Membrane bioreactor followed by solar photo-Fenton oxidation: Bacterial community structure changes and bacterial reduction.

The removal of antibiotic resistance genes (ARGs) and taxon-specific markers, the bacterial community structure changes and the permanent inactivation of total bacteria including their antibiotic-resistant counterparts (ARB) in actual wastewater during a Membrane BioReactor (MBR) application followed by solar photo-Fenton oxidation at bench- and then pilot-scale under solar irradiation, were investigated. The presence of enterococci- and pseudomonad-specific taxon markers and of sul1 and ampC ARGs in the MBR effluent was confirmed, indicating the challenge of such processes, for the removal of biological molecules. On the other hand, >99 % reduction of all types of cultivable bacteria examined was observed after MBR treatment, with a 5-log reduction of E. coli and 6-log reduction of P. aeruginosa and Klebsiella spp. There was a shift in the bacterial community structure in the MBR effluent after the bench- and pilot-scale solar photo-Fenton oxidation. Notably, thermotolerant bacterial genera like Ignavibacterium and Thermomonas were prevalent during the pilot-scale process operated at a high ambient temperature, while the most prevalent genera were Mycobacterium, Nocardioides and Mesorhizobium, which are primarily not pathogenic and plant-related. In agreement, a different bacterial community structure according to the G-C content after DGGE analysis was noted between the MBR and solar photo-Fenton oxidation-treated effluents, but interestingly also between the bench- and pilot-scale oxidation-treated effluents. There was complete absence of ARGs after the bench-scale solar photo-Fenton oxidation application but not after the pilot-scale treatment (1.56 and 1.53 log10 CE 100 ng-1 DNA, of sul and ermB, respectively). Taxon-specific markers were found in both oxidation setups. Inactivation of cultivable Escherichia coli, Pseudomonas aeruginosa and Klebsiella spp. (including ARB) was achieved during both oxidation setups, with no further re-activation observed.

Simultaneous inactivation of multidrug-resistant Escherichia coli and enterococci by peracetic acid in urban wastewater: Exposure-based kinetics and comparison with chlorine.

The presence of antibiotic resistance in wastewater sparked a great interest in investigating the inactivation of antibiotic-resistant bacteria by disinfecting agents. In this study, the inactivation kinetics of multidrug-resistant E. coli and enterococci by an emerging environmentally-friendly disinfectant, peracetic acid (PAA), in wastewater and phosphate buffer at pH 6.5 and pH 7.5, were characterized. It was demonstrated that the inactivation of the studied multidrug-resistant bacteria was governed by their exposure to PAA, i.e., integral of the PAA concentration over time (integral CT or ICT). Both regimes of the PAA inactivation of bacteria, i.e., initial resistance followed by a faster inactivation, were described well by an ICT-based Chick-Watson inactivation kinetic model. In wastewater at pH 7.5, the model-predicted ICT requirements showed that the multidrug-resistant enterococci were less susceptible to PAA than E. coli, e.g., to achieve a 3-log reduction, an ICT of 32.7 mg min/L and 23.4 mg min/L was needed, respectively. No regrowth of the studied bacteria was observed after 72 h from PAA disinfection at 25 ± 1 °C. Soluble constituents of wastewater decreased the PAA inactivation of both multidrug-resistant bacteria, i.e., higher inactivation was observed in phosphate buffer than wastewater at the same pH of 7.5. In phosphate buffer, a lower pH of 6.5 resulted in higher inactivation of multidrug-resistant E. coli compared with pH 7.5, but it did not affect the PAA inactivation of multidrug-resistant enterococci. A comparison with the most commonly used chemical disinfectant, chlorine, showed higher inactivation of both multidrug-resistant bacteria by chlorine and higher chlorine decay than PAA. The results of the present study may have implications in designing a PAA disinfection process, aiming at controlling antibiotic resistance, in terms of selecting a suitable fecal indicator and optimizing disinfectant dosing.

Shotgun metagenomics assessment of the resistome, mobilome, pathogen dynamics and their ecological control modes in full-scale urban wastewater treatment plants.

