Sown Diversity Effects on Yield and Resistance to Weed Invasion : Clues to Improve Mixture Design Under Climatic Change in the Mediterranean

Altres ajuts: acords transformatius de la UABWith the aim to improve mixture design, particularly in regions vulnerable to climate change, we tested several forage communities following the biodiversity-ecosystem function (BEF) framework. We sowed monocultures and 4-species mixtures from a pool of 7 forage species in a sub-Mediterranean region (Eastern Pyrenees) and assessed the diversity effects on yield and resistance to weed invasion. The tested species included two grasses and five legumes with contrasting temporal patterns and different climatic amplitudes. The communities differed in their specific composition (mixture types) and the relative abundance of the components, following a simplex design, which allowed us to estimate separately the two components of the diversity effect: the individual species effects and that due to species interactions. Whereas monocultures performed in a highly variable way within and across harvests, both in relation to yield and weed suppression, mixture variability was narrower. Both functions increased in mixtures(with significant interaction effects between 24% and 57% for yield and 13% and 96% for weed suppression), especially in those mixtures including Mediterranean species, which showed the highest diversity effects that persisted over the three experimental years. Extreme climatic events during the experimental period might have affected not only the species' individual performances but also the strength of species interactions. Both components of diversity, identities and interactions, were key in maintaining high performances. We conclude that, under the current climate change scenario, it is important to include species in mixtures that increase resistance or resilience not only at the species level but also at the community level, through enhanced interaction effects

Introducing automation to the molecular diagnosis of Trypanosoma cruzi infection: A comparative study of sample treatments, DNA extraction methods and real-time PCR assays

Background Polymerase chain reaction (PCR) has become a useful tool for the diagnosis of Trypanosoma cruzi infection. The development of automated DNA extraction methodologies and PCR systems is an important step toward the standardization of protocols in routine diagnosis. To date, there are only two commercially available Real-Time PCR assays for the routine laboratory detection of T. cruzi DNA in clinical samples: TCRUZIDNA.CE (Diagnostic Bioprobes Srl) and RealCycler CHAG (Progenie Molecular). Our aim was to evaluate the RealCycler CHAG assay taking into account the whole process. Methodology/Principal findings We assessed the usefulness of an automated DNA extraction system based on magnetic particles (EZ1 Virus Mini Kit v2.0, Qiagen) combined with a commercially available Real-Time PCR assay targeting satellite DNA (SatDNA) of T. cruzi (RealCycler CHAG), a methodology used for routine diagnosis in our hospital. It was compared with a well-known strategy combining a commercial DNA isolation kit based on silica columns (High Pure PCR Template Preparation Kit, Roche Diagnostics) with an in-house Real-Time PCR targeting SatDNA. The results of the two methodologies were in almost perfect agreement, indicating they can be used interchangeably. However, when variations in protocol factors were applied (sample treatment, extraction method and Real-Time PCR), the results were less convincing. A comprehensive fine-tuning of the whole procedure is the key to successful results. Guanidine EDTA-blood (GEB) samples are not suitable for DNA extraction based on magnetic particles due to inhibition, at least when samples are not processed immediately. Conclusions/Significance This is the first study to evaluate the RealCycler CHAG assay taking into account the overall process, including three variables (sample treatment, extraction method and Real-Time PCR). Our findings may contribute to the harmonization of protocols between laboratories and to a wider application of Real-Time PCR in molecular diagnostic laboratories associated with health centers.Fil: Abras, Alba. Universidad de Barcelona; España. Universidad de Girona; España. Instituto de Salud Global de Barcelona; EspañaFil: Ballart, Cristina. Universidad de Barcelona; España. Instituto de Salud Global de Barcelona; EspañaFil: Llovet, Teresa. Universitat Autònoma de Barcelona; España. Hospital de la Santa Creu I Sant Pau; EspañaFil: Roig, Carme. Hospital de la Santa Creu I Sant Pau; EspañaFil: Gutiérrez, Cristina. Hospital de la Santa Creu I Sant Pau; EspañaFil: Tebar, Silvia. Universidad de Barcelona; España. Instituto de Salud Global de Barcelona; EspañaFil: Berenguer, Pere. Hospital de la Santa Creu I Sant Pau; EspañaFil: Pinazo, María-Jesús. Instituto de Salud Global de Barcelona; EspañaFil: Posada, Elizabeth. Instituto de Salud Global de Barcelona; EspañaFil: Gascón, Joaquim. Instituto de Salud Global de Barcelona; EspañaFil: Schijman, Alejandro Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Instituto de Investigaciones en Ingeniería Genética y Biología Molecular "Dr. Héctor N. Torres"; ArgentinaFil: Gállego, Montserrat. Instituto de Salud Global de Barcelona; España. Universidad de Barcelona; EspañaFil: Muñoz, Carmen. Hospital de la Santa Creu I Sant Pau; España. Universitat Autònoma de Barcelona; Españ

