Use of DMPSA nitrification inhibitor and no-tillage systems to enhance agricultural sustainability: effect on soil nitrogen gaseous emissions and bacterial populations

284 p.El incremento de la presión alimentaria como consecuencia del aumento y desarrollo de la población haobligado a una intensificación de la agricultura. La necesidad de aplicar fertilizantes nitrogenados para obtener mayores producciones ha generado un aumento del nitrógeno reactivo presente en el medio,desencadenando problemas ambientales como el incremento del efecto invernadero, la reducción de labiodiversidad, la acidificación de los suelos y la contaminación de las aguas subterráneas. En un escenario global de emergencia climática, en el que las políticas se están enfocando hacia medidas para reducir el impacto ambiental de la actividad humana, el incremento de la sostenibilidad de la agricultura juega un papel fundamental.El uso de inhibidores sintéticos de la nitrificación permite reducir las perdidas de nitrógeno reactivo,disminuyendo la liberación de N2O, uno de los gases que más contribuye al calentamiento global. Es necesario conocer la eficiencia de estos compuestos en conjunto con otros manejos para mejorar lasostenibilidad como son los sistemas de no laboreo, de gran interés en clima Mediterráneo, así como es imprescindible garantizar la seguridad de su aplicación para la salud del suelo. En esta tesis se analiza la eficacia del inhibidor de la nitrificación 2-(3-4-dimetil- 1H-pirazol-1-yl)-acidosuccínico en mezcla isomérica (DMPSA), en combinación con sistemas de no laboreo para aumentar la sostenibilidad de la agricultura. Los resultados obtenidos demuestran que la aplicación de DMPSA evitala emisión de N2O derivada de la fertilización nitrogenada en suelos no labrados, sin efectos negativos enla producción y sin causar un impacto profundo en la comunidad bacteriana. Sin embargo, su aplicaciónpuede aumentar el riesgo de volatilización de NH3 cuando se fertiliza con urea, lo cual puede ser evitadohaciendo una aplicación conjunta con el inhibidor de la ureasa triamida N-(n-butil) tiofosfórica (NBPT).Nuestros estudios indican que la aplicación del DMPSA es especialmente recomendable cuando el suelotiene contenidos relativamente altos de agua, cobre y/o zinc, condiciones que promueven las pérdidas deN2O. En conjunto, esta tesis concluye que la aplicación del inhibidor DMPSA es una técnica útil y segura paraincrementar la sostenibilidad de la agricultura mediante la mitigación del impacto ambiental derivado dela fertilización nitrogenada

Biological and synthetic approaches to inhibiting nitrification in non-tilled Mediterranean soils

Background: The increasing demand for food production has led to a tenfold increase in nitrogen (N) fertilizer use since the Green Revolution. Nowadays, agricultural soils have been turned into high-nitrifying environments that increase N pollution. To decrease N losses, synthetic nitrification inhibitors (SNIs) such as 3,4-dimethylpyrazole phosphate (DMPP) have been developed. However, SNIs are not widely adopted by farmers due to their biologically limited stability and soil mobility. On the other hand, allelopathic substances from root exudates from crops such as sorghum are known for their activity as biological nitrification inhibitors (BNIs). These substances are released directly into the rhizosphere. Nevertheless, BNI exudation could be modified or even suppressed if crop development is affected. In this work, we compare the performance of biological (sorghum crop) and synthetic (DMPP) nitrification inhibitors in field conditions. Results: Sorghum crop BNIs and DMPP prevented an increase in the abundance of ammonia-oxidizing bacteria (AOB) without affecting the total bacterial abundance. Both nitrification inhibitors maintained similar soil NH4+ content, but at 30 days post-fertilization (DPF), the sorghum BNIs resulted in higher soil NO3− content than DMPP. Even so, these inhibitors managed to reduce 64% and 96%, respectively, of the NO3−-N/NH4+-N ratio compared to the control treatment. Similar to soil mineral N, there were no differences in leaf δ15N values between the two nitrification inhibitors, yet at 30 DPF, δ15N values from sorghum BNI were more positive than those of DMPP. N2O emissions from DMPP-treated soil were low throughout the experiment. Nevertheless, while sorghum BNIs also maintained low N2O emissions, they were associated with a substantial N2O emission peak at 3 DPF that lasted until 7 DPF. Conclusions: Our results indicate that while sorghum root exudates can reduce nitrification in field soil, even at the same efficiency as DMPP for a certain amount of time, they are not able to prevent the N pollution derived from N fertilization as DMPP does during the entire experiment.This project was funded by the Spanish Government (RTI2018-094623-B-C22 MCIU/AEI/FEDER, UE) and by the Basque Government (IT-932-16). Adrián Bozal-Leorri holds a Grant from the Basque Government (PRE-2020-2-0142). Mario Corrochano-Monsalve holds a Grant from the Ministry of Economy and Business of the Spanish Government (BES-2016-076725)