The conventional activated sludge (CAS) process has limited capacity to remove pathogenic microorganisms and antibiotic resistance genes (ARGs), compared to membrane bioreactors (MBRs). However, the full extent of pathogenic microbial fraction, resistome (antibiotic and biocide resistance genes, ARGs and BRGs) and mobilome (mobile genetic elements, MGE) of urban wastewater treatment plant (UWTP) influents and effluents remains unknown. Thus, the fate of putative pathogenic bacteria, ARGs and potential co-occurrence patterns with BRGs, MGEs and bacterial-predatory microorganisms was determined in two full-scale UWTPs, a MBR and a CAS system, using shotgun metagenomics. Both UWTPs significantly reduced the BOD5 (99.4-99.9%), COD (97.6-99.4%) and TSS (98.9-99.9%). MBR was more effective in reducing the abundance and diversity of pathogen-containing taxa, with 4 and 30 taxa enriched in MBR and CAS effluents, respectively. MBR treatment favored resistance genes associated with triclosan, whereas CAS effluents contained ARGs associated with antibiotics of clinical importance. Correlations between putative pathogenic bacteria, ARG/BRGs/MGEs and bacterial-predatory microorganisms suggested that: (i) opportunistic pathogens (Clostridia, Nocardia) may acquire ARGs against first-line treatments and (ii) bacteriophages may act as a biogenic mechanism of pathogen removal. These findings reinforce the MBR capacity to retain pathogenic components, hence reducing potential health risks associated with treated wastewater reuse.

Making Waves: Collaboration in the time of SARS-CoV-2 - rapid development of an international co-operation and wastewater surveillance database to support public health decision-making.

The presence of SARS-CoV-2 RNA in wastewater was first reported in March 2020. Over the subsequent months, the potential for wastewater surveillance to contribute to COVID-19 mitigation programmes has been the focus of intense national and international research activities, gaining the attention of policy makers and the public. As a new application of an established methodology, focused collaboration between public health practitioners and wastewater researchers is essential to developing a common understanding on how, when and where the outputs of this non-invasive community-level approach can deliver actionable outcomes for public health authorities. Within this context, the NORMAN SCORE "SARS-CoV-2 in sewage" database provides a platform for rapid, open access data sharing, validated by the uploading of 276 data sets from nine countries to-date. Through offering direct access to underpinning meta-data sets (and describing its use in data interpretation), the NORMAN SCORE database is a resource for the development of recommendations on minimum data requirements for wastewater pathogen surveillance. It is also a tool to engage public health practitioners in discussions on use of the approach, providing an opportunity to build mutual understanding of the demand and supply for data and facilitate the translation of this promising research application into public health practice.

A global multinational survey of cefotaxime-resistant coliforms in urban wastewater treatment plants.

The World Health Organization Global Action Plan recommends integrated surveillance programs as crucial strategies for monitoring antibiotic resistance. Although several national surveillance programs are in place for clinical and veterinary settings, no such schemes exist for monitoring antibiotic-resistant bacteria in the environment. In this transnational study, we developed, validated, and tested a low-cost surveillance and easy to implement approach to evaluate antibiotic resistance in wastewater treatment plants (WWTPs) by targeting cefotaxime-resistant (CTX-R) coliforms as indicators. The rationale for this approach was: i) coliform quantification methods are internationally accepted as indicators of fecal contamination in recreational waters and are therefore routinely applied in analytical labs; ii) CTX-R coliforms are clinically relevant, associated with extended-spectrum ß-lactamases (ESBLs), and are rare in pristine environments. We analyzed 57 WWTPs in 22 countries across Europe, Asia, Africa, Australia, and North America. CTX-R coliforms were ubiquitous in raw sewage and their relative abundance varied significantly (<0.1% to 38.3%), being positively correlated (p < 0.001) with regional atmospheric temperatures. Although most WWTPs removed large proportions of CTX-R coliforms, loads over 103 colony-forming units per mL were occasionally observed in final effluents. We demonstrate that CTX-R coliform monitoring is a feasible and affordable approach to assess wastewater antibiotic resistance status.

Antibiotic resistance genes in treated wastewater and in the receiving water bodies: A pan-European survey of urban settings.

There is increasing public concern regarding the fate of antibiotic resistance genes (ARGs) during wastewater treatment, their persistence during the treatment process and their potential impacts on the receiving water bodies. In this study, we used quantitative PCR (qPCR) to determine the abundance of nine ARGs and a class 1 integron associated integrase gene in 16 wastewater treatment plant (WWTP) effluents from ten different European countries. In order to assess the impact on the receiving water bodies, gene abundances in the latter were also analysed. Six out of the nine ARGs analysed were detected in all effluent and river water samples. Among the quantified genes, intI1 and sul1 were the most abundant. Our results demonstrate that European WWTP contribute to the enrichment of the resistome in the receiving water bodies with the particular impact being dependent on the effluent load and local hydrological conditions. The ARGs concentrations in WWTP effluents were found to be inversely correlated to the number of implemented biological treatment steps, indicating a possible option for WWTP management. Furthermore, this study has identified blaOXA-58 as a possible resistance gene for future studies investigating the impact of WWTPs on their receiving water.

