The rapid emergence and dissemination of plasmid-mediated mcr genes have raised significant public health concerns. Given the spread of colistin resistance across the natural environment, humans, and animals, the water environment serves as a crucial connecting medium. To address critical gaps in the literature on the dissemination of mcr variants in environmental samples, we prepared the first report on the prevalence of mcr genes in wastewater and river water samples in Poland. We designed the study at the river-basin scale to provide comprehensive insight into colistin resistance mediated by mobile mcr genes in the aquatic environment. Moreover, this is the first study to examine seasonal variation in mcr genes, conducted simultaneously on wastewater and river water samples. The research involved collecting wastewater from 17 wastewater treatment plants (including facilities that receive hospital wastewater) and river water at 8 points along the Pilica River across four seasons. We demonstrated that mcr-1, mcr-4, and mcr-5 were detected in all samples, both wastewater and river water, with mcr-5 the most abundant. The wastewater treatment process was completely ineffective at eliminating mcr-5 genes. In addition, significant differences in the prevalence of mcr-1, mcr-4, and mcr-5 were observed between warmer and colder seasons, indicating a seasonal effect on the dissemination of colistin resistance genes in environmental samples. The present findings help clarify the role of wastewater and rivers in the spread of last-resort drug resistance genes and their significance for public health.

Due to increasing antimicrobial resistance threatening social health stability, the World Health Organization classified anti-infectives into three groups, Access, Watch, and Reserve (AWaRe). The reserve list of antimicrobials, which are mainly used in hospitals as a last-resort option for the most severe infections, includes the carbapenems with enzyme inhibitors selected for this study. The purpose of this paper is to present the results of a comprehensive study of the prevalence of meropenem, cilastatin, and vaborbactam in the wastewater from 64 hospitals collected during two sampling seasons. Pharmaceuticals were present in the wastewater of more than 90% of the hospitals, in concentrations ranging between 1.20 and 17,315 ng/L. The findings of the statistical analyses revealed relationships between meropenem concentrations, annual intake, and hospital characteristics. A link was also visible between cilastatin wastewater levels and yearly imipenem consumption. Additionally, the impact of in situ chlorination was assessed, proving the ineffectiveness of the process regarding the possible removal of analytes. Preliminary calculations of the environmental risk assessment revealed a negligible impact of residues. A different outcome was seen for the risk evaluation for antimicrobial resistance selection. In more than half of the samples in which meropenem was determined, risk quotients reflected medium or high potential for development of antimicrobial resistance. Our results confirm the need for broader, systematic research on last-chance antimicrobials and show possibility of discovering more connections at the initial point of environmental introduction, which could help implement solutions to mitigate further spread of drug resistance.

Plastispheres, microbial biofilms formed on plastic surfaces, are increasingly recognised as ecological niches capable of transporting pollutants and antibiotic-resistant microorganisms. However, mechanistic insights into antimicrobial resistance (AMR) dynamics in natural plastispheres remain limited, particularly for priority pathogens such as carbapenem-resistant Enterobacterales (CRE). Here, we evaluated plastispheres as environmental reservoirs and vectors of carbapenem-resistant bacteria, comparing wastewater (secondary settling tanks, representing the final stage before environmental discharge) and riverine environments. Using a combined SEM-CFM approach, we resolved plastic surface topography and the spatial organisation of biofilm-associated bacteria. Although CRE were not detected, carbapenem-resistant bacteria constituted a stable fraction of heterotrophic communities in both environments and were primarily associated with intrinsic resistance mechanisms. Carbapenem-resistant isolates included Aeromonas spp. (blaCphA), Stenotrophomonas maltophilia (blaL1), and Pseudomonas putida (efflux-based resistance). Microscopy revealed dense bacterial clusters on plastic surfaces, suggesting microenvironments that may facilitate cell-cell interactions, including horizontal gene transfer. These findings highlight plastispheres not only as vectors of AMR but also as potential evolutionary hotspots shaping resistance persistence and dissemination in aquatic systems. Future integrating metagenomic and genomic data on resistance gene mobility with spatially resolved microbial community structure will provide critical insights into the mechanisms and risks of AMR dissemination in plastisphere environments.

Carbapenem-resistant Pseudomonas aeruginosa (CRPA) and Acinetobacter baumannii (CRAB) represent a major clinical and epidemiological challenge and pose a growing threat to public health and the environment. Accordingly, CRPA and CRAB were investigated in hospital wastewater (HWW) collected during winter and summer 2024 from 64 healthcare facilities across all 16 Polish voivodeships. To our knowledge, this study constitutes the first nationwide, large-scale assessment in Poland of carbapenem resistance in these high-risk pathogens in hospital wastewater. The study aimed to determine the prevalence of carbapenem-resistant bacteria (CRB) in HWW discharged into the public sewer system and municipal wastewater treatment plants (WWTPs). In addition, associations between CRB prevalence, hospital geographic location, and sampling season were analyzed to identify spatial and temporal patterns of carbapenem resistance (CR). Carbapenem-resistant P. aeruginosa were predominant in all studied regions. Carbapenem-resistant A. baumannii were identified in a smaller percentage of samples and were characterized by greater genotypic diversity. The ERIC-PCR assay confirmed the presence of both closely related strains and unique genetic profiles, which suggests that CRB emissions into the environment have a complex character. The statistical analysis revealed significant relationships between CRB counts, the physicochemical parameters of HWW, and antibiotic concentrations in HWW samples. In addition, the tested samples harbored many antibiotic resistance genes (ARGs), which confirms that HWW is a significant reservoir of mobile genetic elements (MGEs) involved in the spread of antibiotic resistance. The results of the study indicate that HWW should be rigorously monitored and managed to minimize risks to public health and environment.

Municipal wastewater treatment plants (WWTPs) are crucial for protecting the environment and public health, yet the discharge of treated wastewater can influence the biodiversity of aquatic microbial communities. Enterobacterales are reliable indicators of sanitary risk. Contamination with Enterobacterales often reflects wastewater treatment inefficiency, and pathogenic strains such as E. coli, Klebsiella pneumoniae, and Enterobacter pose significant public health threats. This study assessed bacterial diversity in the wastewater treatment process and evaluated how treated wastewater affects the microbiome of the Pilica River. Its added value lies in the use of an integrated catchment-scale approach, involving an analysis of the Pilica River from its source to its mouth (including eight sampling sites), all seasons, and inflows from 17 WWTPs. The abundance of Enterobacterales was strongly correlated with environmental factors, but not with pH. WWTP size influenced the relative abundance of ASVs of Yersinia, Escherichia-Shigella, and total Enterobacterales, while influent composition had no significant effect on microbial communities. Seasonal variations had the greatest impact on river microbiota, particularly Yersinia, Rahnella, and Providencia. Escherichia-Shigella dominated across wastewater and river samples, confirming its role as an indicator of water quality. The study demonstrated that treated wastewater can modify river microbiomes, thereby increasing sanitary and epidemiological risks.