Evaluation of a crop rotation with biological inhibition potential to avoid N2O emissions in comparison with synthetic nitrification inhibition

Agriculture has increased the release of reactive nitrogen to the environment due to crops' low nitrogen-use efficiency (NUE) after the application of nitrogen-fertilisers. Practices like the use of stabilized-fertilisers with nitrification inhibitors such as DMPP (3,4-dimethylpyrazole phosphate) have been adopted to reduce nitrogen losses. Otherwise, cover crops can be used in crop-rotation-strategies to reduce soil nitrogen pollution and benefit the following culture. Sorghum (Sorghum bicolor) could be a good candidate as it is drought tolerant and its culture can reduce nitrogen losses derived from nitrification because it exudates biological nitrification inhibitors (BNIs). This work aimed to evaluate the effect of fallow-wheat and sorghum cover crop-wheat rotations on N2O emissions and the grain yield of winter wheat crop. In addition, the suitability of DMPP addition was also analyzed. The use of sorghum as a cover crop might not be a suitable option to mitigate nitrogen losses in the subsequent crop. Although sorghum–wheat rotation was able to reduce 22% the abundance of amoA, it presented an increment of 77% in cumulative N2O emissions compared to fallow–wheat rotation, which was probably related to a greater abundance of heterotrophic-denitrification genes. On the other hand, the application of DMPP avoided the growth of ammonia-oxidizing bacteria and maintained the N2O emissions at the levels of unfertilized-soils in both rotations. As a conclusion, the use of DMPP would be recommendable regardless of the rotation since it maintains NH4+ in the soil for longer and mitigates the impact of the crop residues on nitrogen soil dynamics.This work was supported by the Spanish Government (RTI2018-094623-B-C21 and C22 MCIU/AEI/FEDER, UE) and the Basque Government (IT-932-16). Dr. Adrián Bozal-Leorri held a grant from the Basque Government (PRE-2020-2-0142). Dr. Mario Corrochano-Monsalve held a grant from the Ministry of Economy and Business of the Spanish Government (BES-2016-076725)

Biological and synthetic approaches to inhibiting nitrification in non‑tilled Mediterranean soils

