Anticancer drugs impact the performance and prokaryotic microbiome of an aerobic granular sludge system operated in a sequential batch reactor

Increased concerns exist about the presence of anticancer drugs in wastewater. However, knowledge of the impacts of anticancer drugs on the performance of the system and microbial communities during wastewater treatment processes is limited. We examined the effect of three anticancer drugs commonly detected in influents of wastewater treatment plants applied at three different concentration levels on the performance, efficiency of anticancer drug removal, and prokaryotic microbiome in an aerobic granular sludge system (AGS) operated in a sequential batch reactor (SBR). We showed that an AGS can efficiently remove anticancer drugs, with removal rates in the range of 53–100% depending on the type of drug and concentration level. Anticancer drugs significantly decreased the abundance of total bacterial and archaeal communities, an effect that was linked to reduced nitrogen removal efficiency. Anticancer drugs also reduced the diversity, altered the prokaryotic community composition, reduced network complexity, and induced a decrease of a wide range of predicted bacterial functions. Specific bacterial taxa responsive to the addition of anticancer drugs with known roles in nitrification and denitrification were identified. This study shows anticancer drugs should be monitored in the future as they can induce changes in the performance and microbiome of wastewater treatment technologies

Application of full-scale aerobic granular sludge technology for removing nitrate from groundwater intended for human consumption

The dependence on groundwater for human consumption has increased worldwide in the last decades. Nitrate (NO3−) often reaches groundwater and causes significant degradation in groundwater quality. In an effort to address this issue, a full-scale water treatment plant using aerobic granular sludge (AGS) technology was built to remove NO3− from nitrate-polluted groundwater intended for human consumption in a rural village. The impact of changes in the operational conditions of hydraulic retention time (HRT) and organic matter loading (OML) rate on NO3− removal, overall system performance, and the granule microbiome were studied. Regardless of the HRT, the AGS technology was successful in removing NO3− with removal rates >50 % with an optimal OML rate of 75 mg L−1. No significant variations in the total abundance of any of the denitrification genes were observed. The composition of prokaryotic and eukaryotic communities was affected by changes in the HRT and OML rate. Specific prokaryotic taxa were identified as responsive to changes in operational parameters and their abundances were linked to the removal of NO3−, confirming that the microbes are critical to the NO3− removal process. This study demonstrates that the AGS technology can be successfully implemented to treat nitrate-polluted groundwater in rural villages to produce water of drinking quality. In addition, the reported hydraulic retention times and organic matter loading rate can be used to further improve the system performance to remove nitrate from groundwater

Analysis of the denitrification pathway and greenhouse gases emissions in Bradyrhizobium sp. strains used as biofertilizers in South America

Aims Greenhouse gases are considered potential atmospheric pollutants, with agriculture being one of the main emission sources. The practice of inoculating soybean seeds with Bradyrhizobium sp. might contribute to nitrous oxide (N2O) emissions. We analyzed this capacity in five of the most used strains of Bradyrhizobium sp. in South America. Methods and Results We analyzed the denitrification pathway and N2O production by B. japonicum E109 and CPAC15, B. diazoefficiens CPAC7, and B. elkanii SEMIA 587 and SEMIA 5019, both in free‐living conditions and symbiosis with soybean. The in silico analysis indicated the absence of nosZ genes in B. japonicum and the presence of all denitrification genes in B. diazoefficiens strains, as well as the absence of nirK, norC and nosZ genes in B. elkanii. The in planta analysis confirmed the N2O production under saprophytic conditions or symbiosis with soybean roots nodules. In the last case up to 26·1 and 18·4 times higher in plants inoculated with SEMIA5019 and E109 respectively, than in those inoculated with USDA110. Conclusions The strains E109, SEMIA 5019, CPAC15 and SEMIA 587 showed the highest N2O production both as free‐living cells and in symbiotic conditions in comparison with USDA110 and CPAC7, which do have the nosZ gene. Although norC and nosZ could not be identified in silico or in vitro in SEMIA 587 and SEMIA 5019, these strains showed capacity to produce N2O in our experimental conditions. Significance and Impact of Study This is the first report to analyze and confirm the incomplete denitrification capacity and N2O production in four of the five most used strains of Bradyrhizobium sp. for soybean inoculation in South America.Fil: Obando Castellanos, Dolly Melissa. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Ciencias Naturales. Laboratorio de Fisiología Vegetal y de la Interacción Planta-microorganismo; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Correa-Galeote, David. Consejo Superior de Investigaciones Científicas. Estación Experimental del Zaidín; EspañaFil: Castellano-Hinojosa, Antonio. Consejo Superior de Investigaciones Científicas. Estación Experimental del Zaidín; EspañaFil: Gualpa, José Luis. Universidad Nacional de Río Cuarto. Facultad de Ciencias Exactas Fisicoquímicas y Naturales. Departamento de Ciencias Naturales. Laboratorio de Fisiología Vegetal y de la Interacción Planta-microorganismo; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Hidalgo, Alba. Consejo Superior de Investigaciones Científicas. Estación Experimental del Zaidín; EspañaFil: de Dios Alché, Juan. Consejo Superior de Investigaciones Científicas. Estación Experimental del Zaidín; EspañaFil: Bedmar, Eulogio. Consejo Superior de Investigaciones Científicas. Estación Experimental del Zaidín; EspañaFil: Cassan, Fabricio Dario. Universidad Nacional de Rio Cuarto. Facultad de Cs.exactas Fisicoquimicas y Naturales. Instituto de Investigaciones Agrobiotecnologicas. - Consejo Nacional de Investigaciones Cientificas y Tecnicas. Centro Cientifico Tecnologico Conicet - Cordoba. Instituto de Investigaciones Agrobiotecnologicas.; Argentin

