Effect of cover crops on greenhouse gas emissions in an irrigated field under integrated soil fertility management

Abstract

Agronomical and environmental benefits are associated with replacing winter fallow by cover crops (CCs). Yet, the effect of this practice on nitrous oxide (N₂O) emissions remains poorly understood. In this context, a field experiment was carried out under Mediterranean conditions to evaluate the effect of replacing the traditional winter fallow (F) by vetch (Vicia sativa L.; V) or barley (Hordeum vulgare L.; B) on greenhouse gas (GHG) emissions during the intercrop and the maize (Zea mays L.) cropping period. The maize was fertilized following integrated soil fertility management (ISFM) criteria. Maize nitrogen (N) uptake, soil mineral N concentrations, soil temperature and moisture, dissolved organic carbon (DOC) and GHG fluxes were measured during the experiment. Our management (adjusted N synthetic rates due to ISFM) and pedo-climatic conditions resulted in low cumulative N₂O emissions (0.57 to 0.75kgN₂O-Nha^-1yr^-1), yield-scaled N₂O emissions (3-6gN₂O-Nkg aboveground N uptake^-1) and N surplus (31 to 56kgNha^-1) for all treatments. Although CCs increased N₂O emissions during the intercrop period compared to F (1.6 and 2.6 times in B and V, respectively), the ISFM resulted in similar cumulative emissions for the CCs and F at the end of the maize cropping period. The higher C:N ratio of the B residue led to a greater proportion of N₂O losses from the synthetic fertilizer in these plots when compared to V. No significant differences were observed in CH₄ and CO₂ fluxes at the end of the experiment. This study shows that the use of both legume and nonlegume CCs combined with ISFM could provide, in addition to the advantages reported in previous studies, an opportunity to maximize agronomic efficiency (lowering synthetic N requirements for the subsequent cash crop) without increasing cumulative or yield-scaled N₂O losses.

Biogeosciences, 13, 52455257, 2016 www.biogeosciences.net/13/5245/2016/ doi:10.5194/bg-13-5245-2016 Author(s) 2016. CC Attribution 3.0 License.

Guillermo Guardia1, Diego Abalos2, Sonia Garca-Marco1, Miguel Quemada1, Mara Alonso-Ayuso1, Laura M. Crdenas3, Elizabeth R. Dixon3, and Antonio Vallejo1

1ETSI Agronomos, Technical University of Madrid, Ciudad Universitaria, 28040 Madrid, Spain

2School of Environmental Sciences, University of Guelph, Guelph, Ontario, N1G 2W1, Canada

3Rothamsted Research, North Wyke, Devon, EX20 2SB, UK

Correspondence to: Guillermo Guardia ([email protected])

Received: 27 January 2016 Published in Biogeosciences Discuss.: 29 March 2016 Revised: 30 August 2016 Accepted: 2 September 2016 Published: 20 September 2016

Abstract. Agronomical and environmental benets are associated with replacing winter fallow by cover crops (CCs). Yet, the effect of this practice on nitrous oxide (N2O) emissions remains poorly understood. In this context, a eld experiment was carried out under Mediterranean conditions to evaluate the effect of replacing the traditional winter fallow (F) by vetch (Vicia sativa L.; V) or barley (Hordeum vulgare L.; B) on greenhouse gas (GHG) emissions during the intercrop and the maize (Zea mays L.) cropping period. The maize was fertilized following integrated soil fertility management (ISFM) criteria. Maize nitrogen (N) up-take, soil mineral N concentrations, soil temperature and moisture, dissolved organic carbon (DOC) and GHG uxes were measured during the experiment. Our management (adjusted N synthetic rates due to ISFM) and pedo-climatic conditions resulted in low cumulative N2O emissions (0.57 to0.75 kg N2O-N ha1 yr1), yield-scaled N2O emissions (3 6 g N2O-N kg aboveground N uptake1) and N surplus (31 to 56 kg N ha1) for all treatments. Although CCs increased N2O emissions during the intercrop period compared to F(1.6 and 2.6 times in B and V, respectively), the ISFM resulted in similar cumulative emissions for the CCs and F at the end of the maize cropping period. The higher C : N ratio of the B residue led to a greater proportion of N2O losses from the synthetic fertilizer in these plots when compared toV. No signicant differences were observed in CH4 and CO2 uxes at the end of the experiment. This study shows that the use of both legume and nonlegume CCs combined with ISFM could provide, in addition to the advantages reported in previous studies, an opportunity to maximize agronomic

Effect of cover crops on greenhouse gas emissions in an irrigated eld under integrated soil fertility management

efciency (lowering synthetic N requirements for the subsequent cash crop) without increasing cumulative or yield-scaled N2O losses.

1 Introduction

Improved resource-use efciencies are pivotal components of sustainable agriculture that meets human needs and protects natural resources (Spiertz, 2010). Several strategies have been proposed to improve the efciency of intensive irrigated systems, where nitrate (NO3) leaching losses are of major concern, during both cash crop and winter fallow periods (Quemada et al., 2013). In this sense, replacing winter intercrop fallow with cover crops (CCs) has been reported to decrease NO3 leaching via retention of post-harvest surplus inorganic nitrogen (N) (Wagner-Riddle and Thurtell, 1998), consequently improving N use efciency of the cropping system (Gabriel and Quemada, 2011). Furthermore, the use of CCs as green manure for the subsequent cash crop may further increase soil fertility and N use efciency (Tonitto et al., 2006; Veenstra et al., 2007) through slow release of N and other nutrients from the crop residues, leading to a saving in synthetic fertilizer.

From an environmental point of view, N fertilization is closely related to the production and emission of nitrous oxide (N2O) (Davidson and Kanter, 2014), a greenhouse gas (GHG) with a molecular global warming potential ca. 300 times that of carbon dioxide (CO2) (IPCC, 2007). Nitrous oxide released from agricultural soils is mainly generated

Published by Copernicus Publications on behalf of the European Geosciences Union.

5246 G. Guardia et al.: Effect of cover crops on greenhouse gas emissions

by nitrication and denitrication processes, which are inuenced by several soil variables (Firestone and Davidson, 1989). Thereby, modifying these parameters through agricultural management practices (e.g., fertilization, crop rotation, tillage or irrigation) aiming to optimize N inputs can lead to strategies for reducing the emission of (N2O) (Ussiri and

Lal, 2013). In order to identify the most effective GHG mitigation strategies, side effects of methane (CH4) uptake and

CO2 emission (i.e., respiration) from soils, which are also inuenced by agricultural practices (Snyder et al., 2009), need to be considered.

To date, the available information linking GHG emission and maizewinter CC rotation in the scientic literature is scarce. The most important knowledge gaps include effects of plant species selection and CC residue management (i.e., retention, incorporation or removal) (Basche et al., 2014).Cover crop species may affect N2O emissions in contrasting ways by inuencing abiotic and biotic soil factors. These factors include mineral N availability in soil and the availability of carbon (C) sources for the denitrier bacterial communities, soil pH, soil structure and microbial community composition (Abalos et al., 2014). For example, nonlegume CCs such as winter cereals could contribute to a reduction of N2O emissions due to their deep roots, which allow them to extract soil N more efciently than legumes (Kallenbach et al., 2010). Conversely, it has been suggested that the higher C : N ratio of their residues as compared to those of legumes may provide energy (C) for denitriers, thereby leading to higher N2O losses in the presence of mineral N-NO3 from fertilizers (Sarkodie-Addo et al., 2003). In this sense, the presence of cereal residues can increase the abundance of denitrifying microorganisms (Gao et al., 2016), thus enhancing denitrication losses when soil conditions are favorable (e.g., high NO3 availability and soil moisture after rainfall or irrigation events, particularly in ne-textured soils) (Stehfest and Bouwman 2006; Baral et al., 2016). Furthermore, winter CCs can also abate indirect gaseous N losses through the reduction of leaching and subsequent emissions from water resources (Feyereisen et al., 2006). Thus, the estimated N2O mitigation potential for winter CCs ranges from 0.2 to1.1 kg N2O ha1 yr1 according to Ussiri and Lal (2013).

In a CCmaize rotation system, mineral fertilizer applica

tion to the cash crop could have an important effect on N use efciency and N losses from the agro-ecosystem. Different methods for calculating the N application rate (e.g., conventional or integrated) can be employed by farmers, affecting the amount of synthetic N applied to soil and the overall effect of CCs on N2O uxes. Integrated soil fertility management (ISFM) (Kimani et al., 2003) provides an opportunity to optimize the use of available resources, thereby reducing pollution and costs from overuse of N fertilizers (conventional management). ISFM involves the use of inorganic fertilizers and organic inputs, such as green manure, and aims to maximize agronomic efciency (Vanlauwe et al., 2011).When applying this technique to a CCmaize crop rotation,

the N fertilization rate for maize is calculated taking into account the background soil mineral N and the expected available N from mineralization of CC residues, which depends on residue composition. Differences in soil mineral N during the cash crop phase may be signicantly reduced if ISFM practices are employed, affecting the GHG balance of the CCcash crop cropping system.

