Role of endogenous soil microorganisms in controlling antimicrobial resistance after the exposure to treated wastewater.

The reuse of treated wastewater (TWW) for irrigation appears to be a relevant solution to the challenges of growing water demand and scarcity. However, TWW contains not only micro-pollutants including pharmaceutical residues but also antibiotic resistant bacteria. The reuse of TWW could contribute to the dissemination of antimicrobial resistance in the environment. The purpose of this study was to assess if exogenous bacteria from irrigation waters (TWW or tap water-TP) affect endogenous soil microbial communities (from 2 soils with distinct irrigation history) and key antibiotic resistance gene sul1 and mobile genetic elements intl1 and IS613. Experiments were conducted in microcosms, irrigated in one-shot, and monitored for three months. Results showed that TP or TWW exposure induced a dynamic response of soil microbial communities but with no significant increase of resistance and mobile gene abundances. However, no significant differences were observed between the two water types in the current experimental design. Despite this, the 16S rDNA analysis of the two soils irrigated for two years either with tap water or TWW resulted in soil microbial community differentiation and the identification of biomarkers from Xanthomonadaceae and Planctomycetes families for soils irrigated with TWW. Low-diversity soils were more sensitive to the addition of TWW. Indeed, TWW exposure stimulated the growth of bacterial genera known to be pathogenic, correlating with a sharp increase in the copy number of selected resistance genes (up to 3 logs). These low-diversity soils could thus enable the establishment of exogenous bacteria from TWW which was not observed with native soils. In particular, the emergence of Planctomyces, previously suggested as a biomarker of soil irrigated by TWW, was here demonstrated. Finally, this study showed that water input frequency, initial soil microbial diversity and soil history drive changes within soil endogenous communities and the antibiotic resistance gene pool.

Evaluation of different QuEChERS-based methods for the extraction of 48 wastewater-derived organic contaminants from soil and lettuce root using high-resolution LC-QTOF with MRMHR and SWATH acquisition modes.

The reuse of treated wastewater in agriculture is an important route of introducing a large number of organic contaminants into the agroecosystem. In this study, a modified QuEChERS-based approach was developed for rapid, simple, and simultaneous extraction of 48 organic wastewater-derived contaminants from soil and lettuce root. Twenty-two different (modification) scenarios of the known (or original) QuEChERS method have been tested, in order to obtain best and well-compromised recoveries for all target compounds for soil and roots. Finally, a common method was chosen for both matrices consisting of a single extraction step using EDTA-Mcllvaine buffer and the unbuffered Original QuEChERS salts. Method performance was accomplished by liquid chromatography coupled with high-resolution mass spectrometry on a QToF-MS system using two different acquisition modes, the ultra-fast high-resolution multiple reaction monitoring (MRMHR) mode and the innovative Sequential Window Acquisition of All Theoretical Fragment-Ion (SWATH) mode. Performance characterization was evaluated in terms of recovery, linearity, intra-day precision, method detection limits (MDLs), method quantification limits (MQLs), and matrix effect (ME). Recoveries in MRMHR mode ranged from 63 to 111% and 54 to 104% for lettuce root and soil, respectively, for most of compounds in MRMHR mode and from 56 to 121% and 54 to 104% for lettuce root and soil, respectively, for most of compounds in SWATH. Whereas, MQLs ranged from 0.03 to 0.92 ng g-1 in MRMHR and from 0.03 to 82 ng g-1 in SWATH for lettuce root, and from 0.02 to 0.44 ng g-1 in MRMHR and 0.02 to 0.14 ng g-1 in SWATH for soil. The method was then applied to follow the target compounds in soil and lettuce root, where the system lettuce-soil was irrigated with treated wastewater under real greenhouse conditions. Five and 17 compounds were detected in lettuce root and soil, respectively.

Relevance of photocatalytic redox transformations of selected pharmaceuticals in a copper- and iron-rich Mediterranean intermittent river.

