State of the art on the contribution of water to antimicrobial resistance

The antimicrobial resistance (AMR) is defined as the ability of microorganisms to withstand the effects of antibiotics and is considered a universal threat to humans, animals and the environment. The resistance mechanisms developed by bacteria originate from either the overuse of antibiotics in medical care and animal farming or from the spread of resistant genes among microorganisms. Worldwide, 700.000 people die annually from resistant infections and this means that if no action is taken, the estimated annual deaths attributable to AMR will be 10 million by 2050. The aim of this report is to discuss the mechanisms of antibiotic action and antibiotic resistance focusing on potential effects in water. Waterbodies have been indeed recognised as a significant reservoir of antibiotics and antibiotic resistance genes (ARG). They contribute to the interchange of resistance genes between pathogenic and non-pathogenic bacteria and they may favour the maintenance of the resistance in the environment. In this report, a review of the global scientific literature was conducted to show the levels of antibiotics in waste water treatment plants (WWTP), surface waters, agricultural runoff and drinking waters The most frequently monitored antibiotics in WWTP were sulfamethoxazole, ciprofloxacin and trimethoprim, while the most important substances in surface waters were erythromycin, sulfamethoxazole, trimethoprim and sulfamethazine.In parallel, a European database has been consulted to identify the antibiotics monitored in inland surface waters and the co-occurrence of heavy metals and antibiotic resistance in bacteria is discussed. It is indeed known that the chemical environmental pollution caused by heavy metals like silver (Ag), copper (Cu) or zinc (Zn) can co-select for antibiotic resistance. Antibiotics have been frequently detected in different aquatic environments within urban water cycles (waste, surface and drinking water) and, even if the levels are low (in the range of ng/L to µg/L), they could promote the acquisition of resistance by gene transfer between bacteria. The spread of AMR may be however constrained if general safety measures are taken to improve the effectiveness of wastewater treatment processes and to control the use of antibiotics in animal husbandry and in human medical practices. Additionally, new research must be conducted to understand the relationship between antibiotics’ concentration and the selection of resistance determinants in order to experimentally define the minimal concentration of antibiotics (as single and mixture) that induces resistance in bacteria. This factor should be then also considered in the evaluation of the risk assessment of antibiotics in water in order to define their environmental impactJRC.D.2-Water and Marine Resource

Selection of substances for the 3rd Watch List under the Water Framework Directive

The 1st Watch List (WL) was established by the Commission Implementing Decision (EU) 2015/4951 in March 2015. The list was updated in June 2018 by Commission Implementing Decision (EU) 2018/840. During that update, the insecticide metaflumizone and the antibiotics amoxicillin and ciprofloxacin were added to the list. The purpose of this report is to propose candidate substances for the 3rd WL. Three pillars of information were used to select the candidate substances. The first pillar is the outcome of the last prioritisation exercise, the second includes the outcome of the review of the 1st WL and recommendations for the 3rd WL, and the third is based on a literature search and/or other sources (for instance information from Member States). The selection of candidate substances took into consideration their hazard properties as well as the availability of reliable safety thresholds (including the contribution to antimicrobial resistance for antibiotics) and the availability of relevant analytical methods for monitoring in the appropriate environmental matrix. The proposed substances take into account comments and information received from Member-State and stakeholder-group experts. The candidate substances, fulfilling the selection criteria and identified by the JRC as most suitable for inclusion in the next WL, include two antibiotics, which are often prescribed together to overcome antimicrobial resistance; ten azole substances, used as either pharmaceuticals or pesticides; an anti-depressant pharmaceutical and its metabolite; and two additional pesticides used as fungicides.JRC.D.2-Water and Marine Resource

Safety and preliminary efficacy on cognitive performance and adaptive functionality of epigallocatechin gallate (EGCG) in children with Down syndrome. A randomized phase Ib clinical trial (PERSEUS study)

Purpose: Although some caregivers are using epigallocatechin gallate (EGCG) off label in hopes of improving cognition in young adults with Down syndrome (DS), nothing is known about its safety, tolerability, and efficacy in the DS pediatric population. We aimed to evaluate safety and tolerability of a dietary supplement containing EGCG and if EGCG improves cognitive and functional performance. Methods: A total of 73 children with DS (aged 6-12 years) were randomized. Participants received 0.5% EGCG (10 mg/kg daily dose) or placebo for 6 months with 3 months follow up after treatment discontinuation. Results: In total, 72 children were treated and 66 completed the study. A total of 38 participants were included in the EGCG group and 35 in the placebo group. Of 72 treated participants, 62 (86%) had 229 treatment-emergent adverse events (AEs). Of 37 participants in the EGCG group, 13 (35%) had 18 drug-related treatment-emergent AEs and 12 of 35 (34%) from the placebo group had 22 events. In the EGCG group, neither severe AEs nor increase in the incidence of AEs related to safety biomarkers were observed. Cognition and functionality were not improved compared with placebo. Secondary efficacy outcomes in girls point to a need for future work. Conclusion: The use of EGCG is safe and well-tolerated in children with DS, but efficacy results do not support its use in this population. (C) 2022 The Authors. Published by Elsevier Inc. on behalf of American College of Medical Genetics and Genomics

