Antibiotic residues in final effluents of European wastewater treatment plants and their impact on the aquatic environment

A comprehensive monitoring of a broad set of antibiotics in the final effluent of wastewater treatment plants (WWTPs) of 7 European countries (Portugal, Spain, Ireland, Cyprus, Germany, Finland, and Norway) was carried out in two consecutive years (2015 and 2016). This is the first study of this kind performed at an international level. Within the 53 antibiotics monitored 17 were detected at least once in the final effluent of the WWTPs, i.e.: ciprofloxacin, ofloxacin, enrofloxacin, orbifloxacin, azithromycin, clarithromycin, sulfapyridine, sulfamethoxazole, trimethoprim, nalidixic acid, pipemidic acid, oxolinic acid, cefalexin, clindamycin, metronidazole, ampicillin, and tetracycline. The countries exhibiting the highest effluent average concentrations of antibiotics were Ireland and the southern countries Portugal and Spain, whereas the northern countries (Norway, Finland and Germany) and Cyprus exhibited lower total concentration. The antibiotic occurrence data in the final effluents were used for the assessment of their impact on the aquatic environment. Both, environmental predicted no effect concentration (PNEC-ENVs) and the PNECs based on minimal inhibitory concentrations (PNEC-MICs) were considered for the evaluation of the impact on microbial communities in aquatic systems and on the evolution of antibiotic resistance, respectively. Based on this analysis, three compounds, ciprofloxacin, azithromycin and cefalexin are proposed as markers of antibiotic pollution, as they could occasionally pose a risk to the environment. Integrated studies like this are crucial to map the impact of antibiotic pollution and to provide the basis for designing water quality and environmental risk in regular water monitoring programs.Peer reviewe

Towards harmonized laboratory methodologies in veterinary clinical bacteriology: outcomes of a European survey

IntroductionVeterinary clinical microbiology laboratories play a key role in antimicrobial stewardship, surveillance of antimicrobial resistance and prevention of healthcare associated-infections. However, there is a shortage of international harmonized guidelines covering all steps of veterinary bacterial culture from sample receipt to reporting.MethodsIn order to gain insights, the European Network for Optimization of Veterinary Antimicrobial Treatment (ENOVAT) designed an online survey focused on the practices and interpretive criteria used for bacterial culture and identification (C&amp;amp;ID), and antimicrobial susceptibility testing (AST) of animal bacterial pathogens.ResultsA total of 241 microbiology laboratories in 34 European countries completed the survey, representing a mixture of academic (37.6%), governmental (27.4%), and private (26.5%) laboratories. The C&amp;amp;ID turnaround varied from 1 to 2 days (77.8%) to 3–5 days (20%), and 6– 8 days (1.6%), with similar timeframes for AST. Individual biochemical tests and analytical profile index (API) biochemical test kits or similar were the most frequent tools used for bacterial identification (77% and 56.2%, respectively), followed by PCR (46.6%) and MALDI-TOF MS (43.3%). For AST, Kirby-Bauer disk diffusion (DD) and minimum inhibitory concentration (MIC) determination were conducted by 43.8% and 32.6% of laboratories, respectively, with a combination of EUCAST and CLSI clinical breakpoints (CBPs) preferred for interpretation of the DD (41.2%) and MIC (47.6%) results. In the absence of specific CBPs, laboratories used human CBPs (53.3%) or veterinary CBPs representing another body site, organism or animal species (51.5%). Importantly, most laboratories (47.9%) only report the qualitative interpretation of the result (S, R, and I). As regards testing for AMR mechanisms, 48.5% and 46.7% of laboratories routinely screened isolates for methicillin resistance and ESBL production, respectively. Notably, selective reporting of AST results (i.e. excluding highest priority critically important antimicrobials from AST reports) was adopted by 39.5% of laboratories despite a similar proportion not taking any approach (37.6%) to guide clinicians towards narrower-spectrum or first-line antibiotics.DiscussionIn conclusion, we identified a broad variety of methodologies and interpretative criteria used for C&amp;amp;ID and AST in European veterinary microbiological diagnostic laboratories. The observed gaps in veterinary microbiology practices emphasize a need to improve and harmonize professional training, innovation, bacterial culture methods and interpretation, AMR surveillance and reporting strategies.</jats:sec