This work aimed at investigating specific attenuation pathways of pharmaceuticals in copper- and iron-rich Mediterranean intermittent and sunlit rivers by combining lab- and field-scale studies. Poorly photodegradable and biodegradable compounds such as fluconazole, oxazepam and venlafaxine attenuated in two river stretches with short hydraulic residence times (<3 h). This result was assumed to be related to their capacity to interact with photoreactive free Cu2+ and Fe3+ or their associated oxides. Lab-scale photodegradation experiments under simulated solar irradiation revealed the beneficial impact of a mixture Cu2+ and colloidal iron hydroxides at environmental concentrations and at neutral pH on the pharmaceuticals photodegradation kinetic rate constants. These latter were consistent with the in-stream attenuation rate constants of targeted contaminants which ranged from 0.104 to 0.154 h-1. Further identification of phototransformation products by LC-HRMS highlighted reductive transformation pathways including reductive dehalogenation and hydrogenation reactions. Several TPs were found to be stable under irradiation and were detected in field monitoring, accordingly. This was ascribed to the formation of a Cu/Fe composite material under solar irradiation with photocatalytic properties. The role of Cu was to trap the electron in the conduction band of the iron-based photocatalyst, which promoted separation efficiency of electron-hole pairs as well as enhanced photoreduction processes at the expense of oxidation ones. Even though, these mechanisms have been reported in water treatment field for organic micropollutants removal, their significance was demonstrated for the first time in natural settings.

Investigating the Transformation Products of Selected Antibiotics and 17 α-Ethinylestradiol under Three In Vitro Biotransformation Models for Anticipating Their Relevance in Bioaugmented Constructed Wetlands.

The degradation of three antibiotics (sulfamethoxazole, trimethoprim, and ofloxacin) and one synthetic hormone (17 α-ethinylestradiol) was investigated in three in-vitro biotransformation models (i.e., pure enzymes, hairy root, and Trichoderma asperellum cultures) for anticipating the relevance of the formation of transformation products (TPs) in constructed wetlands (CWs) bioaugmented with T. asperellum fungus. The identification of TPs was carried out employing high-resolution mass spectrometry, using databases, or by interpreting MS/MS spectra. An enzymatic reaction with ß-glucosidase was also used to confirm the presence of glycosyl-conjugates. The results showed synergies in the transformation mechanisms between these three models. Phase II conjugation reactions and overall glycosylation reactions predominated in hairy root cultures, while phase I metabolization reactions (e.g., hydroxylation and N-dealkylation) predominated in T. asperellum cultures. Following their accumulation/degradation kinetic profiles helped in determining the most relevant TPs. Identified TPs contributed to the overall residual antimicrobial activity because phase I metabolites can be more reactive and glucose-conjugated TPs can be transformed back into parent compounds. Similar to other biological treatments, the formation of TPs in CWs is of concern and deserves to be investigated with simple in vitro models to avoid the complexity of field-scale studies. This paper brings new findings on the emerging pollutants metabolic pathways established between T. asperellum and model plants, including extracellular enzymes.

Use of Passive and Grab Sampling and High-Resolution Mass Spectrometry for Non-Targeted Analysis of Emerging Contaminants and Their Semi-Quantification in Water.

Different groups of organic micropollutants including pharmaceuticals and pesticides have emerged in the environment in the last years, resulting in a rise in environmental and human health risks. In order to face up and evaluate these risks, there is an increasing need to assess their occurrence in the environment. Therefore, many studies in the past couple of decades were focused on the improvements in organic micropollutants' extraction efficiency from the different environmental matrices, as well as their mass spectrometry detection parameters and acquisition modes. This paper presents different sampling methodologies and high-resolution mass spectrometry-based non-target screening workflows for the identification of pharmaceuticals, pesticides, and their transformation products in different kinds of water (domestic wastewater and river water). Identification confidence was increased including retention time prediction in the workflow. The applied methodology, using a passive sampling technique, allowed for the identification of 85 and 47 contaminants in the wastewater effluent and river water, respectively. Finally, contaminants' prioritization was performed through semi-quantification in grab samples as a fundamental step for monitoring schemes.

Peroxydisulfate activation by CuO pellets in a fixed-bed column, operating mode and assessments for antibiotics degradation and urban wastewater disinfection.