Hospital wastewater (HWW) is a critical hotspot for the dissemination of antibiotic resistance genes (ARGs) and pathogens. This study provides the first comprehensive metagenomic characterization of HWW across Poland, analyzing 64 medical facilities across two seasons via Nanopore long-read sequencing (total of 128 HWW samples). The HWW microbiome was mostly dominated by Proteobacteria, Bacteroidota, and Firmicutes. Multivariate analysis confirmed a significant seasonal shift in the resistome. Winter samples exhibited geographic regionalization, with localized hotspots of specific ARGs, including vancomycin resistance (operon van) and carbapenemase genes (blaOXA, blaNDM). Conversely, summer samples showed a significant trend toward nationwide homogenization, characterized by a uniform distribution of ESBL genes (blaTEM, blaCTX-M) and multidrug resistance (MDR) determinants, alongside the persistence of localized clinical hotspots. Klebsiella pneumoniae emerged as a central network hub, particularly in summer, showing strong correlations with ESBLs. Quantitative genomic co-occurrence analysis revealed a functional division within dominant taxa: while environmental species like Acinetobacter johnsonii comprised the general background microbiome, clinical pathogens such as Acinetobacter baumannii served as primary vectors, showing frequent associations with high-risk ARGs. Environmental and opportunistic bacteria, such as Aeromonas spp. and Citrobacter spp., were identified as putative ‘bridge hosts’ associated with mobile resistance determinants and potentially contributing to HGT. The findings indicate that seasonal factors, such as increased temperature and sub-inhibitory antibiotic concentrations, may contribute to the transition from regionalized to homogenized resistance profiles, demonstrating that background resistome convergence can coexist with point-source clinical outbreaks. This seasonal "blurring" of regional boundaries positions HWW as an active vector for large-scale antimicrobial resistance (AMR) dissemination. These results underscore the urgent need for nationwide metagenomic surveillance and advanced wastewater treatment strategies within the "One Health" framework to mitigate the environmental spread of WHO priority pathogens.

Overuse and inadequate legal regulation of the use of antibiotics have led to antimicrobial resistance, which threatens healthcare by reducing the effectiveness of treatments and promoting multidrug-resistant "superbugs". The World Health Organization created the AWaRe (Access, Watch, Reserve) classification to categorize antimicrobials according to their therapeutic value, emphasizing prudent use. The most important group in terms of protecting effectiveness is the Reserve group. The aim of the study was to develop a method for the determination of the following Reserve antibiotics in hospital wastewater: aztreonam, linezolid, meropenem, tigecycline, vaborbactam, cilastatin, fosfomycin, and ceftazidime. Twenty-six extraction procedures were tested and the one with the best recoveries for all antimicrobials (31.5-103.7%) was selected. The extracts were analyzed by liquid chromatography-tandem mass spectrometry. The method was validated and applied to the determination of the selected pharmaceuticals in effluents from 16 hospitals in Poland, with Reserve antibiotics detected in each sample at concentrations ranging from 1.89 ng L- 1 to 22.49 µg L- 1. This is the first country-scale methodological study on Reserve antibiotics in hospital wastewater, emphasizing the need for early environmental monitoring and the potential integration of such strategies into healthcare systems to mitigate the global antimicrobial resistance crisis.

The aquatic environment is a major pathway for the spread of antibiotic resistance (AR) among microorganisms. Among these, Klebsiella pneumoniae reveals high genome plasticity, adaptability, and the ability to colonize humans, animals, and the natural environment, awarding it a significant role in the spread of AR. This work presents an in-depth analysis of the whole sequences of 149 K. pneumoniae genomes isolated from surface waters available in databases. The sequences were obtained from 20 countries in five continents. The analyses showed a high genomic diversity of isolates, classifying them into 94 unique sequence types. The isolates carried numerous virulence and drug resistance determinants in their genomes, including genes for carbapenem and colistin resistance. The critical resistance genes were located on plasmids, indicating their high mobility and ease of access in water environments. Sublineage 258 members, in particular ST11, have been identified as important carriers of both important drug resistance determinants and key virulence factors, thus posing a substantial threat to human health. Our analysis revealed the direct transmission of drug-resistant and virulent clinical strains to the natural environment, highlighting the role of K. pneumoniae in the dissemination of drug resistance within the "One Health" framework. Surface waters represent an environment conducive to the spread and evolution of drug resistance, and K. pneumoniae plays a significant role in this process by providing clinically-significant antibiotic resistance genes to environmental recipients.

Heavy metals can enter rivers i.a. from point sources such as wastewater treatment plants (WWTPs) and industrial plants with inadequately-treated wastewater. The present article examines (i) the impact of WWTPs and industrial facilities on the pollution of the Pilica River, (ii) heavy metal loads along its continuum, and (iii) the identification of common pollution patterns in wastewater and river water, and the effect of their physicochemical properties. A novel aspect of this study is the use of two distinct data calculation approaches: Variant 1 -values below the quantification limit (LOQ) were replaced with LOQ/2, Variant 2- LOQ replaced with zero. Analyses have shown that pollution scenarios differ depending on the calculation method used. Nevertheless, regardless of the calculation method, the general conclusions that can be drawn are: i) among WWTPs, the highest heavy metal concentrations are generated by the smallest plants, ii) the highest loads by the largest ones, which is related i.a. to the size of their flow, iii) industrial plants are significant sources of point pollution of rivers, with arsenic, tin, zinc, cobalt, copper, molybdenum, nickel, lead, chromium, mercury and barium being detected in wastewater. The most common heavy metal in the Pilica was found to be Barium, with the highest loads observed in winter (66.29-216.98 kg/day), and depending on the season, arsenic, copper and nickel were also detected. The innovative aspect of the work is an example of modification of the sedimentation-biofiltration system as a sustainable solution for removing heavy metals from treated wastewater.

Global population growth generates problems relating to increasing demand for sustainable energy and waste treatment. Proper solid waste management promotes material reuse, maximizes recovery and reduces anthropological pressure on natural resources. Anaerobic digestion (AD) is an alternative method of stabilizing organic substrates and generating biogas as a source of environmentally friendly energy. In addition, digestate is not only a waste product of that process but also a renewable resource with many potential applications. The circular economy concept encourages the use of digestate as a source of nutrients that promotes plant growth and improves soil properties. However, the stabilized substrates often contain various contaminants, including heavy metals (HMs) and antibiotics that are also detected in digestate. Therefore, the agricultural use of digestate obtained by AD could increase the pool of these pollutants in soil and water environments and contribute to their circulation in these ecosystems. Moreover, digestate may also increase the co-selection of genes determining resistance to HMs and antibiotics in environmental microorganisms. This article comprehensively reviews published data on the residues of various HMs and antimicrobial substances in different digestates around the world and maps the scope of the problem. Moreover, the potential risk of residual levels of these contaminants in digestate has also been evaluated. The review highlights the lack of legal standards regulating the concentrations of drugs introduced into the soil with digestate. The results of the ecological risk assessment indicate that the presence of medically important antimicrobials in digestate products, especially those used in agriculture, should be limited.

