Every fifth published metagenome is not available to science.

Have you ever sought to use metagenomic DNA sequences reported in scientific publications? Were you successful? Here, we reveal that metagenomes from no fewer than 20% of the papers found in our literature search, published between 2016 and 2019, were not deposited in a repository or were simply inaccessible. The proportion of inaccessible data within the literature has been increasing year-on-year. Noncompliance with Open Data is best predicted by the scientific discipline of the journal. The number of citations, journal type (e.g., Open Access or subscription journals), and publisher are not good predictors of data accessibility. However, many publications in high-impact factor journals do display a higher likelihood of accessible metagenomic data sets. Twenty-first century science demands compliance with the ethical standard of data sharing of metagenomes and DNA sequence data more broadly. Data accessibility must become one of the routine and mandatory components of manuscript submissions-a requirement that should be applicable across the increasing number of disciplines using metagenomics. Compliance must be ensured and reinforced by funders, publishers, editors, reviewers, and, ultimately, the authors.

Toward a Universal Unit for Quantification of Antibiotic Resistance Genes in Environmental Samples.

Surveillance of antibiotic resistance genes (ARGs) has been increasingly conducted in environmental sectors to complement the surveys in human and animal sectors under the "One-Health" framework. However, there are substantial challenges in comparing and synthesizing the results of multiple studies that employ different test methods and approaches in bioinformatic analysis. In this article, we consider the commonly used quantification units (ARG copy per cell, ARG copy per genome, ARG density, ARG copy per 16S rRNA gene, RPKM, coverage, PPM, etc.) for profiling ARGs and suggest a universal unit (ARG copy per cell) for reporting such biological measurements of samples and improving the comparability of different surveillance efforts.

Ranking of crop plants according to their potential to uptake and accumulate contaminants of emerging concern.

The reuse of treated wastewater (TWW) for irrigation and the use of biosolids and manures as soil amendment constitute significant pathways for the introduction of the contaminants of emerging concern (CECs) to the agricultural environment. Consequently, CECs are routinely detected in TWW-irrigated agricultural soils and runoff from such sites, in biosolids- and manure-amended soils, and in surface and groundwater systems and sediments receiving TWW. Crop plants grown in such contaminated agricultural environments have been found to uptake and accumulate CECs in their tissues, constituting possible vectors of introducing CECs into the food chain; an issue that is presently considered of high priority, thus needing intensive investigation. This review paper aims at highlighting the responsible mechanisms for the uptake of CECs by plants and the ability of each crop plant species to uptake and accumulate CECs in its edible tissues, thus providing tools for mitigating the introduction of these contaminants into the food chain. Both biotic (e.g. plants' genotype and physiological state, soil fauna) and abiotic factors (e.g. soil pore water chemistry, physico-chemical properties of CECs, environmental perturbations) have been proven to influence the ability of crop plants to uptake and accumulate CECs. According to authors' estimates, based on the thorough elaboration of knowledge produced by existing relevant studies, the ability of crop plants to uptake and accumulate CECs decrease in the order of leafy vegetables > root vegetables > cereals and fodder crops > fruit vegetables; though, the uptake of CECs by important crop plants, such as fruit trees, is not yet evaluated. Overall, further studies must be performed to estimate the potential of crop plants to uptake and accumulate CECs in their edible tissues, and to characterize the risk for human health represented by their presence in human and livestock food products.

Novel approach to fast determination of cholesterol oxidation products in Cypriot foodstuffs using ultra-performance liquid chromatography-tandem mass spectrometry.

This paper reports the development and validation of a new method based on ultra-performance LC coupled to MS/MS for the simultaneous determination of four cholesterol oxidation products (COPs) in foodstuffs in only 4.1 min. The COPs were detected by ESI in positive-ion mode with multiple reaction monitoring, and the mass spectrometric conditions were optimized in order to increase sensitivity. The developed method was validated in terms of linearity, precision, LODs, and LOQs. Recoveries of the extraction process ranged from 86 to 98.5% when the samples were fortified at 100, 500, and 1500 ng/mL. The applicability of the method was confirmed by analyzing different food samples. Considering the paucity of data regarding the content of COPs in Cypriot foods, particular attention was devoted, for the first time, to the determination of the profile of the main COPs in widely consumed, traditional Cypriot foodstuffs (halloumi cheese, hiromeri, snails, etc.).

High Throughput Analysis of Integron Gene Cassettes in Wastewater Environments.

