Antibiotic Resistance in Agricultural Soil and Crops Associated to the Application of Cow Manure-Derived Amendments from Conventional and Organic Livestock Farms

he application of organic amendments to agricultural soil can enhance crop yield, while improving the physicochemical and biological properties of the recipient soils. However, the use of manure-derived amendments as fertilizers entails environmental risks, such as the contamination of soil and crops with antibiotic residues, antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs). In order to delve into these risks, we applied dairy cow manure-derived amendments (slurry, fresh manure, aged manure), obtained from a conventional and an organic farm, to soil. Subsequently, lettuce and wheat plants were grown in the amended soils. After harvest, the abundance of 95 ARGs and MGE-genes from the amended soils and plants were determined by high-throughput qPCR. The structure of soil prokaryotic communities was determined by 16S rRNA amplicon sequencing and qPCR. The absolute abundance of ARGs and MGE-genes differed between treatments (amended vs. unamended), origins of amendment (conventional vs. organic), and types of amendment (slurry vs. fresh manure vs. aged manure). Regarding ARG-absolute abundances in the amendments themselves, higher values were usually found in slurry vs. fresh or aged manure. These abundances were generally higher in soil than in plant samples, and higher in wheat grain than in lettuce plants. Lettuce plants fertilized with conventional amendments showed higher absolute abundances of tetracycline resistance genes, compared to those amended with organic amendments. No single treatment could be identified as the best or worst treatment regarding the risk of antibiotic resistance in soil and plant samples. Within the same treatment, the resistome risk differed between the amendment, the amended soil and, finally, the crop. In other words, according to our data, the resistome risk in manure-amended crops cannot be directly inferred from the analysis of the amendments themselves. We concluded that, depending on the specific question under study, the analysis of the resistome risk should specifically focus on the amendment, the amended soil or the cropThis work has been financially supported by the Basque Government (projects: URAGAN and KONTRAE-Elkartek-KK-2020-00007) and the Spanish Ministry of Science and Innovation (project: PRADA PID2019-110055RB-C21). LJ was the recipient of a predoctoral fellowship from the Department of Economic Development and Infrastructure of the Basque Governmen

Agricultural Soils Amended with Thermally-Dried Anaerobically-Digested Sewage Sludge Showed Increased Risk of Antibiotic Resistance Dissemination

The application of sewage sludge (SS) to agricultural soil can help meet crop nutrient requirements and enhance soil properties, while reusing an organic by-product. However, SS can be a source of antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs), resulting in an increased risk of antibiotic resistance dissemination. We studied the effect of the application of thermally-dried anaerobically-digested SS on (i) soil physicochemical and microbial properties, and (ii) the relative abundance of 85 ARGs and 10 MGE-genes in soil. Soil samples were taken from a variety of SS-amended agricultural fields differing in three factors: dose of application, dosage of application, and elapsed time after the last application. The relative abundance of both ARGs and MGE-genes was higher in SS-amended soils, compared to non-amended soils, particularly in those with a more recent SS application. Some physicochemical parameters (i.e., cation exchange capacity, copper concentration, phosphorus content) were positively correlated with the relative abundance of ARGs and MGE-genes. Sewage sludge application was the key factor to explain the distribution pattern of ARGs and MGE-genes. The 30 most abundant families within the soil prokaryotic community accounted for 66% of the total variation of ARG and MGE-gene relative abundances. Soil prokaryotic alpha-diversity was negatively correlated with the relative abundance of ARGs and MGE-genes. We concluded that agricultural soils amended with thermally-dried anaerobically-digested sewage sludge showed increased risk of antibiotic resistance dissemination.This work has been financially supported by the Basque Government (projects: URAGAN and KONTRAE-Elkartek-KK2020-00007) and the Spanish Ministry of Science and Innovation (project: PRADA PID2019-110055RB-C21). LJ was the recipient of a predoctoral fellowship from the Department of Economic Development and Infrastructure of the Basque Government