Uptake and bioaccumulation of three widely prescribed pharmaceutically active compounds in tomato fruits and mediated effects on fruit quality attributes.

Pharmaceutically active compounds (PhACs) released in agroecosystems have been found to be taken up by and accumulated in the edible parts of crop plants. By employing simulated hydroponic cultivation under controlled conditions, the present study aimed at evaluating 1) the uptake and bioaccumulation of three common PhACs (diclofenac, DCF; sulfamethoxazole, SMX; trimethoprim, TMP), either applied individually (10 µg L-1) or as mixture (10 µg L-1 each), in tomato fruits harvested from the first three fruit sets, and 2) the PhACs-mediated effects on fruit quality attributes. DCF was not detected in tomato fruits, whereas both SMX and TMP were detected in varying concentrations in fruits, depending on the time of harvest, the mode of application and the allocation of plants in the greenhouse. The studied PhACs applied at environmentally relevant concentrations did not significantly affect plant productivity. Nevertheless, important fruit quality attributes, such as soluble solids and carbohydrate (fructose, glucose, sucrose, total sugars) content were significantly impacted by all studied PhACs applied individually, suggesting that for DCF, potentially its transformation products (TPs) (not examined in this study) may exert significant effects on fruits quality attributes. In addition, no additive or synergistic effects of the mixture of PhACs on studied fruits quality attributes were revealed. Gene expression analysis showed that the PhACs-mediated effects on the carbohydrate content of fruits can be attributed, at least to some extent, to the significant modulation of the abundance of transcripts related to the biosynthesis and catabolism of sucrose, such as SlSuSys, SlLin5 and SlLin7. To our knowledge, this is the first report highlighting the potential effects of PhACs released in agroecosystems on the quality of widely consumed agricultural products. In any case, further studies are warranted for the overall assessment of the potential impacts of PhACs on the quality of agricultural products under conventional agricultural conditions.

Performance of secondary wastewater treatment methods for the removal of contaminants of emerging concern implicated in crop uptake and antibiotic resistance spread: A review.

Contaminants of emerging concern (CEC) discharged in effluents of wastewater treatment plants (WWTPs), not specifically designed for their removal, pose serious hazards to human health and ecosystems. Their impact is of particular relevance to wastewater disposal and re-use in agricultural settings due to CEC uptake and accumulation in food crops and consequent diffusion into the food-chain. This is the reason why the chemical CEC discussed in this review have been selected considering, besides recalcitrance, frequency of detection and entity of potential hazards, their relevance for crop uptake. Antibiotic-resistant bacteria (ARB) and antibiotic resistance genes (ARGs) have been included as microbial CEC because of the potential of secondary wastewater treatment to offer conditions favourable to the survival and proliferation of ARB, and dissemination of ARGs. Given the adverse effects of chemical and microbial CEC, their removal is being considered as an additional design criterion, which highlights the necessity of upgrading conventional WWTPs with more effective technologies. In this review, the performance of currently applied biological treatment methods for secondary treatment is analysed. To this end, technological solutions including conventional activated sludge (CAS), membrane bioreactors (MBRs), moving bed biofilm reactors (MBBRs), and nature-based solutions such as constructed wetlands (CWs) are compared for the achievable removal efficiencies of the selected CEC and their potential of acting as reservoirs of ARB&ARGs. With the aim of giving a picture of real systems, this review focuses on data from full-scale and pilot-scale plants treating real urban wastewater. To achieve an integrated assessment, technologies are compared considering also other relevant evaluation parameters such as investment and management costs, complexity of layout and management, present scale of application and need of a post-treatment. Comparison results allow the definition of design and operation strategies for the implementation of CEC removal in WWTPs, when agricultural reuse of effluents is planned.

Consolidated vs new advanced treatment methods for the removal of contaminants of emerging concern from urban wastewater.

Urban wastewater treatment plants (WWTPs) are among the main anthropogenic sources for the release of contaminants of emerging concern (CECs) into the environment, which can result in toxic and adverse effects on aquatic organisms and consequently on humans. Unfortunately, WWTPs are not designed to remove CECs and secondary (e.g., conventional activated sludge process, CAS) and tertiary (such as filtration and disinfection) treatments are not effective in the removal of most CECs entering WWTP. Accordingly, several advanced treatment methods have been investigated for the removal of CECs from wastewater, including consolidated (namely, activated carbon (AC) adsorption, ozonation and membranes) and new (such as advanced oxidation processes (AOPs)) processes/technologies. This review paper gathers the efforts of a group of international experts, members of the NEREUS COST Action ES1403 who for three years have been constructively discussing the state of the art and the best available technologies for the advanced treatment of urban wastewater. In particular, this work critically reviews the papers available in scientific literature on consolidated (ozonation, AC and membranes) and new advanced treatment methods (mainly AOPs) to analyse: (i) their efficiency in the removal of CECs from wastewater, (ii) advantages and drawbacks, (iii) possible obstacles to the application of AOPs, (iv) technological limitations and mid to long-term perspectives for the application of heterogeneous processes, and (v) a technical and economic comparison among the different processes/technologies.