Only one study has investigated the effect of CCs on N2O emissions in Mediterranean cropping systems (Sanz-Cobena et al., 2014). These authors found an effect of CC species on N2O emissions during the intercrop period. After 4 years of

CC (vetch, barley or rape)maize rotation, vetch was the only CC species that signicantly enhanced N2O losses compared to fallow, mainly due to its capacity to x atmospheric N2 and because of higher N surplus from the previous cropping phases in these plots. In this study a conventional fertilization (same N synthetic rate for all treatments) was applied during the maize phase; how ISFM practices may affect these ndings remains unknown. Moreover, the relative contribution of mineral N fertilizer, CC residues and/or soil mineral N to N2O losses during the cash crop has not been assessed yet. In this sense, stable isotope analysis (i.e., 15N) represents a way to identify the source and the dominant processes involved in N2O production (Arah, 1997). Stable isotope techniques have been used in eld studies evaluating N leaching and/or plant recovery in systems with cover crops (Bergstrm et al., 2001; Gabriel and Quemada, 2011; Gabriel et al., 2016). Furthermore, some laboratory studies have evaluated the effect of different crop residues on N2O losses using 15N techniques (Baggs et al., 2003; Li et al., 2016), but to date, no previous studies have evaluated the relative contribution of cover crops (which include the aboveground biomass and the decomposition of root biomass) and N synthetic fertilizers to N2O emissions under eld conditions. A comprehensive understanding of the N2O biochemical production pathways and nutrient sources is crucial for the development of effective mitigation strategies.

The objective of this study was to evaluate the effect of two different CC species (barley and vetch) and fallow on GHG emissions during the CC period and during the following maize cash crop period in an ISFM system. An additional objective was to study the contribution of the synthetic fertilizer and other N sources to N2O emissions using

15N-labeled fertilizer. We hypothesized that (1) the presence of CCs instead of fallow would affect N2O losses, leading to higher emissions in the case of the legume CC (vetch) in accordance with the studies of Basche et al. (2014) and Sanz-Cobena et al. (2014) and (2) in spite of the ISFM during the maize period, which theoretically would lead to similar soil N availability for all plots, the distinct composition of the CC residues would affect N2O emissions. In order to test these hypotheses, a eld experiment was carried out using the same management system for 8 years, measuring GHGs during the 8th year. To gain a better understanding of the effect of the management practices tested on the overall GHG budget of a

Biogeosciences, 13, 52455257, 2016 www.biogeosciences.net/13/5245/2016/

G. Guardia et al.: Effect of cover crops on greenhouse gas emissions 5247

cropping system, CH4, CO2 and yield-scaled N2O emissions were also analyzed during the experimental period. The relative contribution of each N source (synthetic fertilizer or soil endogenous N, including N mineralized from the CCs) to N2O emissions was also evaluated by 15N-labeled ammonium nitrate in a parallel experiment.

2 Materials and methods

2.1 Site characteristics

The study was conducted at La Chimenea eld station (40 03[prime] N, 03 31[prime] W; 550 m a.s.l.), located in the central

Tajo River basin near Aranjuez (Madrid, Spain), where an experiment involving cover-cropping systems and conservation tillage has been carried out since 2006. Soil at the eld site is a silty clay loam (Typic Calcixerept; Soil Survey Staff, 2014). Some of the physicochemical properties of the top 010 cm soil layer, as measured by conventional methods, were as follows: pHH2O, 8.16; total organic

C, 19.0 g kg1; CaCO3, 198 g kg1; clay, 25 %; silt, 49 %; and sand, 26 %. Bulk density of the topsoil layer determined in intact core samples (Grossman and Reinsch, 2002) was 1.46 g cm 3. Average ammonium (NH+4) content at the beginning of the experiment was 0.42 [notdef] 0.2 mg N kg soil1

(without differences between treatments). Nitrate concentrations were 1.5 [notdef] 0.2 mg N kg soil1 in fallow and barley and

0.9 [notdef] 0.1 mg N kg soil1 in vetch. Initial dissolved organic C

(DOC) contents were 56.0 [notdef] 7 mg C kg soil1 in vetch and

fallow and 68.8 [notdef] 5 mg C kg soil1 in barley. The area has a

Mediterranean semiarid climate, with a mean annual air temperature of 14 C. The coldest month is January, with a mean temperature of 6 C, and the hottest month is August, with a mean temperature of 24 C. During the last 30 years, the mean annual precipitation has been approximately 350 mm (17 mm from July to August and 131 mm from September to November).

Hourly rainfall and air temperature data were obtained from a meteorological station located at the eld site (CR10X, Campbell Scientic Ltd., Shepshed, UK). A temperature probe inserted 10 cm into the soil was used to measure soil temperature. Mean hourly temperature data were stored on a data logger.

2.2 Experimental design and agronomic management

Twelve plots (12 m [notdef] 12 m) were randomly distributed in

four replications of three cover-cropping treatments, including a cereal and a legume: (1) barley (B) (Hordeum vulgare L. Vanessa), (2) vetch (V) (Vicia sativa L. Vereda), and (3) traditional winter fallow (F). Cover crop seeds were broadcast by hand over the stubble of the previous crop and covered with a shallow cultivator (5 cm depth) on 10 October 2013, at a rate of 180 and 150 kg ha1 for B and V, respectively. The cover-cropping phase nished on 14 March 2014

following local practices, with an application of glyphosate (N-phosphonomethyl glycine) at a rate of 0.7 kg a.e. ha1.

Even though the safe use of glyphosate has been under discussion for many years (Chang and Delzell, 2016), it was used in order to preserve the same killing method in all the campaigns in this long-term experiment under conservation tillage management. All of the CC residues were left on top of the soil. Thereafter, a new set of N fertilizer treatments was set up for the maize cash crop phase. Maize (Zea mays L., Pioneer P1574, FAO Class 700) was directly drilled on 7 April 2014 in all plots, resulting in a plant population density of7.5 plants m2; harvesting took place on 25 September 2014. The fertilizer treatments consisted of ammonium nitrate applied on 2 June at three rates: 170, 140 and 190 kg N ha1 in F, V and B plots, respectively, according to ISFM practices. For the calculation of each N rate, the N available in the soil (which was calculated following soil analysis as described below), the expected N uptake by maize crop, and the estimated N mineralized from V and B residues were taken into account, assuming that crop requirements were 236.3 kg N ha1 (Quemada et al., 2014). Estimated N use efciency of maize plants for calculating N application rate was 70 % according to the N use efciency obtained during the previous years in the same experimental area. Each plot received P as triple superphosphate (45 % P2O5, Fertiberia,

Madrid, Spain) at a rate of 69 kg P2O5 ha1, and K as potassium chloride (60 % K2O, Fertiberia, Madrid, Spain) at a rate of 120 kg K2O ha1 just before sowing maize. All N, P and K fertilizers were broadcast by hand, and immediately after N fertilization the eld was irrigated to prevent ammonia volatilization. The main crop previous to sowing CCs was sunower (Helianthus annuus L. Sambro). Neither the sunower nor the CCs were fertilized.

In order to determine the amount of N2O derived from the N fertilizers, double-labeled ammonium nitrate (15NH154NO3, 5 at.%

15N, from Cambridge Isotope Laboratories, Inc., Massachusetts, USA) was applied on 2 m [notdef] 2 m subplots established within each plot at a rate of

130 kg N ha1. In order to reduce biases due to the use of different N rates (e.g., apparent priming effects or different mixing ratios between the added and resident soil N pools) the same amount of N was applied for all treatments. In each subplot, the CC residue was also left on top of the soil. This application took place on 26 May by spreading the fertilizer homogenously with a hand sprayer, followed by an irrigation event.

Sprinkler irrigation was applied to the maize crop at a total amount of 688.5 mm in 31 irrigation events. Sprinklers were installed in a 12 m [notdef] 12 m framework. The water doses

to be applied were estimated from the crop evapotranspiration (ETc) of the previous week (net water requirements).This was calculated daily as ETc = Kc [notdef] ETo, where ETo is

reference evapotranspiration calculated by the FAO Penman

Monteith method (Allen et al., 1998) using data from the meteorological station located in the experimental eld. The

www.biogeosciences.net/13/5245/2016/ Biogeosciences, 13, 52455257, 2016

5248 G. Guardia et al.: Effect of cover crops on greenhouse gas emissions

crop coefcient (Kc) was obtained using the relationship for maize in semiarid conditions (Martnez-Cob, 2008).

Two different periods were considered for data reporting and analysis: Period I (from CC sowing to N fertilization of the maize crop) and Period II (from N fertilization of maize to the end of the experimental period, after maize harvest).

