Antibiotics and antibiotic resistance genes in anaerobic digesters and predicted concentrations in agroecosystems.

In recent decades, the innovative practice of management and valorization of agrozootechnical waste as energy through anaerobic digestion (AD) has been rapidly growing. However, whether applying digestate to soil as biofertilizer can be a source of antibiotics (ABs) and antibiotic resistance genes (ARGs) has not been fully investigated so far. In this work the ARGs responsible for sulfamethoxazole (SMX) resistance (sul1, sul2), ciprofloxacin (CIP) resistance (qnrS, qepA, aac-(6')-Ib-cr) and the mobile genetic element intl1, together with the concentrations of the antibiotics SMX and CIP, were measured in several anaerobic digesters located in Central Italy. Based on these results, the concentrations of antibiotics and ARGs which can potentially reach soil through amendment with digestate were also estimated. The highest CIP and SMX concentrations were found during winter and spring in anaerobic digesters. The highest ARG abundances were found for the aac-(6')-Ib-cr and sul2 genes. The overall results showed that application of digestate to soil does not exclude AB contamination and spread of ARGs in agroecosystems, especially in the case of ciprofloxacin, owing to its high intrinsic persistence.

Toxic response of the bacterium Vibrio fischeri to sodium lauryl ether sulphate residues in excavated soils.

Sodium lauryl ether sulphate (SLES) is the main chemical component in several lubricant products used for soil conditioning in the mechanized excavation industry using Earth Pressure Balance-Tunnel Boring Machines. During the tunnelling process, huge amounts of excavated soil are produced and the SLES presence can affect the subsequent re-use of this material as a by-product. Currently, there is still no regulatory indication of reliable and sensitive bioassays for monitoring soil quality during the excavation process. The main objective of this work was to verify if the Vibrio fischeri screening test was suitable as a consistent and precautionary tool for this specific purpose. Firstly, the ecotoxicity (EC20 and EC50) of the SLES standard solution and three commercial products (SLES content from 10 to 50%) were evaluated to select the most environmental friendly product. Subsequently, soil samples from about 2 years of tunnelling in a real construction site, conditioned with the selected product, were evaluated for their environmental compatibility with the prescriptions of an Italian site-specific protocol. The latter established 2 mg/L as a threshold value for SLES concentration in soil water extracts and a no toxic response (≤20%) for the Vibrio fischeri test. The comparison of the bacterium bioluminescence inhibition values (%) with analytical determinations showed an ecotoxicity when SLES was >2 mg/L. The toxicity was directly related to SLES concentration, indicating that the V. fischeri test and the SLES analyses are suitable tools for assessing excavated soil as a by-product, ensuring its safe reuse in accordance with a green production process (circular economy).

Gasification treatment of poplar biomass produced in a contaminated area restored using plant assisted bioremediation.

Remediation of polluted soils using phytoremediation techniques is an effective strategy. However, the use of the biomass from these soils for energy purposes may raise efficiency and pollution emission problems and there is currently little research on this issue. In this work, the main results of a fluidized-bed gasification treatment conducted on poplar biomass pruning residues from a multi-contaminated area are presented. The samples were collected from an experimental site in which a plant-assisted bioremediation (PABR) technology has been applied since 2013 to reduce the soil heavy metal (HM) and polychlorinated biphenyl (PCB) contents. The main goal of this study was to identify the specific treatment necessary, in addition to conventional tar reforming, for trapping possible residues of HMs and PCBs in ashes during the gasification process. In our study, we demonstrate that gasification of contaminated biomass coming from PABR (where contaminant residues are concentrated mainly in the roots and are insignificant in the shoots) produces syngas whose characteristics are similar to those obtained using non-contaminated biomass. The results showed that contaminant concentrations in the prunings were negligible; the total amount of PCBs was 1.63 ng/g, while HMs ranged from 0.01 to 0.70 mg/kg, except for Cu and Zn (∼20 mg/kg). Furthermore, the presence in the biomass of Ca and traces of other metals showed a possible catalytic effect with an improvement in the tar conversion in the gasifier leading to a reduction of 5-10% in tar content. The overall results suggest that a specific treatment for pollutant capture is necessary only when the roots, the part of the plants where these contaminants are concentrated, are sampled and used for the gasification process. Although energy from biomass produced on a contaminated site is currently considered waste and involves disposal costs, this paper shows that the poplar biomass grown on a multi-contaminated soil can be used for energy purposes without any impact on the environment.