Integrons are extensively targeted as a proxy for anthropogenic impact in the environment. We developed a novel high-throughput amplicon sequencing pipeline that enables characterization of thousands of integron gene cassette-associated reads, and applied it to acquire a comprehensive overview of gene cassette composition in effluents from wastewater treatment facilities across Europe. Between 38 100 and 172 995 reads per-sample were generated and functionally characterized by screening against nr, SEED, ARDB and ß-lactamase databases. Over 75% of the reads were characterized as hypothetical, but thousands were associated with toxin-antitoxin systems, DNA repair, cell membrane function, detoxification and aminoglycoside and ß-lactam resistance. Among the reads characterized as ß-lactamases, the carbapenemase blaOXA was dominant in most of the effluents, except for Cyprus and Israel where blaGES was also abundant. Quantitative PCR assessment of blaOXA and blaGES genes in the European effluents revealed similar trends to those displayed in the integron amplicon sequencing pipeline described above, corroborating the robustness of this method and suggesting that these integron-associated genes may be excellent targets for source tracking of effluents in downstream environments. Further application of the above analyses revealed several order-of-magnitude reductions in effluent-associated ß-lactamase genes in effluent-saturated soils, suggesting marginal persistence in the soil microbiome.

Antibiotic resistance in urban aquatic environments: can it be controlled?

Over the last decade, numerous evidences have contributed to establish a link between the natural and human-impacted environments and the growing public health threat that is the antimicrobial resistance. In the environment, in particular in areas subjected to strong anthropogenic pressures, water plays a major role on the transformation and transport of contaminants including antibiotic residues, antibiotic-resistant bacteria, and antibiotic resistance genes. Therefore, the urban water cycle, comprising water abstraction, disinfection, and distribution for human consumption, and the collection, treatment, and delivery of wastewater to the environment, is a particularly interesting loop to track the fate of antibiotic resistance in the environment and to assess the risks of its transmission back to humans. In this article, the relevance of different transepts of the urban water cycle on the potential enrichment and spread of antibiotic resistance is reviewed. According to this analysis, some gaps of knowledge, research needs, and control measures are suggested. The critical rationale behind the measures suggested and the desirable involvement of some key action players is also discussed.

Ultraviolet-activated persulfate oxidation of methyl orange: a comparison between artificial neural networks and factorial design for process modelling.

In this work, the degradation of the azo dye methyl orange in model aqueous solutions by UVC light-induced persulfate oxidation was studied. Five operating parameters that may influence the decolorization kinetics were evaluated, namely, methyl orange (MO) (5-50 mg L(-1)) and sodium persulfate (SPS) (50-150 mg L(-1)) concentration, reaction time (up to 60 min), (un-buffered) solution pH (3-9) and the addition of NaCl (0-500 mg L(-1)). The process was simulated, applying and comparing two methodologies, namely two-level factorial design and an artificial neural network (ANN). It was found that MO concentration is the most influential parameter, followed by the reaction time and SPS concentration, while the effects of solution pH and the addition of sodium chloride are statistically less significant; this order of significance was predicted by both methodologies. The ANN can simulate the process more accurately (i.e. in terms of R(2), mean square error (MSE) and residuals) than factorial design, although it needs significantly larger sets of data and longer computational time.

Assessment of long-term wastewater irrigation impacts on the soil geochemical properties and the bioaccumulation of heavy metals to the agricultural products.

An extensive field survey was employed for assessing the impacts of long-term wastewater irrigation of forage crops and orange orchards in three suburban agricultural areas in Cyprus (areas I, II, and III), as compared to rainfed agriculture, on the soil geochemical properties and the bioaccumulation of heavy metals (Zn, Ni, Mn, Cu, Co) to the agricultural products. Both ryegrass fields and orange orchards in areas I and II were continuously wastewater irrigated for 10 years, whereas clover fields in area III for 0.5, 4, and 8 years. The results revealed that wastewater reuse for irrigation caused a slight increase in soil salinity and Cl(-) content in areas I and II, and a remarkable increase, having strong correlation with the period in which wastewater irrigation was practiced, in area III. Soil salinization in area III was due to the high electrical conductivity (EC) of the wastewater applied for irrigation, attributed to the influx of seawater to the sewage collection network in area III. In addition, the wastewater irrigation practice resulted in a slight decrease of the soil pH values in area III, while a subtle impact was identified regarding the CaCO3, Fe, and heavy metal content in the three areas surveyed. The heavy metal content quantified in the forage plants' above-ground parts was below the critical levels of phytotoxicity and the maximum acceptable concentration in dairy feed, whereas heavy metals quantified in orange fruit pulp were below the maximum permissible levels (MPLs). Heavy metal phytoavailability was confined due to soil properties (high pH and clay content), as evidenced by the calculated low transfer factor (TF).