2.3 GHG emissions sampling and analyzing

Fluxes of N2O, CH4 and CO2 were measured from October 2013 to October 2014 using opaque, manually operated circular static chambers as described in detail by Abalos et al. (2013). One chamber (diameter 35.6 cm, height 19.3 cm) was located in each experimental plot. The chambers were hermetically closed (for 1 h) by tting them into stainless steel rings, which were inserted at the beginning of the study into the soil to a depth of 5 cm in order to minimize the lateral diffusion of gases and to avoid the soil disturbance associated with the insertion of the chambers in the soil. The rings were only removed during management events. Each chamber had rubber sealing tape to guarantee an airtight seal between the chamber and the ring and was covered with a radiant barrier reective foil to reduce temperature gradients between inside and outside. A rubber stopper with a three-way stopcock was placed in the wall of each chamber to take gas samples.Greenhouse gas measurements were always made with barley/vetch plants inside the chamber. During the maize period, gas chambers were set up between maize rows.

During Period I, GHGs were sampled weekly or every 2 weeks. During the rst month after maize fertilization, gas samples were taken twice per week. Afterwards, gas sampling was performed weekly or fortnightly, until the end of the cropping period. To minimize any effects of diurnal variation in emissions, samples were always taken at the same time of day (10:0012:00), which is reported as a representative time (Reeves and Wang, 2015).

Measurements of N2O, CO2 and CH4 emissions were made at 0, 30 and 60 min to test the linearity of gas accumulation in each chamber. Gas samples (100 mL) were removed from the headspace of each chamber by syringe and transferred to 20 mL gas vials sealed with a gastight neoprene septum. The vials were previously ushed in the eld using 80 mL of the gas sample. Samples were analyzed by gas chromatography using a HP-6890 gas chromatograph equipped with a headspace autoanalyzer (HT3), both from Agilent Technologies (Barcelona, Spain). Inert gases were separated by HP Plot-Q capillary columns. The gas chromatograph was equipped with a 63Ni electron-capture detector (micro-ECD) to analyze N2O concentrations, and with a ame ionization detector (FID) connected to a methanizer to measure CH4 and CO2 (previously reduced to CH4). The temperatures of the injector, oven and ECD were 50, 50 and 350 C, respectively. The accuracy of the gas chromatographic data was 1 % or better. Two gas standards comprising a mixture of gases (high standard with 1500 [notdef] 7.50 ppm

CO2, 10 [notdef] 0.25 ppm CH4 and 2 [notdef] 0.05 ppm N2O and low

standard with 200 [notdef] 1.00 ppm CO2, 2 [notdef] 0.10 ppm CH4 and

200 [notdef] 6.00 ppb N2O) were provided by Carburos Metlicos

S.A. and Air Products SA/NV, respectively, and used to determine a standard curve for each gas. The response of the GC was linear within 2001500 ppm for CO2 and 210 ppm

CH4 and quadratic within 2002000 ppb for N2O.

The increases in N2O, CH4 and CO2 concentrations within the chamber headspace were generally (80 % of cases) linear (R2 > 0.90) during the sampling period (1 h). Therefore, emission rates of uxes were estimated as the slope of the linear regression between concentration and time (after corrections for temperature) and from the ratio between chamber volume and soil surface area (MacKenzie et al., 1998). Cumulative N2O, CH4 and CO2 emissions per plot during the sampling period were estimated by linear interpolations between sampling dates, multiplying the mean ux of two successive determinations by the length of the period between sampling and adding that amount to the previous cumulative total (Sanz-Cobena et al., 2014). The measurement of CO2 emissions from soil, including plants in opaque chambers, only includes ecosystem respiration and not photosynthesis (Meijide et al., 2010).

2.4 15N isotope analysis

Gas samples from the subplots receiving double-labeled AN fertilizer were taken after 60 min of static chamber closure 1, 4, 9, 11, 15, 18, 22 and 25 days after fertilizer application. Stable 15N isotope analysis of N2O contained in the gas samples was carried out on a cryo-focusing gas chromatography unit coupled to a 20/20 isotope ratio mass spectrometer (both from SerCon Ltd., Crewe, UK). Ambient samples were taken occasionally as required for the subsequent isotopic calculations. Solutions of 6.6 and 2.9 at. % ammonium sulfate [(NH4)2SO4] were prepared and used to generate 6.6 and 2.9 at. % N2O (Laughlin et al., 1997), which were used as reference and quality control standards. In order to calculate the atom percent excess (APE) of the N2O emitted in the subplots, the mean natural abundance of atmospheric N2O from the ambient samples (0.369 at. % 15N) was subtracted from the measured enriched gas samples. To obtain the N2O ux that was derived from fertilizer (N2ONdff), the follow

ing equation was used (Senbayram et al., 2009):

N2ONdff = N2ON [notdef]

N2O_APEsample APEfertilizer

, (1)

in which N2O-N is the N2O emission from soil, N2O_APEsample is the 15N at. % excess of emitted N2O, and APEfertilizer is the 15N at. % excess of the applied fertilizer (Senbayram et al., 2009).

2.5 Soil and crop analyses

In order to relate gas emissions to soil properties, soil samples were collected at 010 cm depth during the growing sea-

Biogeosciences, 13, 52455257, 2016 www.biogeosciences.net/13/5245/2016/

G. Guardia et al.: Effect of cover crops on greenhouse gas emissions 5249

son on almost all gas-sampling occasions, particularly after each fertilization event. Three soil cores (2.5 cm diameter and 15 cm length) were randomly sampled close to the ring in each plot, and then mixed and homogenized in the laboratory. Soil NH+4 and NO3 concentrations were analyzed using 8 g of soil extracted with 50 mL of KCl (1 M), and measured by automated colorimetric determination using a ow injection analyzer (FIAS 400 Perkin Elmer) provided with a UV-visible spectrophotometer detector. Soil (DOC) was determined by extracting 8 g of homogeneously mixed soil with 50 mL of deionized water (and subsequently ltered) and analyzed with a total organic C analyzer (multi N/C 3100 Anal-ityk Jena) equipped with an IR detector. The water-lled pore space (WFPS) was calculated by dividing the volumetric water content by total soil porosity. Total soil porosity was calculated according to the following relationship: soil porosity

= (1 soil bulk density/2.65), assuming a particle density of

2.65 g cm3 (Danielson and Sutherland, 1986). Gravimetric water content was determined by oven-drying soil samples at 105 C with a Sartorius MA30.

Four 0.5 m [notdef] 0.5 m squares were randomly harvested from

each plot before killing the CC by applying glyphosate.

Aerial biomass was cut by hand at soil level, dried, weighed and ground. A subsample was taken for determination of total N content. From these samples the CC biomass and N contribution to the subsequent maize were determined.

At maize harvest, two 8 m central rows in each plot were collected and weighed in the eld following separation of grain and straw. For aboveground N uptake calculations, N content was determined in subsamples of grain and biomass.Total N content of maize and CC subsamples was determined with an elemental analyzer (TruMac CN, Leco).

2.6 Calculations and statistical analysis

Yield-scaled N2O emissions and N surplus in the maize cash crop were calculated as the amount of N2O emitted (considering the emissions of the whole experiment, i.e., Period I and Period II) per unit of aboveground N uptake and taking the difference between N application and aboveground N up-take, respectively (van Groenigen et al., 2010).

Statistical analyses were carried out with Statgraphics Plus 5.1. Analyses of variance were performed for all variables during the experiment (except climatic ones), for both periods indicated in Sect. 2.2. Data distribution normality and variance uniformity were previously assessed by the ShapiroWilk test and Levenes statistic, respectively, and transformed (log10, root square, arcsin or inverse) before analysis when necessary. Means of soil parameters were separated by Tukeys honest signicance test at P < 0.05, while cumulative GHG emissions, yield-scaled N2O emissions and

N surplus were compared by the orthogonal contrasts method at P < 0.05. For non-normally distributed data, the KruskalWallis test was used on non-transformed data to evaluate differences at P < 0.05. Linear correlations were carried out to

determine relationships between gas uxes and WFPS, soil temperature, DOC, NH+4 and NO3. These analyses were performed using the mean/cumulative data of the replicates of the CC treatments (n = 12), and also for all the dates when

soil and GHG were sampled, for Period I (n = 16), Period II

(n = 11) and the whole experimental period (n = 27).

3 Results

3.1 Cover crop (Period I)

3.1.1 Environmental conditions and WFPS

Mean soil temperature during the intercrop period was8.8 C, ranging from 1.8 (December) to 15.5 C (April) (Fig. 1a), which were typical values in the experimental area.Mean soil temperature during maize cropping period was24.6 C, which was also a standard value for this region. The accumulated rainfall during this period was 215 mm, whereas the 30-year mean is 253 mm. WFPS ranged from 40 to 81 % (Fig. 1b). No signicant differences were observed for WFPS mean values between the different treatments (P > 0.05).

3.1.2 Mineral N and DOC and cover crop residues

Topsoil NH+4 content was below 5 mg N kg soil1 most of the time in Period I, although a peak was observed after maize sowing (55 days after CC kill date) (Fig. 2a), with the highest values reached in B (50 mg N kg soil1).

