Phycobiliprotein recovery coupled to the tertiary treatment of wastewater in semi-continuous photobioreactors. Tracking contaminants of emerging concern.

This study evaluated a tertiary wastewater treatment technology using cyanobacteria to recover value-added phycobiliproteins. The presence of contaminants of emerging concern (CECs) in wastewater, cyanobacterial biomass and pigments recovered were also analyzed. For this, a wastewater-borne cyanobacterium (Synechocystis sp. R2020) was used to treat secondary effluent from a municipal wastewater treatment plant, with and without nutrients supplementation. Then, the stability of phycobiliprotein production was assessed by operating the photobioreactor in semi-continuous mode. Results showed similar biomass productivity with and without nutrients supplementation (153.5 and 146.7 mg L-1 d-1, respectively). Upon semi-continuous operation, the phycobiliprotein content was stable and reached up to 74.7 mg gDW-1. The phycocyanin purity ratio ranged from 0.5 to 0.8, corresponding to food grade (>0.7). Out of 22 CECs detected in secondary effluent, only 3 were present in the phycobiliprotein extracts. In order to identify applications, prospective research should focus on CECs removal during pigment purification.

Attenuation and soil biodegradation of fungicides by using vegetated buffer strips in vineyards during a simulated rainfall-runoff event.

Rainfall-runoff events occurring in vineyard fields can result in pesticide ground losses and the subsequent pollution of surface water bodies, derivate from the crop protection spray applications. In this study, the capacity of vegetated buffer strips (BS) to prevent surface water pollution due to the application of five fungicide products typically used in vineyards (copper, dimethomorph, oxathiapiprolin, zoxamide, acibenzolar-s-methyl, and laminarin) following a simulated run-off event has been assessed, and compared to that from a bare ground soil (BG). Two strips (5 m in length, each), one with vegetation and the other without were built up, and two different experiments were performed, a runoff event and a soil fungicide degradation kinetic evaluation. The runoff results show that fungicide mass retention in the strips ranged from 73 to 98% and that the presence of vegetation in BS increased the fungicide mass retention in the strips by almost 10% (on average) in comparison to the unvegetated strip. Moreover, soil degradation studies highlighted that the presence of vegetation reduces significantly the half-time life of almost all the studied fungicides by 55%, on average. Eight fungicide transformation products (TPs) were identified following a runoff event in the soil strips, but the abundance of these TPs was up to 78% lower in vegetated strips. These results highlight the effectiveness of using vegetated buffer zones in vineyards to protect aquatic ecosystem pollution.

The potential of wastewater grown microalgae for agricultural purposes: Contaminants of emerging concern, heavy metals and pathogens assessment.

In the coming years, the use of microalgal biomass as agricultural biofertilizers has shown promising results. The use of wastewater as culture medium has resulted in the reduction of production costs, making microalgae-based fertilizers highly attractive for farmers. However, the occurrence of specific pollutants in wastewater, like pathogens, heavy metals and contaminants of emerging concern (CECs), such as pharmaceuticals and personal care products may pose a risk on human health. This study presents an holistic assessment of the production and use of microalgal biomass grown in municipal wastewater as biofertilizer in agriculture. Results showed that pathogens and heavy metals concentrations in the microalgal biomass were below the threshold established by the European regulation for fertilizing products, except for cadmium. Regarding CECs, 25 out of 29 compounds were found in wastewater. However, only three of them (hydrocinnamic acid, caffeine, and bisphenol A) were found in the microalgae biomass used as biofertilizer. Agronomic tests were performed for lettuce growth in greenhouse. Four treatments were studied, comparing the use of microalgae biofertilizer with a conventional mineral fertilizer, and also a combination of both of them. Results suggested that microalgae can help reducing the mineral nitrogen dose, since similar fresh shoot weights were obtained in the plants grown with the different assessed fertilizers. Lettuce samples revealed the presence of cadmium and CECs in all the treatments including both negative and positive controls, which suggests that their presence was not linked to the microalgae biomass. On the whole, this study revealed that wastewater grown microalgae can be used for agricultural purposes reducing mineral N need and guaranteeing health safety of the crops.

