Fish and Their Role in Regulatory Guidelines

There is a vast array of international guidelines that implement fish and amphibians as models for screening. One of the more relevant is the Guidelines from the Organization for Economic Co-operation and Development (OECD), which provides guidelines and standards for testing and assessing the safety of chemicals, including those about fish and amphibians. The International Organization for Standardization (ISO) guidelines serve as a global reference point for industries and organizations, fostering consistency, interoperability, and reliability in the context of environmental science, where ISO standards play a crucial role in ensuring that research methodologies are standardized, reliable, and comparable across different studies. The US Environmental Protection Agency (EPA), whose guidelines play a crucial role in safeguarding ecosystems and human health. Understanding the impact of pollutants on aquatic life is a vital aspect of these guides.

Guidelines for Fish and Amphibians Testing

OECD Guidelines

The OECD has established specific guidelines for fish and amphibian testing to evaluate the effects of chemicals on aquatic organisms. These guidelines outline standardized protocols for toxicity testing using various fish species, ensuring consistency and comparability of data across different laboratories and countries. Some of the most interesting ones in environmental toxicology are:

  • OECD Test Guideline 203, Fish Acute Toxicity Test: This guideline describes a test method for determining the acute toxicity of chemicals in fish. It involves exposing fish to varying concentrations of a substance to determine lethal effects within a specified timeframe, suggesting different fish species that could be used, including zebrafish,
  • OECD Test Guideline 210, Fish Early-Life Stage Toxicity Test: This guideline focuses on the effects of chemicals on the early life stages of fish, including embryo and larval development. It assesses endpoints such as hatching success, abnormalities, and growth, including the evaluation of them in zebrafish embryos/larvae.
  • OECD Test Guideline 215, Fish Juvenile Growth Test: This guideline evaluates the chronic effects of chemicals on fish growth over an extended period, providing information on long-term exposure effects in different fish species, including juvenile zebrafish.
  • OECD Test Guideline 236, Fish Embryo Acute Toxicity (FET) Test: This guideline is used to determine acute toxicity of chemicals in the embryonic stages of zebrafish. At the end of the exposure period of 96 hours, the acute toxicity is determined by phenotypical screening, and lethality analysis.


ISO Guidelines

The ISO guidelines applied for environmental science determined the tests to be done for standardized, reliable, and comparable water quality controls. The most relevant one with fish is:

  • ISO 7346-3 - Water Quality - Determination of the Acute Lethal Toxicity of Substances to a Freshwater Fish: The use of zebrafish in acute toxicity testing exemplifies adherence to ISO 7346-3, providing standardized protocols for evaluating the lethal effects of substances on freshwater fish.

EPA Guidelines

  • Ambient Water Quality Criteria (AWQC) are specific water quality standards established by the EPA to determine the maximum concentrations tolerated for good water quality, regarding different contaminants such as metals (like mercury and arsenic) and organic compounds (like organochlorine pesticides, aroclors, and polychlorinated biphenyl congeners). 

Fish Species Recommended by OECD, ISO, and EPA  

The regulatory guidelines recommend specific fish species for conducting toxicity tests due to their suitability for various types of assessments:

Zebrafish (Danio rerio): Widely used in developmental and genetic studies due to their embryo transparency and genetic homology with humans (more than 80% homology in genetic variations associated with human diseases). Zebrafish are especially valuable for assessing the effects of chemicals on embryonic development and toxicity screenings

Rainbow Trout (Oncorhynchus mykiss): Often used for acute and chronic toxicity testing due to their sensitivity to contaminants. They are particularly valuable for assessing the effects of chemicals on freshwater ecosystems.

Fathead Minnow (Pimephales promelas): Utilized in toxicity studies for their adaptability and relevance to North American freshwater environments. They serve as indicators of chemical impacts on aquatic ecosystems. 

Japanese Medaka (Oryzias latipes): Known for their fast reproduction, adaptability to laboratory settings, and sensitivity to environmental changes. Japanese medaka are used in developmental and genetic studies, as well as for assessing chemical effects on aquatic organisms. 

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Role of OECD, ISO, and EPA Guidelines in Regulatory Compliance 

Adherence to OECD guidelines ensures that toxicity testing is conducted using standardized methods, facilitating the acceptance and comparison of data across countries and regulatory entities. Data generated through OECD-compliant studies form the basis for risk assessments and regulatory decisions concerning the approval, use, and management of chemicals to protect both the environment and human health.

Overall, the OECD guidelines for fish and amphibian testing and the recommended fish species play a crucial role in providing scientifically robust data that supports regulatory compliance and decision-making, contributing to the protection of aquatic ecosystems and human well-being.

The use of fish as scientific models in environmental research aligns harmoniously with the principles and guidelines set forth by the ISO. These aquatic organisms not only offer standardized research platforms but also contribute to the ecological relevance and ethical considerations championed by international standards.

The EPA is responsible for setting standards and guidelines to regulate pollutants and protect the environment. These guidelines are essential for maintaining ecological balance, ensuring the sustainability of ecosystems, and safeguarding human health. To establish effective guidelines, it is imperative to conduct comprehensive scientific studies, and fish, as a biological model, provides valuable insights into the effects of pollutants on aquatic environments.

Therefore, regulatory guidelines are crucial for ensuring the security of chemical compounds by setting the standards of the testing conditions. The use of fish is vital for ecotoxicity as they are representative organisms of aquatic ecosystems, present high sensitivity to chemicals, have similar physiology to other vertebrates including humans, and there is the availability of standardized High-Content Assays with them, decreasing the time and cost to develop them.

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OECD (2019), Test No. 203: Fish, Acute Toxicity Test, OECD Guidelines for the Testing of Chemicals, Section 2, OECD Publishing, Paris,

OECD (2013), Test No. 210: Fish, Early-life Stage Toxicity Test, OECD Guidelines for the Testing of Chemicals, Section 2, OECD Publishing, Paris,

OECD (2000), Test No. 215: Fish, Juvenile Growth Test, OECD Guidelines for the Testing of Chemicals, Section 2, OECD Publishing, Paris,

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

Burden, N., Benstead, R., Benyon, K., Clook, M., Green, C., Handley, J., Harper, N., Maynard, S. K., Mead, C., Pearson, A., Ryder, K., Sheahan, D., Egmond, R., Wheeler, J. R., & Hutchinson, T. H. (2020). Key Opportunities to Replace, Reduce, and Refine Regulatory Fish Acute Toxicity Tests. Environmental Toxicology and Chemistry, 39(10), 2076–2089.

Pryshlakivsky, J., & Searcy, C. (2013). Fifteen years of ISO 14040: a review. Journal of Cleaner Production, 57, 115–123.

Church, B. G., Van Sprang, P. A., Chowdhury, M. J., & DeForest, D. K. (2017). Updated species sensitivity distribution evaluations for acute and chronic lead toxicity to saltwater aquatic life. Environmental Toxicology and Chemistry, 36(11), 2974–2980.


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