A fixed-bed column packed with copper oxide pellets (FBC-CuO) combined with peroxydisulfate (PDS) as a primary oxidant was assessed as an option for simultaneously wastewater decontamination (antibiotics) and disinfection (bacteria, viruses, and protozoa). Preliminary to these experiments, phenol was used as the target molecule to investigate the working mode of FBC-CuO under various operating conditions, such as varying flow rates, initial persulfate, and phenol concentrations. Then, the removal of a mix of five representative antibiotics (amoxicillin (AMX), cefalexin (CFX), ofloxacin (OFL), sulfamethoxazole (SMX), and clarithromycin (CLA)) in secondary treated urban wastewater (STWW) was evaluated. AMX, CFX, and OFL were effectively removed by simply flowing through the FBC-CuO, and the addition of PDS (500 µM) systematically enhanced the degradation of all targeted antibiotics, which is also the necessary condition for the removal of SMX and CLA. Urban wastewater disinfection was evaluated by monitoring targeted pathogens originally in the STWW. A significant reduction of Escherichia coli, Enterococcus, F-specific RNA bacteriophages was observed after the treatment by FBC-CuO with 500 µM PDS. X-ray diffraction measurement and scanning electron microscopy performed on CuO pellets before and after treatment confirmed that the structure of the catalyst was preserved without any phase segregation. Finally, quantification of Cu(II) at the outlet of FBC-CuO indicate a non-negligible but limited released. All these results underline the potential of the FBC-CuO combined with PDS at the field scale for the degradation of micropollutants and inactivation of pathogens in wastewater.

Comparison of sunlight-AOPs for levofloxacin removal: kinetics, transformation products, and toxicity assay on Escherichia coli and Micrococcus flavus.

Levofloxacin (LFX) is a widely used antibiotic medication. Persistent traces of LFX in water and wastewater may induce bacterial resistance. Photon-driven advanced oxidation processes (AOPs) can assist in attaining complete abatement of LFX for environmental protection. This work benchmarks different solar AOPs based on hydroxyl radical (OH•) and sulphate radical (SO4•-) chemistry. Other oxidant precursors, as radical sources, were used to selectively control the generation of either hydroxyl radical (i.e., H2O2), sulphate radical (i.e., peroxydisulphate (PDS)), or a controlled mixture ratio of both OH•/SO4•- (i.e., peroxymonosulphate (PMS)). The influence of pH on degradation performance was evaluated using unbuffered and buffered solutions. Simulated irradiation/PMS process exhibited a strong pH-dependence attaining partial degradation of ca. 56% at pH 5 up to complete degradation at pH 7. Despite the similitudes on the abatement of target pollutant LFX in pristine solutions, only simulated irradiation/PDS treatment achieved effective abatement of LFX in wastewater samples given the higher selectivity of SO4•-. Toxicity tests were conducted with Escherichia coli (LMG2092) and Micrococcus flavus (DSM1790), demonstrating successful inhibition of the antibiotic character of polluted waters, which would contribute to preventing the development of resistant bacterial strains. Finally, a degradative pathway was suggested from the by-products and intermediates identified by LC-MS. Results demonstrate that the degradation of specific functional groups (i.e., piperazine ring) is associated with the loss of antibacterial character of the molecule.

Impact of repeated irrigation of lettuce cultures with municipal wastewater on soil bacterial community diversity and composition.

The effect of wastewater irrigation on the diversity and composition of bacterial communities of soil mesocosms planted with lettuces was studied over an experiment made of five cultivation campaigns. A limited effect of irrigation with either raw or treated wastewater was observed in both α-diversity and ß-diversity of soil bacterial communities. However, the irrigation with wastewater fortified with a complex mixture of fourteen relevant chemicals at 10 µg/L each, including pharmaceutical, biocide, and pesticide active substances, led to a drift in the composition of soil bacterial community. One hundred operational taxonomic units (OTUs) were identified as responsible for changes between treated and fortified wastewater irrigation treatments. Our findings indicate that under a realistic agronomical scenario, the irrigation of vegetables with domestic (treated or raw) wastewater has no effect on soil bacterial communities. Nevertheless, under the worst-case scenario tested here (i.e., wastewater fortified with a mixture of chemicals), non-resilient changes were observed suggesting that continuous/repeated irrigation with wastewater could lead to the accumulation of contaminants in soil and induce changes in bacterial communities with unknown functional consequences.

Copper oxide/peroxydisulfate system for urban wastewater disinfection: Performances, reactive species, and antibiotic resistance genes removal.