Linking the use of reclaimed water to indicators of crop stress by metabolomic and transcriptomic analyses. A tool to compare water irrigation quality.

The occurrence of contaminants of emerging concern (CECs) or heavy metals in reclaimed water used for agricultural irrigation may affect crop morphology and physiology. Here, we analyzed lettuce (Lactuca sativa) grown in outdoor lysimeters and irrigated with either tap water, used as a control, or reclaimed water: CAS-reclaimed water, an effluent from a conventional activated sludge system (CAS) followed by chlorination and sand filtration, or MBR-reclaimed water, an effluent from a membrane biological reactor (MBR). Chemical analyses identified seven CECs in the reclaimed waters, but only two of them were detected in lettuce (carbamazepine and azithromycin). Metabolomic and transcriptomic analyses revealed that irrigation with reclaimed water increased the concentrations of several crop metabolites (5-oxoproline, leucine, isoleucine, and fumarate) and of transcripts codifying for the plant stress-related genes Heat-Shock Protein 70 (HSP70) and Manganese Superoxide Dismutase (MnSOD). In both cases, MBR-water elicited the strongest response in lettuce, perhaps related to its comparatively high sodium adsorption ratio (4.5), rather than to its content in CECs or heavy metals. Our study indicates that crop metabolomic and transcriptomic profiles depend on the composition of irrigating water and that they could be used for testing the impact of water quality in agriculture.

Fate and transport modelling for evaluating antibiotic resistance in aquatic environments: Current knowledge and research priorities.

Antibiotics have revolutionised medicine in the last century and enabled the prevention of bacterial infections that were previously deemed untreatable. However, in parallel, bacteria have increasingly developed resistance to antibiotics through various mechanisms. When resistant bacteria find their way into terrestrial and aquatic environments, animal and human exposures increase, e.g., via polluted soil, food, and water, and health risks multiply. Understanding the fate and transport of antibiotic resistant bacteria (ARB) and the transfer mechanisms of antibiotic resistance genes (ARGs) in aquatic environments is critical for evaluating and mitigating the risks of resistant-induced infections. The conceptual understanding of sources and pathways of antibiotics, ARB, and ARGs from society to the water environments is essential for setting the scene and developing an appropriate framework for modelling. Various factors and processes associated with hydrology, ecology, and climate change can significantly affect the fate and transport of ARB and ARGs in natural environments. This article reviews current knowledge, research gaps, and priorities for developing water quality models to assess the fate and transport of ARB and ARGs. The paper also provides inputs on future research needs, especially the need for new predictive models to guide risk assessment on AR transmission and spread in aquatic environments.

Making waves: The NORMAN antibiotic resistant bacteria and resistance genes database (NORMAN ARB&ARG)-An invitation for collaboration to tackle antibiotic resistance.

With the global concerns on antibiotic resistance (AR) as a public health issue, it is pivotal to have data exchange platforms for studies on antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARGs) in the environment. For this purpose, the NORMAN Association is hosting the NORMAN ARB&ARG database, which was developed within the European project ANSWER. The present article provides an overview on the database functionalities, the extraction and the contribution of data to the database. In this study, AR data from three studies from China and Nepal were extracted and imported into the NORMAN ARB&ARG in addition to the existing AR data from 11 studies (mainly European studies) on the database. This feasibility study demonstrates how the scientific community can share their data on AR to generate an international evidence base to inform AR mitigation strategies. The open and FAIR data are of high potential relevance for regulatory applications, including the development of emission limit values / environmental quality standards in relation to AR. The growth in sharing of data and analytical methods will foster collaboration on risk management of AR worldwide, and facilitate the harmonization in the effort for identification and surveillance of critical hotspots of AR. The NORMAN ARB&ARG database is publicly available at: https://www.norman-network.com/nds/bacteria/.

Workflow to facilitate the detection of new psychoactive substances and drugs of abuse in influent urban wastewater.

The complexity around the dynamic markets for new psychoactive substances (NPS) forces researchers to develop and apply innovative analytical strategies to detect and identify them in influent urban wastewater. In this work a comprehensive suspect screening workflow following liquid chromatography - high resolution mass spectrometry analysis was established utilising the open-source InSpectra data processing platform and the HighResNPS library. In total, 278 urban influent wastewater samples from 47 sites in 16 countries were collected to investigate the presence of NPS and other drugs of abuse. A total of 50 compounds were detected in samples from at least one site. Most compounds found were prescription drugs such as gabapentin (detection frequency 79%), codeine (40%) and pregabalin (15%). However, cocaine was the most found illicit drug (83%), in all countries where samples were collected apart from the Republic of Korea and China. Eight NPS were also identified with this protocol: 3-methylmethcathinone 11%), eutylone (6%), etizolam (2%), 3-chloromethcathinone (4%), mitragynine (6%), phenibut (2%), 25I-NBOH (2%) and trimethoxyamphetamine (2%). The latter three have not previously been reported in municipal wastewater samples. The workflow employed allowed the prioritisation of features to be further investigated, reducing processing time and gaining in confidence in their identification.

