The Fish Embryo Acute Toxicity (FET) Test was adopted by the Organisation for Economic Co-operation and Development as OECD TG 236 in 2013. The test has been designed to determine acute toxicity of chemicals on embryonic stages of fish and proposed as an alternative method to the Fish Acute Toxicity Test performed according to OECD TG 203. In recent years fish embryos were used not only in the assessment of toxicity of chemicals but also for environmental and wastewater samples. In our study we investigated the acute toxicity of treated wastewater from seven hospitals in the Czech Republic. Our main purpose was to compare the suitability and sensitivity of zebrafish embryos with the sensitivity of two other aquatic organisms commonly used for wastewater testing - Daphnia magna and Aliivibrio fischeri. For the aim of this study, in addition to the lethal endpoints of the FET test, sublethal effects such as delayed heartbeat, lack of blood circulation, pericardial and yolk sac edema, spinal curvature and pigmentation failures were evaluated. The comparison of three species demonstrated that the sensitivity of zebrafish embryos is comparable or in some cases higher than the sensitivity of D. magna and A. fischeri. The inclusion of sublethal endpoints caused statistically significant increase of the FET test efficiency in the range of 1-12 %. Based on our results, the FET test, especially with the addition of sublethal effects evaluation, can be considered as a sufficiently sensitive and useful additional tool for ecotoxicity testing of the acute toxicity potential of hospital effluents.

National Institute of Public Health Praha 10 Czech Republic

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ABBAS M, ADIL M, EHTISHAM-UL-HAQUE S, MUNIR B, YAMEEN M, GHAFFAR A, SHAR GA, TAHIR MA, IQBAL M. Vibrio fischeri bioluminescence inhibition assay for ecotoxicity assessment: A review. Sci Total Environ. 2018;626:1295–1309. doi: 10.1016/j.scitotenv.2018.01.066. PubMed DOI

BABIĆ S, BARIŠIĆ J, VIŠIĆ H, KLOBUČAR RS, TOPIĆ POPOVIĆ N, STRUNJAK-PEROVIĆ I, ČOŽ-RAKOVAC R, KLOBUČAR G. Embryotoxic and genotoxic effects of sewage effluents in zebrafish embryo using multiple endpoint testing. Water Res. 2017;115:9–21. doi: 10.1016/j.watres.2017.02.049. PubMed DOI

BOILLOT C, BAZIN C, TISSOT-GUERRAZ F, DROGUET J, PERRAUD M, CETRE JC, TREPO D, PERRODIN Y. Daily physicochemical, microbiological and ecotoxicological fluctuations of a hospital effluent according to technical and care activities. Sci Total Environ. 2008;403:113–129. doi: 10.1016/j.scitotenv.2008.04.037. PubMed DOI

BRANDHOF EJ, MONTFORTS M. Fish embryo toxicity of carbamazepine, diclofenac and metoprolol. Ecotoxicol Environ Saf. 2010;73:1862–1866. doi: 10.1016/j.ecoenv.2010.08.031. PubMed DOI

BRAUNBECK T, KAIS B, LAMMER E, OTTE J, SCHNEIDER K, STENGEL D, STRECKER R. The fish embryo test (FET): origin, applications, and future. Environ Sci Pollut Res. 2015;22:16247–16261. doi: 10.1007/s11356-014-3814-7. PubMed DOI

CEDRON VP, WEINER AMJ, VERA M, SANCHEZ L. Acetaminophen affects the survivor, pigmentation and development of craniofacial structures in zebrafish (Danio rerio) embryos. Biochem Pharmacol. 2020;174:113816. doi: 10.1016/j.bcp.2020.113816. PubMed DOI

CLEUVERS M. Aquatic ecotoxicity of pharmaceuticals including the assessment of combination effects. Toxicol Lett. 2003;142:185–194. doi: 10.1016/S0378-4274(03)00068-7. PubMed DOI

CLEUVERS M. Initial risk assessment for three beta-blockers found in the aquatic environment. Chemosphere. 2005;59:199–205. doi: 10.1016/j.chemosphere.2004.11.090. PubMed DOI

ČSN 75 6406: Wastewater discharge and treatment from health facilities. 2020

DANG Z, Van der VEN LTM, KIENHUIS AS. Fish embryo toxicity test, threshold approach, and moribund as approaches to implement 3R principles to the acute fish toxicity test. Chemosphere. 2017;186:677–685. doi: 10.1016/j.chemosphere.2017.08.047. PubMed DOI