Impact of fertilization with pig slurry on the isotopic composition of nitrate retained in soil and leached to groundwater in agricultural areas

The isotopic composition of N and O of nitrate (NO3−) is usually employed to trace its sources of pollution in groundwater. In agricultural areas, the amount of NO3− that reaches the aquifers after fertilization is controlled by different transformation processes that can affect the nitrogen species isotopic composition. Aiming to address the reliability of using isotope tools to trace sources of groundwater NO3−, the goal of this study was to check the effect of fertilization on the isotopic composition of N compounds retained and leached from soils. The concentration and isotopic composition (δ15N and δ18O) of ammonium (NH4+), NO3− and nitrite (NO2−) was characterized after the application of pig slurry in lysimeters containing either soil under fallow (LF) or the same soil continuously cropped and fertilized (LC) during the previous six years. Results showed that the leached NO3− isotopic signature did not directly reflect the isotopic composition of the applied pig slurry. Just after fertilization, nitrification led to lower δ15NNO3 values in soil extracts and leachates (e.g. from +5.9 ± 0.9¿ to +3.8 ± 3.1¿ in soil extracts of LF lysimeters). These values increased after complete nitrification (+11.5 ± 1.3¿) towards the δ15Nbulk of pig slurry (+19.6 ± 0.5¿). Later on, due to soil organic matter and plant debris mineralization and subsequent nitrification, values decreased towards the initial δ15NNO3 of soil but remained above them (+8.6 ± 1.0¿). Both LF and LC experiments showed a similar trend and the latter ones allowed to reinforce that long-term fertilization with pig slurry can increase the soil δ15NNO3. Concerning the δ18O of NO3− from soil extracts and leachates, it mainly depended on the δ18O of irrigation water and oxygen, after nitrification of NH4+ from pig slurry. Therefore, studies aiming to trace groundwater NO3− pollution sources in rural areas by using an isotopic approach should consider the fertilization history of each setting. Also, analyzing the δ15Nbulk of soil is recommended, since it could mask the isotopic signature of the N applied through fertilization

Usefulness of Matrix-Assisted Laser Desorption/Ionization-Time of Flight Mass Spectrometry in the Characterization of Leishmania Strains Causing Tegumentary Leishmaniasis in Bolivia versus hsp70 Gene Sequencing

Matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) is a proteomic technique with proven efficiency in the identification of microorganisms, such as bacteria, fungi, and parasites. The present study aimed to evaluate the usefulness of MALDI-TOF MS for the characterization of Leishmania species circulating in Bolivia using hsp70 gene sequencing as a reference technique. 55 Leishmania strains that were isolated from patients with tegumentary leishmaniasis were analyzed. MALDI-TOF MS identified two species of the L. braziliensis complex (L. braziliensis, n = 26; L. braziliensis outlier, n = 18), one species of the L. guyanensis complex (L. guyanensis, n = 1), one species of the L. lainsoni complex (L. lainsoni, n = 2), and two species of the complex (, n = 5; and L. garnhami, n = 3). All of the strains were correctly identified at the subgenus, genus, and complex level, but 10 of them (18%) were misidentified as other species within the same complex by the hsp70 gene sequencing, with 7 of these corresponding to possible hybrids. Thus, one L. braziliensis corresponded to L. peruviana, two L. braziliensis corresponded to L. braziliensis / L. peruviana possible hybrids, two corresponded to , and three L. garnhami and two corresponded to / possible hybrids. Accordingly, MALDI-TOF MS could be used as an alternative to molecular techniques for the identification of Leishmania spp., as it is low cost, simple to apply, and able to quickly produce results. In Bolivia, its application would allow for the improvement of the management of patient follow-ups, the updating of the epidemiological data of the Leishmania species, and a contribution to the control of tegumentary leishmaniasis. IMPORTANCE The objective of the study was to evaluate the usefulness of MALDI-TOF MS for the characterization of Leishmania species circulating in Bolivia, in comparison with the sequencing of the hsp70 gene. In our study, all of the isolates could be identified, and no misidentifications were observed at the complex level. Although the equipment implies a high initial investment in our context, MALDI-TOF MS can be used in different areas of microbiology and significantly reduces the cost of testing. Once the parasite culture is obtained, the technique quickly yields information by accessing a free database that is available online. This would allow for the improvement of the management of patients and follow-ups, the updating of the epidemiological data of the species, and a contribution to the control of tegumentary leishmaniasis in Bolivia. Likewise, it can be used to determine a specific treatment to be given, according to the causal species of Leishmania, when there are protocols in this regard in the area

Serological diagnosis of chronic Chagas disease: Is it time for a change?

Chagas disease has spread to non-endemic areas with human migration. Since no single reference standard test is available, serological diagnosis of chronic Chagas disease requires at least two tests. New generation techniques have significantly improved the accuracy of Chagas disease diagnosis by the use of a large mixture of recombinant antigens with different detection systems, such as chemiluminescence. The aim of the present study was to assess the overall accuracy of a new generation kit, Architect Chagas (cut-off ≥ 1 S/CO, sample relative light units/cut-off value), as a single technique in the diagnosis of chronic Chagas disease. Architect Chagas showed a sensitivity of 100% (95% confidence interval, CI = 99.5-100) and a specificity of 97.6% (95% CI = 95.2-99.9). Five out of six false-positive sera were a consequence of cross-reactivity with Leishmania spp. and all of them achieved results < 5 S/CO. We propose Architect Chagas as a single technique for screening in blood banks and for routine diagnosis in clinical laboratories. Only grey zone and positive sera with a result ≤ 6 S/CO would need to be confirmed by a second serological assay, thus avoiding false-positive sera and the problem of cross-reactivity with Leishmania spp. The application of this proposal would result in important savings in the cost of Chagas disease diagnosis and therefore in the management and control of the disease

Long-term effects of gasification biochar application on soil functions in a Mediterranean agroecosystem: Higher addition rates sequester more carbon but pose a risk to soil faunal communities

17 Pág. Departamento de Medio Ambiente y Agronomía​ (INIA)Biochar applications can have important implications for many of the soil functions upon which agroecosystems rely, particularly regarding organic carbon storage. This study evaluated the impacts of adding a highly aromatic gasification biochar at different rates (0, 12 and 50 t ha-1) to a barley crop on the provision of crucial soil functions (carbon sequestration, water content, greenhouse gas emissions, nutrient cycling, soil food web functioning, and food production). After natural ageing in the field for six years, a wide range of soil properties representative of the studied soil functions were measured and integrated into a soil quality index. Results showed that C sequestration increased with biochar rate (23 and 68% higher than in the control for the 12 and 50 t biochar ha-1 treatments, respectively). Water content was enhanced at the 50 t ha-1 treatment depending on the sampling date. Despite biochar additions neither abating nor increasing CO2 equivalent emissions (carbon dioxide plus nitrous oxide and methane), the system shifted from being a methane sink (-0.017 ± 0.01 mg CH4-C m-2 h-1 at the 12 t ha-1 treatment), to a net source (0.025 ± 0.02 mg CH4-C m-2 h-1 at the 50 t ha-1 treatment). In addition, biochar ageing provoked a loss of nitrate mitigation potential, and indeed ammonium production was stimulated at the 50 t ha-1 rate. The 50 t ha-1 treatment also adversely affected nematode and collembolan functional diversity. Lastly, biochar did not affect barley yield. The results of the soil quality index indicated that no biochar treatment provided more benefits to our agricultural soil, and, although the 50 t ha-1 treatment increased C sequestration, this was potentially offset by its harmful effects on soil faunal communities. Therefore, application of this biochar at high rates should be avoided to prevent risks to soil biological communities.We gratefully acknowledge the funding by the project FERTICHAR (AGL2015-70393-R) of the Spanish Ministry of Economy and Competitiveness. We are also grateful to Dr. van den Brink, P. J., and Dr. Šmilauer, P. for their assistance on PRC analysis interpretation and proper use of CANOCO 5, respectively.Peer reviewe