Sunlight, iron and radicals to tackle the resistant leftovers of biotreated winery wastewater.

Winery wastewater is characterized by high organic content consisting of alcohols, acids and recalcitrant high-molecular-weight compounds (e.g. polyphenols, tannins and lignins). So far, biological treatment constitutes the best available technology for such effluents that are characterized by high seasonal variability; however the strict legislation applied on the reclamation and reuse of wastewaters for irrigation purposes introduces the need for further treatment of the bioresistant fraction of winery effluents. In this context, the use of alternative treatment technologies, aiming to mineralize or transform refractory molecules into others which could be further biodegraded, is a matter of great concern. In this study, a winery effluent that had already been treated in a sequencing batch reactor was subjected to further purification by homogeneous and heterogeneous solar Fenton oxidation processes. The effect of various operating variables such as catalyst and oxidant concentration, initial pH, temperature and lamp power on the abatement of chemical oxygen demand (COD), dissolved organic carbon (DOC), color, total phenolics and ecotoxicity has been assessed in the homogeneous solar Fenton process. In addition, a comparative assessment between homogeneous and heterogeneous solar Fenton processes was performed. In the present study the homogeneous solar Fenton process has been demonstrated to be the most effective process, yielding COD, DOC and total phenolics removal of about 69%, 48% and 71% in 120 min of the photocatalytic treatment, respectively.

Looking into the effects of co-contamination by micro(nano)plastics in the presence of other pollutants on irrigated edible plants.

To address water shortage challenges, treated wastewater is used to meet the demand for irrigation water in several countries worldwide. Considering the presence of pollutants in treated wastewater, its use for land irrigation might have an impact in the environment. This review article focuses on the combined effects (or potential joint toxicity) of microplastics (MPs)/nanoplastics (NPs) and other environmental contaminants present in treated wastewater on edible plants after irrigation. Initially, the concentrations of MPs/NPs in wastewater treatment plant effluents and surface waters are summarized, indicating the presence of MPs/NPs in both water matrices (i.e., wastewater after receiving treatment and lakes/rivers). Then, the results of 19 studies related to joint toxicity of MPs/NPs and co-contaminants (e.g., heavy metals and pharmaceuticals) on edible plants, are reviewed and discussed. This concurrent presence may result in several combined effects on edible plants, e.g., rapid root growth, increase in antioxidant enzymes, decrease in photosynthetic rate and increased production of ROS. These effects, as discussed in the various studies on which this review is based, can generate antagonistic or even neutral impact on plants, depending on the size of MPs/NPs and their mixing ratio with the co-contaminants. However, a combined exposure of edible plants to MPs/NPs and co-contaminants can also lead to hormetic adaptive responses. The data reviewed and discussed herein may relieve overlooked environmental impacts of treated wastewater reuse and may be useful to address challenges related to the combined effects of MPs/NPs and co-contaminants on edible plants after irrigation. The conclusions drawn in this review article are relevant to both direct (i.e., treated wastewater irrigation) and indirect (i.e., discharging treated wastewater in surface waters used for irrigation purposes) reuse, and may contribute to the implementation of the European Regulation 2020/741 on the minimum requirements for water reuse.

Mitigating risks and maximizing sustainability of treated wastewater reuse for irrigation.

Scarcity of freshwater for agriculture has led to increased utilization of treated wastewater (TWW), establishing it as a significant and reliable source of irrigation water. However, years of research indicate that if not managed adequately, TWW may deleteriously affect soil functioning and plant productivity, and pose a hazard to human and environmental health. This review leverages the experience of researchers, stakeholders, and policymakers from Israel, the United-States, and Europe to present a holistic, multidisciplinary perspective on maximizing the benefits from municipal TWW use for irrigation. We specifically draw on the extensive knowledge gained in Israel, a world leader in agricultural TWW implementation. The first two sections of the work set the foundation for understanding current challenges involved with the use of TWW, detailing known and emerging agronomic and environmental issues (such as salinity and phytotoxicity) and public health risks (such as contaminants of emerging concern and pathogens). The work then presents solutions to address these challenges, including technological and agronomic management-based solutions as well as source control policies. The concluding section presents suggestions for the path forward, emphasizing the importance of improving links between research and policy, and better outreach to the public and agricultural practitioners. We use this platform as a call for action, to form a global harmonized data system that will centralize scientific findings on agronomic, environmental and public health effects of TWW irrigation. Insights from such global collaboration will help to mitigate risks, and facilitate more sustainable use of TWW for food production in the future.

