Removal of selected sulfonamides and sulfonamide resistance genes from wastewater in full-scale constructed wetlands.

Sulfonamides are high-consumption antibiotics that reach the aquatic environment. The threat related to their presence in wastewater and the environment is not only associated with their antibacterial properties, but also with risk of the spread of drug resistance in bacteria. Therefore, the aim of this work was to evaluate the occurrence of eight commonly used sulfonamides, sulfonamide resistance genes (sul1-3) and integrase genes intI1-3 in five full-scale constructed wetlands (CWs) differing in design (including hybrid systems) and in the source of wastewater (agricultural drainage, domestic sewage/surface runoff, and animal runs runoff in a zoo). The CWs were located in low-urbanized areas in Poland and in Czechia. No sulfonamides were detected in the CW treating agricultural tile drainage water. In the other four systems, four sulfonamide compounds were detected. Sulfamethoxazole exhibited the highest concentration in those four CWs and its highest was 12,603.23 ± 1000.66 ng/L in a CW treating a mixture of domestic sewage and surface runoff. Despite the high removal efficiencies of sulfamethoxazole in the tested CWs (86 %-99 %), it was still detected in the treated wastewater. The sul1 genes occurred in all samples of raw and treated wastewater and their abundance did not change significantly after the treatment process and it was, predominantly, at the level 105 gene copies numbers/mL. Noteworthy, sul2 genes were only found in the influents, and sul3 were not detected. The sulfonamides can be removed in CWs, but their elimination is not complete. However, hybrid CWs treating sewage were superior in decreasing the relative abundance of genes and the concentration of SMX. CWs may play a role in the dissemination of sulfonamide resistance genes of the sul1 type and other determinants of drug resistance, such as the intI1 gene, in the environment, however, the magnitude of this phenomenon is a matter of further research.

Adsorption of common greywater pollutants and nutrients by various biochars as potential amendments for nature-based systems: Laboratory tests and molecular dynamics.

Spruce wood and Typha (wetland plant) derived biochars pyrolyzed at 350 °C and 600 °C were tested for their sorption affinity for organic pollutants (diclofenac, methylparaben, benzotriazole and sodium 1-decanesulfonate) and nutrients (nitrate, ammonium, phosphate and boron) commonly found in greywater. Batch and column studies combined with molecular dynamics modelling determined the sorption capacity, kinetics, and described the underlying mechanisms. The spruce biochar (600 °C) exhibited the highest sorption capacity mainly for the tested organics. The dynamic test performed for spruce biochar (600 °C) showed that the magnitude of desorption was low, and the desorbed amount ranged between 3 and 11 %. Molecular dynamics modelling (a computational tool for elucidating molecular-level interactions) indicated that the increased sorption of nitrate and boron on spruce biochar (600 °C) could be attributed to hydrophobic interactions. The molecular dynamics shows that predominant adsorption of organic pollutants was governed by π-π stacking, with a minor role of hydrogen-bonding on the biochar surface. In summary, higher pyrolysis temperature biochar yielded greater adsorption capacity greywater borne contaminants and the reaction temperature (10-34 °C) and presence of anionic surfactant had a limited effect on the adsorption of organic pollutants, suggesting efficacious application of biochar in general for greywater treatment in nature-based systems.

Constructed wetlands for the treatment of household organic micropollutants with contrasting degradation behaviour: Partially-saturated systems as a performance all-rounder.