Electroactive Bacteria in Natural Ecosystems and Their Applications in Microbial Fuel Cells for Bioremediation: A Review.

Electroactive bacteria (EAB) are natural microorganisms (mainly Bacteria and Archaea) living in various habitats (e.g., water, soil, sediment), including extreme ones, which can interact electrically each other and/or with their extracellular environments. There has been an increased interest in recent years in EAB because they can generate an electrical current in microbial fuel cells (MFCs). MFCs rely on microorganisms able to oxidize organic matter and transfer electrons to an anode. The latter electrons flow, through an external circuit, to a cathode where they react with protons and oxygen. Any source of biodegradable organic matter can be used by EAB for power generation. The plasticity of electroactive bacteria in exploiting different carbon sources makes MFCs a green technology for renewable bioelectricity generation from wastewater rich in organic carbon. This paper reports the most recent applications of this promising technology for water, wastewater, soil, and sediment recovery. The performance of MFCs in terms of electrical measurements (e.g., electric power), the extracellular electron transfer mechanisms by EAB, and MFC studies aimed at heavy metal and organic contaminant bioremediationF are all described and discussed.

Overview of Direct and Indirect Effects of Antibiotics on Terrestrial Organisms.

Antibiotics (ABs) have made it possible to treat bacterial infections, which were in the past untreatable and consequently fatal. Regrettably, their use and abuse among humans and livestock led to antibiotic resistance, which has made them ineffective in many cases. The spread of antibiotic resistance genes (ARGs) and bacteria is not limited to nosocomial environments, but also involves water and soil ecosystems. The environmental presence of ABs and ARGs is a hot topic, and their direct and indirect effects, are still not well known or clarified. A particular concern is the presence of antibiotics in agroecosystems due to the application of agro-zootechnical waste (e.g., manure and biosolids), which can introduce antibiotic residues and ARGs to soils. This review provides an insight of recent findings of AB direct and indirect effects on terrestrial organisms, focusing on plant and invertebrates. Possible changing in viability and organism growth, AB bioaccumulation, and shifts in associated microbiome composition are reported. Oxidative stress responses of plants (such as reactive oxygen species production) to antibiotics are also described.

Application of the Aliivibrio fischeri bacterium bioassay for assessing single and mixture effects of antibiotics and copper.

The Aliivibrio fischeri bioassay was successfully applied in order to evaluate the acute effect of sulfamethoxazole (SMX), ciprofloxacin (CIP), chlortetracycline (CTC) and copper (Cu), alone or in binary, ternary, and overall mixture. The toxicity results are reported in terms of both effective concentrations, which inhibited 50% of the bacterium bioluminescence (EC50%), and in Toxic Units (TUs). The TUs were compared with predicted values obtained using the Concentration Addition model (CA). Finally, the toxicity of water extracts from a soil contaminated by the three antibiotics (7 mg Kg-1 each) in the presence/absence of copper (30 mg Kg-1) was also evaluated. Copper was the most toxic chemical (EC50: 0.78 mg L-1), followed by CTC (EC50: 3.64 mg L-1), CIP (96 mg L-1) and SMX (196 mg L-1). Comparing the TU and CA values of the mixtures, additive effects were generally found. However, a synergic action was recorded in the case of the CIP+Cu co-presence and antagonistic effects in the case of CTC+Cu and the ternary mixture (containing each antibiotic at 0.7 mg L-1), were identified. Soil water extracts did not show any toxicity, demonstrating the buffering ability of the soil to immobilize these chemicals.

Effects of municipal waste compost on microbial biodiversity and energy production in terrestrial microbial fuel cells.