Antibiotikoen erresistentziak agroekosistemetan

Antibiotics have become an indispensable tool to combat bacterial infections in medicine and veterinary medicine. The abuse and misuse of antibiotics has led to the emergence of one of the most important health challenges, as bacteria have developed widespread resistance to antibiotics. This global problem must be analysed from a One Health perspective, as antibiotic resistance is shared, spread and multiplied between humans, animals and the environment. For example, the application of organic fertilizers to soil can improve soil health and crop yields. However, this practice carries risks, as antibiotics not fully metabolised by humans or animals can find their way to soil. In addition to the antibiotics, the use of such organic fertilisers involves the spread of antibiotic-resistant genes, antibiotic-resistant bacteria and mobile genetic elements. Indeed, the continuous introduction of antibiotics into soils creates a potential threat to indigenous bacteria; antibi-otic-resistant bacteria multiply and alter the biodiversity of indigenous bacterial communities. This wide dispersal makes the use of antibiotics increasingly ineffective in the treatment of pathogenic resistant bacterial infections. This review first tries to explain the origin and mechanisms of antibiotic-resistant genes and antibiotics. Next, the use of organic fertilisers of human or animal origin in soils is mentioned, as they are a major source for the spread of antibiotics and antibiotic resistance in the environment. The environmental impact of antibiotic resistance is then explained, following the exposure pathway from crops to humans. Finally, we suggest using some management processes that could be useful to reduce the risk of antibiotic resistance in livestock-derived organic fertilisers.; Antibiotikoak ezinbesteko tresna bilakatu dira medikuntza eta albaitaritzan bakterio-infekzioei aurre egiteko. Antibiotikoen gehiegizko erabilerak eta erabilera okerrak osasun-erronka garrantzitsuenetako bat bizkortu dute, bakte-rioek antibiotikoekiko erresistentzia zabala garatu baitute. Mundu mailako arazo hau Osasun Bakarraren ikuspegitik aztertu behar da, antibiotikoekiko erresistentziak gizakien, animalien eta ingurumenaren artean partekatu, barreiatu eta ugaritzen baitira. Adibidez, lurzoruetan ongarri organikoak aplikatzeak lurren osasuna eta uzten etekina hobetu ditzake. Hala ere, praktika honek arriskuak dakartza, lurzorura iritsi baitaitezke gizaki nahiz abereek guztiz metabolizatu ez dituzten antibiotikoak. Antibiotikoa hedatzeaz gain, antibiotikoekiko erresistenteak diren geneak, bakterioak eta elementu genetiko mugikorrak ba-rreiatzea dakar mota honetako ongarri organikoen erabilerak. Izan ere, lurzoruetan etengabe antibiotikoak sartzeak mehatxu potentziala sortzen du bertako bakterioentzat; bakterio antibiotikoekiko erresistenteak ugaritu egiten dira eta bertako bakterio-komunitateen bioaniztasuna aldatzen dute. Barreiadura zabal honen ondorioz, gero eta gehiago, antibiotikoen erabilera ez da eraginkorra izaten ari erresistente diren patogenoen infekzioen tratamenduetan. Berrikuspen honetan, lehenik, antibiotikoen nahiz antibiotikoekiko erresistente diren geneen jatorria eta mekanismoak azaltzen dira. Jarraian, lurzoruetan egiten den giza edo animalia-jatorriko ongarri organikoen erabilera aipatzen da, antibiotikoak eta erresistentziak ingurumenean barreiatzeko sarbide garrantzitsuak direlako. Ondoren, antibiotikoekiko erresistentziek ingurumenean duten eragina azaltzen da, esposizio-bidea jarraituz kultiboetatik gizakietara iritsi arte. Azkenik, abereetatik eratorritako ongarri organikoetan anti-biotikoekiko erresistentzien arriskua gutxitzeko erabilgarriak izan litezkeen kudeaketa-prozesuak proposatzen ditugu

Zero-valent iron nanoparticles and organic amendment assisted rhizoremediation of mixed contaminated soil using Brassica napus

Soil is one of our most important natural resources. Regrettably, the expansion of human activities has resulted in the degradation of the soil resource due to contamination with a myriad of organic and inorganic compounds. The remediation of mixed contaminated soils, i.e. soils contaminated with both organic compounds and metals, is challenging as it requires actions to simultaneously decrease metal-induced risks and organic contaminant concentrations. Here, we evaluated the effect of the addition of zero-valent iron nanoparticles (nanoremediation) and organic amendments (biostimulation) on the rhizoremediation, using Brassica napus plants, of soil simultaneously contaminated with zinc (2500 mg kg(-1)) and lindane (100 mg kg(-1)). We used a factorial design with three factors (amendment, nZVI, plant) to evaluate the impact of the applied remediation actions on lindane and extractable Zn concentrations, as well as on soil health recovery as manifested by the values of different soil microbial indicators. The studied microbial indicators were not negatively affected by nZVI application. The application of nZVI was the most effective factor regarding the targeted reduction in lindane concentration (51% average reduction in nZVI treated soils). The highest reduction in extractable Zn was achieved in the presence of B. napus, nZVI and organic amendments (99 and 95% reduction in horse manure-amended and sewage sludge-amended soils, respectively). The combination of the three factors led to the highest values of soil microbial indicators (although a significant triple interaction was not observed for all parameters), especially when combined with horse manure amendment: in this case, prokaryotic richness increased by 64%, respiration by 376%, eukaryotic abundance by 333%, and prokaryotic abundance by 437%, compared to untreated soils. The combination of remediation approaches (rhizoremediation with B. napus, nanoremediation with nZVI, biostimulation with organic amendments) can help overcome the limitations of each individual strategy.This work was supported by the European Union through Interreg SUDOE Program (Project Phy2SUDOE SOE4/P5/E1021), the Spanish Ministry of Economy, Industry, and Competitiveness through NANORRIZORREM-2 (AGL2016-76592-R) and PRADA projects (PID2019-110055RB-C21 and PID2019-110055RB-789 C22), MCIN/AEI/10.13039/501100011033/FEDER, UE, and Consolidated Research Group of the Basque Government (GV ITO18-16). JH is the recipient of a predoctoral fellowship from the Spanish Ministry of Science and Innovation. We sincerely thank Dr. Fernando Blanco for technical assistance