In this study, copper oxide (CuO) catalyzed peroxydisulfate (PDS) system was investigated for the inactivation of a broad range of pathogenic microorganisms from urban wastewater. Complete inactivation of Escherichia coli, Enterococcus, F-specific RNA bacteriophages from secondary treated wastewater was achieved after a short time (15-30 min) treatment with CuO (10 g/L)/PDS (1 mM) system, but spores of sulfite-reducing bacteria took 120 min. No bacterial regrowth occurred during storage after treatment. Significant reduction of the pathogens was explained by the generation of the highly selective Cu(III) oxidant, as the predominant reactive species, which could quickly oxidize guanine through a one-electron oxidation pathway. Additionally, the potential of the CuO (10 g/L)/PDS (1 mM) system to inactivate antibiotic-resistant bacteria and antibiotic resistance genes (ARB&Gs) was explored. Sulfamethoxazole-resistant E. coli was used as the model ARB and a 3.2 log of reduction was observed after 10 min of treatment. A considerable reduction (0.7-2.3 log) of selected ARGs including blaTEM, qnrS, emrB, sul1, and genes related to the dissemination of antibiotic resistance, including the Class 1 integron-integrase (intI1), and the insertion sequence (IS613) was achieved after 60 min treatment. All these findings indicated the promising applicability of the CuO/PDS system as a disinfection technology for wastewater reuse in agriculture.

Impact of long-term irrigation with municipal reclaimed wastewater on the uptake and degradation of organic contaminants in lettuce and leek.

A two years drip irrigation of lettuce and leek crops with treated municipal wastewater without and with spiking with fourteen wastewater relevant contaminants at 10 µg/L concentration level was conducted under greenhouse cultivation conditions to investigate their potential accumulation in soil and leaves and to assess human health related risks. Lettuce and leek crops were selected as a worse-case scenario since leafy green vegetable has a high potential for organic contaminants uptake. The results revealed limited accumulation of contaminants in soil and plant leaves, their concentration levels being in the range of 1-30 ng/g and 1-660 ng/g range in soil and leaves, respectively. This was likely related to abiotic and biotic transformation or simply binding processes in soil, which limited contaminants plant uptake. This assumption was underpinned by studies of the enantiomeric fractionation of chiral compounds (e.g. climbazole and metoprolol) in soil as pieces of evidence of biodegradation and by the identification of transformation products or metabolites in leaves by means of liquid chromatography - high resolution - mass spectrometry using a suspect screening workflow. The high bioconcentration factors were not limited to compounds with intermediate Dow (100 to 1000) such as carbamazepine but also observed for hydrophilic compounds such as clarithromycin, hydrochlorothiazide and the food additives acesulfame and sucralose. This result assumed that accumulation was not only driven by passive processes (e.g. lipoidal diffusion through lipid bilayer cell membranes or Casparian strip) but might be supported by carrier-mediated transporters. As a whole, this study confirmed earlier reports on the a de minimis human health risk related to the consumption of raw leafy green vegetable irrigated with domestic TWW containing organic contaminants residues.

Nitric oxide reactivity accounts for N-nitroso-ciprofloxacin formation under nitrate-reducing conditions.

The formation of N-nitroso-ciprofloxacin (CIP) was investigated both in wastewater treatment plants including nitrification/denitrification stages and in sludge slurry experiments under denitrifying conditions. The analysis of biological wastewater treatment plant effluents by Kendrick mass defect analysis and liquid chromatography - high resolution - mass spectrometry (LCHRMS) revealed the occurrence of N-nitroso-CIP and N-nitroso-hydrochlorothiazide at concentration levels of 34 ± 3 ng/L and 71 ± 6 ng/L, respectively. In laboratory experiments and dark conditions, produced N-nitroso-CIP concentrations reached a plateau during the course of biodegradation experiments. A mass balance was achieved after identification and quantification of several transformation products by LCHRMS. N-nitroso-CIP accounted for 14.3% of the initial CIP concentration (20 µg/L) and accumulated against time. The use of 4,5-diaminofluorescein diacetate and superoxide dismutase as scavengers for in situ production of nitric oxide and superoxide radical anion respectively, revealed that the mechanisms of formation of N-nitroso-CIP likely involved a nitrosation pathway through the formation of peroxynitrite and another one through codenitrification processes, even though the former one appeared to be prevalent. This work extended the possible sources of N-nitrosamines by including a formation pathway relying on nitric oxide reactivity with secondary amines under activated sludge treatment.

Biodegradation of fluoroquinolone antibiotics and the climbazole fungicide by Trichoderma species.