DATEL JV, HRABANKOVA A. Pharmaceuticals load in the Svihov water reservoir (Czech Republic) and impacts on quality of treated drinking water. Water. 2020;12:1387. doi: 10.3390/w12051387. DOI

ELLEPOLA N, OGAS T, TURNER DN, GURUNG R, MALDONADO-TORRES S, TELLO-ABURTO R, PATIDAR PL, ROGELJ S, PIYASENA ME. A toxicological study on photo-degradation products of environmental ibuprofen: Ecological and human health implications. Ecotoxicol Environ Saf. 2020;188:109892. doi: 10.1016/j.ecoenv.2019.109892. PubMed DOI PMC

EMMANUEL E, HANNA K, BAZIN C, KECK G, CLÉMENT B, PERRODIN Y. Fate of glutaraldehyde in hospital wastewater and combined effects of glutaraldehyde and surfactants on aquatic organisms. Environ Int. 2005;31:99–406. doi: 10.1016/j.envint.2004.08.011. PubMed DOI

EUROPEAN PARLIAMENT AND COUNCIL. DIRECTIVE 2000/60/EC of the European Parliament and of the Council establishing a framework for Community action in the field of water policy (EU Water Framework Directive) Official Journal of the European Union L. 2000;327:1–73.

EUROPEAN PARLIAMENT AND COUNCIL. DIRECTIVE 2010/63/EU of the European Parliament and of the Council of 22 September 2010 on the protection of animals used for scientific purposes. Official Journal of the European Union L. 2010;276:33–79.

GODOY AA, OLIVEIRA ÁC, SILVA JGM, JESUS AZEVEDO CC, DOMINGUES I, NOGUEIRA AJA, KUMMROW F. Single and mixture toxicity of four pharmaceuticals of environmental concern to aquatic organisms, including a behavioural assessment. Chemosphere. 2019;235:73–382. doi: 10.1016/j.chemosphere.2019.06.200. PubMed DOI

HALLING-SØRENSEN B, HOLTEN LŰTZHØFT H-C, ANDERSEN HR, INGERSLEV F. Environmental risk assessment of antibiotics: comparison of mecillinam, trimethoprim and ciprofloxacin. J Antimicrob Chemother. 2000;46:53–58. doi: 10.1093/jac/46.suppl_1.53. PubMed DOI

HAMJINDA NS, CHIEMCHAISRI W, WATANABE T, HONDA R, CHIEMCHAISRI CH. Toxicological assessment of hospital wastewater in different treatment processes. Environ Sci Pollut Res. 2015;25:7271–7279. doi: 10.1007/s11356-015-4812-0. PubMed DOI

ISO 11348-2. Water quality – Determination of the inhibitory effect of water samples on the light emission of Vibrio fischeri (Luminescent bacteria test) - Part 2: Method using liquid-dried bacteria. 2007

ISO 15088. Water quality – Determination of the acute toxicity of waste water to zebrafish eggs (Danio rerio) 2007

ISO 6341. Water quality – Determination of the inhibition of the mobility of Daphnia magna Straus (Cladocera, Crustacea) – Acute toxicity test. 2012

ISO 7346-2. Water quality – Determination of the acute lethal toxicity of substances to a freshwater fish [Brachydanio rerio Hamilton-Buchanan (Teleostei, Cyprinidae)]-Part 2: Semi-static method. 1999

ISO 8692. Water quality – Fresh water algal growth inhibition test with unicellular green algae. 2012

JIROVA G, VLKOVA A, WITTLEROVA M, DVORAKOVA M, KASPAROVA L, CHRZ J, KEJLOVA K, WITTLINGEROVA Z, ZIMOVA M, HOSIKOVA B, JIRAVOVA J, KOLAROVA H. Toxicity of wastewater from health care facilities assessed by different bioassays. Neuro Endocrinol Lett. 2018;39:441–453. PubMed

JIROVA G, WITTLINGEROVA Z, ZIMOVA M, VLKOVA A, WITTLEROVA M, DVORAKOVA M, JIROVA D. Bioindicators of wastewater ecotoxicity. Neuro Endocrinol Lett. 2016;37:17–24. PubMed