The degradability of specific organic micropollutants in constructed wetlands (CWs) may differ depending on the prevalence of oxic or anoxic conditions. These conditions are governed, among other factors, by the water saturation level in the system. This study investigated the removal of three environmentally-relevant organic micropollutants: bisphenol-group plasticizer bisphenol S (BPS), household-use insecticide fipronil (FPN) and non-steroidal anti-inflammatory drug ketoprofen (KTP) in the model CWs set up in an outdoor column system. BPS and KTP, in contrast to FPN, exhibit higher biodegradability potential under oxic conditions. The experimental CWs were operated under various saturation conditions: unsaturated, partially saturated and saturated, and mimicked the conditions occurring in unsaturated, partially-saturated intermittent vertical-flow CWs and in horizontal-flow CWs, respectively. The CWs were fed with synthetic household wastewater with the concentration of the micropollutants at the level of 30-45 µg/L. BPS and KTP exhibited contrasting behaviour against FPN in the CWs in the present experiment. Namely, BPS and KTP were almost completely removed in the unsaturated CWs without a considerable effect of plants, but their removal in saturated CWs was only moderate (approx. 50%). The plants had only a pronounced effect on the removal of BPS in saturated systems, in which they enhanced the removal by 46%. The removal of FPN (approx. 90%) was the highest in the saturated and partially-saturated CWs, with moderate removal (66.7%) in unsaturated systems. Noteworthy, partially-saturated CWs provided high or very high removal of all three studied substances despite their contrasting degradability under saturated and unsaturated conditions. Namely, their removal efficiencies in planted CWs were 95.9%, 94.5% and 81.6%, for BPS, KTP and FPN, respectively. The removal of the micropollutants in partially-saturated CWs was comparable or only slightly lower than in the best treatment option making it the performance all-rounder for the compounds with contrasting biodegradability properties.

Towards Sustainable Wastewater Treatment: Bioindication as a Technique for Supporting Treatment Efficiency Assessment.

Constructed wetlands (CWs) are a promising alternative for conventional methods of wastewater treatment. However, the biggest challenge in wastewater treatment is the improvement of the technology used so that it is possible to remove micropollutants without additional costs. The impact of wastewater treatment in CWs on toxicity towards Aliivibrio fischeri, Daphnia magna and Lemna minor was investigated. The effects of feeding regime (wastewater fed in five batches per week at a batch volume of 1 L, or twice per week at a batch volume of 2.5 L) and the presence of pharmaceuticals (diclofenac and sulfamethoxazole), as well as the presence of Miscantus giganteus plants in CW columns (twelve of the 24 columns that were planted) were analyzed. A reduction in toxicity was observed in all experimental setups. The effluents from constructed wetlands were classified as moderately toxic (average TU for A. fischeri, D. magna and L. minor was 0.9, 2.5 and 5.5, respectively). The feeding regime of 5 days of feeding/2 days of resting resulted in a positive impact on the ecotoxicological and chemical parameters of wastewater (removal of TOC, N-NH4 and pharmaceuticals). Extended exposure of Miscantus giganteus to the wastewater containing pharmaceuticals resulted in elevated activity of antioxidant enzymes (catalase and superoxide dismutase) in leaf material.

Removal and transformation of sulfamethoxazole in acclimated biofilters with various operation modes - Implications for full-scale application.

The knowledge gaps regarding the degradation of sulfamethoxazole (SMX) in biofilters include the effect of aeration, constant feeding with readily biodegradable organic carbon and the presence of reactive media such as manganese oxides (MnOx). Thus, the goal of this study was to assess the removal of SMX in lab-scale biofilters with various operation variables: aeration, presence of MnOx as an amendment of filtering medium and the presence of readily biodegradable organic carbon (acetate). The sand used in the experiment as a filtering medium was previously exposed to the presence of SMX and acetate, which provided acclimation of the biomass. The removal of SMX was complete (>99%) with the exception of the unaerated columns fed with the influent containing acetate, due to apparent slower rate of SMX degradation. The obtained results suggest that bacteria were able to degrade SMX as a primary substrate and the degradation of this compound was subsequent to the depletion of acetate. The LC-MS/MS analysis of the effluents indicated several biotransformation reactions for SMX: (di)hydroxylation, acetylation, nitrosation, deamonification, S-N bond cleavage and isoxazole-ring cleavage. The relative abundance of transformation products was decreased in the presence of MnOx or acetate. Based on the Microtox assay, only the effluents from the unaerated columns filled with MnOx were classified as non-toxic. The results offer important implications for the design of biofilters for the elimination of SMX, namely that biofilters offer the greatest performance when fed with secondary wastewater and operated as non-aerated systems with a filtering medium containing MnOx.