Filamentous fungi Trichoderma have been able to efficiently degrade fluoroquinolone antibiotics namely ciprofloxacin (CIP) and ofloxacin (OFL) as well as the fungicide climbazole (CLB) that are persistent in conventional activated sludge processes. All targeted compounds were biotransformed by whole cells of Trichoderma spp., exactly T. harzanium and T. asperellum, and biosorption played a limited role in their elimination. However, contrasting results were obtained with the two strains. T. asperellum was more efficient against CIP, with a 81% degradation rate in 13 days of incubation, while T. harzianum was more efficient against CLB, with a 91% degradation rate. While in the case of OFL, both strains showed same efficiency with degradation rate around 40%. Adding a cytochrome P450 enzyme inhibitor hardly resulted in the modification of degradation kinetics supporting the implication of extracellular enzymes in chemical biotransformation. Transformation products were identified by liquid chromatography-high resolution-mass spectrometry and transformation pathways were proposed. Biotransformation of selected compounds included hydroxylation, oxidation/reduction and N- and O-dealkylation reactions, similarly to those reported with white rot fungi. CIP underwent transformations at the piperazinyl ring through oxidation and conjugation reactions, while OFL mainly underwent hydroxylation processes and CLB carbonyl reduction into alcohol. Consequently, Trichoderma spp. likely possess a machinery of unspecific enzymes, which makes their application in removal of pharmaceutical and personal care products attractive.

Relevance of N-nitrosation reactions for secondary amines in nitrate-rich wastewater under UV-C treatment.

This study investigated the transformation of secondary amine pharmaceuticals in UV-C/NO3- and in nitrate-rich wastewater at 254 nm by taking diclofenac, diphenylamine, mefenamic acid and furosemide as probe compounds. The degradation of targeted compounds were positively related to nitrate concentration and mainly caused by the formation of peroxynitrite and related reactive nitrogen species (e.g., nitrogen oxide and nitrogen dioxide radicals). Major transformation products were identified to provide fundamental understanding of the selective oxidation of secondary amine with reactive nitrogen species. UV photolysis, hydroxyl radical oxidation, nitration and nitrosation processes were found to be the most significant transformation pathways. In case of diphenylamine, for which most of the identified intermediates were available as standard, the relative significance of each transformation route could be established, highlighting for the first time the important role of N-nitrosation processes in UV/NO3- treatment followed by the decomposition of the resulting N-nitroso compounds by an alpha hydroxylation mechanism. This specific transformation pathway was of concern because it constitutes the molecular basis of N-nitrosamine carcinogenicity and may contribute to the increase in effluent genotoxicity under UV-C treatment in addition to the formation of nitrophenols. Hydrogenocarbonate ions at concentration values higher than 300 mg/L appeared to be a protective specie against nitrosation processes due to the formation of carbamate adducts but H2O2 in UV-C/H2O2 could be responsible for an exacerbation of the N-nitrosation pathway due to an addition source of hydroxyl radical in the system. The occurrence of major transformation products of diclofenac was confirmed in nitrate-rich wastewater under UV-C treatment at pilot-scale operation.

Analysis and fate of 14 relevant wastewater-derived organic pollutants in long-term exposed soil.

The growing use of reclaimed water in agriculture worldwide calls for developing high-sensitivity methods to quantify wastewater-derived organic contaminants in soils so that the potential risk of this irrigation practice can be properly assessed. This work describes an analytical method for the determination of trace levels of 14 drugs that are known to be poorly removed during conventional wastewater treatment in soil. The analytes selected for investigation included ten pharmaceuticals from different therapeutic classes (carbamazepine, diclofenac, cis-diltiazem, lamotrigine, methadone, midazolam, oxcarbazepine, sulfamethoxazole, trimethoprim, valsartan), one illicit drug (cocaine), and three transformation products/metabolites (acridone, 4'-hydroxydiclofenac, and valsartan acid), thereby covering a broad range of physical-chemical properties. The methodology developed was based on ultrasonic solvent extraction (USE) of the analytes from the soil matrix, and subsequent clean-up and analysis of the USE extracts with a fully automated approach by means of solid-phase extraction and liquid chromatography-tandem mass spectrometry detection (online SPE-LC-MS/MS). The method was fully validated with affording method detection and quantification limits ranging from 0.03 to 1 ng g-1 and from 0.09 to 3.3 ng g-1, respectively. This method was applied to investigate the fate of the selected drugs in potting soil irrigated for a long term (60 days) either with water containing the target compounds at a concentration of 200 µg L-1 or with wastewater treatment plant effluent and thus, at real environmental concentrations. All investigated compounds were found to accumulate in soil irrigated with artificially fortified water. The highest accumulation potential was observed for cis-diltiazem followed by methadone and midazolam that presented average concentrations of 1517 ng g-1, 1041 ng g-1, and 962 ng g-1 d.w., respectively. On the contrary, oxcarbazepine (5.8 ng g-1) and sulfamethoxazole (22 ng g-1) were the target drugs presenting the lowest accumulation potential. Only trace levels of ten drugs were measured in soil irrigated with regenerated water (average concentrations between 1.6 and 4.7 ng g-1 d.w.). Graphical abstract.

