like silt- and clay-sized particles, increasing the fine fraction while proportionally reducing sand content (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). This shift towards finer texture likely reduced macroporosity in sandy Arenosol (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ee), thereby increasing WFPS. In contrast, in the already finer-textured Luvisols, ASi slightly lowers WFPS, potentially due to changes in pore structure, such as improved pore volume (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ef) that lowers WFPS under fixed water input. The resulting aeration shifts match the observed decrease (Arenosols) or increase (Luvisols) in CO\u003csub\u003e2\u003c/sub\u003e and N\u003csub\u003e2\u003c/sub\u003eO emissions observed during experiment II. We found that 1% ASi addition to Arenosols caused an increase in bulk density, likely due to ASi clogging meso- and macropores, thereby enhancing bulk density. In contrast, Luvisols showed a decrease in bulk density, which may be attributed to the low density and high internal porosity of ASi particles reducing packing efficiency and increasing total pore volume. Neither the increase nor the decrease in bulk density at 1% ASi treatment was significant (Table\u0026nbsp;\u003cspan refid=\"Tab1\" class=\"InternalRef\"\u003e1\u003c/span\u003e). However, at increased ASi levels (10%), both Arenosols and Luvisols showed a significant reduction in both bulk density and WFPS, suggesting that higher ASi levels may have increased the proportion of finer pores or enhanced internal porosity within soil aggregates, leading to a decrease in bulk density and WFPS.\u003c/p\u003e\u003cp\u003eIn our second hypothesis, we proposed that ASi-induced WFPS changes via bulk density alteration would affect N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emission dynamics. In experiment II, contrasting N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions responses in Arenosols and Luvisols following ASi treatment were found, confirming the second hypothesis. The variations can be attributed to differences in soil texture that influence how ASi alters pore structure and bulk density, subsequently modulating the WFPS and oxygen availability.\u003c/p\u003e\u003cp\u003eFor example, the incorporation of ASi in Arenosols resulted in an increase in WFPS, from 67.5\u0026ndash;70.4% in Arenosol I and from 82.2\u0026ndash;94.5% in Arenosol II (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ec \u0026amp; d). This rise in WFPS may be attributed to a reduction in mesopore and macropore volume due to ASi addition. These changes likely limit oxygen availability and diffusion within the soil, thereby inhibiting microbial respiration, essential oxygen-dependent pathways for CO\u003csub\u003e2\u003c/sub\u003e production, supporting the decrease in CO\u003csub\u003e2\u003c/sub\u003e emissions in Arenosol (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eb \u0026amp; d). These findings are in line with established relationships indicating that microbial activity and GHG emissions decrease at high WFPS in response to oxygen limitation [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e]. We showed that N\u003csub\u003e2\u003c/sub\u003eO emissions were highest at elevated WFPS (~\u0026thinsp;80%) compared to those at lower WFPS (~\u0026thinsp;60), irrespective of treatment (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ec). This aligns with expectations that denitrification is enhanced under high WFPS, a condition that typically generates more N\u003csub\u003e2\u003c/sub\u003eO than nitrification processes occurring at lower WFPS [\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e]. This could explain the overall higher N\u003csub\u003e2\u003c/sub\u003eO emissions at elevated WFPS in our study for both control and ASi-treated soil, particularly in Arenosol II (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea \u0026amp; c). However, with ASi treatment, further increases in WFPS beyond ~\u0026thinsp;80% (up to ~\u0026thinsp;95%) were associated with a slight reduction in N\u003csub\u003e2\u003c/sub\u003eO emissions compared to the control (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea \u0026amp; c), even though emissions at these high WFPS remained greater than at lower WFPS. This pattern suggests that while denitrification was a key contributor to the high N\u003csub\u003e2\u003c/sub\u003eO flux at elevated WFPS, the more extreme oxygen-limited condition induced by ASi at \u0026gt;\u0026thinsp;80% WFPS may have promoted complete denitrification, thereby shifting nitrogen transformations from N\u003csub\u003e2\u003c/sub\u003eO toward dinitrogen (N\u003csub\u003e2\u003c/sub\u003e) production [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e, \u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e]. This shift in WFPS probably played a significant role in substantially reducing overall N\u003csub\u003e2\u003c/sub\u003eO emissions found in Arenosols over time. This suggests that beyond a certain threshold, an increase in WFPS could limit oxygen-dependent microbial processes, thereby reducing both CO\u003csub\u003e2\u003c/sub\u003e and N\u003csub\u003e2\u003c/sub\u003eO emissions. Maillet, et al. [\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e] reported that soils with lower meso-and macroporosity, associated with WFPS, showed higher N\u003csub\u003e2\u003c/sub\u003eO emissions. In our study, this