Emission of greenhouse gases and microbial biodiversity in soils of agricultural interest. Effect of nitrogen fertilisation

En los suelos agrícolas, la aplicación de fertilizantes inorgánicos nitrogenados conduce a la interacción de múltiples factores y procesos que están asociados principalmente con cambios en las propiedades fisicoquímicas del suelo, la emisión de gases de efecto invernadero y la ecología microbiana. Aunque el nitrógeno (N) es un nutriente esencial para el crecimiento de las plantas, el aumento de la fertilización nitrogenada ha alterado el ciclo natural de N en la biosfera, lo que resulta en diversos impactos ambientales, ecológicos y sobre la salud humana. Entre ellos, el aumento de la emisión de gases de efecto invernadero, de la lluvia ácida y eutrofización. Después de la aplicación de un fertilizante nitrogenado al suelo, los procesos microbianos de nitrificación y desnitrificación son los principales responsables de las reacciones que conducen a la conversión de amonio (NH4+) y nitrato (NO3-), respectivamente, a la liberación a la atmósfera del gas de efecto invernadero óxido nitroso (N2O). Combatir los impactos negativos del aumento de los flujos de N2O plantea desafíos considerables y será ineficaz sin incorporar los procesos microbianos que intervienen en la emisión de N2O en las estrategias de mitigación. Aunque previos estudios han mostrado la existencia de relaciones individuales entre la fertilización con N y cambios en las propiedades bióticas y abióticas del suelo, un estudio integrado relacionando la forma del fertilizante N con diferencias en la emisión de N2O, cambios en las propiedades fisicoquímicas del suelo, alteraciones en la abundancia de los genes involucrados en la producción y reducción de N2O y los efectos sobre la diversidad bacteriana no se ha realizado. La investigación de los efectos de la aplicación simple y combinada del inhibidor de la ureasa N-(n-butil) triamida tiofosfórica (NBPT) y del inhibidor de la nitrificación 3,4 dimetilpirazol fosfato (DMPP) sobre la volatilización de amoniaco (NH3) y la abundancia de las comunidades nitrificantes y desnitrificantes también se abordó en este estudio. La aplicación de NBPT redujo la volatilización de NH3 y no afectó la abundancia de bacterias y arqueas totales, ni la de los genes de la nitrificación, pero redujo la abundancia de genes de la desnitrificación al 80% de WFPS. El DMPP, solo y en combinación con NBPT, aumentó la volatilización del NH3 y la abundancia de bacterias y arqueas totales así como de la comunidad nitrificante en el suelo. Independientemente de las condiciones de humedad, DMPP y, en menor medida, DMPP + NBPT, aumentaron el número de copias los genes norB y nosZ, lo que indica que DMPP, de alguna manera, induce la expresión de, al menos, estos dos genes de la desnitrificación.In agricultural soils, the application of inorganic nitrogen (N)-fertilisers leads to the interaction of multiple factors and processes which are mainly associated with changes in soil physicochemical properties, emission of greenhouse gases and microbial ecology. Although N is an essential nutrient for plant growth, increased application of N-fertilisers in agriculture has altered the natural N cycle, which result in many environmental, ecological and human health impacts. Among them, N-fertilisation may lead to an increase in the emission of greenhouse gases, acidic deposition and eutrophication. After application of an N-fertiliser, the microbial processes of nitrification and denitrification are the main esponsible of the reactions driving the conversion of ammonia (NH4+) and nitrate (NO3-) to the release of the greenhouse gas nitrous oxide (N2O) into the atmosphere, respectively. Combating the negative impacts of increasing N2O fluxes poses considerable challenges and will be ineffective without incorporating microbial regulated N2O processes into mitigation strategies. Although previous studies had shown individual relationships between N-fertilisation and soil biotic and abiotic parameters, an integrated study relating the form of the N-fertiliser with differences in N2O emission, changes in soil physicochemical properties, alterations in the abundance of the genes involved in N2O production and reduction, and effects on bacterial diversity had not been reported. Investigation of the effects of the single and combined application of the urease inhibitor N-(n-butyl) thiophosphoric triamide (NBPT) and the nitrification inhibitor 3,4 dimethylpyrazole phosphate (DMPP) on ammonia (NH3) volatilisation and the abundance of the nitrifier and denitrifier communities have also pursued in this study. The application of the urease inhibitor NBPT reduced NH3 volatilisation and did not affect the bacterial and archaeal abundance, nor that of the nitrifiers, but reduced the abundance of denitrifiers at 80% WFPS. DMPP, alone and in combination with NBPT, increases NH3 volatilisation and the abundance of bacteria, archaea and nitrifiers in the soil. Regardless of the moisture conditions, DMPP and to a lower extent DMPP + NBPT, increases the gene copy number of the norB- and nosZ-bearing denitrifying communities, which indicates that DMPP, somehow, induces the expression of the, at least, the norB and nosZ denitrification genes.Tesis Univ. Granada.Esta investigación forma parte de los Proyectos de Investigación "Emisión de óxido nitroso por suelos cultivados con leguminosas y hortalizas de interés agrícola", referencia Proyecto de Excelencia AGR-1968, de la Consejería de Economía, Innovación y Ciencia de la Junta de Andalucía, y RECUPERA 2020, "Emisión de óxido nitroso en suelos y biodiversidad bacteriana asociada a la fertilización nitrogenada", referencia 20134R070, financiado por el Ministerio de Economía y Competitivida