Long-term wastewater irrigation of vegetables in real agricultural systems: Concentration of pharmaceuticals in soil, uptake and bioaccumulation in tomato fruits and human health risk assessment.

Wastewater (WW) reuse for vegetable crops irrigation is regularly applied worldwide. Such a practice has been found to allow the uptake of pharmaceutical active compounds (PhACs) by plants and their subsequent entrance to the food web, representing an important alternative pathway for the exposure of humans to PhACs, with potential health implications. Herein we report the impacts of the long-term (three consecutive years) WW irrigation of a tomato crop with two differently treated effluents under real agricultural conditions, on (1) the soil concentration of selected PhACs (i.e. diclofenac, DCF; sulfamethoxazole, SMX; trimethoprim, TMP), (2) the bioaccumulation of these PhACs in tomato fruits, and (3) the human risks associated with the consumption of WW-irrigated fruits. Results revealed that the concentration of the studied PhACs in both the soil and tomato fruits varied depending on the qualitative characteristics of the treated effluent applied and the duration of WW irrigation. The PhAC with the highest soil concentration throughout the studied period was SMX (0.98 µg kg-1), followed by TMP (0.62 µg kg-1) and DCF (0.35 µg kg-1). DCF was not found in tomato fruits harvested from WW-irrigated plants during the first year of the study. However, DCF displayed the highest fruit concentration (11.63 µg kg-1) throughout the study (as a result of prolonged WW irrigation), followed by SMX (5.26 µg kg-1) and TMP (3.40 µg kg-1). The calculated fruit bioconcentration factors (BCFF) were extremely high for DCF in the 2nd (108) and 3rd year (132) of the experimental period, with the respective values for SMX (0.5-5.4) and TMP (0.2-6.4) being significantly lower. The estimated threshold of toxicity concern (TTC) and hazard quotients (HQ) values revealed that the consumption of fruits harvested from tomato plants irrigated for long period with the WW applied for irrigation under field conditions in this study represent a de minimis risk to human health. However, more studies need to be performed in order to obtain more solid information on the safety of WW reuse for irrigation.

Reduction of clarithromycin and sulfamethoxazole-resistant Enterococcus by pilot-scale solar-driven Fenton oxidation.

The presence of pathogenic antibiotic-resistant bacteria in aquatic environments has become a health threat in the last few years. Their presence has increased due to the presence of antibiotics in wastewater effluents, which are not efficiently removed by conventional wastewater treatments. As a result there is a need to study the possible ways of removal of the mixtures of antibiotics present in wastewater effluents and the antibiotic-resistant bacteria, which may also spread the antibiotic resistance genes to other bacterial populations. In this study the degradation of a mixture of antibiotics i.e. sulfamethoxazole and clarithromycin, the disinfection of total enterococci and the removal of those resistant to: a) sulfamethoxazole, b) clarithromycin and c) to both antibiotics have been examined, along with the toxicity of the whole effluent mixture after treatment to the luminescent aquatic bacterium Vibrio fischeri. Solar Fenton treatment (natural solar driven oxidation) using Fenton reagent doses of 50 mg L(-1) of hydrogen peroxide and 5 mg L(-1) of Fe(3+) in a pilot-scale compound parabolic collector plant was used to examine the disinfection and antibiotic resistance removal efficiency in different aqueous matrices, namely distilled water, simulated and real wastewater effluents. There was a faster complete removal of enterococci and of antibiotics in all aqueous matrices by applying solar Fenton when compared to photolytic treatment of the matrices. Sulfamethoxazole was more efficiently degraded than clarithromycin in all three aqueous matrices (95% removal of sulfamethoxazole and 70% removal of clarithromycin in real wastewater). The antibiotic resistance of enterococci towards both antibiotics exhibited a 5-log reduction with solar Fenton in real wastewater effluent. Also after solar Fenton treatment, there were 10 times more antibiotic-resistant enterococci in the presence of sulfamethoxazole than in the presence of clarithromycin. Finally, the toxicity of the treated wastewater to V. fischeri remained very low throughout the treatment time.