pattern was not uniform: in Luvisol, at lower WFPS under ASi treatment coincided with higher N\u003csub\u003e2\u003c/sub\u003eO emission compared to the untreated control (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eA \u0026amp; C), whereas in Arenosol, under higher WFPS, due to ASi likely clogging meso- and macropores, coincided with lower N\u003csub\u003e2\u003c/sub\u003eO emission. These differences highlight that although pore structure strongly affects emissions, the direction of response depends on the initial texture and how ASi alters the pore network or structure\u003c/p\u003e\u003cp\u003eIn contrast, the addition of ASi to Luvisol soils resulted in a decrease in WFPS, from 65.7\u0026ndash;62.7% in Luvisol I and from 64.6\u0026ndash;58.8% in Luvisol II (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea \u0026amp; c). This adjustment in WFPS moved the conditions closer to the optimal WFPS range (40\u0026ndash;60%) for microbial nitrification and aerobic respiration [\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e]. However, N\u003csub\u003e2\u003c/sub\u003eO emissions were generally lower in Luvisols at these lower WFPS compared to Arenosols (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ec), which, under higher WFPS, produced the highest N\u003csub\u003e2\u003c/sub\u003eO emission irrespective of the treatment. The reduction in WFPS found for Luvisols (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003ea \u0026amp; c) may have improved soil pore volume and enhanced aeration. These changes probably facilitated oxygen diffusion, which may have promoted aerobic microbial activity, thereby increasing N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions. For Luvisols, no significant increase in N\u003csub\u003e2\u003c/sub\u003eO emissions was observed; the small sample size (n\u0026thinsp;=\u0026thinsp;4) may have reduced statistical power; increasing the sample size could demonstrate differences, through increased sensitivity, however. While ASi may contribute to lower N\u003csub\u003e2\u003c/sub\u003eO emissions by altering bulk density and WFPS, improved plant nitrogen uptake [\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e] could further reduce available mineral N for microbial nitrification and denitrification, being a potential way to make agriculture more sustainable [\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e].\u003c/p\u003e\u003cp\u003eIn conclusion, our study particularly highlights ASi's effect on soil bulk density and WFPS, which in turn affects the N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions by shifting towards or from optimal conditions for microbial respiration, nitrification, and denitrification. Under our experimental conditions, the highest N\u003csub\u003e2\u003c/sub\u003eO emissions coincided with the highest WFPS (\u0026gt;\u0026thinsp;80%), indicating that denitrification contributed substantially to N\u003csub\u003e2\u003c/sub\u003eO production under high WFPS. It is still not clear, however, what the observed effect of ASi on WFPS and associated N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions would mean under field conditions. ASi may decrease or increase the WFPS in the field depending on the initial soil texture, and this may either raise or lower N\u003csub\u003e2\u003c/sub\u003eO emissions. Although denitrification was evident under high WFPS in our study, its magnitude and relevance could vary under field conditions, where additional factors such as high organic matter availability may further impact this pathway. In addition, other ASi-induced processes, such as enhanced crop growth, improved N uptake, and interactions with variable water availability and precipitation, may dominate or counteract the mechanisms observed under controlled conditions. These interconnected processes highlight the complexity of ASi's role in real-world settings and the need for further field-based research, since optimizing the use of ASi as a soil amendment requires an understanding of the relationships among microbial activity, structural changes caused by ASi, and GHG emission dynamics.\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe thank Oscar Monzon for help with analysis and Bj\u0026ouml;rn Gustav Wang and Felix Erbe for help during soil sampling and map visualization.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe overall project was funded by the Deutsche Forschungsgemeinschaft for funding (SCHA 1822/14-1 and KA 1737/21-1). Funding for the infrastructure of the patchCROP experiment was provided by the Leibniz Centre for Agricultural Landscape Research (ZALF) and the German Research Foundation under Germany\u0026rsquo;s Excellence Strategy (EXC-2070\u0026ndash;390732324 \u0026ndash; PhenoRob). KG acknowledges support from BMBF for the Junior Research Group SoilRob (Grant 031B1391).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics, Consent to Participate, and Consent to Publish declarations\u003c/strong\u003e: Not applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eClinical trial number:\u003c/strong\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthor contributions:\u0026nbsp;\u003c/strong\u003eP.U. was responsible for conceptualization, investigation, data curation, data analysis, visualization, and writing (original draft). M.H. contributed to experimental design, data validation, visualization, and writing (review and editing). M.L. contributed to experimental monitoring. K.G. contributed to visualization and writing (review and editing). K.K. contributed to writing (review and editing). J.S. contributed to experimental design and writing (review and editing). All authors reviewed the manuscript and approved the final version for submission.