Introducing automation to the molecular diagnosis of Trypanosoma cruzi infection: A comparative study of sample treatments, DNA extraction methods and real-time PCR assays

Background: Polymerase chain reaction (PCR) has become a useful tool for the diagnosis of Trypanosoma cruzi infection. The development of automated DNA extraction methodologies and PCR systems is an important step toward the standardization of protocols in routine diagnosis.To date, there are only two commercially available Real-Time PCR assays for the routine laboratory detection of T. cruzi DNA in clinical samples: TCRUZIDNA.CE (Diagnostic Bioprobes Srl) and RealCycler CHAG (Progenie Molecular). Our aim was to evaluate the RealCycler CHAG assay taking into account the whole process. Methodology/Principal findings: We assessed the usefulness of an automated DNA extraction system based on magnetic particles (EZ1 Virus Mini Kit v2.0, Qiagen) combined with a commercially available Real- Time PCR assay targeting satellite DNA (SatDNA) of T. cruzi (RealCycler CHAG), a methodology used for routine diagnosis in our hospital. It was compared with a well-known strategy combining a commercial DNA isolation kit based on silica columns (High Pure PCR Template Preparation Kit, Roche Diagnostics) with an in-house Real-Time PCR targeting SatDNA. The results of the two methodologies were in almost perfect agreement, indicating they can be used interchangeably. However, when variations in protocol factors were applied (sample treatment, extraction method and Real-Time PCR), the results were less convincing. A comprehensive fine-tuning of the whole procedure is the key to successful results. Guanidine EDTA-blood (GEB) samples are not suitable for DNA extraction based on magnetic particles due to inhibition, at least when samples are not processed immediately. Conclusions/Significance: This is the first study to evaluate the RealCycler CHAG assay taking into account the overall process, including three variables (sample treatment, extraction method and Real-Time PCR). Our findings may contribute to the harmonization of protocols between laboratories and to a wider application of Real-Time PCR in molecular diagnostic laboratories associated with health centers

Pilot study of living donor liver transplantation for patients with HCC exceeding Milan criteria

BACKGROUND & AIMS: A subset of patients with hepatocellular carcinoma (HCC) beyond Milan criteria might obtain acceptable survival outcomes after liver transplantation. Living donor liver transplantation (LDLT) has emerged as a feasible alternative to overcome the paucity of donors. METHODS: In 2001 we started a protocol for LDLT in Child A-B patients with HCC fulfilling a set of criteria - the BCLC expanded criteria- that expanded the conventional indications of transplantation: 1 tumor ≤ 7cm, 5 tumors ≤ 3cm, 3 tumors ≤ 5cm without macrovascular invasion or down-staging to Milan after loco-regional therapies. RESULTS: We present a prospective cohort of 22 patients with BCLC extended indications based on size/number (17) or down-staging (5) treated with LDLT between 2001 and 2014. Characteristics of the patients were as follows: median age: 57yr old; males/female: 20/2, Child-Pugh A/B: 16/6, AFP <100ng/mL: 21. Twelve patients received neo-adjuvant loco-regional therapies. At the time of transplantation, 12 patients had HCC staging beyond Milan criteria and 10 within. Pathological reports showed that 50% exceeded BCLC expanded criteria. Perioperative mortality was 0%. After a median follow up of 81 months, the 1-, 3-, 5- and 10-year survival was 95.5%, 86.4%, 80.2% and 66.8%, respectively. Overall, seven patients recurred (range 9-108 mo), and the 5-y and 10-yr actuarial recurrence rate was of 23.8% and 44,4%, respectively. CONCLUSION: A proper selection of candidates for extended indications of living donor liver transplantation for HCC patients provide survival outcomes comparable