Toxicogenomic analysis of environmental impact in aquatic systems

Field biomonitoring based on molecular biomarkers detects early warning signals of stress suffered by organisms exposed to contamination. The lower part of the Ebro River basin has a long pollution history related to the presence of a chlor-alkali plant adjacent to Flix. In addition, the Ebro River has been affected by the invasion of alien species that damage its natural ecosystem. In this thesis, Cyprinus carpio (common carp) an Dreissena polymorpha (zebra mussel) were used as a model species in laboratory and field studies intended to the development of a multimolecular biomarker approach to identify the most relevantly effects of pollutants in the field. Gene expression techniques were used to study specific physiological defensive mechanism. The application and improvement of a well-established markers in different aquatic species and following different approaches allowed to analyze modes of action and to outline effects of persistent organic and inorganic pollutants found in the field. The results showed that chronic exposure to mercury of common carp results in increased levels of metallothionein in kidney, in scales (albeit at lower extent), but not in liver, considered as the primary detoxification organ. The measure of gene expression in scales provides the possibility of a new non-lethal method of study. Studies of toxicant effects in zebra mussel revealed that this bivalve could be used as potential sentinel specie for freshwater monitoring. Analysis of its detoxicfication mechanisms, and particularly of its ABC membrane transporters in adult and the early life stages could help understanding the survival of this species in highly contaminated areas.Postprint (published version

Decontamination trends in the aquacultured fish gilthead seabream (Sparus aurata) after feeding long-term a PCDD/F spiked feed

8 pages, 2 figures, 5 tablesAquacultured fish gilthead seabream (Sparusaurata), previously exposed to low levels of polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) trough the diet for a 13 months period, were fed on a clean feed for another 22 months. Gilthead seabream is a protandrous hermaphrodite species and this “decontamination” period coincided with the stages of sex differentiation, maturity and reproduction of the fish. PCDD/F levels in the fish tissues (i.e. muscle, liver, perivisceral fat and gonads), expressed in pg WHO-TEQ g−1 fresh weight, showed a general decreasing trend during the “decontamination” period. However, this general trend varied among tissues and was also dependent on sex and lipid contents. Toxicological effects affecting fish behaviour and hepatic marker responses were also evaluated. The results pointed out that exposure to PCDD/Fs did not have an impact on fish development and reproduction, since the proportion of sexes found after the sex reversal process was within the normal range described for this species. In addition, long-term exposure to low PCDD/F levels did not significantly affect the response of most of the biochemical markers considered. On the contrary, some of them (e.g. EROD activity) showed variations in their responses during the sex differentiation process and onwards. Finally, the hepatic AhR mRNA levels increased during dioxin exposure but they returned to values typical for non-exposed fish after the “decontamination” periodThis project was supported by the Spanish National Plan for Research PN2003 ref. AGL2003 02283-ALI and ‘‘Proyectos Intramurales de 30 de Marzo de 2004. Consejo Superior de Investigaciones Científicas (CSIC), Ministerio de Educación (200680I106)”. Additional funding from Ministerio Ciencia e Innovación (CGL2008-01898) is also acknowledged. M. Ábalos acknowledges an I3PDR contract from CSIC in the framework of the I3P projectPeer reviewe

A noninvasive test of exposition to toxicants: quantitative analysis of cytochrome P4501A expression in fish scales