Mean NH+4 content was signicantly higher in B than in F (P < 0.05), but daily NH+4 concentrations between treatments were only signicantly different between treatments on one sampling date (210 days after CC sowing). Nitrate content increased after CC killing, reaching values above 25 mg N kg soil1 in the V treatment (Fig. 2c). Mean NO3 content during Period I was signicantly higher in the V plots than in the B and F plots (P < 0.001). Dissolved organic C ranged from 60 to 130 mg C kg soil1 (Fig. 2e). Average topsoil DOC content was signicantly higher in B than in V and F (10 and 12 %, respectively, P < 0.01) but differences were only observed on some sampling dates. The total amount of cover crop biomass left on the ground was 540.5 [notdef] 26.5 and 1106.7 [notdef] 93.6 kg dry matter ha1 in B and

V, respectively. Accordingly, the total N content of these residues was 11.0 [notdef] 0.6 and 41.3 [notdef] 4.5 kg N ha1 in B and

V, respectively.

3.1.3 GHG uxes

Nitrous oxide uxes ranged from 0.06 to

0.22 mg N m2 d1 (Fig. 3a) in Period I. The soil acted as a sink for N2O at some sampling dates, especially for the F plots. Cumulative uxes at the end of Period I were significantly greater in CC treatments compared to F (1.6 and 2.6 higher in B and V, respectively) (P < 0.05; Table 1). Net CH4

www.biogeosciences.net/13/5245/2016/ Biogeosciences, 13, 52455257, 2016

5250 G. Guardia et al.: Effect of cover crops on greenhouse gas emissions

0

3.2 Maize crop (Period II)

3.2.1 Environmental conditions and WFPS

Mean soil temperature ranged from 19.6 (reached in September) to 32.3 C (reached in August) with a mean value of27.9 C (Fig. 1a). Total rainfall during the maize crop period was 57 mm. WFPS ranged from 19 to 84 % (Fig. 1c). Higher mean WFPS values (P < 0.01) were measured in B during some sampling dates.

3.2.2 Mineral N and DOC

Topsoil NH+4 content increased rapidly after N fertilization (Fig. 2b), decreasing to values below 10 mg N kg soil1 from 15 days after fertilization to the end of the experimental period. Nitrate concentrations (Fig. 2d) also peaked after AN addition, reaching the highest value (170 mg N kg soil1)

15 days after fertilization in B (P < 0.05). No signicant differences (P > 0.05) between treatments were observed in average soil NH+4 or NO3 during the maize phase. Dissolved organic C ranged from 56 to 138 mg C kg soil1 (Fig. 2f).Average topsoil DOC content was 26 and 44 % higher in B than in V and F, respectively (P < 0.001).

3.2.3 GHG uxes, yield-Scaled N2O emissions and N surplus

Nitrous oxide uxes ranged from 0.0 to 5.6 mg N m2 d1 (Fig. 3b). The highest N2O emission peak was observed 1 4 days after fertilization for all plots. Other peaks were subsequently observed until 25 days after fertilization, particularly in B plots, where N2O emissions 23 and 25 days after fertilization were higher (P < 0.05) than those of F and V (Fig. 3b). No signicant differences in cumulative N2O uxes were observed between treatments throughout or at the end of the maize crop period (Table 1), although uxes were numerically higher in B than in V (0.05 < P < 0.10). Daily N2O emissions were signicantly correlated with NH+4 top-soil content (P < 0.05, n = 12, r = 0.84).

As in the previous period, all treatments were CH4 sinks, without signicant differences between treatments (P > 0.05; Table 1). Respiration rates ranged from 0.15 to3.0 g C m2 d1; no signicant differences (P > 0.05; Table 1) were observed among the CO2 values for the different treatments. Yield-scaled N2O emissions and N surplus are shown in Table 1. No signicant differences were ob-served between treatments, although these values were generally lower in V than in B (0.05 < P < 0.15).

Considering the whole cropping period (Period I and Period II), N2O uxes signicantly correlated with

WFPS (P < 0.05, n = 12, r = 0.61) NH+4 (P < 0.05, n = 27,

r = 0.84) and NO3 (P < 0.05, n = 27, r = 0.50).

Biogeosciences, 13, 52455257, 2016 www.biogeosciences.net/13/5245/2016/

(a)

Irrigation Rainfall Soil temp

Period I Period II

40

35

30

Irrigation/rainfall (mm)

Temperature (C)

30

25

20

20

15

10

10

5

0

Date

(b)

F V B

100

90

80

70

60

WFPS (%)

50

40

30

20

10

0 0 20 40 60 80 100 120 140 160 180 200 220 240

Days after CC sowing

(c)

fig01

F V B

100

90

80

70

WFPS (%)

60

50

40

30

20

10

0 -10 10 30 50 70 90 110 130

Days after N fertilization

Figure 1. Daily mean soil temperature ( C) rainfall and irrigation (mm) (a) and soil WFPS (%) in the three cover crop (CC) treatments (fallow, F; vetch, V; and barley, B) during Period I (b) and II (c). Vertical lines indicate standard errors.

uptake was observed in all intercrop treatments, and daily uxes ranged from 0.60 to 0.25 mg C m2 d1 (data not

shown). No signicant differences were observed between treatments in cumulative CH4 uxes at the end of Period I (P > 0.05; Table 1). Carbon dioxide uxes (data not shown) remained below 1 g C m2 d1 during the intercrop period.

The greatest uxes were observed in B, although differences in cumulative uxes were not signicant (P > 0.05; Table 1) in the whole intercrop period, but soil respiration was increased in B, with respect to F, from mid-February to the end of Period I. Nitrous oxide emissions were signicantly correlated to CO2 uxes (P < 0.01, n = 17, r = 0.69) and

soil temperature (P < 0.05, n = 17, r = 0.55).

G. Guardia et al.: Effect of cover crops on greenhouse gas emissions 5251

(a)

F V B

(b)

F V B

70

70

60

60

mg NH 4+- N kg soil-1

mg NH 4+-N kg soil-1

50

50

40

40

30

30

20

20

10

10

0 0 20 40 60 80 100 120 140 160 180 200 220 240

0 -10 10 30 50 70 90 110 130

(c)

F V B

(d)

F V B

35

250

30

mg NO 3- -N kg soil-1

mg NO 3- -N kg soil-1

200

25

20

150

15

100

10

50

5

0 0 20 40 60 80 100 120 140 160 180 200 220 240

0 -10 10 30 50 70 90 110 130

(e)

F V B

(f)

F V B

180

180

160

160

mg DOC kg soil-1

mg DOC kg soil-1

140

140

120

120

100

100

80

80

60

60

40

40

20

20

0 0 20 40 60 80 100 120 140 160 180 200 220 240

0 -10 10 30 50 70 90 110 130

Days after CC sowing

Days after N fertilization

fig02

Figure 2. (a, b) NH+

4 -N, (c, d) NO

3 -N and (e, f) DOC concentrations in the 010 cm soil layer for the three CC treatments (fallow, F; vetch, V; and barley, B) during both cropping periods. The black arrows indicate the time of spraying glyphosate over the CCs. The dotted arrows indicate the time of maize sowing. Vertical lines indicate standard errors.

3.2.4 Fertilizer-derived N2O emissions

The proportion (%) of N2O losses from ammonium nitrate, calculated by isotopic analyses, is represented in Fig. 4. The highest percentages of N2O uxes derived from the synthetic fertilizer were observed 1 day after fertilization, ranging from 34 % (V) to 67 % (B). On average, almost 50 % of N2O emissions in the rst sampling event after N synthetic fertilization came from other sources (i.e., soil endogenous N, including N mineralized from the CCs). The mean percentage of N2O losses from synthetic fertilizer throughout all sampling dates was 2.5 times higher in B compared to V (P < 0.05) and was positively correlated with DOC concentrations (P < 0.05, n = 12, r = 0.71). There were no signi-

cant differences between V and F (P > 0.05).

4 Discussion

4.1 Role of CCs in N2O emissions: Period I

Cover crop treatments (V and B) increased N2O losses compared to F, especially in the case of V (Table 1). These results are consistent with the meta-analysis of Basche et al. (2014), which showed that, overall, CCs increase N2O uxes (compared to bare fallow), with highly signicant increments in the case of legumes and a lower effect in the case of non-legume CCs. In the same experimental area, Sanz-Cobena et al. (2014) found that V was the only CC signicantly affecting N2O emissions. The greatest differences between treatments were observed at the beginning (1340 days after CC sowing) and at the end of this period (229 days after CC sowing) (Fig. 3a). On these dates, the mild soil temperatures and the relatively high moisture content were more suitable for soil biochemical processes, which may trigger

www.biogeosciences.net/13/5245/2016/ Biogeosciences, 13, 52455257, 2016

5252 G. Guardia et al.: Effect of cover crops on greenhouse gas emissions

Table 1. Total cumulative N2O-N, CH4-C and CO2-C uxes; N surplus; and yield-scaled N2O emissions in the three CC treatments (fallow, F; vetch, V; and barley, B) at the end of both cropping periods. P values were calculated with Students t test and df = 9.