5 Role of biochar in N cycling in a Mediterranean agroecosystem : potential benefits and trade-offs

Les aportacions de nitrogen reactiu (N) mitjançant el procés Haber-Bosch (fixació industrial de N2 per produir fertilitzants) i el cultiu de lleguminoses són responsables de més de la meitat de l'aportació de N als ecosistemes terrestres. Aquests inputs excessius provinents dels agroecosistemes han augmentat els fluxos d'òxid nitrós (N2O), amoníac (NH3) i nitrat (NO3−), provocant efectes tals com l'acceleració de l'escalfament global, l'eutrofització i la reducció de l'ozó estratosfèric, entre d'altres. L'esmena del sòl amb biochar, el sòlid ric en carboni (C) produït per la conversió termoquímica de biomassa en condicions limitants d'oxigen, podria ser una eina valuosa per fer front als fluxos elevats de N. No obstant, la implementació a gran escala de biochar es veu compromesa per les incerteses sobre possibles conseqüències adverses, així com pels seus efectes a llarg termini sobre la qualitat del sòl, atesa la seva persistència al sòl. Per tal d'escurçar aquests buits de coneixement, es va avaluar en condicions d'hivernacle (Capítol 1) i en condicions de camp (Capítol 2) un biochar de gasificació que va demostrar prèviament poder reduir el nivell de NO3− soluble al sòl. El Capítol 1 examinà la possible mitigació de la lixiviació de nitrats tant a curt com a llarg termini i a dues dosis d'aplicació contrastades (12 i 50 t ha-1). Es van examinar columnes de sòl amb biochar fresc i biochar envellit 6 anys al camp. Al cap de 8 mesos, es va trobar una reducció significativa de NO3− en la solució del sòl, així com d'altres ions (incloent clorur, magnesi, sodi i calci), per a les dues dosis d'aplicació de biochar fresc. Aquest fenòmen s'atribuí a la formació d'un recobriment organo-mineral sobre els porus del biochar, tot atrapant aigua i nutrients dins els porus del biochar. Tot i la disminució de NO3− en la solució del sòl, la lixiviació no va veure's afectada pel biochar, posant en dubte la seva capacitat de mitigació. Al Capítol 2, s'avaluà la qualitat del sòl mitjançant el seguiment de diverses propietats del sòl en mesocosmos de camp establerts fa 6 anys (d'on provenen les mescles sòl-biochar envellit utilitzats al Capítol 1). La dosi de 50 t ha-1 fou la més eficaç en el segrest de C i va millorar el contingut d'humitat al sòl puntualment. No obstant, aquesta dosi representà un trade-off ja que va impactar negativament les comunitats de fauna del sòl (nematodes i col·lèmbols), i també va augmentar les emissions de metà (CH4). Encara que la dosi de 12 t ha-1 no presentà aquests efectes negatius, va mostrar indicis de metabolisme de compostos recalcitrants de C, que, si es mantingués en el temps, podria afectar el potencial de segrest de C del biochar. En conjunt, es pot concloure que al nostre agroecosistema la dosi de 12 t ha-1 representa un escenari amb menys riscos. Finalment, el Capítol 3 va abordar els possibles beneficis agronòmics d'incloure un biochar de piròlisi lenta en tres formulacions de fertilitzants organo-minerals (NPK, NP i K). La lixiviació de nutrients es va alentir en la formulació amb biochar i NPK (NPK + B). Probablement, el C làbil incorporat amb el biochar i l'input simultani de diversos nutrients podria haver estimulat el creixement microbià causant un emmagatzematge temporal de nutrients a la biomassa microbiana. Tot i que el fertilitzant NPK + B només va millorar significativament la biomassa de palla d'ordi i no el gra, tots els fertilitzants formulats amb biochar van provocar un millor estat nutricional vegetal ( en relació al potassi, sofre, calci i manganès). Aquests resultats indiquen que les formulacions investigades són prometedores per al desenvolupament de fertilitzants de nova generació.Las aportaciones de nitrógeno reactivo (N) mediante el proceso Haber-Bosch (fijación