Antibiotikaresistens i kloakk

Antibiotic resistance is a characteristic possessed by some bacteria that enables them to withstand the effects of antibiotic treatment. Antibiotic use and abuse may result in development of antibiotic resistant bacteria. After an antibiotic treatment, antibiotics are partially excreted in urine and feces, and wastewater is therefore an important source of both bacteria and antibiotics from healthy and sick individuals. Due to the close contact between large numbers of bacteria and diverse types of antibiotics over prolonged periods, wastewater has the potential to select for and disseminate antibiotic resistance into the environment. Previous studies of hospital wastewater effluents have shown that these outlets discharge more antibiotic resistance factors than the general population. This study was initiated to map the occurrence of antibiotic resistance in wastewater under Norwegian conditions to identify whether local treatment of hospital wastewater could contribute to limit the dissemination of antibiotic resistance to the environment. Samples of untreated wastewater were collected monthly during one year from a hospital, a community, and from the wastewater treatment plant that treats the wastewater from the Greater Oslo region. The samples were analyzed for the occurrence of antibiotic resistant E. coli, 53 different antibiotics, heavy metals, and 144 bacterial genes encoding antibiotic resistance and mechanisms for the transfer of these genes between bacteria. During this study, higher levels of antibiotics and antibiotic resistant bacteria were found in wastewater from the hospital than from the general population. Broad-spectrum antibiotics were also present at much higher concentrations in the hospital effluents. These results indicate that local treatment of the hospital wastewater system may help limit the spread of resistance to the environment. A cost-benefit analysis should be performed to determine whether localized hospital wastewater treatment may be more effective than large-scale implementation at the wastewater treatment plants. This study also identified variants of multi-resistant E. coli producing ESBL-enzymes that can degrade broad-spectrum β-lactam antibiotics in the wastewater from the studied community. Some of these bacteria were exclusively and repeatedly found in the community wastewater. These belonged to two types of highly infectious E. coli (ST131 and ST648) that have been found all over the world. The finding of these bacterial clones in the community wastewater indicates that the dissemination of multi-resistant and pathogenic bacteria through wastewater is not exclusively attributable to hospital effluents. Hence, it is recommended to investigate additional as well as other types of outlets to identify potential high-risk sources for the spread of antibiotic resistance. High abundances of genes encoding mechanisms for lateral transfer of antibiotic resistance were found in this study, as well as clinically relevant antibiotic resistance genes like ndm-1 that have only scarcely been reported in association with clinical disease in Norway. These results thus verified a previous study of wastewater from European countries, wherein Norway had the highest levels of lateral transfer genes. Even though Norway has a relatively low prevalence of antibiotic resistance and a moderate consumption of antibiotics due to strict regulation, these results indicate the underlying potential for a substantial mobilization and dissemination in the future. This study has illuminated important aspects of antibiotic resistance that have previously not been described. The results of this study contribute to a broader knowledge base for decision-making regarding wastewater management in Norway and internationally.Antibiotikaresistens er en egenskap hos noen bakterier som gjør at de kan stå imot effektene av en antibiotikakur. Både fornuftig bruk og misbruk av antibiotika kan føre til at bakteriene blir mer motstandsdyktige. Etter en antibiotikakur skilles antibiotikumet delvis ut i urin og avføring, og avløpsvann er derfor en viktig kilde til både bakterier og antibiotika fra friske og syke mennesker. På grunn av den tette kontakten mellom store mengder bakterier og ulike typer antibiotika over lengre tid har avløpsvann potensialet for å spre antibiotikaresistens videre ut i miljøet. Tidligere studier av avløpsvann fra sykehus har vist at disse skiller ut større mengder antibiotikaresistens-faktorer enn resten av befolkningen. Denne studien ble påbegynt for å kartlegge forekomsten av antibiotikaresistens i avløpsvann under norske forhold for å se om lokal rensing av avløpsvann fra sykehus kan bidra til å begrense spredningen av antibiotikaresistens til miljøet. Prøver av avløpsvann ble samlet inn månedlig gjennom ett år fra et sykehus, et boligområde og fra Stor-Oslos hovedrenseanlegg på Slemmestad. Prøvene ble analysert for forekomsten av antibiotikaresistente E. coli-bakterier i tillegg til 53 ulike antibiotikastoffer, tungmetaller, og 144 bakteriegener som koder for antibiotikaresistens samt for mekanismer for overføring av disse genene mellom bakterier