Combining micelle-clay sorption to solar photo-Fenton processes for domestic wastewater treatment.

A tertiary treatment of effluent from a biological domestic wastewater treatment plant was tested by combining filtration and solar photocatalysis. Adsorption was carried out by a sequence of two column filters, the first one filled with granular activated carbon (GAC) and the second one with granulated nano-composite of micelle-montmorillonite mixed with sand (20:100, w/w). The applied solar advanced oxidation process was homogeneous photo-Fenton photocatalysis using peroxymonosulfate (PMS) as oxidant agent. This combination of simple, robust, and low-cost technologies aimed to ensure water disinfection and emerging contaminants (ECs, mainly pharmaceuticals) removal. The filtration step showed good performances in removing dissolved organic matter and practically removing all bacteria such as Escherichia coli and Enterococcus faecalis from the secondary treated water. Solar advanced oxidation processes were efficient in elimination of trace levels of ECs. The final effluent presented an improved sanitary level with acceptable chemical and biological characteristics for irrigation.

Oxidative degradation of pentachlorophenol by permanganate for ISCO application.

Potassium permanganate (KMnO4) has been an effective technology for the in situ chemical oxidation (ISCO) of many organic compounds including chlorinated alkanes and alkenes, but it has rarely been applied for oxidizing aromatic organochlorines. This study confirms the ability of permanganate to oxidize an aromatic chlorinated compound, pentachlorophenol (PCP), in an efficient manner at neutral pH. The rate of the reaction between KMnO4 and PCP was calculated and the results indicated that the reaction between PCP and permanganate is relatively fast with a second-order rate constant k″ ∼ 30 M-1 s-1. Besides the kinetic aspect, the authors identified the main reaction by-products, and proposed a possible reaction mechanism scheme. The general pathway shows the formation of chlorinated intermediates, and ultimately, the complete mineralization to chloride, water, and CO2 confirmed by total organic carbon and chloride measurement in solution. Flow-through column experiments, consisting of flushing a PCP-contaminated sandy or natural soil with oxidant, showed the good ability of permanganate to eliminate the pollutant. After 24 h of treatment, 77% and 56% of PCP abatement were obtained for sandy and natural soil, respectively. These findings show the high potential of permanganate for the in situ remediation of aromatic organochlorine contaminated soils.

Levels and sources of heavy metals and PAHs in sediment of Djibouti-city (Republic of Djibouti).

Selected heavy metals and polycyclic aromatic hydrocarbons (PAHs) were determined in marine sediment from 28 sites Djibouti city. The concentrations of trace elements varied from 0 to 288.1mg/kg with relative abundance of trace metals in sediments was in the order of Zn>Cu>Ni>Cr>Co>Pb>Cd. Zn, Cu and Ni exceeded consensus based sediment quality guideline values 7, 14, 15 sites respectively. Enrichment factor and pollution load index showed relatively low to moderate contamination. The concentrations of total 16 PAHs varied widely depending on the sample location and ranged from 2.65 to 3760.11ng·g-1, with the mean concentration value of 387.87ng·g-1. Compositions and relative abundance of individual PAH using molecular diagnostic ratio using congener's m/z 178 and 202 indicated pyrolytic origin and reflecting a petroleum combustion, grass/wood and coal combustion and a petrogenic source. This study represents the first pollution baseline and a reference for future studies in Djibouti.

4-nitroso-sulfamethoxazole generation in soil under denitrifying conditions: Field observations versus laboratory results.