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Learn More CiteCitationCitation and abstractCitation and referencesMore citation options ShareShare onFacebookX (Twitter)WeChatLinkedInRedditEmailJump toExpandCollapse ViewpointJanuary 28, 2025Strengthening Policy Relevance of Wastewater-Based Surveillance for Antimicrobial ResistanceClick to copy article linkArticle link copied!Sheena Conforti*Sheena ConfortiEawag, Swiss Federal Institute of Aquatic Science and Technology, Dübendorf 8600, Switzerland*[email protected]More by Sheena ConfortiView Biographyhttps://orcid.org/0000-0002-0173-6170Amy PrudenAmy PrudenCivil and Environmental Engineering, Virginia Polytechnic Institute and State University, 418 Durham Hall, Blacksburg, Virginia 24061, United StatesMore by Amy Prudenhttps://orcid.org/0000-0002-3191-6244Nicole AcostaNicole AcostaUniversity of Calgary, Cumming School of Medicine, Calgary, AB T2N 1N4, CanadaMore by Nicole AcostaChristopher AndersonChristopher AndersonWest Virginia University, Morgantown, West Virginia 26506-6201, United StatesMore by Christopher AndersonHelmut BuergmannHelmut BuergmannEawag, Swiss Federal Institute of Aquatic Science and Technology, Kastenienbaum 6047, SwitzerlandMore by Helmut Buergmannhttps://orcid.org/0000-0002-5651-5906Juliana Calabria De AraujoJuliana Calabria De AraujoFederal University of Minas Gerais, Belo Horizonte, MG 31270-901, BrazilMore by Juliana Calabria De AraujoJudith R. CristobalJudith R. CristobalDepartment of Chemistry, University at Buffalo - The State University of New York, 633 Natural Science Complex, Buffalo, New York 14260, United StatesMore by Judith R. CristobalBarbara DrigoBarbara DrigoUniversity of South Australia, Adelaide, SA 5001, AustraliaMore by Barbara DrigoClaire EllisonClaire EllisonQueen's University, Beaty Water Research Center, Kingston, ON K7L 3N6, CanadaMore by Claire EllisonZanah FrancisZanah FrancisU.S. Department of Health and Human Services, Washington, D.C. 20201-0004, United StatesMore by Zanah FrancisDominic FrigonDominic FrigonMcGill University, Civil Engineering and Applied Mechanics, 817 Sherbrooke Street West, Montreal, QC H3A 0C3, CanadaMore by Dominic Frigonhttps://orcid.org/0000-0003-1587-8943Markus GaenzleMarkus GaenzleUniversity of Alberta, Edmonton, AB T6G 2R3, CanadaMore by Markus GaenzleJulia VierheiligJulia VierheiligTU Wien, Institute of Water Quality and Resource Management, ICC Water & Health, 1040 Wien, AustriaMore by Julia VierheiligTimothy R. JulianTimothy R. JulianEawag, Swiss Federal Institute of Aquatic Science and Technology, Dübendorf 8600, SwitzerlandMore by Timothy R. Julianhttps://orcid.org/0000-0003-1000-0306Uli KlümperUli KlümperInstitute for Hydrobiology, TU Dresden, Dresden 01062, GermanyMore by Uli Klümperhttps://orcid.org/0000-0002-4169-6548Liping MaLiping MaEast China Normal University, Dongchuan Road 500, Shanghai 200241, ChinaMore by Liping Mahttps://orcid.org/0000-0002-1646-6767Chand MangatChand MangatPublic Health Agency of Canada, Wastewater Surveillance Unit, National Microbiology Laboratory, Winnipeg, MB R3E 3R2, CanadaMore by Chand MangatMaya NadimpalliMaya NadimpalliGangarosa Department of Environmental Health, Emory University, Atlanta, Georgia 30322, United StatesMore by Maya Nadimpallihttps://orcid.org/0000-0002-6526-116XManami NakashitaManami NakashitaNational Institute of Infectious Diseases, Shinjuku-ku, Tokyo 162-8640, JapanMore by Manami NakashitaGilbert OsenaGilbert OsenaUniversity of Gothenburg, Goteborg, Västra Götaland 405 30, SwedenMore by Gilbert OsenaSasikaladevi RathinaveluSasikaladevi RathinaveluEawag, Swiss Federal Institute of Aquatic Science and Technology, Kastenienbaum 6047, SwitzerlandMore by Sasikaladevi RathinaveluRichard Reid-SmithRichard Reid-SmithPublic Health Agency of Canada Foodborne, Waterborne and Zoonotic Infections Division, Guelph, ON N1G 5B2, CanadaMore by Richard Reid-SmithMichael SaldanaMichael SaldanaSonny Astani Civil and Environmental Engineering, University of Southern California, 920 Downey Way, BHE 201, Los Angeles, California 90089-0001, United StatesMore by Michael SaldanaHeike SchmittHeike SchmittNational Institute for Public Health and the Environment, Bilthoven 3720 BA, The NetherlandsDelft University of Technology, Delft, Zuid-Holland 2600 AA, NetherlandsMore by Heike SchmittShuxian LiShuxian LiDepartment of Civil Engineering, The University of Hong Kong, Hong Kong 999077, ChinaMore by Shuxian LiAndrew C. SingerAndrew C. SingerCentre for Ecology & Hydrology, Mansfield Road, Oxford OX1 3SR, United KingdomMore by Andrew C. Singerhttps://orcid.org/0000-0003-4705-6063Tam T. TranTam T. TranNORCE Norwegian Research Centre AS, Tromso, Troms og Finnmark 9019, NorwayMore by Tam T. TranKadir YanacKadir YanacUniversity of Manitoba, Department of Civil Engineering, Winnipeg, MB CR3T 5V6, CanadaMore by Kadir YanacGustavo YbazetaGustavo YbazetaHealth Sciences North Research Institute, Sudbury, ON P3E 2H2, CanadaMore by Gustavo YbazetaMonika HarniszMonika HarniszUniversity of Warmia and Mazury in Olsztyn, Department of Water Protection Engineering and Environmental Microbiology, Prawochenskiego 1, Olsztyn 10-790, PolandMore by Monika HarniszOpen PDFEnvironmental Science & TechnologyCite this: Environ. Sci. Technol. 2025, XXXX, XXX, XXX-XXXClick to copy citationCitation copied!https://pubs.acs.org/doi/10.1021/acs.est.4c09663https://doi.org/10.1021/acs.est.4c09663Published January 28, 2025 Publication History Received 11 September 2024Published online 28 January 2025article-commentary© 2025 The Authors. Published by American Chemical Society. This publication is licensed under CC-BY 4.0 . License Summary*You are free to share (copy and redistribute) this article in any medium or format and to adapt (remix, transform, and build upon) the material for any purpose, even commercially within the parameters below:Creative Commons (CC): This is a Creative Commons license.Attribution (BY): Credit must be given to the creator.View full license*DisclaimerThis summary highlights only some of the key features and terms of the actual license. It is not a license and has no legal value. Carefully review the actual license before using these materials. This publication is licensed underCC-BY 4.0 . License Summary*You are free to share(copy and redistribute) this article in any medium or format and to adapt(remix, transform, and build upon) the material for any purpose, even commercially within the parameters below: Creative Commons (CC): This is a Creative Commons license. Attribution (BY): Credit must be given to the creator.View full license *DisclaimerThis summary highlights only some of the key features and terms of the actual license. It is not a license and has no legal value. Carefully review the actual license before using these materials. License Summary*You are free to share(copy and redistribute) this article in any medium or format and to adapt(remix, transform, and build upon) the material for any purpose, even commercially within the parameters below: Creative Commons (CC): This is a Creative Commons license. Attribution (BY): Credit must be given to the creator. View full license *DisclaimerThis summary highlights only some of the key features and terms of the actual license. It is not a license and has no legal value. Carefully review the actual license before using these materials. License Summary*You are free to share(copy and redistribute) this article in any medium or format and to adapt(remix, transform, and build upon) the material for any purpose, even commercially within the parameters below: Creative Commons (CC): This is a Creative Commons license. Attribution (BY): Credit must be given to the creator. View full license *DisclaimerThis summary highlights only some of the key features and terms of the actual license. It is not a license and has no legal value. Carefully review the actual license before using these materials. ACS Publications© 2025 The Authors. Published by American Chemical SocietySubjectswhat are subjectsArticle subjects are automatically applied from the ACS Subject Taxonomy and describe the scientific concepts and themes of the article.Antibiotic resistanceAntimicrobial agentsImmunologyPeptides and proteinsWastewaterAntimicrobial resistance (AMR) is among the top 10 public health threats, with nearly 5 million deaths in 2019 linked to AMR-related bacterial infections. (1) A One Health approach is needed to combat AMR.Healthcare-based surveillance (HBS) of AMR provides incomplete information about the scope of the AMR threat. HBS screens only patients seeking medical attention, lacking community-level representativeness, and suffers from underreporting. (2) Consequently, researchers are turning to wastewater-based surveillance (WBS) to complement HBS. (3) WBS can provide information about AMR circulating within communities and hospitals, offering a comprehensive understanding of AMR prevalence. However, the surveillance targets and data obtained from WBS are distinct from those derived from HBS, creating uncertainty regarding their utility to the public health sector and ability to yield policy relevant information. In May 2024, participants in a workshop during the 7th Environmental Dimension of Antimicrobial Resistance (EDAR7) conference (Montréal, Canada) sought to answer four questions aimed at advancing the policy relevance of AMR data generated by WBS.What Public Health Relevant Indicators Are Currently Used to Drive Antimicrobial Stewardship Policy?Click to copy section linkSection link copied!There is a pressing need to integrate available information across One Health sectors (human health, agriculture, and environment) to inform policy and practice aimed at mitigating AMR (Figure 1). HBS aims to guide antibiotic prescriptions by generating antibiograms and provides data on AMR prevalence and trends by prescreening inpatients for carriage, assessing resistance of pathogens responsible for infections, and tracking in- and out-patient antibiotic prescription patterns. Tracking trends of multidrug-resistant (MDR) organisms in healthcare facilities helps to identify units experiencing high rates of nosocomial infections and informs the selection of appropriate treatment options. Well-established surveillance programs can result in public health reports used to define strategies to regulate antibiotic stewardship and to monitor and evaluate interventions.Figure 1Figure 1. Integration of data across One Health sectors to inform AMR policy and the potential role of WBS. Clinical settings, environmental studies, and agricultural surveillance can provide indicators used for antimicrobial stewardship and antimicrobial resistance (AMR) management. Indicators include prevalence of AMR and prescription patterns in clinical settings, geographic distribution and emerging threats in the environment to identify horizontal gene transfer (HGT) hot spots, and antibiotic/biocide residues and zoonotic pathways in agriculture. Wastewater-based surveillance (WBS) programs can be designed to integrate within and across all sectors.High Resolution ImageDownload MS PowerPoint SlideSurveillance in agriculture tends to be more focused on antibiotic use than tracking resistant infections, although some monitoring programs track resistant isolates in meat, produce, and other food products. Measuring antibiotic consumption in animals and crops can help identify hot spots for selective pressure and potential for AMR to spread. Monitoring residues in food of animal origin and biocides in vegetables can also contribute. Surveillance of AMR in livestock, wild animals, and the food chain can inform transmission pathways between animals and humans, providing insights into interventions to curb foodborne and zoonotic spread. Agricultural surveillance supports the establishment and enforcement of antibiotic stewardship