The Application of Nanoscale Zero-Valent Iron Promotes Soil Remediation While Negatively Affecting Soil Microbial Biomass and Activity

The use of nanoscale zero-valent iron (nZVI) particles for soil remediation is gaining increased attention. However, there are concerns about the potential adverse effects of nZVI on soil microbial communities and, hence, soil quality. The objective of this study was to assess the impact of the application of nZVI on soil microbial parameters (as bioindicators of soil quality) during the nanoremediation of soil artificially contaminated with lindane (10 mg gamma-HCH kg(-1) DW soil) and zinc (1,500 mg Zn kg(-1) DW soil). nZVI particles were also applied to non-contaminated soil. The following nZVI doses were applied twice: 0, 0.25, 0.5, 1, and 2 mg nZVI g(-1) DW soil. Nine weeks after nZVI application, the following parameters were determined in soil samples: lindane concentration, extractable Zn concentration, microbial biomass carbon (C-MB), bacterial and fungal abundance (gene copy numbers by qPCR), enzyme activities (beta-glucosidase, beta-glucosaminidase, xylosidase, acid phosphatase, arylsulphatase, and Leu-aminopeptidase) and bacterial richness by ARISA profiles. The application of nZVI reduced lindane and extractable Zn concentrations following a dose-dependent pattern. The presence of contaminants reduced soil microbial biomass and activity. The application of nZVI negatively affected the microbial quality of the contaminated soil but not of the non-contaminated soil. This observation might reflect a "stress-on-stress" effect, i.e., the already stressed microbial populations present in the contaminated soil were more sensitive to the application of nZVI (a second stress) than those present in the non-contaminated soil.This work was supported by the Spanish Ministry of Economy, Industry and Competitiveness through NANORRIZORREM-2 Project (AGL2016-76592-R)

