Adverse effects in the fish embryo acute toxicity (FET) test: a catalogue of unspecific morphological changes versus more specific effects in zebrafish (Danio rerio) embryos

Abstract

Background The Fish Embryo Acute Toxicity (FET) test with the zebrafish (Danio rerio) embryo, the OECD test guideline (TG) 236, has been designed as an alternative for acute fish toxicity testing such as the OECD Acute Fish Toxicity Test (TG 203). To provide equivalent sensitivity to the acute fish test, the original FET test was designed to use only four morphological core endpoints: coagulation of the embryo, lack of somite formation, lack of heart beat, and non-detachment of the tail. These endpoints were selected due to (1) their association with mortality, directly or indirectly, (2) improve the practicality for screening by well-trained technical staff, and (3) the endpoints being relatively simple morphological alterations. Results With the growing need to understand the developmental toxicity of compounds found in the environment, the FET protocol has repeatedly been extended to a multitude of additional morphological endpoints that also allow the monitoring of teratogenicity. As the extensive use of the FET test has generated a multitude of observations in the scientific literature, a harmonisation of the terminology used for the description of the morphological effects seen after chemical exposure has become necessary. Conclusion For this end, the present communication provides an overview of both common and selected more specific morphological effects seen in zebrafish embryos after exposure to a wide variety of chemical substances together with suggestions for a harmonised nomenclature.

Specificity of time- and dose-dependent morphological endpoints in the fish embryo acute toxicity (FET) test for substances with diverse modes of action: the search for a “fingerprint”

“…In contrast, the concentration-response relationship of the insecticide carbaryl displayed a much steeper slope, i.e., sublethal and lethal toxicity data were much closer (EC 10 , 2.2 ± 0.26 mg/L, and LC 50 , 12.2 ± 0.72 mg/L). For further details and analyses of LC and EC data in zebrafish embryos, see von Hellfeld et al ( 2020 ), where FET observations were discussed in the context of a catalog of FET endpoints.…”

“…In contrast to a previous communication, which aimed at compiling a comprehensive and standardized catalog of observations in the FET test (von Hellfeld et al 2020 ), the present study was designed to also analyze the onset and frequency (severity) of observations. Table 5 summarizes observations made at test concentrations ≤ EC 10 .…”

“…In addition to the endpoints specified by OECD TG 236, namely (1) coagulation of fertilized eggs, (2) lack of somite formation, (3) non-detachment of tail bud, and (4) lack of heartbeat (OECD 2013 ), any other observation was recorded as further lethal or sublethal morphological endpoints: Common examples were reduced heartbeat or reduced blood flow, inhibited or missing pigmentation, delayed or altered development, modified movement(s), distortion of the spine, craniofacial deformations, eye development, delayed hatching and fin formation, and formation of various types of edema (von Hellfeld et al 2020 ). In the case of evidence for delayed toxicity, the standard exposure duration of 96 h specified by OECD TG 236 (OECD 2013 ) was extended to 120 h. In any case, the developmental stage at the end of the experiments never exceeded the limits for unprotected developmental stages set by the current EU animal welfare legislation (EU 2010 ; Strähle et al 2012 ).…”

“…Since tests with vertebrates are an integral part of environmental hazard identification and risk assessment of chemicals, plant protection products, pharmaceuticals, biocides, feed additives, and effluents (Scholz et al 2013 ), this increase in testing requirements has raised increasing concern about animal welfare (Braunbeck et al 2005 , 2015 ; Paparella et al 2020 ; von Hellfeld et al 2020 ). In order to meet these concerns, in 2003, Germany replaced whole effluent acute fish toxicity (AFT) testing according to OECD TG 203 (OECD 1992 , 2019 ) with the zebrafish ( Danio rerio ) fish egg test (Bundesgesetzblatt 2005 ; ISO 2007 ), and, in 2013, the OECD adopted the fish embryo acute toxicity (FET) test (TG 236; (OECD 2013 )) as an alternative method for the AFT test.…”

Comparative Toxicity Assessment of Kratom Decoction, Mitragynine and Speciociliatine Versus Morphine on Zebrafish (Danio rerio) Embryos

“…An example, drugs such as alcohol and nicotine that are known to affect human fetal development have been reported to induce morphological defects in the zebrafish embryos ( Lantz-McPeak et al, 2015 ). Spinal curvature can be further specified into three types: lordosis (spine curved inward), kyphosis (spine curved outward), and scoliosis (spine curved sideways) ( von Hellfeld et al, 2020 ). In this study, spinal curvature (scoliosis) was observed in mitragynine (50 μg/ml) and speciociliatine (25 and 50 μg/ml) exposed groups.…”