Use of wood and cork in biofilters for the simultaneous removal of nitrates and pesticides from groundwater.

About 13% and 7% of monitored groundwater stations in Europe exceed the permitted levels of nitrates (50 mg NO3- L-1) or pesticides (0.1 µg L-1), respectively. Although slow sand filtration can remove nitrates via denitrification when oxygen is limited, it requires an organic carbon source. The present study evaluates the performance of the use of wood pellets and granulated cork as carbon sources in bench-scale biofilters operated under water-saturated and water-unsaturated conditions for more than 400 days. The biofilters were monitored for nitrate (200 mg L-1) and pesticide (mecoprop, diuron, atrazine, and bromacil, each at a concentration of 5 µg L-1) attenuation, as well as for the formation of nitrite and pesticide transformation products. Microbiological characterization of each biofilter was also performed. The water-saturated wood biofilter achieved the best nitrate removal (>99%), while the cork biofilters lost all denitrification power over time (from 38% to no removal). The unsaturated biofilter columns were not effective for removing nitrates (20-30% removal). As for pesticides, all the biofilters achieved high removal rates of mecoprop and diuron (>99% and >75%, respectively). Atrazine removal was better in the wood-pellet biofilters than the cork ones (68-96% vs. 31-38%). Bromacil was only removed in the water-unsaturated cork biofilter (67%). However, a bromacil transformation product was formed there. The water-saturated wood biofilter contained the highest number of denitrifying microorganisms, with Methyloversatilis as the characteristic genus. Microbial composition could explain the high removal of pesticides and nitrates achieved in the wood-pellet biofilter. Overall, the results indicate that wood-pellet biofilters operated under water-saturated conditions are a good solution for treating groundwater contaminated with nitrates and pesticides.

Assessment of a novel microalgae-cork based technology for removing antibiotics, pesticides and nitrates from groundwater.

Groundwater pollution has increased in recent years due to the intensification of agricultural and livestock activities. This results in a significant reduction in available freshwater resources. Here, we have studied the long term assessment of a green technology (1-4 L/day) based on a photobioreactor (PBR) containing immobilised microalgae-bacteria in polyurethane foam (PF) followed by a cork filter (CF) for removing nitrates, pesticides (atrazine and bromacil), and antibiotics (sulfamethoxazole and sulfacetamide) from groundwater. The prototype was moderately effective for removing nitrates (58%) at an HRT of 8 days, while its efficiency decreased at a HRT of 4 and 2 days (<20% removal). The combined use of PBR-CF enabled antibiotics and pesticides to be attenuated by up to 95% at an HRT of 8 days, but their attenuation decreased with shorter HRT, with pesticides being the compounds most affected (reducing from 97 to 98% at an HRT of 8 days to 23-45% at an HRT of 2 days). Pesticide transformation products were identified after the CF, supporting biodegradation as the main attenuation process. A gene-based metataxonomic assessment linked the attenuation of micropollutants to the presence of specific pesticide biodegradation species (e.g. genus Phenylobacterium, Sphingomonadaceae, and Caulobacteraceae). Therefore, the results highlighted the potential use of microalgae and cork to treat polluted groundwater.

Exploring the usage of artificial root exudates to enhance the removal of contaminants of emerging concern in slow sand filters: Synthetic vs. real wastewater conditions.