Microbial Fuel Cells (MFCs) transform organic matter into electricity through microbial electrochemical reactions catalysed on anodic and cathodic half-cells. Terrestrial MFCs (TMFCs) are a bioelectrochemical system for bioelectricity production as well as soil remediation. In TMFCs, the soil is the ion-exchange electrolyte, whereas a biofilm on the anode oxidises organic matter through electroactive bacteria. Little is known of the overall microbial community composition in a TMFC, which impedes complete exploitation of the potential to generate energy in different soil types. In this context, an experiment was performed to reveal the prokaryotic community structure in single chamber TMFCs with soil in the presence and absence of a municipal waste compost (3% w/v). The microbial community was assessed on the anode and cathode and in bulk soil at the end of the experiment (54 days). Moreover, TMFC electrical performance (voltage and power) was also evaluated over the experimental period, varying the external resistance to improve performance. Compost stimulated soil microbial activity, in line with a general increase in voltage and power. Significant differences were observed in the microbial communities between initial soil conditions and TMFCs, and between the anode, cathode and bulk soil in the presence of the compost. Several electroactive genera (Bacillus, Fulvivirga, Burkholdeira and Geobacter) were found at the anode in the presence of compost. Overall, the use of municipal waste compost significantly increased the performance of the MFCs in terms of electrical power and voltage generated, not least thanks to the selective pressure towards electroactive bacteria on the anode.

Mix-Tool: An Edge-of-Field Approach to Predict Pesticide Mixtures of Concern in Surface Water From Agricultural Crops.

Current regulation on the authorization of plant protection products (PPPs) in the European Union is limited to the evaluation of ecological risks for the single active substances they contain. However, plant protection treatments in agriculture often consist of PPPs already containing more than one active substance; moreover, each cropped field receives multiple applications per year, leading to complex pesticide mixtures in the environment. Different transport processes lead to a multitude of heterogeneous and potentially toxic substances that, for example, may reach water bodies and act simultaneously on natural freshwater ecosystems. In this context, the development of methodologies and tools to manage risks of pesticides mixtures is imperative to improve the current ecological risk assessment procedures and to avoid further deterioration of ecological quality of natural resources. The present study suggests new procedures for identifying pesticide mixtures of potential concern released from agricultural crops in surface water. The approach follows the European Union regulatory context for the authorization of PPPs in the market (edge-of field risk assessment) and requires the use of Forum for the Co-ordination of pesticide fate models and their Use (FOCUS) models (Step 3 and 4) for calculating the concentrations in surface water of mixture components on a daily basis. Moreover, it uses concentration addition models to calculate the toxic potency of the pesticide mixtures released by a treated crop. To implement this procedure, we developed a simple Microsoft-Excel-based tool. We also considered two case studies (maize and apple tree), representative of Italian agricultural scenarios for annual and perennial crops. Moreover, we compared results with 3 years of monitoring data of surface water bodies of the Lombardia region (northern Italy) where the two crops are largely present. Environ Toxicol Chem 2022;41:2028-2038. © 2022 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

Use of large datasets of measured environmental concentrations for the ecological risk assessment of chemical mixtures in Italian streams: A case study.

A method to evaluate the ecological risk of chemical mixtures in water bodies is here presented. In the first phase, the approach considered routine chemical monitoring data (MEC: measured environmental concentrations) obtained from the Italian National Institute for Environmental Protection and Research, which were georeferenced to a single coordinate system for each monitoring station. The overall mixture toxicity were then evaluated for three representative aquatic organisms (algae, Daphnia, fish) using the concentration addition model to combine exposure with ecotoxicological data (from different databases). A database management system was used to facilitate the creation, organisation, and management of the large datasets of this study. The outputs were obtained as GIS-based mixture risk maps and tables (listing the toxic unit of mixtures and individual substances) useful for further analysis. The method was applied to an Italian watershed (Adda River) as a case study. In the first phase, the mixture toxicity was calculated using two scenarios: best- and worst-case; wherein the former included only those compounds that were be detected, while the latter involved also substances with concentrations below the limit of quantification. The ratio between the two scenarios indicated the range within which mixture toxicity should ideally vary. The method demonstrates that these ratios were very small when the calculated toxicity using the best case indicated a potential risk and vice versa, indicating that the worst-case scenario could not be appropriate (extremely conservative). Consequently, in the successive phase, we focused exclusively on the best-case scenario. Finally, this approach allowed the priority mixture identification (those most likely occurring in the analysed water samples), algae as the organism at the highest risk, and the substances that contributed the most to the overall mixture toxicity (terbuthylazine and s-metolachlor for algae, and chlorpyrifos and chlorpyrifos-CH3 for Daphnia and fish).