industrial de N2 para producir fertilizantes) y el cultivo de leguminosas son responsables de más de la mitad del input de N a los ecosistemas terrestres. Estos insumos excesivos provenientes de los agroecosistemas han aumentado los flujos de óxido nitroso (N2O), amoníaco (NH3) y nitrato (NO3-), provocando efectos tales como la aceleración del calentamiento global, la eutrofización y la reducción de la ozono estratosférico, entre otros. La enmienda del suelo con biochar, el sólido rico en carbono (C) producido por la conversión termoquímica de biomasa en condiciones limitantes de oxígeno, podría ser una herramienta valiosa para hacer frente a los flujos elevados de N. Sin embargo, la implementación a gran escala del biochar se ve comprometida por la incertidumbre sobre posibles efectos adversos, así como por sus efectos a largo plazo sobre la calidad del suelo, debido a su persistencia en el suelo. Para reducir estas brechas de conocimiento, se evaluó en condiciones de invernadero (Capítulo 1) y en condiciones de campo (Capítulo 2) un biochar de gasificación que demostró previamente poder reducir el nivel de NO3- soluble del suelo. El Capítulo 1 examinó la posible mitigación de lixiviación de nitratos a corto y largo plazo en dos dosis de aplicación contrastadas (12 y 50 t ha-1). Se examinaron columnas de suelo con biochar fresco y biochar envejecido durante 6 años en el campo. Al cabo de 8 meses, se encontró una reducción significativa de NO3- en la solución del suelo, así como de otros iones (incluyendo cloruro, magnesio, sodio y calcio), para las dos dosis de aplicación de biochar fresco. Este fenómeno se atribuyó a la formación de un recubrimiento organo-mineral sobre los poros del biochar, que atraparía agua y nutrientes dentro de los poros del biochar. A pesar de la disminución de NO3- en la solución del suelo, la lixiviación no se redujo, poniendo en duda la capacidad de mitigación del biochar. En el Capítulo 2, se evaluó la calidad del suelo mediante el seguimiento de varias propiedades del suelo en mesocosmos de campo establecidos hace 6 años (de donde provienen las mezclas suelo-biochar envejecido utilizados en el Capítulo 1). La dosis de 50 t ha-1 fue la más eficaz en el secuestro de C y mejoró el contenido de humedad en el suelo puntualmente. Sin embargo, esta dosis representó un trade-off, ya que impactó negativamente las comunidades de fauna del suelo (nematodos y colémbolos), y también aumentó las emisiones de metano (CH4). Aunque la dosis de 12 t ha-1 no presentó estos efectos negativos, mostró indicios de metabolismo de compuestos recalcitrantes de C, que, si se mantuviera en el tiempo, podría afectar el potencial de secuestro de C del biochar. En conjunto, se puede concluir que en nuestro agroecosistema la dosis de 12 t ha-1 representa un escenario con menos riesgos. Finalmente, el Capítulo 3 abordó los posibles beneficios agronómicos incluir un biochar de pirólisis lenta en tres formulaciones de fertilizantes organo-minerales (NPK, NP y K). La lixiviación de nutrientes se ralentizó en la formulación con biochar y NPK (NPK + B). Probablemente, el C lábil incorporado con el biochar y el input simultáneo de varios nutrientes podría haber estimulado el crecimiento microbiano causando un almacenamiento temporal de nutrientes en la biomasa microbiana. Aunque el fertilizante NPK + B solo mejoró significativamente la biomasa de paja de cebada y no el grano, todos los fertilizantes formulados con biochar provocaron un mejor estado nutricional vegetal (en relación al potasio, azufre, calcio y manganeso). Estos resultados indican que las formulaciones investigadas son prometedoras para el desarrollo de fertilizantes de nueva generación.Anthropogenic inputs of reactive nitrogen (N) via the Haber-Bosch process (synthetic N2 fixation for fertiliser production) and legume cultivation are responsible for