Antibiotic resistance in wastewater

Antibiotic resistance is a characteristic possessed by some bacteria that enables them to withstand the effects of antibiotic treatment. Antibiotic use and abuse may result in development of antibiotic resistant bacteria. After an antibiotic treatment, antibiotics are partially excreted in urine and feces, and wastewater is therefore an important source of both bacteria and antibiotics from healthy and sick individuals. Due to the close contact between large numbers of bacteria and diverse types of antibiotics over prolonged periods, wastewater has the potential to select for and disseminate antibiotic resistance into the environment. Previous studies of hospital wastewater effluents have shown that these outlets discharge more antibiotic resistance factors than the general population. This study was initiated to map the occurrence of antibiotic resistance in wastewater under Norwegian conditions to identify whether local treatment of hospital wastewater could contribute to limit the dissemination of antibiotic resistance to the environment. Samples of untreated wastewater were collected monthly during one year from a hospital, a community, and from the wastewater treatment plant that treats the wastewater from the Greater Oslo region. The samples were analyzed for the occurrence of antibiotic resistant E. coli, 53 different antibiotics, heavy metals, and 144 bacterial genes encoding antibiotic resistance and mechanisms for the transfer of these genes between bacteria. During this study, higher levels of antibiotics and antibiotic resistant bacteria were found in wastewater from the hospital than from the general population. Broad-spectrum antibiotics were also present at much higher concentrations in the hospital effluents. These results indicate that local treatment of the hospital wastewater system may help limit the spread of resistance to the environment. A cost-benefit analysis should be performed to determine whether localized hospital wastewater treatment may be more effective than large-scale implementation at the wastewater treatment plants. This study also identified variants of multi-resistant E. coli producing ESBL-enzymes that can degrade broad-spectrum β-lactam antibiotics in the wastewater from the studied community. Some of these bacteria were exclusively and repeatedly found in the community wastewater. These belonged to two types of highly infectious E. coli (ST131 and ST648) that have been found all over the world. The finding of these bacterial clones in the community wastewater indicates that the dissemination of multi-resistant and pathogenic bacteria through wastewater is not exclusively attributable to hospital effluents. Hence, it is recommended to investigate additional as well as other types of outlets to identify potential high-risk sources for the spread of antibiotic resistance. High abundances of genes encoding mechanisms for lateral transfer of antibiotic resistance were found in this study, as well as clinically relevant antibiotic resistance genes like ndm-1 that have only scarcely been reported in association with clinical disease in Norway. These results thus verified a previous study of wastewater from European countries, wherein Norway had the highest levels of lateral transfer genes. Even though Norway has a relatively low prevalence of antibiotic resistance and a moderate consumption of antibiotics due to strict regulation, these results indicate the underlying potential for a substantial mobilization and dissemination in the future. This study has illuminated important aspects of antibiotic resistance that have previously not been described. The results of this study contribute to a broader knowledge base for decision-making regarding wastewater management in Norway and internationally

A unique role of flagellar function in Aliivibrio salmonicida pathogenicity not related to bacterial motility in aquatic environments

publishedVersio

Diversity and antibiotic resistance among Escherichia coli populations in hospital and community wastewater compared to wastewater at the receiving urban treatment plant