Bioaccumulation of antibiotics and resistance genes in lettuce following cattle manure and digestate fertilization and their effects on soil and phyllosphere microbial communities.

The degradation and bioaccumulation of selected antibiotics such as the sulfonamide sulfamethoxazole (SMX) and the fluoroquinolones enrofloxacin (ENR) and ciprofloxacin (CIP) were investigated in soil microcosm experiments where Lactuca sativa was grown with manure or digestate (1%) and spiked with a mixture of the three antibiotics (7.5 mg/kg each). The soil, rhizosphere and leaf phyllosphere were sampled (at 0 and 46 days) from each microcosm to analyze the antibiotic concentrations, main resistance genes (sul1, sul2, qnrS, aac-(6')-Ib-crand qepA), the intI1and tnpA mobile genetic elements and the microbial community structure.Overall results showed that SMX and CIP decreased (70-85% and 55-79%, respectively), and ENR was quite persistent during the 46-day experiment. In plant presence, CIP and ENR were partially up-taken from soil to plant. In fact the bioaccumulation factors were > 1, with higher values in manure than digestate amended soils. The most abundant gene in soil was sul2 in digestate- and aac-(6')-Ib-cr in the manure-amended microcosms. In soil, neither sulfamethoxazole-resistance (sul1 and sul2), nor fluoroquinolone-resistance (aac-(6')-Ib-cr, qepA and qnrS) gene abundances were correlated with any antibiotic concentration. On the contrary, in lettuce leaves, the aac-(6')-Ib-cr gene was the most abundant, in accordance with the fluoroquinolone bioaccumulation. Finally, digestate stimulated a higher soil microbial biodiversity, introducing and promoting more bacterial genera associated with antibiotic degradation and involved in soil fertility and decreased fluoroquinolone bioaccumulation.

Effects of Ciprofloxacin Alone or in Mixture with Sulfamethoxazole on the Efficiency of Anaerobic Digestion and Its Microbial Community.

Some livestock farms rely on anaerobic digestion (AD) technology for manure disposal, thus obtaining energy (biogas) and fertilizer (digestate). Mixtures of antibiotics used for animal health often occur in organic waste and their possible synergistic/antagonistic effects on microorganisms involved in AD are still poorly studied. This work focuses on the effects of adding ciprofloxacin, alone (5 mg L-1) and in combination with sulfamethoxazole (2.5-5-10 mg L-1), on AD efficiency and microbial community structure. The experiment consisted of 90-day cattle manure batch tests and antibiotic removal percentages were assessed. Adding antibiotics always promoted CH4 and H2 production compared to untreated controls; however, CH4 production was lowered with the highest ciprofloxacin (CIP) concentrations. The overall results show antibiotic degradation caused by acidogenic Bacteria, and CH4 was mainly produced through the hydrogenotrophic-pathway by methanogenic Archaea. Shifts in microbial community abundance (DAPI counts) and composition (Illumina-MiSeq and FISH analyses) were observed.

Rhizosphere Microbial Communities and Heavy Metals.

The rhizosphere is a microhabitat where there is an intense chemical dialogue between plants and microorganisms. The two coexist and develop synergistic actions, which can promote plants' functions and productivity, but also their capacity to respond to stress conditions, including heavy metal (HM) contamination. If HMs are present in soils used for agriculture, there is a risk of metal uptake by edible plants with subsequent bioaccumulation in humans and animals and detrimental consequences for their health. Plant productivity can also be negatively affected. Many bacteria have defensive mechanisms for resisting heavy metals and, through various complex processes, can improve plant response to HM stress. Bacteria-plant synergic interactions in the rhizosphere, as a homeostatic ecosystem response to HM disturbance, are common in soil. However, this is hard to achieve in agroecosystems managed with traditional practices, because concentrating on maximizing crop yield does not make it possible to establish rhizosphere interactions. Improving knowledge of the complex interactions mediated by plant exudates and secondary metabolites can lead to nature-based solutions for plant health in HM contaminated soils. This paper reports the main ecotoxicological effects of HMs and the various compounds (including several secondary metabolites) produced by plant-microorganism holobionts for removing, immobilizing and containing toxic elements.