in animals and plants, including antibiotic use regulations.There is increasing recognition of the environmental dimension of One Health approaches to inform AMR policy, but environmental surveillance programs, including WBS, are still limited. Environmental indicators of AMR, such as the presence and concentration of antibiotic resistance genes (ARGs) and resistant organisms in wastewater effluents, rivers, lakes, air, and soils, are primarily derived from spatial and longitudinal studies. These studies identify hot spots of resistance, guide environmental regulations like wastewater management policies, and inform agricultural practices to reduce antibiotic runoff. Environmental surveillance is also uniquely positioned for identifying emerging threats, including new ARGs, mobile genetic elements (MGEs), biocidal resistance genes, and resistant organisms. Monitoring targeted sources, including human, industrial, and agricultural wastewaters, has identified hot spots of resistance and selective pressure. The study of MGEs, in particular, offers insights into the mobility of ARGs and the potential acquisition of new resistance mechanisms in pathogens.What Public Health Relevant Targets and Data Can Be Derived from Wastewater through Monitoring Programs?Click to copy section linkSection link copied!WBS can include monitoring of antimicrobials, resistant organisms, ARGs, and MGEs in human, industrial, or agricultural/food production wastewaters, with monitoring locations selected to integrate across specific sources of interest. However, there is a need to better strategize and coordinate WBS of AMR in a manner that focuses on targets and data that are very likely to provide actionable information. One strategy could be prioritizing low-prevalence resistant bacteria of high clinical relevance, such as carbapenemase-producing Enterobacterales, vancomycin-resistant Enterococcus spp., or other bacteria of the World Health Organization Bacterial Priority Pathogen List. (4) An increase in the level of resistant organisms in wastewater can indicate rising community-level carriage, potential outbreak risks, or intervention failures. WBS may also help determine if outbreaks have ended or if asymptomatic cases persist in the community. However, it is important to be aware of population-scale detection limits and to determine the necessary temporal resolution (e.g., weekly monitoring) to achieve the monitoring goal. In contrast, monitoring pathogens that are already widespread does not necessarily add significant value to inform public health actions.Metagenomic approaches, i.e., sequencing of DNA across microbial populations encountered in wastewater, can offer a comprehensive view of ARGs and MGEs circulating within the corresponding population. Because metagenomics is a nontargeted approach, this perspective could identify emerging ARGs or provide an early warning regarding acquisition of ARGs by pathogens of concern in a community. For example, early detection of the mcr-1 gene conferring resistance to colistin through metagenomics led to the implementation of stricter colistin stewardship and monitoring in high-risk areas, such as units with high rates of MDR. (5)WBS can also target antimicrobials, thus filling knowledge gaps regarding the patterns and prevalence of the use of antimicrobials and other pharmaceuticals. Efforts are needed to improve reporting of antimicrobial use data. Where data are available, they tend to be highly aggregated and costly and with low spatial and temporal resolution. However, antibiotic testing does require sophisticated instrumentation and expertise and works best for antibiotics, such as macrolides and fluoroquinolones, that persist longer in wastewater environments. Fast-degrading antibiotics such as β-lactams might still be detected in the outflow from hospitals with short retention times.A general advantage of WBS is the ability to capture longitudinal and spatial trends across populations and sources of interest. Notably, different sanitation infrastructures and spatial scales of WBS provide distinct opportunities for measurement and interpretation. For example, in hospital wastewater, the indicators reflect carriage of resistant organisms or antibiotic usage within a specific facility. In municipal wastewater, the indicators reflect trends of resistance or antibiotic consumption within the community. Importantly, most of the world is served by nonsewered sanitation, particularly in low- and middle-income countries; surveillance in these settings might focus on tracking emergence and estimating prevalence in specific community settings (e.g., schools, universities, and hospitals). WBS can help to fill critical knowledge gaps in HBS, particularly in countries lacking comprehensive diagnostic capabilities.What Information, Resources, and Contextualization Are Needed to Align Public Health Indicators Derived from Wastewater with Other Public Health Indicators to Better Inform Our Epidemiological Understanding of AMR?Click to copy section linkSection link copied!A key consensus of the workshop was the need to integrate WBS data with HBS to better inform public health strategies.Information needed includes data on AMR prevalence from clinics, meaning the pathogens encountered in the population and corresponding rates of resistance to specific antibiotics obtained through HBS. Such monitoring can reveal clinically relevant targets for WBS and allow the establishment of standard methodologies for consistent data collection and interpretation. Whole genome sequencing of human and animal clinical strains can provide information needed to calibrate WBS data and track persistent pathogens and ARGs of concern in wastewater, potentially indicating ongoing transmission. Information about antibiotic usage in humans, animals, and plants, prescription practices, rates of antibiotic degradation in wastewater, flow data, and transport in sewage systems will help better align WBS and healthcare sector AMR indicators.Resources necessary for advancing WBS of AMR include institutional, financial, and human capital investments. These can support the design, implementation, and continuity of a monitoring plan to yield comprehensive and longitudinal data collection needed to infer AMR dynamics within the community. Initial costs for setting up laboratories, building infrastructure, and establishing workflows among stakeholders such as those who operate wastewater facilities and other monitoring locations are necessary to centralize analyses and build capacity. The investments made in infrastructure and organization for COVID-19 surveillance, and increasingly other pathogens, provide an opportunity to leverage existing resources for AMR monitoring. Establishing publicly accessible databases to collect, visualize, and analyze data from both wastewater and clinical surveillance will enhance collaboration among clinicians, policy makers, researchers, and other stakeholders.WBS indicators for AMR should be contextualized with respect to clinical and agricultural/food sector surveillance through strong collaborations among researchers, clinicians, and communities. While WBS alone may not always generate information about specific targets of interest, it can identify broader trends and emerging hot spots and inform public health strategies like early warnings and antimicrobial stewardship efforts. Notably, transitioning from WBS to wastewater-based epidemiology for AMR poses significant challenges, for example, in predicting the prevalence of AMR within the population. Complications include the dynamics and complexity of pathogen shedding rates and antibiotic resistance mechanisms, and the growth, fate, and transport processes in sewer networks. One key issue is the potential proliferation of resistant organisms within the sewer network, both in the wastewater and in biofilm, which can decouple wastewater-based quantitative estimates from inferences about AMR epidemiology. Indicators from WBS could be developed to help inform progress toward the Sustainable Development Goals or otherwise provide insight into key socioeconomic factors driving overall trends. Geographical and mobility patterns within sewersheds, and connections between industries and hospitals, should be considered to calibrate wastewater indicators and discern community-sourced data from other origins. Research on the fate of resistant bacteria in wastewater systems, along with cohort studies on resistant bacteria in human carriers, may help improve our understanding and interpretation of WBS-derived data.How Can the Information Derived from WBS of AMR Contribute to the Formulation of Effective Public Health Policies or Interventions?Click to copy section linkSection link copied!WBS offers population-integrated data at comparatively low cost and effort relative to monitoring individuals within a population. It provides broader views on population prevalence, independent of screening effort, participation rates, and the likelihood of reporting to health services. Additionally, it enables a comprehensive overview of the microbial genomes circulating in the environment and provides space- and time-resolved data that can be scaled to various needs. As critiqued in the recent 2024 NASEM report, we acknowledge the limitations of WBS for ARG-focused monitoring at the community level, which can be complicated by ARGs from non-human sources and the amplification of ARGs between the human source and the wastewater treatment plant. (6) However, it is important to recognize that WBS of AMR could provide much broader value beyond serving as an early warning system, especially in terms of evaluating long-term trends and effects of policy interventions on shaping these trends. We highlight alternative use cases that are of particular value for aligning WBS data with actionable public health objectives and HBS, such as detection of the emergence of novel resistance genes, or use of culture- and molecular-based methods to track long-term changes in community prevalence rates. (7)Integrating WBS data with existing surveillance methods (Figure 1) is a promising approach to enhance AMR understanding by correlating wastewater findings with clinical data, making policies actionable. WBS data can expand and provide greater resolution to traditional clinical antibiograms while also filling diagnostic gaps and better optimizing the selection of antibiotic treatments in regions with limited spatial and longitudinal AMR data.Public access and education, e.g., via media outlets, can increase AMR awareness, thereby enhancing public support and compliance with AMR policies. To inform effective public health interventions from WBS, it is necessary to have clear objectives and collaborate closely with stakeholders across One Health sectors, which can facilitate the implementation of targeted and efficient measures aimed at limiting the evolution and transmission of antibiotic-resistant pathogens.Author InformationClick to copy section linkSection link copied!Corresponding AuthorSheena Conforti - Eawag, Swiss Federal Institute of Aquatic Science and Technology, Dübendorf 8600, Switzerland; https://orcid.org/0000-0002-0173-6170; Email: [email protected]AuthorsAmy Pruden - Civil and Environmental Engineering, Virginia Polytechnic Institute and State University, 418 Durham Hall, Blacksburg, Virginia 24061, United States; https://orcid.org/0000-0002-3191-6244Nicole Acosta - University of Calgary, Cumming School of Medicine, Calgary, AB T2N 1N4, CanadaChristopher Anderson - West Virginia University, Morgantown, West Virginia 26506-6201, United StatesHelmut Buergmann - Eawag, Swiss Federal Institute of Aquatic Science and Technology, Kastenienbaum 6047, Switzerland; https://orcid.org/0000-0002-5651-5906Juliana Calabria De Araujo - Federal University of