In previous studies we observed that laboratory-scale constructed wetlands exposed to contaminants of emerging concern (CECs) increased the release of certain root metabolites which were probably linked with the enhancement of CEC biodegradation. Based on this, the current study tested if the addition of artificial root exudates in slow sand-filtration systems could also enhance the removal of CECs from wastewater. First, in a preliminary study, twelve laboratory-scale slow sand-filtration columns were operated under synthetic and unrealistic wastewater conditions. Thus, by using synthetic wastewater, high concentration of CECs (100 µg L-1 of benzotriazole, sulfamethoxazole, carbamazepine and diclofenac) and artificial root exudates (2-9 g L-1 of glucose, salicylic acid or arginine) we observed that attenuation was enhanced for most of the studied CECs (up to 400%). This enhancement was attributed to cometabolism because the effects on CEC removal ceased when the supply of root exudates was stopped. A follow-up study was conducted to approach real-wastewater treatment conditions. Sand columns were operated with real wastewater, lower concentrations of the selected CECs (20 µg L-1) and of root exudates (0.2 mg L-1 of salicylic acid and 1.14 mg L-1 TOC of Cyperus alternifolius' root exudates). Under these conditions, the addition of root exudates on CEC removal had no effects. Thus, we conducted another test with three different concentrations of salicylic acid. When the concentration of salicylic acid increased to 200 mg L-1, CEC removal modestly increased (up to 40%). Divergence between synthetic and real wastewater studies might be explained, mainly, by the presence of organic sources of nutrients in wastewater, which probably masked the effect of root exudates addition at lower concentrations. This study demonstrates that the effectiveness of root exudates application on the attenuation of CECs from wastewater should be explored under real wastewater conditions.

Linking plant-root exudate changes to micropollutant exposure in aquatic plants (Lemna minor and Salvinia natans). A prospective metabolomic study.

Recent findings indicate that plant-root exudates can stimulate plant-associated microorganisms to enhance the biodegradation of contaminants in constructed wetlands. To understand this process, we studied the root-exudation changes of two aquatic plants (Lemna minor and Salvinia natans) upon micropollutants exposure (10, 100 and 1000 µg/L mixes containing naproxen, diclofenac, carbamazepine, and benzotriazole). After a 2-day exposure, plant exudates were collected, extracted and non-target analysis was performed with a gas chromatography-high resolution Orbitrap mass-spectrometer. Plants didn't show morphological or growth differences between the control and spiked reactors, but exudation changes were observed in both plants at all concentration levels. Partial least squares discriminant analysis showed that, for Lemna minor, the increase of micropollutants exposure was linked to the reduction of sugar and fatty acid exudation. This may trigger changes in the microbial community living on complex carbon forms. Instead, in Salvinia natans, micropollutants exposure was linked to the release of long-chain compounds such as cuticular waxes and sesquiterpenoids, which might be related to stress signaling. These results demonstrate that plant micropollutant-exposure at environmentally relevant concentration levels triggers changes in root exudates. This may help to design new strategies to enhance micropollutants degradation in nature based solutions such as in constructed wetlands.

Degradation of mecoprop in polluted landfill leachate and waste water in a moving bed biofilm reactor.

Mecoprop is a common pollutant in effluent-, storm- and groundwater as well as in leachates from derelict dumpsites. Thus, bioremediation approaches may be considered. We conducted batch experiments with moving bed biofilm (MBBR)-carriers to understand the degradation of mecoprop. As a model, the carriers were incubated in effluent from a conventional wastewater treatment plant which was spiked to 10, 50 and 100 µg L-1 mecoprop. Co-metabolic processes as well as mineralization were studied. Initial mecoprop concentration and mecoprop degradation impacted the microbial communities. The removal of (S)-mecoprop prevailed over the (R)-mecoprop. This was associated with microbial compositions, in which several operational taxonomic units (OTUs) co-varied positively with (S)-mecoprop removal. The removal-rate constant of (S)-mecoprop was 0.5 d-1 in the 10 µg L-1 set-up but it decreased in the 50 and 100 µg L-1 set-ups. The addition of methanol prolonged the removal of (R)-mecoprop. During mecoprop degradation, 4-chloro-2-methylphenol was formed and degraded. A new metabolite (4-chloro-2-methylphenol sulfate) was identified and quantified.

Transformation products of clindamycin in moving bed biofilm reactor (MBBR).