Treatment N2O CH4 CO2 Surplus Yield-scaled N2O emissions kg N2O-N ha1 kg CH4-C ha1 kg CO2-C ha1 kg N ha1 g N2O-N kg aboveground N uptake1

Estimate 11.48 11.45 134.37

t test 2.5 0.61 1.00

P value 0.03 0.56 0.34

End of Period I

F 0.05 0.30 443.02

V 0.13 -0.28 463.01 B 0.08 0.24 582.13

SE 0.03 0.07 46.33

F vs. CCs

V vs. B

Estimate 5.29 6.23 127.50

t test 1.99 0.57 1.64

P value 0.08 0.58 0.14

Estimate 7.46 23.69 83.36 3.16 0.12

t test 0.30 1.25 0.19 0.08 0.14

P value 0.77 0.24 0.86 0.94 0.89

End of Period II

F 0.57 0.46 2595.07 31.47 4.21

V 0.48 0.33 2778.84 13.72 3.06

B 0.74 0.35 2372.07 55.94 5.64

SE 0.10 0.08 177.35 15.30 0.85

F vs. CCs

Estimate 26.59 2.08 417.8 38.67 2.59

t test 1.90 0.19 1.62 1.79 2.16

P value 0.09 0.85 0.14 0.11 0.06

and SE denote signicant at P < 0.05 and the standard error of the mean, respectively.

V vs. B

through leaching, resulting in low concentrations of soil mineral N in F plots.

Nitrous oxide emissions were low during this period but in the range of those reported by Sanz-Cobena et al. (2014) in the same experimental area. Total emissions during Period I represented 8, 10 and 21 % of total cumulative emissions in F, B and V, respectively (Table 1). The absence of N fertilizer application to the soil combined with the low soil temperatures during winter which were far from the optimum values for nitrication and denitrication (2530 C) processes (Ussiri and Lal, 2013) may have caused these low N2O uxes. The signicant positive correlation between soil temperature and N2O uxes during this period highlights the key role of this parameter as a driver of soil emissions (Schindlbacher et al., 2004; Garca-Marco et al., 2014).

4.2 Role of CCs in N2O emissions: Period II

Isotopic analysis during Period II, in which ISFM was carried out, showed that a signicant proportion of N2O emissions came from endogenous soil N or the mineralization of crop residues, especially after the rst few days following N fertilization (Fig. 4). In this sense, even though an interaction between crop residue and N fertilizer application has been previously described (e.g., in Abalos et al., 2013), the similar proportion of N2O losses coming from fertilizer in B and F (without residue) 1 day after N fertilization revealed the importance of soil mineral N contained in the micropores for

Biogeosciences, 13, 52455257, 2016 www.biogeosciences.net/13/5245/2016/

N2O emissions (Fig. 1a, b) (Firestone and Davidson, 1989). Average topsoil NO3 was signicantly higher in V (Fig. 2b), which was the treatment that led to the highest N2O emissions. Legumes such as V are capable of biologically xing atmospheric N2, thereby increasing soil NO3 content with the potential to be denitried. Furthermore, the mineralization of the most recalcitrant fraction of the previous V residue (which supplies nearly 4 times more N than the B residue, as indicated in Sect. 3.1.2) together with high C-content sunower residue could also explain higher NO3 contents in V plots (Frimpong et al., 2011) and higher N2O losses from denitrication (Baggs et al., 2000). After the CC kill date, N release from decomposition of roots and nodules and faster mineralization of V residue compared to that of B (shown by NO3 in soil in Fig. 2c) are the most plausible explanations for the N2O increases at the end of the intercrop period (Fig. 3a) (Rochette and Janzen, 2005; Wichern et al., 2008).

Some studies (e.g., Justes et al., 1999; Nemecek et al., 2008) have pointed out that N2O losses can be reduced with the use of CCs, due to the extraction of plant-available N unused by previous cash crop. However, in our study lower N2O emissions were measured from F plots without CCs during the intercrop period. This may be a consequence of higher NO3 leaching in F plots (Gabriel et al., 2012; Quemada et al., 2013), limiting the availability of the substrate for denitrication. Frequent rainfall during the intercrop period (Fig. 1a) and the absence of N uptake by CCs may have led to N losses

G. Guardia et al.: Effect of cover crops on greenhouse gas emissions 5253

-0,2

(a)

F V B

F V B

0,4

100

90

mg N 2O -N m-2 d-1

% N 2O from N fertilizer

0,3

80

0,2

70

F vs CCs: NS

V vs. B: * (P=0.04)

60

0,1

50

0,0

40

30

-0,1

20

10

0 20 40 60 80 100 120 140 160 180 200 220 240

0 1 4 9 11 15 18 22 25 Average

Days after CC sowing

Days after N fertilization

(b)

F V B

fig04

Figure 4. Proportion of N2O losses (%) that come from N synthetic fertilizer during Period II, for the three CC treatments (fallow,

F; vetch, V; and barley, B). Vertical lines indicate standard errors. NS and denote not signicant and signicant at P < 0.05, respectively.

8

7

6

mg N 2O -N m-2 d-1

5

4

3

2

1

residue (with low C : N ratio) provided an additional N source for soil microorganisms, thus decreasing the relative amount of N2O derived from the synthetic fertilizer (Baggs et al., 2000; Shan and Yan, 2013); and (iv) V plots were fertilized with a lower amount of immediately available N (i.e., ammonium nitrate) than B plots, which could have resulted in better synchronization between N release and crop needs (Ussiri and Lal, 2013) in V plots. Supporting these ndings, Bayer et al. (2015) recently concluded that partially supplying the maize N requirements with winter legume cover crops can be considered a N2O mitigation strategy in subtropical agroecosystems.

The mineralization of B residues resulted in higher DOC contents for these plots compared to the F or V plots (P < 0.001). This was observed in both Period I (as a consequence of soil C changes after the 8-year cover-cropping management) and Period II (due to the CC decomposition).Although in the present study the correlation between DOC and N2O emissions was not signicant, positive correlations have been previously found in other low-C Mediterranean soils (e.g., Vallejo et al., 2006; Lpez-Fernndez et al., 2007).Some authors have suggested that residues with a high C : N ratio can induce microbial N immobilization (Frimpong and Baggs, 2010; Dendooven et al., 2012). In our experiment, a N2O peak was observed in B plots 2025 days after fertilization (Fig. 3b) after a remarkable increase of NO3 content (Fig. 2d), which may be a result of a remineralization of previously immobilized N in these plots.

The positive correlation of N2O uxes and soil NO3 content and WFPS during the whole cycle further supports the importance of denitrication process for explaining N2O losses in this agro-ecosystem (Davidson et al., 1991; Garca-Marco et al., 2014). However, the strong positive correlation of N2O with NH+4 indicated that nitrication was also a major process leading to N2O uxes, and showed that the continuous drying-wetting cycles during a summer irri-

www.biogeosciences.net/13/5245/2016/ Biogeosciences, 13, 52455257, 2016

0 -10 10 30 50 70 90 110 130

Days after N fertilization

fig03

Figure 3. N2O emissions for the three CC treatments (fallow, F; vetch, V; and barley, B) during Period I (a) and II (b). The black arrows indicate the time of spraying glyphosate over the CCs. The dotted arrows indicate the time of maize sowing. Vertical lines indicate standard errors.

the N2O bursts after the rst irrigation events, with respect to the N released from CC residues.

As we hypothesized, the different CCs played a key role in the N2O emissions during Period II. Barley plots had higher N2O emissions than fallow or V-residue plots (at the 10 % signicance level; Table 1). Further, a higher proportion of N2O emissions was derived from the fertilizer in B-residue than in V-residue plots (Fig. 4). These results are in agreement with those of Baggs et al. (2003), who reported a higher percentage of N2O derived from the 15N-labeled fertilizer using a cereal (ryegrass) as surface mulching instead of a legume (bean), in a eld trial with zero-tillage management. The differences between B and V in terms of cumulative N2O emissions and in the relative contribution of each source to these emissions (fertilizer- or soil-N) could be explained by: (i) the higher C : N residue of B (20.7 [notdef] 0.7

while that of V was 11.1 [notdef] 0.1, according to Alonso-Ayuso

et al., 2014) may have provided an energy source for denitrication (Sarkodie-Addo et al., 2003), favoring the reduction of the NO3 supplied by the synthetic fertilizer and enhancing N2O emissions, as supported by the positive correlation of DOC with the proportion of N2O coming from the synthetic fertilizer; (ii) NO3 concentrations, which tended to be higher in B during the maize cropping phase, could have led to incomplete denitrication and larger N2O / N2 ratios (Yamulki and Jarvis, 2002); (iii) the easily mineralizable V

5254 G. Guardia et al.: Effect of cover crops on greenhouse gas emissions

gated maize crop in a semiarid region can lead to favorable WFPS conditions for both nitrication and denitrication processes (Fig. 1c) (Bateman and Baggs, 2005). Emission factors ranged from 0.2 to 0.6 % of the synthetic N applied, which were lower than the IPCC default value of 1 %. As explained above, ecological conditions during the intercrop period (rainfall and temperature) and maize phase (temperature) could be considered normal (based on the 30-year average) in Mediterranean areas. Aguilera et al. (2013) obtained a higher emission factor for high (1.01 %) and low (0.66 %) water-irrigation conditions in a meta-analysis of Mediterranean cropping systems. We hypothesized that management practices may have contributed to these low emissions, but other inherent factors such as soil pH should also be considered. Indeed, a higher N2O / N2 ratio has been associated with acidic soils, so lower N2O emissions from denitrication could be expected in alkaline soils (Mrkved et al., 2007;Baggs et al., 2010).