Supramolecular Solvent-Based Microextraction of Selected Anticonvulsant and Nonsteroidal Anti-Inflammatory Drugs from Sediment Samples.

The increase in the production and consumption of pharmaceuticals increases their presence in the global environment, which may result in direct threats to living organisms. For this reason, there is a need for new methods to analyze drugs in environmental samples. Here, a new procedure for separating and determining selected drugs (diclofenac, ibuprofen, and carbamazepine) from bottom sediment and water samples was developed. Drugs were determined by ultra-high performance liquid chromatography coupled with an ultraviolet detector (UHPLC-UV). In this work, a universal and single-step sample treatment, based on supramolecular solvents (SUPRAS), was proposed to isolate selected anticonvulsants and nonsteroidal anti-inflammatory drugs (NSAIDs) from sediment samples. The following parameters were experimentally selected: composition of the supramolecular solvent (composition THF:H2O (v/v), amount of decanoic acid), volume of extractant, sample mass, extraction time, centrifugation time, and centrifugation speed. Finally, the developed procedure was validated. A Speedisk procedure was also developed to extract selected drugs from water samples. The recovery of analytes using the SUPRAS procedure was in the range of 88.8-115%, while the recoveries of the Speedisk solid-phase extraction procedure ranged from 81.0-106%. The effectiveness of the sorption of the tested drugs by sediment was also examined.

Antimicrobial pharmaceuticals in the aquatic environment - occurrence and environmental implications.

The environmental occurrence of antimicrobial pharmaceuticals and antibiotic resistant bacteria and antibiotic resistant genes has become a global phenomenon and a multifaceted threat. Integrated actions of many parties are needed to prevent further aggravation of the problem. Well-directed actions require clear understanding of the problem, which can be ensured by frequent revaluation of the existing knowledge and disseminating it among relevant audiences. The goal of this review paper is to discuss the occurrence and abundance of antimicrobial pharmaceuticals in the aquatic environment in context of adverse effects caused directly by these substances and the threat associated with the antibiotics resistance phenomenon. Several classes of antimicrobial pharmaceuticals (aminoglycosides, ß-lactams, glycopeptides, macrolides, fluoroquinolones, sulfonamides and trimethoprim, tetracyclines) have been selected to illustrate their sources, environmental abundance, degradation routes (transformation products) and environmental implications including their ecotoxic effect and the spread of antibiotic resistance within the compartments of the aquatic environment and wastewater treatment plants. Wastewater treatment plants are indeed the main source responsible for the prevalence of these factors in the aquatic environment, since predominantly the plants have not been designed to retain antimicrobial pharmaceuticals. In order to limit the prevalence of these impurities into the environment, better source control is recommended as well as the establishment of stricter environmental quality standards. Counteracting all the above-mentioned threats requires to undertake integrated activities based on cooperation of professionals and scientists from various fields of science or industry, such as environmental sciences, medicine, veterinary, pharmacology, chemical engineering and others.

Removal and transformation pathways of benzothiazole and benzotriazole in membrane bioreactors treating synthetic municipal wastewater.

Lab-scale membrane bioreactors (MBRs), with aerated activated sludge and internal microfiltration module, were used for the treatment of municipal wastewater containing high, yet environmentally relevant, concentrations of benzothiazole (BT) and benzotriazole (BTA). These high production volume compounds are commonly used in the industry and households, and therefore occur ubiquitously in municipal wastewater and the aquatic environment. The aim of this study was to assess the removal of BT and BTA from synthetic municipal wastewater in MBRs and to estimate the contribution of elimination processes and to identify potential biotransformation products. The overall removal of BT and BTA was high, and after the adaptation period, it reached 99.8% and 97.2%, respectively, but recurring periods of unstable BTA removal occurred. The removal due to biotransformation was 88% for BT and 84% for BTA and the disposal with waste sludge accounted for only <1% of the removed load. The remaining fraction of the removed load of BT and BTA was attributed to be retained by phenomena associated with membrane fouling. The adaptation process was reflected in multifold increase in biodegradation kinetic coefficient (kbiol) for BT (reported for the first time) and BTA. Biodegradation was attributed to catabolic mechanism rather than to cometabolism. Hydroxylation was observed to be the main transformation reaction for BT, whereas for BTA hydroxylation, methylation and cleavage of benzene ring were noted. This study has shown the feasibility of treating municipal wastewater with high concentrations of BT and BTA in MBRs and identified potential challenges for the removal of BTA.