Minas Gerais, Belo Horizonte, MG 31270-901, BrazilJudith R. Cristobal - Department of Chemistry, University at Buffalo - The State University of New York, 633 Natural Science Complex, Buffalo, New York 14260, United StatesBarbara Drigo - University of South Australia, Adelaide, SA 5001, AustraliaClaire Ellison - Queen's University, Beaty Water Research Center, Kingston, ON K7L 3N6, CanadaZanah Francis - U.S. Department of Health and Human Services, Washington, D.C. 20201-0004, United StatesDominic Frigon - McGill University, Civil Engineering and Applied Mechanics, 817 Sherbrooke Street West, Montreal, QC H3A 0C3, Canada; https://orcid.org/0000-0003-1587-8943Markus Gaenzle - University of Alberta, Edmonton, AB T6G 2R3, CanadaJulia Vierheilig - TU Wien, Institute of Water Quality and Resource Management, ICC Water & Health, 1040 Wien, AustriaTimothy R. Julian - Eawag, Swiss Federal Institute of Aquatic Science and Technology, Dübendorf 8600, Switzerland; https://orcid.org/0000-0003-1000-0306Uli Klümper - Institute for Hydrobiology, TU Dresden, Dresden 01062, Germany; https://orcid.org/0000-0002-4169-6548Liping Ma - East China Normal University, Dongchuan Road 500, Shanghai 200241, China; https://orcid.org/0000-0002-1646-6767Chand Mangat - Public Health Agency of Canada, Wastewater Surveillance Unit, National Microbiology Laboratory, Winnipeg, MB R3E 3R2, CanadaMaya Nadimpalli - Gangarosa Department of Environmental Health, Emory University, Atlanta, Georgia 30322, United States; https://orcid.org/0000-0002-6526-116XManami Nakashita - National Institute of Infectious Diseases, Shinjuku-ku, Tokyo 162-8640, JapanGilbert Osena - University of Gothenburg, Goteborg, Västra Götaland 405 30, SwedenSasikaladevi Rathinavelu - Eawag, Swiss Federal Institute of Aquatic Science and Technology, Kastenienbaum 6047, SwitzerlandRichard Reid-Smith - Public Health Agency of Canada Foodborne, Waterborne and Zoonotic Infections Division, Guelph, ON N1G 5B2, CanadaMichael Saldana - Sonny Astani Civil and Environmental Engineering, University of Southern California, 920 Downey Way, BHE 201, Los Angeles, California 90089-0001, United StatesHeike Schmitt - National Institute for Public Health and the Environment, Bilthoven 3720 BA, The Netherlands; Delft University of Technology, Delft, Zuid-Holland 2600 AA, NetherlandsShuxian Li - Department of Civil Engineering, The University of Hong Kong, Hong Kong 999077, ChinaAndrew C. Singer - Centre for Ecology & Hydrology, Mansfield Road, Oxford OX1 3SR, United Kingdom; https://orcid.org/0000-0003-4705-6063Tam T. Tran - NORCE Norwegian Research Centre AS, Tromso, Troms og Finnmark 9019, NorwayKadir Yanac - University of Manitoba, Department of Civil Engineering, Winnipeg, MB CR3T 5V6, CanadaGustavo Ybazeta - Health Sciences North Research Institute, Sudbury, ON P3E 2H2, CanadaMonika Harnisz - University of Warmia and Mazury in Olsztyn, Department of Water Protection Engineering and Environmental Microbiology, Prawochenskiego 1, Olsztyn 10-790, PolandAuthor ContributionsS.C. and A.P. contributed equally to this work.NotesThe authors declare no competing financial interest.BiographyClick to copy section linkSection link copied!Sheena ConfortiHigh Resolution ImageDownload MS PowerPoint SlideSheena Conforti is a postdoctoral fellow in the Pathogens and Human Health group at the Department of Environmental Microbiology, Eawag, Switzerland. She earned her Ph.D. in 2024 from the Department of Biosystems Science and Engineering, at ETH Zurich. Her research focuses on antimicrobial resistance surveillance through wastewater and environmental monitoring, combining culture-based methods and whole-genome sequencing to study resistant bacteria and transmission dynamics. Her work supports the One Health framework and aims to inform public health strategies by integrating data from human, animal, and environmental sources.AcknowledgmentsClick to copy section linkSection link copied!The authors thank the 7th Conference on Environmental Dimension of Antimicrobial Resistance for the financial support of this publication. The authors acknowledge the Environmental Dimension of Antimicrobial Resistance (EDAR7) conference, held in May 2024 in Montréal, Canada, for hosting the workshop that contributed to the development of the manuscript. The authors thank all of the workshop participants and Molly Cantrell for their valuable input during the workshop. The authors also thank the Swiss National Science Foundation (Grant 192763) for funding S.C.ReferencesClick to copy section linkSection link copied! This article references 7 other publications. 1Antimicrobial Resistance Collaborators Global burden of bacterial antimicrobial resistance in 2019: a systematic analysis. Lancet 2022, 399 (10325), 629– 655, DOI: 10.1016/S0140-6736(21)02724-0 Google ScholarThere is no corresponding record for this reference.2 Global Antimicrobial Resistance and Use Surveillance System (GLASS) Report 2022. World Health Organization, 2022. https://www.who.int/publications/i/item/9789240062702.Google ScholarThere is no corresponding record for this reference.3Chau, K. K.; Barker, L.; Budgell, E. P.; Vihta, K. D.; Sims, N.; Kasprzyk-Hordern, B.; Harriss, E.; Crook, D. W.; Read, D. S.; Walker, A. S.; Stoesser, N. Systematic review of wastewater surveillance of antimicrobial resistance in human populations. Environ. Int. 2022, 162, 107171, DOI: 10.1016/j.envint.2022.107171 Google ScholarThere is no corresponding record for this reference.4 WHO Bacterial Priority Pathogens List 2024: bacterial pathogens of public health importance to guide research, development and strategies to prevent and control antimicrobial resistance. World Health Organization, 2024. https://www.who.int/publications/i/item/9789240093461.Google ScholarThere is no corresponding record for this reference.5von Wintersdorff, C. J. H.; Wolffs, P. F. G.; van Niekerk, J. M.; Beuken, E.; van Alphen, L. B.; Stobberingh, E. E.; Oude Lashof, A. M. L.; Hoebe, C. J. P. A.; Savelkoul, P. H. M.; Penders, J. Detection of the plasmid-mediated colistin-resistance gene mcr-1 in faecal metagenomes of Dutch travellers. J. Antimicrob. Chemother. 2016, 71 (12), 3416– 3419, DOI: 10.1093/jac/dkw328 Google ScholarThere is no corresponding record for this reference.6National Academies of Sciences, Engineering, and Medicine. Increasing the Utility of Wastewater-based Disease Surveillance for Public Health Action: A Phase 2 Report. The National Academies Press: Washington, DC, 2024. DOI: 10.17226/27516 Google ScholarThere is no corresponding record for this reference.7Conforti, S.; Holschneider, A.; Sylvestre, É; Julian, T. R. Monitoring ESBL-Escherichia coli in Swiss wastewater between November 2021 and November 2022: insights into population carriage. mSphere. 2024, 9 (5), e0076023 DOI: 10.1128/msphere.00760-23 Google ScholarThere is no corresponding record for this reference.Cited By Click to copy section linkSection link copied!This article has not yet been cited by other publications.Download PDFFiguresReferences Get e-AlertsGet e-AlertsEnvironmental Science & TechnologyCite this: Environ. Sci. Technol. 2025, XXXX, XXX, XXX-XXXClick to copy citationCitation copied!https://doi.org/10.1021/acs.est.4c09663Published January 28, 2025 Publication History Received 11 September 2024Published online 28 January 2025© 2025 The Authors. Published by American Chemical Society. This publication is licensed under CC-BY 4.0 . License Summary*You are free to share (copy and redistribute) this article in any medium or format and to adapt (remix, transform, and build upon) the material for any purpose, even commercially within the parameters below:Creative Commons (CC): This is a Creative Commons license.Attribution (BY): Credit must be given to the creator.View full license*DisclaimerThis summary highlights only some of the key features and terms of the actual license. It is not a license and has no legal value. Carefully review the actual license before using these materials. Article Views-Altmetric-Citations-Learn about these metrics closeArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated.Recommended Articles FiguresReferencesAbstractHigh Resolution ImageDownload MS PowerPoint SlideFigure 1Figure 1. Integration of data across One Health sectors to inform AMR policy and the potential role of WBS. Clinical settings, environmental studies, and agricultural surveillance can provide indicators used for antimicrobial stewardship and antimicrobial resistance (AMR) management. Indicators include prevalence of AMR and prescription patterns in clinical settings, geographic distribution and emerging threats in the environment to identify horizontal gene transfer (HGT) hot spots, and antibiotic/biocide residues and zoonotic pathways in agriculture. Wastewater-based surveillance (WBS) programs can be designed to integrate within and across all sectors.High Resolution ImageDownload MS PowerPoint SlideSheena ConfortiHigh Resolution ImageDownload MS PowerPoint SlideSheena Conforti is a postdoctoral fellow in the Pathogens and Human Health group at the Department of Environmental Microbiology, Eawag, Switzerland. She earned her Ph.D. in 2024 from the Department of Biosystems Science and Engineering, at ETH Zurich. Her research focuses on antimicrobial resistance surveillance through wastewater and environmental monitoring, combining culture-based methods and whole-genome sequencing to study resistant bacteria and transmission dynamics. Her work supports the One Health framework and aims to inform public health strategies by integrating data from human, animal, and environmental sources.References This article references 7 other publications. 1Antimicrobial Resistance Collaborators Global burden of bacterial antimicrobial resistance in 2019: a systematic analysis. Lancet 2022, 399 (10325), 629– 655, DOI: 10.1016/S0140-6736(21)02724-0 There is no corresponding record for this reference.2 Global Antimicrobial Resistance and Use Surveillance System (GLASS) Report 2022. World Health Organization, 2022. https://www.who.int/publications/i/item/9789240062702.There is no corresponding record for this reference.3Chau, K. K.; Barker, L.; Budgell, E. P.; Vihta, K. D.; Sims, N.; Kasprzyk-Hordern, B.; Harriss, E.; Crook, D. W.; Read, D. S.; Walker, A. S.; Stoesser, N. Systematic review of wastewater surveillance of antimicrobial resistance in human populations. Environ. Int. 2022, 162, 107171, DOI: 10.1016/j.envint.2022.107171 There is no corresponding record for this reference.4 WHO Bacterial Priority Pathogens List 2024: bacterial pathogens of public health importance to guide research, development and strategies to prevent and control antimicrobial resistance. World Health Organization, 2024. https://www.who.int/publications/i/item/9789240093461.There is no corresponding record for this reference.5von Wintersdorff, C. J. H.; Wolffs, P. F. G.; van Niekerk, J. M.; Beuken, E.; van Alphen, L. B.; Stobberingh, E. E.; Oude Lashof, A. M. L.; Hoebe, C. J. P. A.; Savelkoul, P. H. M.; Penders, J. Detection of the plasmid-mediated colistin-resistance gene mcr-1 in faecal metagenomes of Dutch travellers. J. Antimicrob. Chemother. 2016, 71 (12), 3416– 3419, DOI: 10.1093/jac/dkw328 There is no corresponding record for this reference.6National Academies of Sciences, Engineering, and Medicine. Increasing the Utility of Wastewater-based Disease Surveillance for Public Health Action: A Phase 2 Report. The National Academies Press: Washington, DC, 2024. DOI: 10.17226/27516 There is no corresponding record for this reference.7Conforti, S.; Holschneider, A.; Sylvestre, É; Julian, T. R. Monitoring ESBL-Escherichia coli in Swiss wastewater between November 2021 and November 2022: insights into population carriage. mSphere. 2024, 9 (5), e0076023 DOI: 10.1128/msphere.00760-23 There is no corresponding record for this reference.