Impact of Cover Crops on the Soil Microbiome of Tree Crops

Increased concerns associated with interactions between herbicides, inorganic fertilizers, soil nutrient availability, and plant phytotoxicity in perennial tree crop production systems have renewed interest in the use of cover crops in the inter-row middles or between trees as an alternative sustainable management strategy for these systems. Although interactions between the soil microbiome and cover crops have been examined for annual cropping systems, there are critical differences in management and growth in perennial cropping systems that can influence the soil microbiome and, therefore, the response to cover crops. Here, we discuss the importance of cover crops in tree cropping systems using multispecies cover crop mixtures and minimum tillage and no-tillage to not only enhance the soil microbiome but also carbon, nitrogen, and phosphorus cycling compared to monocropping, conventional tillage, and inorganic fertilization. We also identify potentially important taxa and research gaps that need to be addressed to facilitate assessments of the relationships between cover crops, soil microbes, and the health of tree crops. Additional evaluations of the interactions between the soil microbiome, cover crops, nutrient cycling, and tree performance will allow for more effective and sustainable management of perennial cropping systems

Native Rhizobia Improve Plant Growth, Fix N2, and Reduce Greenhouse Emissions of Sunnhemp More than Commercial Rhizobia Inoculants in Florida Citrus Orchards

Sunnhemp (Crotalaria juncea L.) is an important legume cover crop used in tree cropping systems, where there is increased interest by growers to identify rhizobia to maximize soil nitrogen (N) inputs. We aimed to isolate and identify native rhizobia and compare their capabilities with non-native rhizobia from commercial inoculants to fix atmospheric dinitrogen (N2), produce and reduce nitrous oxide (N2O), and improve plant growth. Phylogenetic analyses of sequences of the 16S rRNA and recA, atpD, and glnII genes showed native rhizobial strains belonged to Rhizobium tropici and the non-native strain to Bradyrhizobium japonicum. Plant nodulation tests, sequencing of nodC and nifH genes, and the acetylene-dependent ethylene production assay confirmed the capacity of all strains to nodulate sunnhemp and fix N2. Inoculation with native rhizobial strains resulted in significant increases in root and shoot weight and total C and N contents in the shoots, and showed greater N2-fixation rates and lower emissions of N2O compared to the non-native rhizobium. Our results suggest that native rhizobia improve plant growth, fix N2, and reduce greenhouse emissions of sunnhemp more than commercial rhizobia inoculants in Florida citrus orchards

Efficiency of reactive nitrogen removal in a model Mediterranean high-mountain lake and its downwater river ecosystem: Biotic and abiotic controls