Germination, root elongation, and photosynthetic performance of plants exposed to sodium lauryl ether sulfate (SLES): an emerging contaminant.

The anionic surfactant SLES (sodium lauryl ether sulfate) is an emerging contaminant, being the main component of foaming agents that are increasingly used by the tunnel construction industry. To fill the gap of knowledge about the potential SLES toxicity on plants, acute and chronic effects were assessed under controlled conditions. The acute ecotoxicological test was performed on Lepidum sativum L. (cress) and Zea mays L. (maize). Germination of both species was not affected by SLES in soil, even at concentrations (1200 mg kg-1) more than twice higher than the maximum realistic values found in contaminated debris, thus confirming the low acute SLES toxicity on terrestrial plants. The root elongation of the more sensitive species (cress) was instead reduced at the highest SLES concentration. In the chronic phytotoxicity experiment, photosynthesis of maize was downregulated, and the photosynthetic performance (PITOT) significantly reduced already under realistic exposures (360 mg kg-1), owing to the SLES ability to interfere with water and/or nutrients uptake by roots. However, such reduction was transient, likely due to the rapid biodegradation of the surfactant by the soil microbial community. Indeed, SLES amount decreased in soil more than 90% of the initial concentration in only 11 days. A significant reduction of the maximum photosynthetic capacity (Pnmax) was still evident at the end of the experiment, suggesting the persistence of negative SLES effects on plant growth and productivity. Overall results, although confirming the low phytotoxicity and high biodegradability of SLES in natural soils, highlight the importance of considering both acute and nonlethal stress effects to evaluate the environmental compatibility of soil containing SLES residues.

Bioaugmentation With a Consortium of Bacterial Sodium Lauryl Ether Sulfate-Degraders for Remediation of Contaminated Soils.

The anionic surfactant sodium lauryl ether sulfate (SLES) is the main component of most commercial foaming agents (FAs) used in the excavation of highway and railway tunnels with Earth pressure balance-tunnel boring machines (EPB-TBMs). Several hundreds of millions of tons of spoil material, consisting of soil mixed with FAs, are produced worldwide, raising the issue of their handling and safe disposal. Reducing waste production and reusing by-products are the primary objectives of the "circular economy," and in this context, the biodegradation of SLES becomes a key question in reclaiming excavated soils, especially at construction sites where SLES degradation on the spot is not possible because of lack of space for temporary spoil material storage. The aim of the present work was to apply a bacterial consortium (BC) of SLES degraders to spoil material excavated with an EPB-TBM and coming from a real construction site. For this purpose, the BC capability to accelerate SLES degradation was tested. Preliminary BC growth, degradation tests, and ecotoxicological evaluations were performed on a selected FA. Subsequently, a bioaugmentation experiment was conducted; and the microbial abundance, viability, and SLES concentrations in spoil material were evaluated over the experimental time (0.5, 3, 6, 24, 48, and 144 h). Moreover, the corresponding aqueous elutriates were extracted from all the soil samples and analyzed for SLES concentration and ecotoxicological evaluations with the bacterium Aliivibrio fischeri. The preliminary experiments showed the BC capability to grow under 14 different concentrations of the FA. The maximum BC growth rates and degradation efficiency (100%) were achieved with initial SLES concentrations of 125, 250, and 500 mg/L. The subsequent bioaugmentation of the spoil material with BC significantly (sixfold) improved the degradation time of SLES (DT50 1 day) compared with natural attenuation (DT50 6 days). In line with this result, neither SLES residues nor toxicity was recorded in the soil extracts showing the spoil material as a by-product promptly usable. The bioaugmentation with BC can be a very useful for cleaning spoil material produced in underground construction where its temporary storage (for SLES natural biodegradation) is not possible.

Anaerobic Digestion and Removal of Sulfamethoxazole, Enrofloxacin, Ciprofloxacin and Their Antibiotic Resistance Genes in a Full-Scale Biogas Plant.