Antibiotic resistance (AR) is one of the greatest public health challenges worldwide. Processes that allow the reduction of AR predictor of hospital wastewater has become crucial process that contributes to the protection of public health and the environment. The aim of this review article was to compare the effectiveness of various methods for treatment hospital wastewater in eliminating antibiotic-resistant bacteria (ARB) and degrading antibiotic resistance genes (ARGs) and antibiotics. A large number of studies dealing with wastewater treatment suggest that this topic is highly relevant and that new solutions are being developed to limit the spread of AR. Some wastewater treatment techniques have been in use for decades. Despite the negative effects of chlorine compounds, chlorination is still applied to eliminate ARB, ARGs, and drug metabolites. Ultraviolet (UV) radiation and ozonation have long been recognized for their treating properties. In the literature, advanced oxidation processes (AOPs) are increasingly often indicated as the most effective alternative to conventional treatment methods. Various methods for disinfecting hospital wastewater were reviewed and their environmental impact was analyzed in this article, and the results provide valuable insights for the further development of effective wastewater management strategies.

The rising incidence of antibiotic resistance poses a significant threat to public health. In recent years the widespread use of antibiotics has led to an increase in the concentration of antibiotic-resistant bacteria also in natural environments. The study was conducted in bathing areas three recreational lakes located in the Zaborski Landscape Park in northern Poland. Water samples were collected in three parallel repetitions in April, June and September 2022. Our study indicates that anthropopressure connected with tourism and recreation promotes the growth of fecal bacteria, including antibiotic-resistant strains, whose significant accumulation was recorded in September, the month marking the end of summer vacation. Antibiotic resistance profiles showed that isolated strains of fecal bacteria were resistant to beta-lactam antibiotics. The highest percentage of Escherichia coli strains showed resistance to cefepime (39.1%), and enterococci to imipenem (26.9%). The amplification of resistance genes confirmed the presence of only selected bla genes in the examined strains of fecal bacteria. The blaTEM gene was found in 14 strains of Enterococcus faecium (82.4%), in all 4 isolates of Enterococcus faecalis, and in 4 out of 5 unspecified strains of fecal streptococci. In Escherichia coli only blaCTX gene was identified in one strain. The presence of blaTEM genes was strongly correlated with the concentration of fecal bacteria, it can therefore be assumed that the presence of resistance genes was caused by direct contamination of the studied lakes with feces containing antibiotic-resistant bacteria, presumably without contamination from other sources. Resistance genes found in the control strains from sewage treatment plants were not identified in the studied isolates. Antibiotic resistance genetic markers found in strains isolated from wastewater may prove helpful in determining the sources of contamination of natural aquatic ecosystems with antibiotic-resistant fecal bacteria and thus ensure efficient management of projects aimed at making these waterbodies available for public use.