High-mountain lakes and rivers are usually oligotrophic and strongly influenced by atmospheric transport processes. Thus, wet deposition of reactive N species (Nr), mainly in the form of nitrate (NO), is a major source of N input in these high-mountain ecosystems. Bacterial denitrifiers are thought to be largely responsible for reduction of NO to nitrous oxide (NO) and molecular dinitrogen (N) as main biological pathway of N removal in these ecosystems. Nitrifiers, through the oxidation of ammonium to NO, can also be a source of NO and NO. However, there is uncertainty regarding the abiotic and biotic factors controlling Nr elimination from aquatic ecosystems at different altitudes and seasons. We examined the efficiency of Nr removal as NO and N (total removal) or N only (clean removal) in a model lake and its downwater river ecosystem (Sierra Nevada, Spain) representative of Mediterranean high-mountain freshwater ecosystems along an altitudinal gradient during the warm period of the year. Denitrification activity and the abundance of nitrifiers and denitrifiers in sediments were measured at thaw, mid ice-free and late ice-free periods. We found the efficiency of total and clean removal of Nr increased from the downwater river to the high-mountain lake. Regardless of the location, the efficiency of total removal of Nr decreased over the ice-free period whereas that of clean removal of Nr peaked at mid ice-free period. The efficiency of total removal of Nr was mainly controlled by the abundance of archaeal nitrifiers and bacterial denitrifiers. Abiotic (ammonium and NO concentration) and biotic (mainly nosZI-type denitrifiers) factors drove changes in the efficiency of clean removal of Nr. Our results suggest that abiotic and biotic factors can control the efficiencies of Nr removal in Mediterranean high-mountain lakes and their downwater rivers, and that these efficiencies increase with altitude and vary over the ice-free period.We thank two anonymous reviewers and Editor (Dr. Sergi Sabater) whose criticism greatly improved the manuscript. This study was funded by FEDER/Junta de Andalucía-Consejería de Economía y Conocimiento/Proyecto (A-RNM-237-UGR18), and partially by PID2020-118872RB-I00 (MICIN/AEI/10.13039/501100011033) projects. This research is also part of the project “Thematic Center on Mountain Ecosystem & Remote sensing, Deep learning-AI e-Services University of Granada-Sierra Nevada” (LifeWatch-2019-10-UGR-01), which has been co-funded by the Ministry of Science and Innovation through the FEDER funds from the Spanish Pluriregional Operational Program 2014-2020 (POPE), LifeWatch-ERIC action line

Effect of nitrogen fertilisation on nitrous oxide emission and the abundance of microbial nitrifiers and denitrifiers in the bulk and rhizosphere soil of Solanum lycopersicum and Phaseolus vulgaris

Aims: To determine the effect of three N-fertilisers on NO emission and abundance of nitrification and denitrification genes in bulk and rhizosphere soil of tomato and common bean, two vegetable crops representative of main horticultural crops in South Spain. Methods: Four consecutive harvests of tomato and common bean fertilised with urea, ammonium or nitrate were carried out. The total abundance of bacteria, archaea, nitrifiers and denitrifiers was estimated by quantitative PCR. Soil physicochemical properties and NO emission were also determined. Results: Regardless of the plant species, the highest NO emission was produced by the soil treated with urea, followed by ammonium and nitrate. Bacteria were more abundant than archaea in the bulk and rhizosphere soil. The abundance of the ammonia-oxidising archaea was greater than the ammonia-oxidising bacteria in the rhizosphere, but lower in the bulk soil. N-fertilisation increased the gene copy number of denitrifiers, which were more abundant in the bulk soil. Conclusions: N-fertilisation decreases NO production due to increased abundance of the nosZ gene. The abundance of nitrification and denitrification genes in bulk and rhizosphere soils is dependent on the type of fertiliser. For both plant species, the ratio of the genes involved in production and reduction of NO by bulk and rhizosphere was similar.This study was supported by the ERDF-cofinanced grant PEAGR2012-1968 from Consejería de Economía, Innovación y Ciencia (Junta de Andalucía, Spain) and the MINECO-CSIC Agreement RECUPERA 2020. ACH is the recipient of a grant of MECD (FPU 2014/01633)

Distinct effect of nitrogen fertilisation and soil depth on nitrous oxide emissions and nitrifiers and denitrifiers abundance