Tarjetas de salud para la evaluación de la sostenibilidad agrícola

Agroecosystem Health Cards are handbooks that provide straightforward, practical information on how to assess agroecosystem health through the analysis of a group of aboveground (crop) and, above all, belowground (soil) indicators. In addition, they allow for the assessment of the impact of specific agricultural practices on agroecosystem health. The main objective of these Agroecosystem Health Cards is to provide farmers and other stakeholders related to agroecosystem management (technical staff, scientists and administration staff) with a practical tool that allows them to evaluate the impact of agricultural practices on agroecosystem health by themselves, so that they can choose those management options that prove to be more sustainable. To this purpose, these Agroecosystem Health Cards explain which health indicators can be measured, how to do it properly, the meaning of the different indicators and, finally, the reference values considered “good”, “medium” and “bad”. Indicators are divided into two categories (basic and advanced) in order to carry out a basic and advanced diagnosis of health, respectively, depending on the interests and possibilities of the user.Os Cartões de Saúde dos Agroecossistemas são manuais que fornecem informaçao de forma prática e simples sobre como avaliar a saúde global de um agroecossistema através da análise de indicadores de superfície (culturas) e, pricipalmente, indicadores subterrâneos (solo). Além disso, os indicadores permitem a avaliação do impacto de práticas agrícolas específicas na saúde dos agroecossistemas. O principal objectivo destes cartões é o de proporcionar aos agricultores e outros intervenientes relacionados com a gestão dos agroecossistemas (pessoal técnico, cientistas e pessoal de administração) uma ferramenta prática que lhes permite avaliar o impacto das práticas agrícolas na saúde dos agroecossistemas por si só, por forma a que possam escolher as opções de gestão que provem ser mais sustentáveis. Para o efeito, estes cartões definem quais os indicadores de saúde que podem ser medidos, como fazê-lo corretamente, o significado dos diferentes indicadores e, finalmente, os valores de referência considerados como "bom", "médio" e "mau". Os indicadores são divididos em duas categorias (básica e avançada), a fim de levar a cabo um diagnóstico básico e avançado da saúde, respectivamente, de acordo com os interesses e possibilidades do utilizador.Las Tarjetas de Salud de los Agroecosistemas son manuales que explican de manera sencilla y práctica cómo puede realizarse un diagnóstico global del estado de salud de los ecosistemas agrícolas, a través del análisis de una serie de indicadores a nivel superficial (cultivo) y, sobre todo, subterráneo (suelo). Asimismo, nos permiten valorar el impacto de una determinada práctica agraria en la salud de los agroecosistemas. El objetivo fundamental de las Tarjetas es dotar a los agricultores y demás agentes implicados en la gestión de los agroecosistemas (técnicos, científicos y administración) de una herramienta práctica que les permita evaluar por sí mismos el impacto de sus prácticas agrícolas sobre la salud de los agroecosistemas, pudiendo así optar por aquellas alternativas de manejo que demuestren ser más sostenibles. Para ello, estas Tarjetas detallan qué indicadores de salud se pueden medir, cómo hacerlo correctamente, el significado de los diferentes indicadores y, finalmente, los valores de referencia considerados “buenos”, “regulares” y “malos”. Los indicadores se dividen en dos categorías (básicos y avanzados) que permiten realizar un diagnóstico de salud “básico” o “avanzado”, respectivamente, en función  de los intereses y posibilidades del usuario

EFECTO DEL ABANDONO DEL PASTOREO SOBRE LA SALUD DE LOS ECOSISTEMAS PASCÍCOLAS = EFFECTS OF GRAZING ABANDONMENT ON THE HEALTH OF PASTURE ECOSYSTEMS

ResumenLa disminución de la actividad pastoral en zonas de fuerte tradición ganadera provoca cambios no sólo a nivel superficial (comunidades vegetales) sino también a nivel subterráneo (propiedades fisicoquímicas y biológicas del suelo). El objetivo principal de este estudio fue la evaluación del efecto del abandono del pastoreo sobre la salud de los ecosistemas pascícolas, con especial interés en su soporte edá- fico. Para ello, se establecieron tres cierres (exclusiones al pastoreo) en la Finca Experimental de NEIKER-Tecnalia (Arkaute, Álava) donde tradicionalmente pasta un rebaño de ovejas de raza latxa. Tras año y medio de exclusión, se realizaron medidas de campo y se recogie- ron muestras para su posterior análisis en el laboratorio para obtener información sobre el estado de las comunidades vegetales y sobre la actividad, biomasa y diversidad de las comunidades edáficas. En el periodo de tiempo estudiado, la diversidad vegetal de los pastos no mostró variación entre las zonas pastoreadas y las zonas de exclusión. Por el contrario, en las zonas excluidas al pastoreo, la diversi- dad de macrofauna fue significativamente mayor que en las zonas pastadas por las ovejas. El resto de parámetros físicos y microbianos analizados también parecieron seguir esta tendencia positiva, si bien las diferencias no fueron estadísticamente significativas debido a la variabilidad entre muestras. Esto pone de manifiesto la necesidad de incrementar el número de réplicas en este tipo de estudios de campo, siendo como es el suelo un sistema altamente heterogéneo.AbstractThe decline in pastoral activity in areas with long lasting livestock raising tradition leads to changes not only at the surface level (plant communities) but also at the belowground level (physicochemical and biological soil properties). The main objective of this study was to evaluate the effect of grazing abandonment on the health of pasture ecosystems, with special emphasis on the soil ecosystem. To this purpose, three grazing exclusion areas were established in Neiker-Tecnalia Experimental Farm (Arkaute, Alava) where traditionally a flock of sheep (latxa breed) graze. After one and a half years, field measurements were performed and samples were taken to the laboratory to obtain information about the status of plant communities and the activity, biomass and diversity of soil communities. In the study pe- riod, plant diversity of pastures showed no variation in grazed areas versus exclusion areas. By contrast, in the grazing exclusion areas, macrofauna diversity was significantly higher than in areas grazed by sheep. The remaining physical and microbial parameters also ap- peared to follow this positive trend, although differences were not statistically significant due to the high variability between samples. This highlights the need to increase the number of replicates in this type of field experiments, since the soil ecosystem is highly heterogeneous