Bacterial diversity and antimicrobial resistance patterns among the indicator organism Escherichia coli were monitored in wastewater samples collected over one year from a hospital (HW), a community (CW) and the receiving urban (UW) wastewater treatment plant (WWTP). We compared levels of antibiotic resistance in the different types of wastewater, and identified whether resistant strains were endemic in the wastewater system. If so, implementation of local treatment at certain resistance hotspots (e.g. hospital outlets) could be used to decrease the amount of resistant bacteria in the wastewater. E. coli from HW (n = 2644), CW (n = 2525) and UW (n = 2693) were analyzed by biochemical phenotyping (PhenePlate System) and antimicrobial susceptibility testing to nine antibiotics (AREB System). The phenotypic diversities of the total E. coli populations were similar for all three sites (Simpson's Diversity index, Di = 0.973), however for individual samples, HW showed low diversities (Median Di = 0.800) and the E. coli flora was often dominated by strains that may have originated from the fecal flora of single individuals. The diversities in CW samples was higher (Median Di = 0.936), and UW samples showed similar diversities as the whole collection of isolates (Median Di = 0.971). Resistance to at least one of the nine antibiotics was observed in 45% of the HW isolates, 44% of CW isolates, and 33% of UW isolates. Resistance to gentamicin and chloramphenicol was uncommon (3.2 and 5.3%, respectively), whereas resistance to tetracycline and ampicillin was most common (24% and 31%, respectively). Extended-spectrum beta-lactamase-producing E. coli (ESBL-EC) were more common in HW (11.5%) and in CW (6.9%) compared to UW (3.7%). A high diversity (Di = 0.974) was observed among ESBL-EC isolates from UW (n = 99), indicating absence of any clonal structure among these isolates. Common PhP types of ESBL-EC often dominated in each HW sample, but were not identified across different samples, whereas ESBL-EC in CW showed low diversity (Di = 0.857) and were dominated by a specific PhP type that was found across almost all CW samples. The antibiotic resistance rates were highest in hospital wastewater, but surprisingly they were also high in the studied community wastewater, compared to the urban wastewater. The relative contribution of HW seemed low in terms of dissemination of antibiotic resistant bacteria to the WWTP.publishedVersio

Escherichia coli ST2797 Is Abundant in Wastewater and Might Be a Novel Emerging Extended-Spectrum Beta-Lactamase E. coli

ABSTRACT The increasing prevalence of antibiotic-resistant bacteria is an emerging threat to global health. The analysis of antibiotic-resistant enterobacteria in wastewater can indicate the prevalence and spread of certain clonal groups of multiresistant bacteria. In a previous study of Escherichia coli that were isolated from a pump station in Norway over 15 months, we found a recurring E. coli clone that was resistant to trimethoprim, ampicillin, and tetracycline in 201 of 3,123 analyzed isolates (6.1%). 11 representative isolates were subjected to whole-genome sequencing and were found to belong to the MLST ST2797 E. coli clone with plasmids carrying resistance genes, including blaTEM-1B, sul2, dfrA7, and tetB. A phenotypic comparison of the ST2797 isolates with the uropathogenic ST131 and ST648 that were repeatedly identified in the same wastewater samples revealed that the ST2797 isolates exhibited a comparable capacity for temporal survival in wastewater, greater biofilm formation, and similar potential for the colonization of mammalian epithelial cells. ST2797 has been isolated from humans and has been found to carry extended spectrum β-lactamase (ESBL) genes in other studies, suggesting that this clonal type is an emerging ESBL E. coli. Collectively, these findings show that ST2797 was more ubiquitous in the studied wastewater than were the infamous ST131 and ST648 and that ST2797 may have similar abilities to survive in the environment and cause infections in humans. IMPORTANCE The incidence of drug-resistant bacteria found in the environment is increasing together with the levels of antibiotic-resistant bacteria that cause infections. The COVID-19 pandemic has shed new light on the importance of monitoring emerging threats and finding early warning systems. Therefore, to mitigate the antimicrobial resistance burden, the monitoring and early identification of antibiotic-resistant bacteria in hot spots, such as wastewater treatment plants, are required to combat the occurrence and spread of antibiotic-resistant bacteria. Here, we applied a PhenePlate system as a phenotypic screening method for genomic surveillance and discovered a dominant and persistent E. coli clone ST2797 with a multidrug resistance pattern and equivalent phenotypic characteristics to those of the major pandemic lineages, namely, ST131 and ST648, which frequently carry ESBL genes. This study highlights the continuous surveillance and report of multidrug resistant bacteria with the potential to spread in One Health settings

Value of MRI and ultrasound for prediction of therapeutic response and erosive progression in patients with early rheumatoid arthritis managed by an aggressive treat-to-target strategy