The formation of 4-nitroso-sulfamethoxazole and 4-nitro-SMX, two transformation products (TPs) of sulfamethoxazole (SMX) was investigated under batch soil slurry experiments and in a field study. Due to their low occurrence levels (ng/L) in environmental waters, a suitable analytical method based on liquid chromatography - high resolution - mass spectrometry was developed. Consequently, field observations revealed, for the first time, the occurrence of 4-nitroso-SMX in groundwater at concentrations as high as 18ng/L.Nitric oxide (NO) steady-state concentrations were determined in soil slurry experiments because this reactive specie accounted for the formation of 4-nitroso-SMX and 4-nitro-SMX. Measurements revealed that environmental SMX concentrations (0.2-2µg/L) at neutral pH induced the accumulation of nitric oxide. Under acidic conditions (pH<6), nitrous acid (HONO) was the major source of nitric oxide while under neutral/basic conditions nitric oxide release was related to the inhibition of denitrification processes. Under laboratory experiments, SMX nitration reaction appeared to be an irreversible transformation pathway, while 4-nitroso-SMX was slowly transformed over time. The occurrence of 4-nitroso-SMX conditions was therefore unexpected in the field study but could be due to its continuous input from soil and/or its relative persistence under anoxic conditions. A mechanism for 4-nitroso-SMX formation was proposed involving a nitrosative desamination pathway through a phenyl radical.

Enantioselective reductive transformation of climbazole: A concept towards quantitative biodegradation assessment in anaerobic biological treatment processes.

An efficient chiral method-based using liquid chromatography-high resolution-mass spectrometry analytical method has been validated for the determination of climbazole (CBZ) enantiomers in wastewater and sludge with quantification limits below the 1 ng/L and 2 ng/g range, respectively. On the basis of this newly developed analytical method, the stereochemistry of CBZ was investigated over time in sludge biotic and sterile batch experiments under anoxic dark and light conditions and during wastewater biological treatment by subsurface flow constructed wetlands. CBZ stereoselective degradation was exclusively observed under biotic conditions, confirming the specificity of enantiomeric fraction variations to biodegradation processes. Abiotic CBZ enantiomerization was insignificant at circumneutral pH and CBZ was always biotransformed into CBZ-alcohol due to the specific and enantioselective reduction of the ketone function of CBZ into a secondary alcohol function. This transformation was almost quantitative and biodegradation gave good first order kinetic fit for both enantiomers. The possibility to apply the Rayleigh equation to enantioselective CBZ biodegradation processes was investigated. The results of enantiomeric enrichment allowed for a quantitative assessment of in situ biodegradation processes due to a good fit (R2 > 0.96) of the anoxic/anaerobic CBZ biodegradation to the Rayleigh dependency in all the biotic microcosms and was also applied in subsurface flow constructed wetlands. This work extended the concept of applying the Rayleigh equation towards quantitative biodegradation assessment of organic contaminants to enantioselective processes operating under anoxic/anaerobic conditions.

Biotic nitrosation of diclofenac in a soil aquifer system (Katari watershed, Bolivia).

Up till now, the diclofenac (DCF) transformation into its nitrogen-derivatives, N-nitroso-DCF (NO-DCF) and 5-nitro-DCF (NO2-DCF), has been mainly investigated in wastewater treatment plant under nitrification or denitrification processes. This work reports, for the first time, an additional DCF microbial mediated nitrosation pathway of DCF in soil under strictly anoxic conditions probably involving codenitrification processes and fungal activities. This transformation pathway was investigated by using field observations data at a soil aquifer system (Katari watershed, Bolivia) and by carrying out soil slurry batch experiments. It was also observed for diphenylamine (DPA). Field measurements revealed the occurrence of NO-DCF, NO2-DCF and NO-DPA in groundwater samples at concentration levels in the 6-68s/L range. These concentration levels are more significant than those previously reported in wastewater treatment plant effluents taking into account dilution processes in soil. Interestingly, the p-benzoquinone imine of 5-OH-DCF was also found to be rather stable in surface water. In laboratory batch experiments under strictly anoxic conditions, the transformation of DCF and DPA into their corresponding N-nitroso derivatives was well correlated to denitrification processes. It was also observed that NO-DCF evolved into NO2-DCF while NO-DPA was stable. In vitro experiments showed that the Fisher-Hepp rearrangement could not account for NO2-DCF formation. One possible mechanism might be that NO-DCF underwent spontaneous NO loss to give the resulting intermediates diphenylaminyl radical or nitrenium cation which might evolve into NO2-DCF in presence of NO2 radical or nitrite ion, respectively.