Three years of wastewater surveillance for new psychoactive substances from 16 countries.

The proliferation of new psychoactive substances (NPS) over recent years has made their surveillance complex. The analysis of raw municipal influent wastewater can allow a broader insight into community consumption patterns of NPS. This study examines data from an international wastewater surveillance program that collected and analysed influent wastewater samples from up to 47 sites in 16 countries between 2019 and 2022. Influent wastewater samples were collected over the New Year period and analysed using validated liquid chromatography - mass spectrometry methods. Over the three years, a total of 18 NPS were found in at least one site. Synthetic cathinones were the most found class followed by phenethylamines and designer benzodiazepines. Furthermore, two ketamine analogues, one plant based NPS (mitragynine) and methiopropamine were also quantified across the three years. This work demonstrates that NPS are used across different continents and countries with the use of some more evident in particular regions. For example, mitragynine has highest mass loads in sites in the United States, while eutylone and 3-methylmethcathinone increased considerably in New Zealand and in several European countries, respectively. Moreover, 2F-deschloroketamine, an analogue of ketamine, has emerged more recently and could be quantified in several sites, including one in China, where it is considered as one of the drugs of most concern. Finally, some NPS were detected in specific regions during the initial sampling campaigns and spread to additional sites by the third campaign. Hence, wastewater surveillance can provide an insight into temporal and spatial trends of NPS use.

A study on the attitudes and behavioural influence of construction waste management in occupied Palestinian territory.

As a step towards comprehending what drives the management of construction waste in the occupied Palestinian territory, this paper quantifies construction waste generation and examines how the local contractors' waste management attitudes and behaviour are influenced. Collection of data was based on a survey, carried out in the southern part of the West Bank between April and May 2010. The survey targeted contractors who specialized in the construction of buildings. A logistic regression model was used to investigate the relationship between various attributes and the attitudes and behaviour that the local contractors demonstrate towards waste management. The results showed that during the construction of buildings, 17 to 81 kg of construction waste are generated per square metre of building floor. Although the area of a building is the key factor determining 74.8% of the variation of construction waste generation, the employment of labour-intensive techniques in the study area means that human factors such as the contractor's attitude and behaviour towards waste management, exert a key influence on waste generation. Attitudes towards the 3Rs of waste minimization and behaviour towards waste disposal are generally positive with smaller contractors exhibiting more positive attitudes and more satisfactory behaviour towards waste management. Overall, while contractors' behaviour towards waste sorting and disposal tends to be more satisfactory among contractors who are more conscious about the potential environmental impacts of construction waste, it was generally observed that in the absence of a regulatory framework, the voluntary attitudes and behaviour among the local contractors are mostly driven by direct economic considerations.

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.

Investigation of the effect of microplastics on the UV inactivation of antibiotic-resistant bacteria in water.

This study investigated the effect of polyethylene and polyvinyl chloride microplastics on the UV fluence response curve for the inactivation of multidrug-resistant E. coli and enterococci in ultrapure water at pH 6.0 ± 0.1. In the absence of microplastics, the UV inactivation of the studied bacteria exhibited an initial resistance followed by a faster inactivation of free (dispersed) bacteria, while in the presence of microplastics, these 2 regimes were followed by an additional regime of slower or no inactivation related to microplastic-associated bacteria (i.e., bacteria aggregated with microplastics resulting in shielding bacteria from UV indicated by tailing at higher UV fluences). The magnitude of the negative effect of microplastics varied with different microplastics (type/particle size) and bacteria (Gram-negative and Gram-positive). Results showed that when the UV transmittance of the microplastic-containing water was not taken into account in calculating UV fluences, the effect of microplastics as protectors of bacteria was overestimated. A UV fluence-based double-exponential microbial inactivation model accounting for both free and microplastic-associated bacteria could describe well the disinfection data. The present study elucidated the effect of microplastics on the performance of UV disinfection, and the approach used herein to prove this concept may guide future research on the investigation of the possible effect of other particles including nanoplastics with different characteristics on the exposure response curve for the inactivation of various microorganisms by physical and chemical disinfection processes in different water and wastewater matrices.