The composition of microbial communities is the key to effective anaerobic digestion (AD). The microbiome driving the AD process has been extensively researched, whereas the influence of specific substrates on the microbiome of digestate remains insufficiently investigated. Digestate has considerable potential for use in soil fertilization and bioremediation, therefore its biological safety should be monitored. Moreover, the knowledge about the composition of microbial communities and their interconnections in digestate should be extended, due to the impact on soil microbiota and its functionality. The aim of this study was a comprehensive assessment of the (1) sanitary quality, (2) core microbiome, and (3) microbial interactions in digestates collected from three full-scale agricultural biogas plants, with particular emphasis on their applicability from the perspective of the resident microbiota. Analyzed samples of digestate were derived from various substrates used for AD, including plant- and animal-based materials, and industrial waste. The study demonstrated that the phyla Bacillota, Bacteroidota, and Cloacimonadota were the most dominant in digestates regardless of the composition of the processed substrates, however, member composition at the genus level differed significantly between samples. In addition, we observed that microbial genera belonging to the less prevalent phyla play an integral role in the forming of microbial community interactions. Dominant microbial taxa with broad metabolic capabilities, potentially improving soil quality and functionality, have been identified. Moreover, we confirmed, that digestate samples were free of analyzed pathogenic bacteria and parasites. The study results indicate that digestate may have an immense fertilizing and bioremediation potential that has not been fully availed of to date.