Anaerobic digestion is one of the best ways to re-use animal manure and agricultural residues, through the production of combustible biogas and digestate. However, the use of antibiotics for preventing and treating animal diseases and, consequently, their residual concentrations in manure, could introduce them into anaerobic digesters. If the digestate is applied as a soil fertilizer, antibiotic residues and/or their corresponding antibiotic resistance genes (ARGs) could reach soil ecosystems. This work investigated three common soil emerging contaminants, i.e., sulfamethoxazole (SMX), ciprofloxacin (CIP), enrofloxacin (ENR), their ARGs sul1, sul2, qnrS, qepA, aac-(6')-Ib-cr and the mobile genetic element intI1, for one year in a full scale anaerobic plant. Six samplings were performed in line with the 45-day hydraulic retention time (HRT) of the anaerobic plant, by collecting input and output samples. The overall results show both antibiotics and ARGs decreased during the anaerobic digestion process. In particular, SMX was degraded by up to 100%, ENR up to 84% and CIP up to 92%, depending on the sampling time. In a similar way, all ARGs declined significantly (up to 80%) in the digestate samples. This work shows how anaerobic digestion can be a promising practice for lowering antibiotic residues and ARGs in soil.

Isolation and Characterization in a Soil Conditioned With Foaming Agents of a Bacterial Consortium Able to Degrade Sodium Lauryl Ether Sulfate.

The anionic surfactant Sodium Lauryl Ether Sulfate (SLES) is the principal component of several commercial foaming products for soil conditioning in the tunneling industry. Huge amounts of spoil material are produced during the excavation process and the presence of SLES can affect its re-use as a by-product. Anionic surfactants can be a risk for ecosystems if occurring in the environment at toxic concentrations. SLES biodegradability is a key issue if the excavated soil is to be reused. The aim of this study was to identify bacteria able to degrade SLES, so that it could potentially be used in bioaugmentation techniques. Enrichment cultures were performed using bacterial populations from spoil material collected in a tunnel construction site as the inoculum. A bacterial consortium able to grow in a few hours with SLES concentrations from 125 mg/L to 2 g/L was selected and then identified by Next Generation Sequencing analysis. Most of bacteria identified belonged to Gamma-Proteobacteria (99%) and Pseudomonas (ca 90%) was the predominant genus. The bacterial consortium was able to degrade 94% of an initial SLES concentration of 250 mg/L in 9 h. A predictive functional analysis using the PICRUSt2 software showed the presence of esterase enzymes, responsible for SLES degradation. The bacterial consortium selected could be useful for its possible seeding (bioaugmentation) on spoil material from tunneling excavation.

Co-presence of the anionic surfactant sodium lauryl ether sulphate and the pesticide chlorpyrifos and effects on a natural soil microbial community.

There is a growing concern about the simultaneous presence in the environment of different kinds of pollutants, because of the possible synergic or additive effects of chemical mixtures on ecosystems. Chlorpyrifos (CPF) is an organophosphate insecticide extensively used in agricultural practices. The anionic surfactant sodium lauryl ether sulphate (SLES) is the main component of several commercial products, including foaming agents used in underground mechanised excavation. Both compounds are produced and sold in high amounts worldwide and can be found in the environment as soil contaminants. The persistence of SLES and CPF in agricultural soils and their possible effects on the natural microbial community was evaluated in microcosms. The experimental set consisted of soil samples containing the autochthonous microbial community and treated with only SLES (70 mg/kg), only CPF (2 mg/kg) or with a mix of both compounds. Control microcosms (without the contaminants) were also performed. Soil samples were collected over the experimental period (0, 7, 14, 21 and 28 days) and analysed for CPF, SLES and the main metabolite of CPF (3, 5, 6-trichloropyridinol, TCP). The half-life time (DT50) of each parent compound was estimated in all experimental conditions. At the same time, the abundance, activity and structure of the microbial community were also evaluated. The results showed that the co-presence of SLES and CPF did not substantially affect their persistence in soil (DT50 of 11 and 9 days with co-presence and 13 and 10 days, respectively, when alone); however, in the presence of SLES, a higher amount of the metabolite TCP was found. Interestingly, some differences were found in the bacterial community structure, abundance and activity among the various conditions.

Impact of bacterial motility on biosorption and cometabolism of pyrene in a porous medium.