over half of the input of N to terrestrial ecosystems. These excessive inputs from agroecosystems have resulted in enhanced fluxes of nitrous oxide (N2O), ammonia (NH3), and nitrate (NO3−), which induce far-reaching adverse effects, including the acceleration of global warming, promoting eutrophication, and depleting stratospheric ozone, among others. Soil-application of biochar, the carbon(C)-rich solid produced by thermochemical conversion of biomass under oxygen-limited conditions, could be a valuable tool to cope with elevated N fluxes. However, broad-scale biochar implementation is hindered by the uncertainties concerning possible unintended consequences as well as its long-term effects on soil quality, given its persistence in soil. In order to fill in some of these knowledge gaps, a pine gasification biochar that was found to reduce nitrate contents in soil solution in a previous study was assessed both under greenhouse conditions (Chapter 1) and under field conditions (Chapter 2). Chapter 1 was aimed at evaluating whether this particular biochar could induce a nitrate leaching mitigation response both in the short and long-term and at two contrasted application rates (12 and 50 t ha-1). To do so, soil columns filled with either fresh biochar or 6-yr field-aged biochar were monitored for 8 months. At the end of the trial, a significant reduction of NO3− in soil solution as well as other ions (including chloride, magnesium, sodium, and calcium) was found for both application rates in the fresh biochar-amended columns but not in the aged biochar scenario. The formation of an organo-mineral coating that entrapped nutrient-enriched water into biochar pores was ascribed as the most plausible explanation. Despite the alleviation of NO3− in soil solution, its leaching was unaffected by biochar treatment, casting doubt about possible environmental effects. In Chapter 2, by monitoring a wide range of soil properties in 6-yr old field mesocosms (from where the soil-aged biochar mixtures of Chapter 1 were taken), a soil quality assessment was conducted. The 50 t ha-1 application rate was the most effective in sequestering C and presented enhanced water contents at some sampling dates. However, an important trade-off emerged, since it exerted negative effects towards soil fauna communities (nematodes and collembolans), and also it boosted methane (CH4) soil emissions. Conversely, the 12 t ha-1 rate did not pose serious risks to soil faunal communities and soils acted as a CH4 sink. On the other hand, this same treatment showed signs of promotion of recalcitrant carbon metabolism, which, if maintained over time, could affect biochar's C sequestration potential and reduce its expected persistence. Taking all into consideration, in our agroecosystem, the 12 t ha-1 rate would be safer to apply. Finally, Chapter 3 addressed the potential agronomical benefits of including biochar into fertiliser formulation. Particularly, a slow pyrolysis biochar was included in three organo-mineral fertiliser formulations (NPK, NP, and K). Nutrient leaching was slowed down in the formulation combining biochar and NPK (NPK+B). This effect was plausibly attributed to the labile-C added with the slow-pyrolysis biochar and the concomitant provision of NPK, which could have promoted microbial growth and caused a temporary storage of nutrients in microbial biomass. While the NPK+B fertiliser only significantly enhanced barley straw biomass and not grain, all biochar-based fertilisers presented improved plant nutrient content and export (regarding potassium, sulphur, calcium, and manganese). Both the improved nutrient release pattern in NPK+B and the enhanced crop nutrient status found for all biochar-based fertilisers indicate that the investigated formulations hold promise for further research and development of new generation fertilisers