[EN] Background: The increasing demand for food production has led to a tenfold increase in nitrogen (N) fertilizer use since the Green Revolution. Nowadays, agricultural soils have been turned into high-nitrifying environments that increase N pollution. To decrease N losses, synthetic nitrification inhibitors (SNIs) such as 3,4-dimethylpyrazole phosphate (DMPP) have been developed. However, SNIs are not widely adopted by farmers due to their biologically limited stability and soil mobility. On the other hand, allelopathic substances from root exudates from crops such as sorghum are known for their activity as biological nitrification inhibitors (BNIs). These substances are released directly into the rhizosphere. Nevertheless, BNI exudation could be modified or even suppressed if crop development is affected. In this work, we compare the performance of biological (sorghum crop) and synthetic (DMPP) nitrification inhibitors in field conditions. Results: Sorghum crop BNIs and DMPP prevented an increase in the abundance of ammonia-oxidizing bacteria (AOB) without affecting the total bacterial abundance. Both nitrification inhibitors maintained similar soil NH4+ content, but at 30 days post-fertilization (DPF), the sorghumBNIs resulted in higher soil NO3- content than DMPP. Even so, these inhibitors managed to reduce 64% and 96%, respectively, of the NO3--N/NH4+-N ratio compared to the control treatment. Similar to soil mineral N, there were no differences in leaf delta N-15 values between the two nitrification inhibitors, yet at 30 DPF, delta N-15 values from sorghum BNI were more positive than those of DMPP. N2O emissions from DMPP-treated soil were low throughout the experiment. Nevertheless, while sorghum BNIs also maintained low N2O emissions, they were associated with a substantial N2O emission peak at 3 DPF that lasted until 7 DPF. Conclusions: Our results indicate that while sorghum root exudates can reduce nitrification in field soil, even at the same efficiency as DMPP for a certain amount of time, they are not able to prevent the N pollution derived from N fertilization as DMPP does during the entire experiment. Graphic AbstractThis project was funded by the Spanish Government (RTI2018-094623-B-C22 MCIU/AEI/FEDER, UE) and by the Basque Government (IT-932-16). Adrian Bozal-Leorri holds a Grant from the Basque Government (PRE-2020-2-0142). Mario Corrochano-Monsalve holds a Grant from the Ministry of Economy and Business of the Spanish Government (BES-2016-076725)

Impact of dimethylpyrazole-based nitrification inhibitors on soil-borne bacteria

[EN] Nitrogen (N) input from fertilizers modifies the properties of agricultural soils as well as bacterial community diversity, composition and relationships. This can lead to negative impacts such as the deterioration of system multifunctionality, whose maintenance is critical to normal nutrient cycling. Synthetic nitrification inhibitors (NIs) can be combined with fertilizers to improve the efficiency of N use by reducing N losses. However, analysis of their effects on non-target bacteria are scarce. This study aimed to analyze the effect of applying the NIs DMPP and DMPSA on the whole bacterial community. Through 16S rRNA amplicon sequencing we determined the differences between samples in terms of microbial diversity, composition and co-occurrence networks. The application ofDMPP and DMPSA exerted little impact on the abundance of the dominant phyla. Nevertheless, several significant shifts were detected in bacterial diversity, co-occurrence networks, and the abundance of particular taxa, where soil water content played a key role. For instance, the application of NIs intensified the negative impact of N fertilization on bacterial diversity under high water-filled pore spaces (WFPS) (>64%), reducing community diversity, whereas alpha-diversity was not affected at low WFPS (<55%). Interestingly, despite NIs are known to inhibit ammonia monooxygenase (AMO) enzyme, both NIs almost exclusively inhibited Nitrosomonas genera among AMO holding nitrifiers. Thus, Nitrosomonas showed abundance reductions of up to 47% (DMPP) and 66% (DMPSA). Nonetheless, non-target bacterial abundances also shifted with NI application. Notably,DMPSA application partially alleviated the negative effect of fertilization on soil multifunctionality. A remarkable increase in populations related to system multifunctionality, such as Armatimonadetes (up to+21%), Cyanobacteria (up to +30%) and Fibrobacteres (up to+25%) was observed when DMPSA was applied. NI application substantially influenced microbial associations by decreasing the complexity of co-occurrence networks, decreasing the total edges and node connectivity, and increasing path distances.This project was funded by the Spanish Government (RTI2018- 094623-B-C21 MCIU/AEI/FEDER, UE), the Basque Government (EJ/GV, IT-932-16 and IT-1213-19) and by EuroChem Agro Iberia S.L.-UPV/ EHU 2018.0612. Mario Corrochano-Monsalve holds a grant from the Ministry of Science and Innovation of the Spanish Government (BES- 2016-076725)

Evidences towards deciphering the mode of action of dimethylpyrazole-based nitrification inhibitors in soil and pure cultures of Nitrosomonas europaea