8 pages, 3 tables, 3 figures.-- PMID: 17867895 [PubMed].Elevated expression of cytochrome P4501A (CYP1A) is an established biomarker for exposition to a wide range of toxicants, particularly for dioxin and structurally similar compounds. Expression of CYP1A usually is analyzed in internal organs, which involves dissection of the specimen. To avoid unnecessary animal killing, we present here an alternative method based on the monitoring of CYP1A expression in fish scales. Using β-naphthoflavone (BNF; 50 mg/kg body wt, intraperitoneal injection) as inducer in goldfish (Carassius auratus), we monitored levels of CYP1A mRNA both in scales and liver of treated and control specimens. Treatment with BNF resulted in a similar induction of CYP1A gene in both tissues, although scales responded faster (at 8 h after treatment) than liver (between 24 and 48 h). The scale-based test has the unique advantage of allowing sequential testing in the same specimen, which facilitates analysis of the time course of CYP1A induction and allows the study of individual variability. The method implies minimal suffering of the animals, because it only requires removal of a moderate (n = 1–3) number of scales for each time point. This nondestructive, fast, and relatively inexpensive test for toxic exposure therefore is suitable for environmental monitoring and food safety control programs in which specimen preservation is required.This work has been supported by the European Union and reflects only the author's views, and the European Community is not liable for any use that may be made of the information contained in it (AQUATERRA, GOCE-CT-2004-505428). The Spanish Ministry of Education and Science project GOCE-CT-2004-505428 (CTM2005-25168-E, CGL-2004-03514/HD, and BIO2005-00840) is also acknowledged for financial support. D. Raldúa acknowledges the "Ramón y Cajal" contract from the Spanish Ministry of Education and Science.Peer reviewe

Evaluation of environmental impact on natural populations of the Mediterranean killifish Aphanius fasciatus by quantitative RNA biomarkers

The Mediterranean killifish, Aphanius fasciatus (Valenciennes, 1821) represents an excellent sentinel species for analysis of environmental impact in coastal areas. Quantitative changes on liver mRNA levels of five stress-related genes – metallothionein (metal exposure), vitellogenin, (estrogenic effects), cytochrome P4501A (CYP1A, dioxin-like compounds), superoxide dismutase (oxidative stress) and HSP70 (general stress) – were tested as markers of exposure to pollutants both in natural populations and in experimental treatments with model effectors. Analysis of fish from a metal-contaminated site in the Tunisian coast (Sfax) indicated that fish presenting spinal deformities (eight times more frequent at this site than in reference sites) showed increased levels of hepatic CYP1A mRNA levels compared to non-deformed fish from the same site or from the reference Louza site, suggesting a combined impact by metals and organic pollutants. Due to the strategy used in their design, these quantitative RNA markers will likely be useful to analyze environmental impacts on other related fish species.This work has been supported by the Spanish Ministry of Research, Science and Innovation (CGL2008-01898) and the Spanish Agency of Cooperation for Development AECID (AECI A/019101/08). We thank also the Tunisian Ministry of Higher Education, Scientific Research and Technology.Peer reviewe

Abcb and Abcc transporter homologs are expressed and active in larvae and adults of zebra mussel and induced by chemical stress

Multixenobiotic resistance (MXR) of aquatic invertebrates has so far been associated with cellular efflux activity mediated by P-glycoprotein (ABCB1) and MRP (multidrug resistance protein; ABCC) type ABC (ATP binding cassette) transporters. Expression and activity of an abcb1/Abcb1 homolog has been shown in eggs and larvae of the zebra mussel Dreissena polymorpha. Here we report identification of a partial cDNA sequence of an abcc/Abcc homolog from zebra mussel that is transcribed and active as a cellular efflux transporter in embryos and gill tissue of adult mussels. Transcript expression levels were comparatively low in eggs and sharply increased after fertilization, then maintaining high expression levels in 1 and 2 dpf (days post fertilization) larvae. MK571, a known inhibitor of mammalian ABCC transporters, blocks efflux of calcein-am in larvae and gill tissue as indicated by elevated calcein fluorescence; this indicates the presence of active Abcc protein in cells of the larvae and gills. Dacthal and mercury used as chemical stressors both induced expression of abcb1 and abcc mRNAs in larvae; accordingly, assays with calcein-am and ABCB1 inhibitor reversin 205 and ABCC inhibitor MK571 indicated enhanced Abcb1 and Abcc efflux activities. Responses to chemicals were different in gills, where abcb1 transcript abundances were enhanced in dacthal and mercury treatments, whereas abcc mRNA was only increased with mercury. Abcb1 and Abcc activities did not in all cases show increases that were according to respective mRNA levels; thus, Abcc activity was significantly higher with dacthal, whereas Abcb1 activity was unchanged with mercury. Our data indicate that abcb1/Abcb1 and abcc/Abcc transporters are expressed and active in larvae and adult stages of zebra mussel. Expression of both genes is induced as cellular stress response, but regulation appears to differ in larvae and tissue of adult stages. © 2012 Elsevier B.V.This study was supported by the bi-lateral German (DAAD PPP) and Spanish project DE2009-0089, by the Spanish MICINN grants (CGL2008-01898/BOS and CTM2011-30471-C02-01) and by FEDER funds.Peer Reviewe