Clindamycin is widely prescribed for its ability to treat a number of common bacterial infections. Thus, clindamycin enters wastewater via human excretion or disposal of unused medication and widespread detection of pharmaceuticals in rivers proves the insufficiency of conventional wastewater treatment plants in removing clindamycin. Recently, it has been discovered that attached biofilm reactors, e.g., moving bed biofilm reactors (MBBRs) obtain a higher removal of pharmaceuticals than conventional sludge wastewater treatment plants. Therefore, this study investigated the capability of MBBRs applied in the effluent of conventional wastewater treatment plants to remove clindamycin. First, a batch experiment was executed with a high initial concentration of clindamycin to identify the transformation products. It was shown that clindamycin can be removed from wastewater by MBBR and the treatment process converts clindamycin into the, possibly persistent, products clindamycin sulfoxide and N-desmethyl clindamycin as well as 3 other mono-oxygenated products. Subsequently, the removal kinetics of clindamycin and the formation of the two identified products were investigated in batch experiments using MBBR carriers from polishing and nitrifying reactors. Additionally, the presence of these two metabolites in biofilm-free wastewater effluent was studied. The nitrifying biofilm reactor had a higher biological activity with k-value of 0.1813 h-1 than the reactor with polishing biofilm (k = 0.0161 h-1) which again has a much higher biological activity for removal of clindamycin than of the suspended bacteria (biofilm-free control). Clindamycin sulfoxide was the main transformation product which was found in concentrations exceeding 10% of the initial clindamycin concentration after 1 day of MBBR treatment. Thus, MBBRs should not necessarily be considered as reactors mineralizing clindamycin as they perform transformation reactions at least to some extent.

Influence of humic acid addition on the degradation of pharmaceuticals by biofilms in effluent wastewater.

The degradation of organic micropollutants in wastewater treatment is suspected to depend on co-degradation i.e. be dependent on concentrations of substrate. This complicates predicting and modelling their fate. The effect of humic acid, as a model for complex organic substrate, was investigated in relation to the biodegradation of pharmaceuticals by suspended biofilm carriers adapted to polishing effluent water from a tertiary sewage treatment plant. Twelve out of 22 investigated pharmaceuticals were significantly biodegradable. The biodegradation rate constants of ten of those compounds were increasing with increased humic acid concentrations. At the highest humic acid concentration (30mgC/L), the biodegradation rate constants were four times higher than the biodegradation rate constants without added humic acid. This shows that the presence of complex substrate stimulates degradation via a co-metabolism-like mechanism and competitive inhibition does not occur. Increases of rate constant per mgC/L are tentatively calculated.

Impact of external carbon dose on the removal of micropollutants using methanol and ethanol in post-denitrifying Moving Bed Biofilm Reactors.

Addition of external carbon sources to post-denitrification systems is frequently used in wastewater treatment plants to enhance nitrate removal. However, little is known about the fate of micropollutants in post-denitrification systems and the influence of external carbon dosing on their removal. In this study, we assessed the effects of two different types and availability of commonly used carbon sources -methanol and ethanol- on the removal of micropollutants in biofilm systems. Two laboratory-scale moving bed biofilm reactors (MBBRs), containing AnoxKaldnes K1 carriers with acclimated biofilm from full-scale systems, were operated in continuous-flow using wastewater dosed with methanol and ethanol, respectively. Batch experiments with 22 spiked pharmaceuticals were performed to assess removal kinetics. Acetyl-sulfadiazine, atenolol, citalopram, propranolol and trimethoprim were easily biotransformed in both MBBRs (biotransformations rate constants kbio between 1.2 and 12.9 L gbiomass-1 d-1), 13 compounds were moderately biotransformed (rate constants between 0.2 and 2 L gbiomass-1 d-1) and 4 compounds were recalcitrant. The methanol-dosed MBBR showed higher kbio (e.g., 1.5-2.5-fold) than in the ethanol-dosed MBBR for 9 out of the 22 studied compounds, equal kbio for 10 compounds, while 3 compounds (i.e., targeted sulfonamides) were biotransformed faster in the ethanol-dosed MBBR. While biotransformation of most of the targeted compounds followed first-order kinetics, removal of venlafaxine, carbamazepine, sulfamethoxazole and sulfamethizole could be described with a cometabolic model. Analyses of the microbial composition in the biofilms using 16S rRNA amplicon sequencing revealed that the methanol-dosed MBBR contained higher microbial richness than the one dosed with ethanol, suggesting that improved biotransformation of targeted compounds could be associated with higher microbial richness. During continuous-flow operation, at conditions representative of full-scale denitrification systems (hydraulic residence time = 2 h), the removal efficiencies of micropollutants were below 35% in both MBBRs, with the exception of atenolol and trimethoprim (>80%). Overall, this study demonstrated that MBBRs used for post-denitrification could be optimized to enhance the biotransformation of a number of micropollutants by accounting for optimal carbon sources and extended residence time.