4.3 Methane and CO2 emissions

As is generally found in non-ooded arable soils, all treatments were net CH4 sinks (Snyder et al., 2009). No significant differences were observed between treatments in any of the two periods (Table 1), which is similar to the pattern observed by Sanz-Cobena et al. (2014). Some authors (Duneld and Knowles, 1995; Tate, 2015) have suggested an inhibitory effect of soil NH+4 on CH4 uptake. Low NH+4 contents during almost all of the CC and maize cycle may explain the apparent lack of this inhibitory effect (Banger et al., 2012). However, during the dates when the highest NH+4 contents were reached in V and B (225 days after CC sowing) (Fig. 3a), CH4 emissions were signicantly higher for these plots (0.12 and 0.16 mg CH4-C m2 d1 for V and B, respectively) than for F (0.01 mg CH4-C m2 d1) (data

not shown). Similarly, the NH+4 peak observed 2 days after fertilization (Fig. 3b) decreased in the order V > F > B, the same trend as CH4 emissions (which were 0.03, 0.04

and 0.63 mg CH4-C m2 d1 in V, F and B, respectively;

data not shown). Contrary to Sanz-Cobena et al. (2014), the presence of CCs did not increase CO2 uxes (Table 1) during the whole of Period I (which was longer than the period considered by these authors), even though higher uxes were associated with B (but not V) with respect to F plots in the last phase of the intercrop. This was probably as a consequence of higher root biomass and plant respiration rates in the cereal (B) than in the legume (V). Differences from fall to early winter were not signicant, since low soil temperatures limited respiration activity. The decomposition of CC residues and the growth of the maize rooting system resulted in an increase in CO2 uxes during Period II (Oorts et al., 2007; Chirinda et al., 2010), although differences between treatments were not observed.

4.4 Yield-scaled emissions, N surplus and general assessment

Yield-scaled N2O emissions ranged from 1.74 to 7.15 g N2ON kg aboveground N uptake1, which is about 14 times lower than those reported in the meta-analysis of van Groenigen et al. (2010) for a fertilizer N application rate of 150 200 kg ha1. Mean N surpluses of V and F (Table 1) were in the range (050 kg N ha1) recommended by van Groenigen et al. (2010), while the mean N surplus in B (55 kg N ha1)

was also close to optimal. In spite of higher N2O emissions in V during Period I (which accounted for a low proportion of total cumulative N2O losses during the experiment), these plots did not emit greater amounts of N2O per kg of N taken up by the maize plants, and even tended to decrease yield-scaled N2O emissions and N surplus (Table 1).

Adjusting fertilizer N rate to soil endogenous N led to lower N2O uxes than previous experiments where conventional N rates were applied (e.g., Adviento-Borbe et al., 2007; Hoben et al., 2011; Sanz-Cobena et al., 2012; Li et al., 2015), in agreement with the study by Migliorati et al. (2014). Moreover, CO2 equivalent emissions associated with manufacturing and transport of N synthetic fertilizers (Lal, 2004) can be reduced when low synthetic N input strategies, such as ISMF, are employed. Our results highlight the critical importance of the cash crop period on total N2O emissions and demonstrate that the use of nonlegume and particularly legume CCs combined with ISFM may provide an optimum balance between GHG emissions from crop production and agronomic efciency (i.e., lowering synthetic N requirements for a subsequent cash crop, and leading to similar yield-scaled N2O emissions as fallow).

The use of CCs has environmental implications beyond effects on direct soil N2O emissions. For instance, CCs can mitigate indirect N2O losses (from NO3 leaching). In the study by Gabriel et al. (2012), conducted in the same experimental area, NO3 leaching was reduced (on average)

by 30 and 59 % in V and B, respectively. Considering an emission factor of 0.075 from N leached (De Klein et al., 2006), indirect N2O losses from leaching could be mitigated by 0.23 [notdef] 0.16 and 0.45 [notdef] 0.17 kg N ha1 yr1 if V and B

are used as CCs, respectively. Furthermore, the recent meta-analysis of Poeplau and Don (2015) revealed a C sequestration potential of 0.32 [notdef] 0.08 Mg C ha1 yr1 with the in

troduction of CCs. These environmental factors, together with CO2 emissions associated with CC sowing and killing, should be assessed in future studies in order to conrm the potential in CCs for increasing both the agronomic and the environmental efciency of irrigated cropping areas.

Biogeosciences, 13, 52455257, 2016 www.biogeosciences.net/13/5245/2016/

G. Guardia et al.: Effect of cover crops on greenhouse gas emissions 5255

5 Conclusions

Our study conrmed that the presence of CCs (particularlyV) during the intercrop period increased N2O losses, but the contribution of this phase to cumulative N2O emissions, considering the whole cropping cycle (intercropcash crop), was low (821 %). The high inuence of the maize crop period over total N2O losses was due to not only N synthetic fertilization, but also CC residue mineralization and especially endogenous soil N. The type of CC residue determined the N synthetic rate in an ISFM system and affected the percentage of N2O losses coming from N fertilizer/soil N, as well as the pattern of N2O losses during the maize phase (through changes in soil NH+4, NO3 and DOC concentrations). By employing ISFM, similar N2O emissions were measured from CCs and F treatments at the end of the whole cropping period, resulting in low yield-scaled N2O emissions (36 g N2O-N kg aboveground N uptake1) and N surplus (31 to 56 kg N ha1). Replacing winter F with CCs did not signicantly affect CH4 uptake or respiration rates, during either intercrop or maize cropping periods. Our results highlight the critical importance of the cash crop period on total N2O emissions, and demonstrate that the use of nonlegume and particularly legume CCs combined with ISFM could be considered an efcient practice from both environmental and agronomic points of view, leading to similar N2O losses per kilogram of aboveground N uptake to bare fallow.

6 Data availability

Our row data will be accessible through the repository of the Technical University of Madrid (UPM) http://oa.upm.es/contact/

Web End =http://oa.upm.es/ contact/.

Acknowledgements. The authors are grateful to the Spanish Ministry of Economy and Innovation and the Community of Madrid for their economic support through projects AGL2012-37815-C05-01-AGR and the Agrisost-CM project (S2013/ABI- 2717).We also thank the technicians and researchers at the Department of Chemistry and Agricultural Analysis of the Agronomy Faculty (Technical University of Madrid, UPM). Rothamsted Research is grant funded by the Biotechnology and Biological Sciences Research Council (BBSRC), UK.

Edited by: E. VeldkampReviewed by: two anonymous referees

Abalos, D., Deyn, G. B., Kuyper, T. W., and van Groenigen, J. W.: Plant species identity surpasses species richness as a key driver of N2O emissions from grassland, Glob. Change Biol., 20, 265

275, 2014.

Adviento-Borbe, M. A. A., Haddix, M. L., Binder, D. L., Walters,D. T., and Dobermann, A.: Soil greenhouse gas uxes and global warming potential in four high-yielding maize systems, Glob.Change Biol., 13, 19721988, 2007.

Aguilera, E., Lassaletta, L., Sanz-Cobena, A., Garnier, J., and Vallejo, A.: The potential of organic fertilizers and water management to reduce N2O emissions in Mediterranean climate cropping systems. A review, Agr. Ecosyst. Environ., 164, 3252, 2013.

Allen, R. G., Raes, L. S., and Smith, D. M.: Crop evapotranspiration, Guidelines for computing crop water requirements, Irrigation and Drainage, Paper 56, Rome, Italy, FAO, 1998.Alonso-Ayuso, M., Gabriel, J. L., and Quemada, M.: The kill date as a management tool for cover cropping success, Plos One, 9, e109587, doi:http://dx.doi.org/10.1371/journal.pone.0109587

Web End =10.1371/journal.pone.0109587 http://dx.doi.org/10.1371/journal.pone.0109587

Web End = , 2014.

Arah, J. R. M.: Apportioning nitrous oxide uxes between nitrication and denitrication using gas-phase mass spectrometry, Soil Biol. Biochem., 29, 12951299, 1997.

Baggs, E. M., Rees, R. M., Smith, K. A., and Vinten, A. J.A.: Nitrous oxide emission from soils after incorporating crop residues, Soil Use Manage., 16, 8287, 2000.

Baggs, E. M., Stevenson, M., Pihlatie, M., Regar, A., Cook, H., and Cadisch, G.: Nitrous oxide emissions following application of residues and fertiliser under zero and conventional tillage, Plant Soil, 254, 361370, 2003.