Removal and transformation of benzotriazole in manganese-oxide biofilters with Mn(II) feeding.

The aim of this study was to evaluate the treatment of organic-carbon-deficient wastewater containing benzotriazole (BTA) in lab-scale aerated biofilters filled with natural manganese oxide ore, sand coated with synthetic manganese oxides and sand (as a control material) in terms of BTA removal efficiency, its transformation products and ecotoxicological impact of the treated wastewater. Additionally, the effect of Mn(II) feeding was tested. The removal of BTA in all the biofilters was ≥97%. The contribution of the biotic removal of this compound was 15%, 50%, and 75% in the systems filled with sand, synthetic and natural manganese oxides, respectively. Only the columns filled with natural manganese oxides provided significant removal of DOC and decrease of UV254 and SUVA254, with even more pronounced effect with Mn(II) feeding. The presence of Mn(II) was also found to enhance the removal of NNH4 in the systems filled with either form of manganese oxides, otherwise the removal of NNH4 was negligible or negative. The transformation reactions of BTA were methylation, hydroxylation, and triazole ring cleavage. Based on the number of compounds and their relative abundance, the methylated transformation products were predominant in the effluent. The reduction of the ecotoxicity (Microtox bioassay) of the effluents was positively correlated with the decrease of UV254, SUVA and DOC and only moderately with the removal of BTA. This study has shown that the natural manganese oxide ore provides the broadest set of services as a filtering material for aerated biofilters treating carbon-deficient wastewater containing BTA.

The treatment of wastewater containing pharmaceuticals in microcosm constructed wetlands: the occurrence of integrons (int1-2) and associated resistance genes (sul1-3, qacEΔ1).

The aim of this study was to analyze the occurrence of sulfonamide resistance genes (sul1-3) and other genetic elements as antiseptic resistance gene (qacEΔ1) and class 1 and class 2 integrons (int1-2) in the upper layer of substrate and in the effluent of microcosm constructed wetlands (CWs) treating artificial wastewater containing diclofenac and sulfamethoxazole (SMX), which is a sulfonamide antibiotic. The bacteria in the substrate and in the effluents were equipped with the sul1-2, int1, and qacEΔ1 resistance determinants, which were introduced into the CW system during inoculation with activated sludge and with the soil attached to the rhizosphere of potted seedlings of Phalaris arundinacea 'Picta' roots (int1). By comparing the occurrence of the resistance determinants in the upper substrate layer and the effluent, it can be stated that they neither were lost nor emerged along the flow path. The implications of the presence of antibiotic resistance genes in the effluent may entail a risk of antibiotic resistance being spread in the receiving environment. Additionally, transformation products of SMX were determined. According to the obtained results, four (potential) SMX transformation products were identified. Two major metabolites of SMX, 2,3,5-trihydroxy-SMX and 3,5-dihydroxy-SMX, indicated that SMX may be partly oxidized during the treatment. The remaining two SMX transformation products (hydroxy-glutathionyl-SMX and glutathionyl-SMX) are conjugates with glutathione, which suggests the ability of CW bacterial community to degrade SMX and resist antimicrobial stress.

Degradation of benzotriazole and benzothiazole in treatment wetlands and by artificial sunlight.

