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      Brazilian silverside, Atherinella brasiliensis (Quoy & Gaimard,1825) embryos as a test-species for marine fish ecotoxicological tests

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          The fish embryo test (FET) is an alternative to the classic freshwater toxicity test used to assess environmental hazards and risks to fish. This test has been standardized and adopted by the Organization for Economic and Cooperation and Development (OECD). As salinity may affect the substances’ toxicity, we describe the development of an alternative euryhaline test species for embryonic ecotoxicological tests: the Brazilian silverside Atherinella brasiliensis (Quoy & Gaimard, 1825). This species is broadly distributed along the coast of South America and is able to inhabit a broad range of environmental and saline conditions. Ours is the first study on the maintenance of a native South American species for natural reproduction and the generation of embryos for tests. The embryos used are transparent and possess fluorescent cells which have only been seen in a few species and which may be used as markers, making it an alternative assessment tool for the lethal and sublethal substances in marine and estuarine environments. We provide a detailed description and analysis of embryonic development under different salinities and temperatures. The embryos and larvae developed in similar ways at different salinities, however as temperatures increased, mortality also increased. We considered the effects of the reference toxicants Zn 2+ and SDS using a protocol similar to the FET that was standardized for zebrafish. Brazilian silverside embryos are as sensitive as freshwater, or euryhaline fish, to the surfactant but are more resistant to metals prior to hatching. We were able to show the advantages of the Brazilian silverside as a model for a marine fish embryo test (FETm) with high levels of reproducibility and little contaminated waste.

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          Most cited references 58

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          Stages of embryonic development of the zebrafish.

          We describe a series of stages for development of the embryo of the zebrafish, Danio (Brachydanio) rerio. We define seven broad periods of embryogenesis--the zygote, cleavage, blastula, gastrula, segmentation, pharyngula, and hatching periods. These divisions highlight the changing spectrum of major developmental processes that occur during the first 3 days after fertilization, and we review some of what is known about morphogenesis and other significant events that occur during each of the periods. Stages subdivide the periods. Stages are named, not numbered as in most other series, providing for flexibility and continued evolution of the staging series as we learn more about development in this species. The stages, and their names, are based on morphological features, generally readily identified by examination of the live embryo with the dissecting stereomicroscope. The descriptions also fully utilize the optical transparancy of the live embryo, which provides for visibility of even very deep structures when the embryo is examined with the compound microscope and Nomarski interference contrast illumination. Photomicrographs and composite camera lucida line drawings characterize the stages pictorially. Other figures chart the development of distinctive characters used as staging aid signposts.
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            Stages of normal development in the medaka Oryzias latipes.

            Unfertilized eggs of Oryzias latipes were artificially inseminated and incubated at 26+/-1 degrees C. Careful observation of the process of embryonic development by light microscopy allowed division of the process into 39 stages based on diagnostic features of the developing embryos. The principal diagnostic features are the number and size of blastomeres, form of the blastoderm, extent of epiboly, development of the central nervous system, number and form of somites, optic and otic development, development of the notochord, heart development, blood circulation, the size and movement of the body, development of the tail, membranous fin (fin fold) development, and development of such viscera as the liver, gallbladder, gut tube, spleen and swim (air) bladder. After hatching, development of the larvae (fry) and young can be divided into six stages based on such diagnostic features as the fins, scales and secondary sexual characteristics.
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              Is the fish embryo toxicity test (FET) with the zebrafish (Danio rerio) a potential alternative for the fish acute toxicity test?

              The fish acute toxicity test is a mandatory component in the base set of data requirements for ecotoxicity testing. The fish acute toxicity test is not compatible with most current animal welfare legislation because mortality is the primary endpoint and it is often hypothesized that fish suffer distress and perhaps pain. Animal alternative considerations have also been incorporated into new European REACH regulations through strong advocacy for the reduction of testing with live animals. One of the most promising alternative approaches to classical acute fish toxicity testing with live fish is the fish embryo toxicity (FET) test. The FET has been a mandatory component in routine whole effluent testing in Germany since 2005 and has already been standardized at the international level. In order to analyze the applicability of the FET also in chemical testing, a comparative re-evaluation of both fish and fish embryo toxicity data was carried out for a total of 143 substances, and statistical approaches were developed to evaluate the correlation between fish and fish embryo toxicity data. Results confirm that fish embryo tests are neither better nor worse than acute fish toxicity tests and provide strong scientific support for the FET as a surrogate for the acute fish toxicity test.

                Author and article information

                PeerJ Inc. (San Diego, USA )
                14 April 2021
                : 9
                [1 ]Laboratório Integrado de Ciências Morfofuncionais, Instituto de Biodiversidade e Sustentabilidade (NUPEM), Universidade Federal do Rio de Janeiro , Macaé, RJ, Brazil
                [2 ]Laboratório Integrado de Biociências Translacionais, Instituto de Biodiversidade e Sustentabilidade (NUPEM), Universidade Federal do Rio de Janeiro , Macaé, RJ, Brazil
                [3 ]Petrobras Research and Development Center (CENPES), PETROBRAS , Rio de Janeiro, RJ, Brazil
                [4 ]LABTOX—Laboratório de Análise Ambiental Ltda , Rio de Janeiro, RJ, Brazil
                [5 ]Programa Pós-Graduação em Ciências Ambientais e Conservação (PPG-CiAC), Universidade Federal do Rio de Janeiro , Macaé, RJ, Brazil
                © 2021 Feitosa et al.

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ) and either DOI or URL of the article must be cited.

                Funded by: PETROBRAS
                Award ID: 0050.0088225.14.9
                Funded by: CAPES
                The work received financial support of PETROBRAS (Research Project n° 0050.0088225.14.9), and the Coordination for the Improvement of Higher Education Personnel (CAPES) for the fellowship given to Emiliano Nicolas Calderon. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Aquaculture, Fisheries and Fish Science
                Developmental Biology
                Marine Biology


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