Background: Agriculture relies on the intensive use of synthetic nitrogen (N) fertilizers to maximize crop yields, which has led to the transformation of agricultural soils into high-nitrifying environments. Nevertheless, nitrification inhibitors (Nis) have been developed to suppress soil-nitrifier activity and decrease N losses. The Nis 3,4-dimethylpyrazole phosphate (DMPP) and 2-(3,4-dimethyl-1H-pyrazol-1-yl) succinic acid isomeric mixture (DMPSA) are able to reduce N2O emissions and maintain soil NH4+ for a longer time. Although both Nls have been proven to be effective to inhibit soil nitrification, their exact mode of action has not been confirmed. We aimed to provide novel insights to further understand the mode of action of DMP-based Nis. We evaluated the performance of DMPP and DMPSA in soil and pure cultures of nitrifying bacteria Nitrosomonas europaea. Results: DMPSA did not inhibit nitrification in pure cultures of N. europaea. In the soil, we evidenced that DMPSA needs to be broken into DMP to achieve the inhibition of nitrification, which is mediated by a soil biological process that remains to be identified. Moreover, both DMPP and DMPSA are thought to inhibit nitrification due to their ability to chelate the Cu2+ cations that the ammonia monooxygenase enzyme (AMO) needs to carry on the first step of NH4+ oxidation. However, the efficiency of DMPP was not altered regardless the Cu2+ concentration in the medium. In addition, we also showed that DMPP targets AMO but not hydroxylamine oxidoreductase enzyme (HAO). Conclusions: The inability of DMPSA to inhibit nitrification in pure cultures together with the high efficiency of DMPP to inhibit nitrification even in presence of toxic Cu2+ concentration in the medium, suggest that the mode of action of DMP-based Nis does not rely on their capacity as metal chelators.This project was funded by the Spanish Government (RTI2018-094623-B-C21 MCIU/AEI/FEDER, UE), by the Basque Government (IT-932-16), and by EuroChem Agro Iberia S.L.U. Dr. Adrian Bozal-Leorri held a grant from the Basque Government (PRE-2020-2-0142)

Relationship between tillage management and DMPSA nitrification inhibitor efficiency

Agricultural sustainability is compromised by nitrogen (N) losses caused by soil microbial activity. Nitrous oxide (N2O) is a potent greenhouse gas (GHG) produced as consequence of nitrification and denitrification processes in soils. Nitrification inhibitors (NI) as 3,4-dimethylpyrazole-succinic acid (DMPSA) are useful tools to reduce these N losses from fertilization. The objective of this work was to test the efficiency of DMPSA in two different tillage management systems, conventional tillage (CT) and no-tillage (NT), in a winter wheat crop under Humid Mediterranean conditions. N fertilizer was applied as ammonium sulphate (AS) with or without DMPSA in a single or split application, including an unfertilized treatment. GHG fluxes N2O, CO2 and CH4) were measured by the closed chamber method. amoA and nosZl genes were quantified by qPCR as indicators of nitrifying and denitrifying populations. Nitrification was inhibited by DMPSA in both CT and NT, while the higher water filled pore space (WFPS) in NT promoted a better efficiency of DMPSA in this system. This higher efficiency might be due to a greater N2O reduction to N-2 as result of the nosZl gene induction. Consequently, DMPSA was able to reduce N2O emissions down to the unfertilized levels in NT. Provided that NT reduced CO2 emissions and maintained crop yield compared to CT, the application DMPSA under NT management is a promising strategy to increase agro-systems sustainability under Humid Mediterranean conditions. (C) 2019 The Author(s). Published by Elsevier B.V.This project was funded by de Spanish Government (AGL2015-64582-C3-2-R MINECO/FEDER and RTI2018-094623-B-C21 MCIU/AEI/FEDER, UE), by the Basque Government (IT-932-16) and by EuroChem Agro Iberia S.L.-UPV/EHU 2017.0016. Mario Corrochano-Monsalve held a grant from the Ministry of Economy and Business of the Spanish Government and Ximena Huerfano received a specialization fellowship for PhD researches from the UPV/EHU