The risks of pollution by polycyclic aromatic hydrocarbons (PAHs) may increase in bioremediated soils as a result of the formation of toxic byproducts and the mobilization of pollutants associated to suspended colloids. In this study, we used the motile and chemotactic bacterium Pseudomonas putida G7 as an experimental model for examining the potential role of bacterial motility in the cometabolism and biosorption of pyrene in a porous medium. For this purpose, we conducted batch and column transport experiments with 14C-labelled pyrene loaded on silicone O-rings, which acted as a passive dosing system. In the batch experiments, we observed concentrations of the 14C-pyrene equivalents well above the equilibrium concentration observed in abiotic controls. This mobilization was attributed to biosorption and cometabolism processes occurring in parallel. HPLC quantification revealed pyrene concentrations well below the 14C-based quantifications by liquid scintillation, indicating pyrene transformation into water-soluble polar metabolites. The results from transport experiments in sand columns revealed that cometabolic-active, motile cells were capable of accessing a distant source of sorbed pyrene. Using the same experimental system, we also determined that salicylate-mobilized cells, inhibited for pyrene cometabolism, but mobilized due to their tactic behavior, were able to sorb the compound and mobilize it by biosorption. Our results indicate that motile bacteria active in bioremediation may contribute, through cometabolism and biosorption, to the risk associated to pollutant mobilization in soils. This research could be the starting point for the development of more efficient, low-risk bioremediation strategies of poorly bioavailable contaminants in soils.

Effects of Soil Fertilization on Terpenoids and Other Carbon-Based Secondary Metabolites in Rosmarinus officinalis Plants: A Comparative Study.

Rosmarinus officinalis is an evergreen aromatic plant with important commercial interest as it contains numerous essential oils (composed of terpenoid compounds) and phenolic constituents (natural antioxidant compounds). This work aims at evaluating the concomitant effects of different inorganic and organic fertilization treatments and the subsequent increases in soil nutrient availability on terpenoids and other carbon-based secondary metabolites, e.g., flavonoids and phenolic compounds, in Rosmarinus officinalis leaves. The results showed that, as expected, the structural carbohydrate content (lignocellulosic compounds) in stems was higher in fertilized plants than in controls. Additionally, positive correlations were observed of the absolute amounts of total terpenoids and some single terpenoid compounds with N or P contents in leaves. On the contrary, the phenolic and flavonoid concentrations in all the rosemary plant parts were lower with the fertilization treatments. Indeed, negative correlations between the phenolic compounds (and flavonoids) and N in rosemary leaves were also found. Overall, the results suggest that the terpenoid production's response to fertilization was due to N, which is essential for protein synthesis and terpene synthase activity, and to P, which is necessary for the synthesis of both terpenoid precursors and ATP and NADPH, also needed for terpenoid synthesis. On the other hand, the basis for the fertilization's effects on the production of phenolic compounds is the direct nitrogen trade-off between growth and the shikimic acid pathway by which phenolics compounds are synthesized.

Characterization of the Belowground Microbial Community in a Poplar-Phytoremediation Strategy of a Multi-Contaminated Soil.

Due to their widespread use in industrial applications in recent decades, Polychlorobiphenyls (PCBs) and heavy metals (HMs) are the most common soil contaminants worldwide, posing a risk for both ecosystems and human health. In this study, a poplar-assisted bioremediation strategy has been applied for more than 4 years to a historically contaminated area (PCBs and HMs) in Southern Italy using the Monviso poplar clone. This clone was effective in promoting a decrease in all contaminants and an increase in soil quality in terms of organic carbon and microbial abundance. Moreover, a significant shift in the structure and predicted function of the belowground microbial community was also observed when analyzing both DNA and cDNA sequencing data. In fact, an increase in bacterial genera belonging to Proteobacteria able to degrade PCBs and resist HMs was observed. Moreover, the functional profiling of the microbial community predicted by PICRUSt2 made it possible to identify several genes associated with PCB transformation (e.g., bphAa, bphAb, bphB, bphC), response to HM oxidative stress (e.g., catalase, superoxide reductase, peroxidase) and HM uptake and expulsion (e.g., ABC transporters). This work demonstrated the effectiveness of the poplar clone Monviso in stimulating the natural belowground microbial community to remove contaminants and improve the overall soil quality. It is a practical example of a nature based solution involving synergic interactions between plants and the belowground microbial community.