Biodegradation of pharmaceuticals in hospital wastewater by a hybrid biofilm and activated sludge system (Hybas).

Hospital wastewater contributes a significant input of pharmaceuticals into municipal wastewater. The combination of suspended activated sludge and biofilm processes, as stand-alone or as hybrid process (hybrid biofilm and activated sludge system (Hybas™)) has been suggested as a possible solution for hospital wastewater treatment. To investigate the potential of such a hybrid system for the removal of pharmaceuticals in hospital wastewater a pilot plant consisting of a series of one activated sludge reactor, two Hybas™ reactors and one moving bed biofilm reactor (MBBR) has been established and adapted during 10 months of continuous operation. After this adaption phase batch and continuous experiments were performed for the determination of degradation of pharmaceuticals. Removal of organic matter and nitrification mainly occurred in the first reactor. Most pharmaceuticals were removed significantly. The removal of pharmaceuticals (including X-ray contrast media, ß-blockers, analgesics and antibiotics) was fitted to a single first-order kinetics degradation function, giving degradation rate constants from 0 to 1.49 h(-1), from 0 to 7.78 × 10(-1)h(-1), from 0 to 7.86 × 10(-1)h(-1) and from 0 to 1.07 × 10(-1)h(-1) for first, second, third and fourth reactors respectively. Generally, the highest removal rate constants were found in the first and third reactors while the lowest were found in the second one. When the removal rate constants were normalized to biomass amount, the last reactor (biofilm only) appeared to have the most effective biomass in respect to removing pharmaceuticals. In the batch experiment, out of 26 compounds, 16 were assessed to degrade more than 20% of the respective pharmaceutical within the Hybas™ train. In the continuous flow experiments, the measured removals were similar to those estimated from the batch experiments, but the concentrations of a few pharmaceuticals appeared to increase during the first treatment step. Such increase could be attributed to de-conjugation or formation from other metabolites.

Can those organic micro-pollutants that are recalcitrant in activated sludge treatment be removed from wastewater by biofilm reactors (slow sand filters)?

The degradation of seven compounds which are usually recalcitrant in classical activated sludge treatment (e.g., diclofenac, propranolol, iopromide, iohexol, iomeprol tebuconazole and propiconazole) was studied in a biofilm reactor (slow sand filtration). This reactor was used to treat real effluent-wastewater at different flow rates (hydraulic loadings) under aerobic conditions so removal and degradation kinetics of these recalcitrant compounds were calculated. With the hydraulic loading rate of 0.012 m(3)m(2)h(-1) the reactor removed 41, 94, 58, 57 and 85% of diclofenac, propranolol, iopromide, iohexol and iomeprol respectively. For these compounds the removal efficiency was dependent on hydraulic residence-times. Only 59 and 21% of the incoming tebuconazole and propiconazole respectively were removed but their removal did not depend on hydraulic residence time. Biofilm reactors are thus efficient in removing micro-pollutants and could be considered as an option for advanced treatment in small wastewater treatment plants.