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting Interests:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll authors certify that they have no affiliations with or involvement in any organization or entity with any financial interest or non-financial interest in the subject matter or materials discussed in this manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eDual Publication:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe results/data/figures in this manuscript have not been published elsewhere, nor are they under consideration (from you or one of your Contributing Authors) by another publisher.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData Availability:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eI do not have any research data outside the submitted manuscript file.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eWedepohl, K H, \u0026quot;The composition of the continental crust,\u0026quot; \u003cem\u003eGeochimica et cosmochimica Acta, \u003c/em\u003evol. 59, no. 7, pp. 1217-1232, 1995; Ma, J F and Yamaji, N, \u0026quot;Silicon uptake and accumulation in higher plants,\u0026quot; \u003cem\u003eTrends in plant science, \u003c/em\u003evol. 11, no. 8, pp. 392-397, 2006.\u003c/li\u003e\n\u003cli\u003eSchaller, J, Puppe, D, Kaczorek, D, Ellerbrock, R, and Sommer, M, \u0026quot;Silicon Cycling in Soils Revisited,\u0026quot; \u003cem\u003ePlants (Basel), \u003c/em\u003evol. 10, no. 2, Feb 4 2021, doi: 10.3390/plants10020295.\u003c/li\u003e\n\u003cli\u003eSchaller, J, Kleber, M, Puppe, D, Stein, M, Sommer, M, and Rillig, M C, \u0026quot;The importance of reactive silica for maintaining soil health,\u0026quot; \u003cem\u003ePlant and Soil, \u003c/em\u003epp. 1-12, 2025.\u003c/li\u003e\n\u003cli\u003eStruyf, E\u003cem\u003e et al.\u003c/em\u003e, \u0026quot;Historical land use change has lowered terrestrial silica mobilization,\u0026quot; \u003cem\u003eNat Commun, \u003c/em\u003evol. 1, p. 129, Nov 30 2010, doi: 10.1038/ncomms1128.\u003c/li\u003e\n\u003cli\u003eMengel, K and Kirkby, E A, \u003cem\u003ePrinciples of plant nutrition\u003c/em\u003e (no. 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\u003cem\u003eBioscience, \u003c/em\u003evol. 50, no. 8, pp. 667-680, 2000.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Amorphous silica, Water-filled-pore space, Greenhouse gas emissions, bulk density, Silicon","lastPublishedDoi":"10.21203/rs.3.rs-7348495/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-7348495/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eSilicon (Si) is abundant in the Earth\u0026rsquo;s crust; however, its amorphous form (ASi) is often depleted in agricultural soils. While ASi benefits plant nutrient uptake and growth, its effects on soil pore characteristics, such as water-filled pore space (WFPS), and regulating greenhouse gas (GHG) emissions remain poorly understood. We investigated the effect of ASi addition on soil bulk density, WFPS, and subsequent N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emission dynamics in two soil types of differing texture: Luvisols (moderate silt and clay) and Arenosols (low silt and clay). In a first experiment, we assessed how varying ASi levels affected soil bulk density and WFPS. A second experiment investigated the impact of 1% ASi on N\u003csub\u003e2\u003c/sub\u003eO and CO\u003csub\u003e2\u003c/sub\u003e emissions. ASi addition altered soil bulk density, leading to a decrease in WFPS, especially at 10% ASi in Luvisols. In Arenosols, WFPS increased at 1% ASi before declining at higher rates. The 1% ASi addition increased CO\u003csub\u003e2\u003c/sub\u003e and N\u003csub\u003e2\u003c/sub\u003eO emissions in Luvisols but reduced both in Arenosols. These contrasting outcomes likely reflect a dual effect of ASi: in finer-textured Luvisols, ASi reduces bulk density and increases pore volume, which lowers WFPS under fixed water input, improves aeration, and enhances microbial respiration and nitrification, resulting in increased CO\u003csub\u003e2\u003c/sub\u003e and N\u003csub\u003e2\u003c/sub\u003eO emissions. In coarser-textured Arenosols, ASi may reduce macroporosity by clogging larger pores, resulting in higher WFPS and oxygen limitation, thereby decreasing emissions. Our findings suggest ASi has texture-dependent effects on soil properties and GHG emissions. These outcomes highlight the need for further field-based investigation under natural conditions.\u003c/p\u003e","manuscriptTitle":"Effects of Amorphous Silica on CO2 and N2O Emissions Mediated by Water-Filled Pore Space in Diverse Agricultural Soils","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-09-11 10:41:35","doi":"10.21203/rs.3.rs-7348495/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"ccc55b88-d300-4661-af8f-3d23d61ca545","owner":[],"postedDate":"September 11th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2025-10-03T06:38:51+00:00","versionOfRecord":[],"versionCreatedAt":"2025-09-11 10:41:35","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-7348495","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-7348495","identity":"rs-7348495","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}