The nitrous oxide and molecular N emissions from 5-cm length subsamples taken from 20-cm length sample corers containing eutric Cambisol soil fertilised either with urea, ammonium or nitrate for 1 year have been examined using gas chromatography. At the beginning of the incubation, the same N rate (260 kg N/ha) was added to the soil and kept constant during the experiment. The total abundance of the soil Bacteria and Archaea and that of nitrifiers and denitrifiers was estimated by quantitative PCR of the corresponding biotic variables 16S rRNA, amoA and napA, narG, nirK, nirS, norB, nosZI and nosZII genes. The abiotic variables dissolved oxygen, pH, exchangeable NH -N and NO -N contents and total C and total N were also analysed. None of the three fertilisers affected the total abundance of Bacteria and Archaea and nitrification was the main driver of nitrous oxide production in the 0- to 5-cm and 5- to 10-cm soil layers while denitrification was in the 10- to 15-cm and 15- to 20-cm soil horizons. Parallel to the reduction in the content of dissolved oxygen along the soil profile, there was a decrease in the total and relative abundance of the bacterial and archaeal amoA gene and an increase in the abundances of the denitrification genes, mainly in the 10- to 15-cm and 15- to 20-cm soil layers. A non-metric multidimensional scaling plot comparing the biotic and abiotic variables examined in each of the four 5-cm soil subsamples and the whole 20-cm sample showed a disparate effect of N fertilisation on N gas emissions and abundance of nitrifiers and denitrifiers bacterial and archaeal communities.This study was supported by the ERDF-cofinanced grant PEAGR2012-1968 from Consejería de Economía, Innovación y Ciencia (Junta de Andalucía, Spain) and the MINECO-CSIC Agreement RECUPERA 2020. ACH is recipient of a grant of MECD (FPU 2014/01633).Peer Reviewe

Zinc-nitrogen co-fertilization influences N2O emissions and microbial communities in an irrigated maize field

Previous studies have shown that the use of zinc (Zn) chelate fertilizers combined with a nitrogen (N) fertilizer (urea) can lead to both agronomic (i.e., yields and Zn and N biofortification due to the synergies between both nutrients) and environmental (i.e., by reducing the emissions of nitrous oxide, NO, derived from N fertilization) benefits under rainfed semi-arid conditions. However, little is known about the effect of Zn-N co-fertilization on greenhouse gas (GHG) emissions or soil microbial processes involved in NO fluxes under non-flooded irrigated conditions (during the dry season). Under these conditions, water-filled pore space continuously fluctuates following a periodic pattern and soil temperatures are in the optimum range for soil microorganisms. In this context, a field experiment was conducted using a maize (Zea mays L.) crop treated with two N levels (no N application and 120 kg N ha as urea), and three Zn sources (no Zn application, Zn sulphate, and Zn applied with a mixture of chelating compounds, DTPA-HEDTA-EDTA). Nitrous oxide, methane (CH) and carbon dioxide (CO) fluxes were measured using opaque chambers, as well as the total abundances of soil bacteria, archaea and nitrifier and denitrifier communities. Zn-N co-fertilization increased cumulative NO emissions from 0.36 kg N-NO ha (for urea combined with Zn chelates) to 0.76 kg N-NO ha (for urea combined with Zn sulphate), with respect to urea without Zn application. The NO emission factors were lower (0.34%–0.72%) than the IPCC default value of 1%. Total abundances of the nosZ denitrification gene, which is involved in the reduction in NO to dinitrogen (N), were reduced by 75% on average in the plots that received Zn fertilizers. This reduction may explain the higher NO emissions in these treatments. In contrast with the case with non-irrigated crops, Zn-N co-fertilization cannot be recommended as a strategy to mitigate NO emissions in irrigated maize under semi-arid conditions, despite of the enhancement of Zn availability in soil.Financial support was provided by an ERDF-cofinanced grant AGL2015-64582-C3-3-R (MINECO-FEDER) from the Ministerio de Economía y Competitividad (Spanish Government). We are grateful to the Comunidad de Madrid (Spain) and Structural Funds 2014-2020 (ERDF and ESF) for the financial support (project AGRISOST-CM S2013/ABI-2717). The authors are also grateful to the SIRENA network (Ref. AGL2015-68881-REDT), funded by MINECO, for supporting the stay at the Department of Microbiology and Symbiotic System, Estación Experimental del Zaidín, CSIC. M. Montoya and A. Castellano-Hinojosa are the recipients of the FPI grant BES-2016-076712 and a grant of MECD (FPU 2014/01633), respectively. Special thanks are given to the field assistants working with us at Centro Nacional de Tecnología de Regadíos (CENTER), particularly to Alejandro Sánchez de Ribera. We also thank the technicians of the Department of Chemistry and Food Technology of the ETSIAAB. This work was done within the framework of the Moncloa Campus of International Excellence (UCM-UPM)