Baggs, E. M., Smales, C. L., and Bateman, E. J.: Changing pH shifts the microbial sourceas well as the magnitude of N2O emission from soil, Biol. Fert. Soils, 46, 793805, 2010.

Banger, K., Tian, H., and Lu, C.: Do nitrogen fertilizers stimulate or inhibit methane emissions from rice elds?, Glob. Change Biol., 18, 32593267, 2012.

Baral, K. R., Arthur, E., Olesen, J. E., and Petersen, S. O.: Predicting nitrous oxide emissions from manure properties and soil moisture: An incubation experiment, Soil Biol. Biochem., 97, 112 120, 2016.

Basche, A. D., Miguez, F. E., Kaspar, T. C., and Castellano, M. J.:

Do cover crops increase or decrease nitrous oxide emissions? A meta-analysis, J. Soil Water Conserv., 69, 471482, 2014.Bateman, E. J. and Baggs, E.M.: Contributions of nitrication and denitrication to N2O emissions from soils at different water-lled pore space, Biol. Fert. Soils, 41, 379388, 2005.

Bayer, C., Gomes, J., Zanatta, J. A., Vieira, F. C. B., de Cssia Piccolo, M., Dieckow, J., and Six, J.: Soil nitrous oxide emissions as affected by long-term tillage, cropping systems and nitrogen fertilization in Southern Brazil, Soil Till. Res., 146, 213222, 2015.

Bergstrm, L. F. and Jokela, W. E.: Ryegrass Cover Crop Effects on Nitrate Leaching in Spring Barley Fertilized with(15)NH4(15)NO3, J. Environ. Qual., 30, 16591667, 2001.

Chang, E. T. and Delzell, E.: Systematic review and meta-analysis of glyphosate exposure and risk of lymphohematopoietic cancers, J. Environ. Sci. Heal. B, 51, 402434, 2016.

Chirinda, N., Olesen, J. E., Porter, J. R., and Schjnning, P.: Soil properties, crop production and greenhouse gas emissions from organic and inorganic fertilizer-based arable cropping systems, Agr. Ecosyst. Environ., 139, 584594, 2010.

www.biogeosciences.net/13/5245/2016/ Biogeosciences, 13, 52455257, 2016

References

Abalos, D., Sanz-Cobena, A., Garcia-Torres, L., van Groenigen, J.W., and Vallejo, A.: Role of maize stover incorporation on nitrogen oxide emissions in a non-irrigated Mediterranean barley eld, Plant Soil, 364, 357371, 2013.

5256 G. Guardia et al.: Effect of cover crops on greenhouse gas emissions

Danielson, R. E. and Sutherland, P. L.: Porosity, in: Methods of Soil Analysis. Part I. Physical and Mineralogical Methods, edited by: Klute, A., Agronomy Monograph 9, American Society of Agronomy, Soil Science Society of America, Madison, WI, 443461, 1986.

Davidson, E. A.: Fluxes of nitrous oxide and nitric acid from terrestrial ecosystem, in: Microbial production and consumption of greenhouse gases: Methane, Nitrous oxide and Halomethane, edited by: Rogers, J. E. and Whitman, W. B., American Society of Microbiology, Washington, 219236, 1991.

Davidson, E. A. and Kanter, D.: Inventories and scenarios of nitrous oxide emissions, Environ. Res. Lett., 9, 105012, doi:http://dx.doi.org/10.1088/1748-9326/9/10/105012

Web End =10.1088/1748-9326/9/10/105012 http://dx.doi.org/10.1088/1748-9326/9/10/105012

Web End = , 2014.

De Klein, C., Novoa, R. S. A., Ogle, S., Smith, K. A., Rochette,P., Wirth, T. C., Mc Conket, B. G., Walsh, M., Mosier, A., Rypdal, K., and Williams, S. A.: IPCC guidelines for national greenhouse gas inventories, Volume 4, Chapter 11: N2O emissions from managed soils, and CO2 emissions from lime and urea application, Technical Report 4-88788-032-4, Intergovernmental

Panel on Climate Change, 2006.

Dendooven, L., Patino-Zniga, L., Verhulst, N., Luna-Guido, M., Marsch, R., and Govaerts, B.: Global warming potential of agricultural systems with contrasting tillage and residue management in the central highlands of Mexico, Agr. Ecosyst. Environ., 152, 5058, 2012.

Duneld, P. and Knowles, R.: Kinetics of inhibition of methane oxidation by nitrate, nitrite, and ammonium in a humisol, Appl.Environ. Microb., 61, 31293135, 1995.

Feyereisen, G. W., Wilson, B. N., Sands, G. R., Strock, J. S., and

Porter, P. M.: Potential for a rye cover crop to reduce nitrate loss in southwestern Minnesota, Agron. J., 98, 14161426, 2006.Firestone, M. K. and Davidson, E. A.: Microbiological basis of NO and N2O production and consumption in soil, in: Exchange of

Trace Gases between Terrestrial Ecosystems and the Atmosphere Chichester, edited by: Andeae, M. O. and Schimel, D. S., Wiley, 721, 1989.

Frimpong, K. A. and Baggs, E. M.: Do combined applications of crop residues and inorganic fertilizer lower emission of N2O from soil?, Soil Use Manage., 26, 412424, 2010.

Frimpong, K. A., Yawson, D. O., Baggs, E. M., and Agyarko, K.: Does incorporation of cowpea-maize residue mixes inuence nitrous oxide emission and mineral nitrogen release in a tropical luvisol?, Nutr. Cycl. Agroecosys., 91, 281292, 2011.

Gabriel, J. L. and Quemada, M.: Replacing bare fallow with cover crops in a maize cropping system: yield, N uptake and fertiliser fate, Eur. J. Agron., 34, 133143, 2011.

Gabriel, J. L., Muoz-Carpena, R., and Quemada, M.: The role of cover crops in irrigated systems: Water balance, nitrate leaching and soil mineral nitrogen accumulation, Agr. Ecosyst. Environ., 155, 5061, 2012.

Gabriel, J. L., Alonso-Ayuso, M., Garca-Gonzlez, I., Hontoria, C., and Quemada, M.: Nitrogen use efciency and fertiliser fate in a long-term experiment with winter cover crops, Eur. J. Agron., 79, 1422, 2016.

Gao, J., Xie, Y., Jin, H., Liu, Y., Bai, X., Ma, D., Zhu, Y., Wang,C., and Guo, T.: Nitrous Oxide Emission and Denitrier Abundance in Two Agricultural Soils Amended with Crop Residues and Urea in the North China Plain, Plos One, 11, e0154773, doi:http://dx.doi.org/10.1371/journal.pone.0154773

Web End =10.1371/journal.pone.0154773 http://dx.doi.org/10.1371/journal.pone.0154773

Web End = , 2016.

Garca-Marco, S., Ravella, S. R., Chadwick, D., Vallejo, A., Gregory, A. S., and Crdenas, L. M.: Ranking factors affecting emissions of GHG from incubated agricultural soils, Eur. J. Soil Sci., 65, 573583, 2014.

Grossman, R. B. and Reinsch, T. G.: 2.1 Bulk density and linear extensibility. Methods of Soil Analysis. Part 4: Physical Methods, Soil Science Society of America, Madison, USA, 201228, 2002.

Hoben, J. P., Gehl, R. J., Millar, N., Grace, P. R., and Robertson, G.P.: Nonlinear nitrous oxide (N2O) response to nitrogen fertilizer in on-farm corn crops of the US Midwest, Glob. Change Biol., 17, 11401152, 2011.

IPCC: Climate change 2007. The Physical Science Basis. Contribution of Working Group I to the Fourth Assessment Report of the Intergovernmental Panel on Climate Change, edited by: Solomon, S., Qin, D., Manning, M., Chen, Z., Marquis, M., Averyt, K. B., Tignor, M., and Miller, H. L., Cambridge University Press, p. 996, 2007.

Justes, E., Mary, B., and Nicolardot, B.: Comparing the effectiveness of radish cover crop, oilseed rape volunteers and oilseed rape residues incorporation for reducing nitrate leaching, Nutr. Cycl. Agroecosys., 55, 207220, 1999.

Kallenbach, C. M., Rolston, D. E., and Horwath, W. R.: Cover cropping affects soil N2O and CO2 emissions differently depending on type of irrigation, Agr. Ecosyst. Environ., 137, 251260, 2010.

Kimani, S. K., Nandwa, S. M., Mugendi, D. N., Obanyi, S. N., Ojiem, J., Murwira, Herbert K., and Bationo, A.: Principles of integrated soil fertility management, in: Soil fertility management in Africa: A regional perspective, edited by: Gichuri, M.P., Bationo, A., Bekunda, M. A., Goma, H. C., Mafongoya, P.L., Mugendi, D. N., Murwuira, H. K., Nandwa, S. M., Nyathi,P., and Swift, M. J., Academy Science Publishers (ASP), Centro Internacional de Agricultura Tropical (CIAT), Tropical Soil Biology and Fertility (TSBF), Nairobi, KE, 5172, 2003.