Objectives To investigate if inflammation detected by MRI or ultrasound at rheumatoid arthritis (RA) onset is predictive of erosive progression or poor response to methotrexate monotherapy, and to investigate if subclinical inflammation in remission is predictive of future treatment escalation or erosive progression.Methods In a 2-year study, 218 patients with disease-modifying antirheumatic drug-naïve early RA were treated by a tight-control treat-to-target strategy corresponding to current recommendations. MRI and ultrasound were performed at regular intervals. Baseline imaging-based inflammation measures were analysed as predictors for early methotrexate failure and erosive progression using univariate and multivariate regression adjusted for clinical, laboratory and radiographic measures. In patients in remission after 1 year, imaging measures were analysed as predictors of treatment escalation and erosive progression during the second year. The added value of imaging in prediction models was assessed using receiver operating characteristic analyses.Results Baseline MRI inflammation was associated with MRI erosive progression and ultrasound with radiographic erosive progression. No imaging inflammation measure was associated with early methotrexate failure. Imaging inflammation was present in a majority of patients in clinical remission. Tenosynovitis was associated with treatment escalation, and synovitis and tenosynovitis with MRI/radiographic erosive progression during the second year. Imaging information did not improve prediction models for any of the outcomes.Conclusions Imaging-detected inflammation, both at diagnosis and in remission, is associated with elements of future disease development. However, the lack of a significant effect on prediction models indicates limited value of systematic MRI and ultrasound in management of early RA

Towards harmonized laboratory methodologies in veterinary clinical bacteriology: outcomes of a European survey

Peer reviewed: TrueIntroductionVeterinary clinical microbiology laboratories play a key role in antimicrobial stewardship, surveillance of antimicrobial resistance and prevention of healthcare associated-infections. However, there is a shortage of international harmonized guidelines covering all steps of veterinary bacterial culture from sample receipt to reporting.MethodsIn order to gain insights, the European Network for Optimization of Veterinary Antimicrobial Treatment (ENOVAT) designed an online survey focused on the practices and interpretive criteria used for bacterial culture and identification (C&amp;amp;ID), and antimicrobial susceptibility testing (AST) of animal bacterial pathogens.ResultsA total of 241 microbiology laboratories in 34 European countries completed the survey, representing a mixture of academic (37.6%), governmental (27.4%), and private (26.5%) laboratories. The C&amp;amp;ID turnaround varied from 1 to 2 days (77.8%) to 3–5 days (20%), and 6– 8 days (1.6%), with similar timeframes for AST. Individual biochemical tests and analytical profile index (API) biochemical test kits or similar were the most frequent tools used for bacterial identification (77% and 56.2%, respectively), followed by PCR (46.6%) and MALDI-TOF MS (43.3%). For AST, Kirby-Bauer disk diffusion (DD) and minimum inhibitory concentration (MIC) determination were conducted by 43.8% and 32.6% of laboratories, respectively, with a combination of EUCAST and CLSI clinical breakpoints (CBPs) preferred for interpretation of the DD (41.2%) and MIC (47.6%) results. In the absence of specific CBPs, laboratories used human CBPs (53.3%) or veterinary CBPs representing another body site, organism or animal species (51.5%). Importantly, most laboratories (47.9%) only report the qualitative interpretation of the result (S, R, and I). As regards testing for AMR mechanisms, 48.5% and 46.7% of laboratories routinely screened isolates for methicillin resistance and ESBL production, respectively. Notably, selective reporting of AST results (i.e. excluding highest priority critically important antimicrobials from AST reports) was adopted by 39.5% of laboratories despite a similar proportion not taking any approach (37.6%) to guide clinicians towards narrower-spectrum or first-line antibiotics.DiscussionIn conclusion, we identified a broad variety of methodologies and interpretative criteria used for C&amp;amp;ID and AST in European veterinary microbiological diagnostic laboratories. The observed gaps in veterinary microbiology practices emphasize a need to improve and harmonize professional training, innovation, bacterial culture methods and interpretation, AMR surveillance and reporting strategies.</jats:sec

A unique role of flagellar function in Aliivibrio salmonicida pathogenicity not related to bacterial motility in aquatic environments

publishedVersio