KERN DI, OLIVEIRA SCHWAICKHARDT R, LUTTERBECK CA, KIST LT, ALCAYAGA EAL, MACHADO ÊL. Arch Environ Contam Toxicol. 2015;68:64–73. doi: 10.1007/s00244-014-0072-0. PubMed DOI

KIMMEL CHB, BALLARD WW, KIMMEL SR, ULLMANN B, SCHILLING TF. Stages of embryonic development of the zebrafish. Dev Dyn. 1995;203:253–310. doi: 10.1002/aja.1002030302. PubMed DOI

KRZYKWA JC, SAEID A, JEFFRIES MKS. Identifying sublethal endpoints for evaluating neurotoxic compounds utilizing the fish embryo toxicity test. Ecotoxicol Environ Saf. 2019;170:521–529. doi: 10.1016/j.ecoenv.2018.11.118. PubMed DOI

LAMMER E, CARR GJ, WENDLER K, RAWLINGS JM, BELANGER SE, BRAUNBECK T. Is the fish embryo toxicity test (FET) with the zebrafish (Danio rerio) a potential alternative for the fish acute toxicity test? Comp Biochem Phys C. 2009;149:196–209. doi: 10.1016/j.cbpc.2008.11.006. PubMed DOI

LAQUAZ M, DAGOT CH, BAZIN CH, BASTIDE T, GASCHET M, PLOY M-C, PERRODIN Y. Ecotoxicity and antibiotic resistance of a mixture of hospital and urban sewage in a wastewater treatment plant. Environ Sci Pollut Res. 2017;25:9243–9253. doi: 10.1007/s11356-017-9957-6. PubMed DOI

LI Q, WANG P, CHEN L, GAO H, WU L. Acute toxicity and histopathological effects of naproxen in zebrafish (Danio rerio) early life stages. Environ Sci Pollut Res. 2016;23:18832–18841. doi: 10.1007/s11356-016-7092-4. PubMed DOI

LI S-W, WANG Y-H, LIN AY-C. Ecotoxicological effect of ketamine: Evidence of acute, chronic and photolysis toxicity to Daphnia magna. Ecotoxicol Environ Saf. 2017;143:173–179. doi: 10.1016/j.ecoenv.2017.05.040. PubMed DOI

MARTINS N, PEREIRA R, ABRANTES N, GONÇALVES F, MARQUES CR. Ecotoxicological effects of ciprofloxacin on freshwater species: data integration and derivation of toxicity thresholds for risk assessment. Ecotoxicology. 2012;21:1167–1176. doi: 10.1007/s10646-012-0871-x. PubMed DOI

MEZA LC, PIOTROWSKI P, FARNAN J, TASKER TL, XIONG B, WEGGLER B, MURRELL K, DORMAN FL, HEUVEL JPV, BURGOS WD. Detection and removal of biologically active organic micropollutants from hospital wastewater. Sci Total Environ. 2020;700:134469. doi: 10.1016/j.scitotenv.2019.134469. PubMed DOI

OECD. OECD Guidelines for Testing of Chemicals, Section 2. Paris, France: 2019. Test Guideline No. 203: Fish, Acute Toxicity Test.

OECD. OECD Guidelines for Testing of Chemicals, Section 2. Paris, France: 2013. Test Guideline No. 236: Fish Embryo Acute Toxicity (FET) Test.

ORIAS F, PERRODIN Y. Characterisation of the ecotoxicity of hospital effluents: A review. Sci Total Environ. 2013;454–455:250–276. doi: 10.1016/j.scitotenv.2013.02.064. PubMed DOI

PÉREZ-ALVAREZ I, ISLAS-FLORES H, GÓMEZ-OLIVÁN LM, BARCELÓ D, De ALDA ML, SOLSONA SP, SÁNCHEZ-ACEVES L, SANJUAN-REYES N, GALAR-MARTÍNEZ M. Determination of metals end pharmaceutical compounds released in hospital wastewater from Toluca, Mexico, and evaluation of their toxic impact. Environ Pollut. 2018;240:30–341. doi: 10.1016/j.envpol.2018.04.116. PubMed DOI

ROGOWSKA J, CIESZYNSKA-SEMENOWICZ M, RATAJCZYK W, WOLSKA L. Micropollutants in treated wastewater. Ambio. 2020;49:487–503. doi: 10.1007/s13280-019-01219-5. PubMed DOI PMC