Lal, R.: Carbon emission from farm operations, Environ. Int. 30,981990, 2004.

Laughlin, R. J., Stevens, R. J., and Zhuo, S.: Determining nitrogen-15 in ammonium by producing nitrous oxide, Soil Sci. Soc. Am.J., 61, 462465, 1997.

Li, N., Ning, T., Cui, Z., Tian, S., Li, Z., and Lal, R.: N2O emissions and yield in maize eld fertilized with polymer-coated urea under subsoiling or rotary tillage, Nutr. Cycl. Agroecosys., 102, 397 410, 2015.

Li, X., Srensen, P., Olesen, J. E., and Petersen, S. O.: Evidence for denitrication as main source of N2O emission from residue-amended soil, Soil Biol. Biochem., 92, 153160, 2016.

Lpez-Fernndez, S., Diez, J. A., Hernaiz, P., Arce, A., Garca-

Torres, L., and Vallejo, A.: Effects of fertiliser type and the presence or absence of plants on nitrous oxide emissions from irrigated soils, Nutr. Cycl. Agroecosys., 78, 279289, 2007. MacKenzie, A. F., Fan, M. X., and Cadrin, F.: Nitrous oxide emission in three years as affected by tillage, corn-soybean-alfalfa rotations, and nitrogen fertilization, J. Environ. Qual., 27, 698703, 1998.

Martnez-Cob, A.: Use of thermal units to estimate corn crop coefcients under semiarid climatic conditions, Irrigation Sci., 26, 335345, 2008.

Biogeosciences, 13, 52455257, 2016 www.biogeosciences.net/13/5245/2016/

G. Guardia et al.: Effect of cover crops on greenhouse gas emissions 5257

Meijide, A., Crdenas, L. M., Snchez-Martn, L., and Vallejo, A.: Carbon dioxide and methane uxes from a barley eld amended with organic fertilizers under Mediterranean climatic conditions, Plant Soil, 328, 353367, 2010.

Migliorati, M. D. A., Scheer, C., Grace, P. R., Rowlings, D. W., Bell, M., and McGree, J.: Inuence of different nitrogen rates and DMPP nitrication inhibitor on annual N2O emissions from a subtropical wheatmaize cropping system, Agr. Ecosyst. Environ., 186, 3343, 2014.

Mrkved, P. T., Drsch, P., and Bakken, L. R.: The N2O product ratio of nitrication and its dependence on long-term changes in soil pH, Soil Biol. Biochem., 39, 20482057, 2007.

Nemecek, T., von Richthofen, J. S., Dubois, G., Casta, P., Charles,R., and Pahl, H.: Environmental impacts of introducing grain legumes into European crop rotations, Eur. J. Agron., 28, 380 393, 2008.

Oorts, K., Merckx, R., Grhan, E., Labreuche, J., and Nicolardot,B.: Determinants of annual uxes of CO2 and N2O in long-term no-tillage and conventional tillage systems in northern France,

Soil Till. Res., 95, 133148, 2007.

Poeplau, C. and Don, A.: Carbon sequestration in agricultural soils via cultivation of cover crops A meta-analysis, Agr. Ecosyst.Environ., 200, 3341, 2015.

Quemada, M., Baranski, M., Nobel-de Lange, M. N. J., Vallejo, A., and Cooper, J. M.: Meta-analysis of strategies to control nitrate leaching in irrigated agricultural systems and their effects on crop yield, Agr. Ecosyst. Environ., 174, 110, 2013.

Quemada, M., Gabriel, J. L., and Zarco-Tejada, P.: Airborne hyper-spectral images and ground-level optical sensors as assessment tools for maize nitrogen fertilization, Remote Sens., 6, 2940 2962, 2014.

Reeves, S. and Wang, W.: Optimum sampling time and frequency for measuring N2O emissions from a rain-fed cereal cropping system, Sci. Total Environ., 530, 219226, 2015.

Rochette, P. and Janzen, H. H.: Towards a revised coefcient for estimating N2O emissions from legumes, Nutr. Cycl. Agroecosys., 73, 171179, 2005.

Sanz-Cobena, A., Snchez-Martn, L., Garca-Torres, L., andVallejo, A.: Gaseous emissions of N2O and NO and NO

3 leach

ing from urea applied with urease and nitrication inhibitors to a maize (Zea mays) crop, Agr. Ecosyst. Environ., 149, 6473, 2012.

Sanz-Cobena, A., Garca-Marco, S., Quemada, M., Gabriel, J. L., Almendros, P., and Vallejo, A.: Do cover crops enhance N2O, CO2 or CH4 emissions from soil in Mediterranean arable systems?, Sci. Total Environ., 466, 164174, 2014.

Sarkodie-Addo, J., Lee, H. C., and Baggs, E. M.: Nitrous oxide emissions after application of inorganic fertilizer and incorporation of green manure residues, Soil Use Manage., 19, 331339, 2003.

Schindlbacher, A., Zechmeister-Boltenstern, S., and Butterbach-Bahl, K.: Effects of soil moisture and temperature on NO, NO2, and N2O emissions from European forest soils, J. Geophys. Res.-

Atmos., 109, D17302, doi:http://dx.doi.org/10.1029/2004JD004590

Web End =10.1029/2004JD004590 http://dx.doi.org/10.1029/2004JD004590

Web End = , 2004.

Senbayram, M., Chen, R., Mhling, K. H., and Dittert, K.: Contribution of nitrication and denitrication to nitrous oxide emissions from soils after application of biogas waste and other fertilizers, Rapid Commun. Mass Sp., 23, 24892498, 2009.

Shan, J. and Yan, X.: Effects of crop residue returning on nitrous oxide emissions in agricultural soils, Atmos. Environ., 71, 170 175, 2013.

Snyder, C. S., Bruulsema, T. W., Jensen, T. L., and Fixen, P. E.: Review of greenhouse gas emissions from crop production systems and fertilizer management effects, Agr. Ecosyst. Environ., 133, 247266, 2009.

Soil Survey Staff: Keys to Soil Taxonomy, 12th Edn., USDA-Natural Resources Conservation Service, Washington DC, USA, 2014.

Spiertz, J. H. J.: Nitrogen, sustainable agriculture and food security.A review, Agron. Sustain. Dev., 30, 4355, 2010.

Stehfest, E. and Bouwman, L.: N2O and NO emission from agricultural elds and soils under natural vegetation: summarizing available measurement data and modeling of global annual emissions, Nutr. Cycl. Agroecosys., 74, 207228, 2006.

Tate, K. R.: Soil methane oxidation and land-use changefrom process to mitigation, Soil Biol. Biochem., 80, 260272, 2015. Tonitto, C., David, M. B., and Drinkwater, L. E.: Replacing bare fallows with cover crops in fertilizer-intensive cropping systems: A meta-analysis of crop yield and N dynamics, Agr. Ecosyst. Environ., 112, 5872, 2006.

Ussiri, D. and Lal, R.: Soil emission of nitrous oxide and its mitigation, Springer, the Netherlands, 2013.

Vallejo, A., Skiba, U. M., Garca-Torres, L., Arce, A., Lpez-

Fernndez, S., and Snchez-Martn, L.: Nitrogen oxides emission from soils bearing a potato crop as inuenced by fertilization with treated pig slurries and composts, Soil Biol. Biochem., 38, 27822793, 2006.van Groenigen, J. W., Velthof, G. L., Oenema, O., van Groenigen,K. J., and van Kessel, C.: Towards an agronomic assessment of N2O emissions: a case study for arable crops, Eur. J. Soil Sci., 61, 903913, 2010.

Vanlauwe, B., Kihara, J., Chivenge, P., Pypers, P., Coe, R., and Six,J.: Agronomic use efciency of N fertilizer in maize-based systems in sub-Saharan Africa within the context of integrated soil fertility management, Plant Soil, 339, 3550, 2011.

Veenstra, J. J., Horwath, W. R., and Mitchell, J. P.: Tillage and cover cropping effects on aggregate-protected carbon in cotton and tomato, Soil Sci. Soc. Am. J., 71, 362371, 2007. Wagner-Riddle, C. and Thurtell, G. W.: Nitrous oxide emissions from agricultural elds during winter and spring thaw as affected by management practices, Nutr. Cycl. Agroecosys., 52, 151163, 1998.

Wichern, F., Eberhardt, E., Mayer, J., Joergensen, R. G., and Mller,T.: Nitrogen rhizodeposition in agricultural crops: methods, estimates and future prospects, Soil Biol. Biochem., 40, 3048, 2008.

Yamulki, S. and Jarvis, S.: Short-term effects of tillage and compaction on nitrous oxide, nitric oxide, nitrogen dioxide, methane and carbon dioxide uxes from grassland, Biol. Fert. Soils, 36, 224231, 2002.

www.biogeosciences.net/13/5245/2016/ Biogeosciences, 13, 52455257, 2016