Laboratory-scale experiments were performed using unsaturated subsurface-flow treatment wetlands and artificial sunlight (with and without TiO2) to study the efficiency of benzotriazole and benzothiazole removal and possible integration of these treatment methods. Transformation products in the effluent from the treatment wetlands and the artificial sunlight reactor were identified by high performance liquid chromatography coupled with tandem mass spectrometry. The removal of benzothiazole in the vegetated treatment wetlands was 99.7%, whereas the removal of benzotriazole was 82.8%. The vegetation positively affected only the removal of benzothiazole. The major transformation products in the effluents from the treatment wetlands were methylated and hydroxylated derivatives of benzotriazole, and hydroxylated derivatives of benzothiazole. Hydroxylation was found to be the main process governing the transformation pathway for both compounds in the artificial sunlight experiment (with and without TiO2). Benzotriazole was not found to be susceptible to photodegradation in the absence of TiO2. The integration of the sunlight-induced processes (with TiO2) with subsurface-flow treatment wetlands caused further elimination of the compounds (42% for benzotriazole and 58% for benzothiazole). This was especially significant for the elimination of benzotriazole, because the removal of this compound was 96% in the coupled processes.

Removal of diclofenac and sulfamethoxazole from synthetic municipal waste water in microcosm downflow constructed wetlands: Start-up results.

The objectives of this study were to investigate the start-up removal of pharmaceutical compounds diclofenac and sulfamethoxazole in microcosm downflow constructed wetlands and their effect on the performance of the studied constructed wetlands, and also to assess the effect of plants on the removal of these compounds. The experimental system that was used in this 86-day experiment consisted of 24 columns filled up to 70 cm with predominantly sandy material. Four types of columns were used (six replicates) depending on the presence of plants (Phalaris arundinacea L. var. picta L.) and the presence of pharmaceutical compounds in the influent. The influent was synthetic municipal waste water to which a mixture of 5 mg/L of diclofenac and 5 mg/L of sulfamethoxazole was added. The observed removal of diclofenac was moderate (approx. 50%) and the removal of sulfamethoxazole was relatively low (24-30%). It was found that the removal of diclofenac and sulfamethoxazole was not affected by the vegetation. The presence of diclofenac and sulfamethoxazole in the influent had significant effect on the effluent concentration of N-NO3 and the water loss in the columns, which in both cases were lower than in the control columns. The scope for further research was discussed.

Accumulation of Metals and Boron in Phragmites australis Planted in Constructed Wetlands Polishing Real Electroplating Wastewater.

The concentration of metals (Al, Cu, Fe, Mn, Ni, Zn) and B were determined in the above- and belowground biomass of Phragmites australis collected from the microcosm constructed wetland system used for the polishing of real electroplating wastewater. Translocation factor and bioconcentration factor were determined. Pearson correlation test was used to determine correlation between metal concentration in substrate and above- and belowground parts of Phragmites australis. The obtained results suggested that Phragmites australis did not play a major role as an accumulator of metals. It was observed also that the substrate could have exerted an effect on the translocation of Ni, Cu, Zn and Mn. The analysed concentrations of metals and B in biomass were in the range or even below the concentrations reported in the literature with the exception of Ni. The aboveground biomass was found suitable as a composting input in terms of metals concentrations.

R&D priorities in the field of sustainable remediation and purification of agro-industrial and municipal wastewater.

This article was presented as a position paper during the Environmental Biotechnology and Microbiology Conference in Bologna, Italy in April 2012. It indicates major and emerging environmental biotechnology research and development (R&D) priorities for EU members in the field of sustainable remediation and purification of agro-industrial and municipal wastewater. The identified priorities are: anaerobic/aerobic microbial treatment, combination of photochemical and biological treatment, phytoremediation and algae-based remediation, as well as innovative technologies currently investigated, such as enzyme-based treatment, bioelectrochemical treatment and recovery of nutrients and reuse of cleaned water. State of the art, research needs and prospective development in these domains are crucially discussed. As a result, goals of the future development of bioremediation and purification processes are defined and the way to achieve them is proposed.