ROMANUCCI V, SICILIANO A, GALDIERO E, GUIDA M, LUONGO G, LIGUORI R, FABIO G, PREVITERA L, ZARRELLI A. Disinfection by-products and ecotoxic risk associated with hypochlorite treatment of tramadol. Molecules. 2019;24:693. doi: 10.3390/molecules24040693. PubMed DOI PMC

SANTOS LHMLM, ARAÚJO AN, FACHINI A, PENA A, DELERUE-MATOS C, MONTENEGRO MCBSM. Ecotoxicological aspects related to the presence of pharmaceuticals in the aquatic environment. J Hazard Mater. 2010;175:45–95. doi: 10.1016/j.jhazmat.2009.10.100. PubMed DOI

SCHOLZ S, FISCHER S, GŰNDEL U, KŰSTER E, LUCKENBACH T, VOELKER D. The zebrafish embryo model in environmental risk assessment-applications beyond acute toxicity testing. Environ Sci Pollut Res. 2008;15:394–404. doi: 10.1007/s11356-008-0018-z. PubMed DOI

SOBANSKA M, SCHOLZ S, NYMAN A-M, CESNAITIS R, ALONSO SG, KLŰVER N, KŰHNE R, TYLE H, KNECHT J, DANG Z, LUNDBERGH I, CARLON C, COEN W. Applicability of the Fish Embryo Acute Toxicity (FET) Test (OECD 236) in the Regulatory Context of Registration, Evaluation, Authorisation, and Restriction of Chemicals (REACH) Environ Toxicol Chem. 2017;37:657–670. doi: 10.1002/etc.4055. PubMed DOI

SOKAL RR, ROHLF FJ. Biometry The Principles and Practice of Statistics in Biological Research. 3rd Edition. W.H Freeman and Co; New York: 1995. p. 887.

STELZER JAA, ROSIN CK, BAUER LH, HARTMANN M, PULGATI FH, ARENZON A. Is Fish Embryo Test (FET) According to OECD 236 sensible enough for delivering quality data for effluent risk assessment? Environ Toxicol Chem. 2018;37:2925–2932. doi: 10.1002/etc.4215. PubMed DOI

TENORIO-CHÁVEZ P, CERRO-LÓPEZ M, CASTRO-PASTRANA LI, RAMÍREZ-RODRIGUES MM, OROZCO-HERNÁNDEZ JM, GÓMEZ-OLIVÁN LM. Effects of effluent from a hospital in Mexico on the embryonic development of zebrafish, Danio rerio. Sci Total Environ. 2020;727:1–11. doi: 10.1016/j.scitotenv.2020.138716. PubMed DOI

VASCONCELOS EC, DALKE CR, OLIVEIRA CMR. Influence of select antibiotics on Vibrio fischeri and Desmodesmus subspicatus at μg L-1 concentrations. J Environ Manage. 2017;60:157–164. doi: 10.1007/s00267-017-0841-4. PubMed DOI

VÄLITALO P, MASSEI R, HEISKANEN I, BEHNISCH P, BRACK W, TINDALL AJ, PASQUIER D, KŰSTER E, MIKOLA A, SCHULZE T, SILLANPÄÄ M. Effect-based assessment of toxicity removal during wastewater treatment. Water Res. 2017;126:153–163. doi: 10.1016/j.watres.2017.09.014. PubMed DOI

VERLICCHI P, AL AUKIDY M, ZAMBELLO E. What have we learned from worldwide experiences on the management and treatment of hospital effluent? - An overview and a discussion on perspectives. Sci Total Environ. 2015;514:467–491. doi: 10.1016/j.scitotenv.2015.02.020. PubMed DOI PMC

WIEST L, CHONOVA T, BERGÉ A, BAUDOT R, BESSUEILLE-BARBIER F, AYOUNI-DEROUICHE L, VULLIET E. Two-year survey of specific hospital wastewater treatment and its impact on pharmaceutical discharges. Environ Sci Pollut Res. 2018;25:9207–9218. doi: 10.1007/s11356-017-9662-5. PubMed DOI

WIGH A, DEVAUX A, BROSSELIN V, GONZALES-OSPINA A, DOMENJOUD B, AÏT-AÏSSA S, CREUSOT N, GOSSET A, BAZIN CH, BONY S. Proposal to optimize ecotoxicological evaluation of wastewater treated by conventional biological and ozonation processes. Environ Sci Pollut Res. 2016;23:008–3017. doi: 10.1007/s11356-015-5419-1. PubMed DOI