The increase in the prevalence of carbapenem-resistant Gram-negative bacteria, in particular Acinetobacter baumannii (CRAB) and Pseudomonas aeruginosa (CRPA), poses a serious threat for public health worldwide. This article reviews the alarming data on the prevalence of infections caused by CRAB and CRPA pathogens and their presence in hospital and municipal wastewater, and it highlights the environmental impact of antibiotic resistance. The article describes the key role of antibiotic resistance genes (ARGs) in the acquisition of carbapenem resistance and sheds light on bacterial resistance mechanisms. The main emphasis was placed on the transfer of ARGs not only in the clinical setting, but also in the environment, including water, soil, and food. The aim of this review was to expand our understanding of the global health risks associated with CRAB and CRPA in hospital and municipal wastewater and to analyze the spread of these micropollutants in the environment. A review of the literature published in the last decade will direct research on carbapenem-resistant pathogens, support the implementation of effective preventive measures and interventions, and contribute to the development of improved strategies for managing this problem.

Antimicrobial resistance (AMR) is a global problem that gives serious cause for concern. Hospital wastewater (HWW) is an important link between the clinical setting and the natural environment, and an escape route for pathogens that cause hospital infections, including urinary tract infections (UTI). Bacteria of the genera Escherichia and Klebsiella are common etiological factors of UTI, especially in children, and they can cause short-term infections, as well as chronic conditions. ESBL-producing Escherichia and Klebsiella have also emerged as potential indicators for estimating the burden of antimicrobial resistance under environmental conditions and the spread of AMR between clinical settings and the natural environment. In this study, whole-genome sequencing and the nanopore technology were used to analyze the complete genomes of ESBL-producing E.coli and Klebsiella spp. and the HWW metagenome, and to characterize the mechanisms of AMR. The similarities and differences in the encoded mechanisms of AMR in clinical isolates (causing UTI) and environmental strains (isolated from HWW and the HWW metagenome) were analyzed. Special attention was paid to the genetic context and the mobility of antibiotic resistance genes (ARGs) to determine the common sources and potential transmission of these genes. The results of this study suggest that the spread of drug resistance from healthcare facilities via HWW is not limited to the direct transmission of resistant clonal lines that are typically found in the clinical setting, but it also involves the indirect transfer of mobile elements carrying ARGs between bacteria colonizing various environments. Hospital wastewater could offer a supportive environment for plasmid evolution through the insertion of new ARGs, including typical chromosomal regions. These results indicate that interlined environments (hospital patients - HWW) should be closely monitored to evaluate the potential transmission routes of drug resistance in bacteria.

The emergence of carbapenem-resistant Enterobacteriaceae in the hospital sector as well as in the natural environment is a problem that affects both high-income countries (HICs) and low-/middle-income countries (LMICs). The observed differences in the prevalence of carbapenem-resistant strains between HICs and LMICs can be attributed mainly to antibiotic consumption in healthcare facilities and the quantity of treated wastewater. Hospital wastewater is a major hotspot for the spread of carbapenem-resistant Enterobacteriaceae (CRE) and carbapenem resistance genes (CRGs) between the hospital sector and the environment. In this review article, attempts were made to describe and compare antibiotic consumption in hospitals, antimicrobial concentrations in both hospital and municipal wastewater, and the prevalence of CRE and CRGs in patients and in hospital and municipal wastewater in HICs and LMICs.A review of the literature has shown that carbapenems are more widely used in LMICs, but Saudi Arabia, an HIC, is a country with the highest carbapenem consumption in the world. The results of research conducted in both groups of countries indicate that Klebsiella sp./K. pneumoniae is the most common CRE in samples isolated from patients. Escherichia coli was the dominant pathogen in hospital and municipal wastewater in HICs, whereas Enterobacter spp. were most prevalent in LMICs. An analysis of the prevalence of CRGs demonstrated that the same genes are present in both groups of countries (blaKPC, blaKPC-2, blaVIM, blaVIM-1,2, blaNDM, blaIMP, blaIMP-8, blaOXA-48,181, blaNDM-1,5, blaGES, blaGES-5,6, blaIMI-1). The fact that the same CRGs are most prevalent in countries with different levels of economic development could suggest that these genes have a high potential to persist in the natural environment. These findings underscore the need for enhanced monitoring, effective control techniques, and a better understanding of carbapenem resistance pathways to mitigate public health hazards, notwithstanding the constraints of data analysis.

Antimicrobials and antibiotic resistance genes (ARGs) in substrates processed during anaerobic digestion in agricultural biogas plants (BPs) can reach the digestate (D), which is used as fertilizer. Antimicrobials and ARGs can be transferred to agricultural land, which increases their concentrations in the environment. The concentrations of 13 antibiotics in digestate samples from biogas plants (BPs) were investigated in this study. The abundance of ARGs encoding resistance to beta-lactams, tetracyclines, sulfonamides, fluoroquinolones, macrolide-lincosamide-streptogramin antibiotics, and the integrase genes were determined in the analyzed samples. The presence of cadmium, lead, nickel, chromium, zinc, and mercury was also examined. Antimicrobials were not eliminated during anaerobic digestion. Their concentrations differed in digestates obtained from different substrates and in liquid and solid fractions (ranging from 62.8 ng/g clarithromycin in the solid fraction of sewage sludge digestate to 1555.9 ng/L doxycycline in the liquid fraction of cattle manure digestate). Digestates obtained from plant-based substrates were characterized by high concentrations of ARGs (ranging from 5.73 × 102 copies/gDcfxA to 2.98 × 109 copies/gDsul1). The samples also contained mercury (0.5 mg/kg dry mass (dm)) and zinc (830 mg/kg dm). The results confirmed that digestate is a reservoir of ARGs (5.73 × 102 to 8.89 × 1010 copies/gD) and heavy metals (HMs). In addition, high concentrations of integrase genes (105 to 107 copies/gD) in the samples indicate that mobile genetic elements may be involved in the spread of antibiotic resistance. The study suggested that the risk of soil contamination with antibiotics, HMs, and ARGs is high in farms where digestate is used as fertilizer.

Escherichia coli bacteria are an essential indicator in evaluations of environmental pollution, which is why they must be correctly identified. This study aimed to determine the applicability of various methods for identifying E. coli strains in environmental samples. Bacterial strains preliminary selected on mFc and Chromocult media as E. coli were identified using MALDI Biotyper techniques, based on the presence of genes characteristic of E. coli (uidA, uspA, yaiO), as well as by 16S rRNA gene sequencing. The virulence and antibiotic resistance genes pattern of bacterial strains were also analyzed to investigate the prevalence of factors that may indicate adaptation to unsupportive environmental conditions and could have any significance in further identification of E. coli. Of the strains that had been initially identified as E. coli with culture-based methods, 36-81% were classified as E. coli with the use of selected techniques. The value of Cohen's kappa revealed the highest degree of agreement between the results of 16S rRNA gene sequencing, the results obtained in the MALDI Biotyper system, and the results of the analysis based on the presence of the yaiO gene. The results of this study could help in the selection of more accurate and reliable methods which can be used in a preliminary screening and more precise identification of E. coli isolated from environmental samples.

The aim of the present study was to analyze changes in the microbiome, resistome, and mobilome of hospital wastewater (HWW) induced by disinfection with chlorine compounds. Changes in bacterial communities and specific antibiotic resistance genes (ARGs) in HWW were determined with the use of a nanopore long-read metagenomic approach. The main hosts of ARGs in HWW were identified, and the mobility of resistance mechanisms was analyzed. Special attention was paid to the prevalence of critical-priority pathogens in the HWW microbiome, which pose the greatest threat to human health. The results of this study indicate that chlorine disinfection of HWW can induce significant changes in the structure of the total bacterial population and antibiotic resistant bacteria (ARB) communities, and that it can modify the resistome and mobilome of HWW. Disinfection favored the selection of ARGs, decreased their prevalence in HWW, while increasing their diversity. The mobility of the HWW resistome increased after disinfection. Disinfection led to the emergence of new drug resistance mechanisms in previously sensitive bacterial taxa. In conclusion, this study demonstrated that HWW disinfected with low (sublethal) concentrations of free chlorine significantly contributes to the mobility and transfer of drug resistance mechanisms (including critical mechanisms) between bacteria (including pathogens).