Pesticide Testing

Pesticides (also known as biocides, agrochemicals, and plant-protection products) undergo a battery of tests to fulfil government regulatory agencies’ data requirements for health and safety. These include tests for eye and skin irritation, skin sensitisation, and acute oral, inhalation, and dermal toxicities—the “six-pack” of acute toxicity tests—as well as studies on reproduction and development, neurotoxicity, immunotoxicity, carcinogenicity, mutagenicity, and ecological effects. Many of the required tests use animals, and in the United States, an estimated 7,100 rats, mice, rabbits, birds, fish, and dogs are used to test a single active ingredient, and additional animals are used to test various formulations.

Eye Irritation/Corrosion

The use of non-animal methods to assess the eye irritation potential of pesticides may be allowed (or even required), depending on the regulatory region. In the US, the Environmental Protection Agency (EPA) Office of Pesticide Programs (OPP) accepts the use of an alternate framework for evaluating eye irritation, but non-animal methods are not widely used. The Science Consortium has proposed a number of actions that can be taken to overcome the identified barriers to their use (Clippinger, Hill et al, 2016). These actions include promoting global regulatory acceptance, increasing regulatory reviewers’ awareness of and experience with alternative methods through training, providing incentives for companies that use the methods by expediting their registration submissions, reconsidering algorithms for predicting hazard category, and making a transition from voluntary to mandatory use.

To help overcome these barriers, the Science Consortium collaborated with the EPA OPP, the European Commission’s Joint Research Centre, and others to review the available test methods (the in vitro, ex vivo, and rabbit tests) and show that the in vitro and ex vivo methods are less variable and as or more human-relevant than the currently used rabbit test. The review also states that the rabbit test is not a reliable reference standard and that new methods should not be compared to the rabbit test to show their validity (Clippinger, Raabe et al, 2021). Working with the NTP Interagency Center for the Evaluation of Alternative Toxicological Methods (NICEATM), the Science Consortium has also coordinated and funded the testing of agrochemical formulations (with existing rabbit data) in the various in vitro and ex vivo methods to better understand how the different methods compare (report forthcoming).

Avian Dietary Toxicity

For decades, the EPA and other regulatory agencies have required data from avian acute oral toxicity (OCSPP 850.2100) and avian dietary toxicity (OCSPP 850.2200) tests for the registration of pesticides. However, trends over the past 20 years have suggested that the avian sub-acute dietary test generally does not drive risk management decisions, and scientists from the Science Consortium and EPA conducted a detailed retrospective analysis to confirm this hypothesis.

One hundred nineteen pesticides registered between 1998 and 2017 were evaluated, and risk quotient (RQ) data from the avian toxicity tests were compared with RQ data from ecological risk assessments. Results from the comparison showed that no pesticides had an avian dietary RQ greater than the level of concern that was not also identified in the acute oral test.

These results demonstrate that the sub-acute dietary test conducted on birds is not used in risk management and can be removed without causing harm to the environment (see Hilton, Odenkirchen et al, 2019). This analysis was used to support the EPA’s guidance for waiving the avian sub-acute dietary test for pesticide registration (see here).


The Science Consortium, government, and industry are working to modernise carcinogenicity testing through the Rethinking Carcinogenicity Assessment for Agrochemicals Project (ReCAAP). More information on this project and carcinogenicity testing can be found here.

Acute Systemic Toxicity

The Science Consortium partnered with NICEATM, the EPA, and others on the implementation of alternative approaches for acute systemic toxicity testing, including a publication outlining current regulatory requirements, data obtained from the currently required tests, data actually used and needed by regulators, and opportunities for the use of non-animal approaches to meet regulatory needs (Strickland, Clippinger et al, 2018). The publication of this paper was an action item proposed at a 2015 workshop co-organized by the Science Consortium, and was followed, in 2016, with a workshop focused on acute inhalation toxicity testing (Clippinger, Allen et al, 2018). A summary of the Science Consortium’s extensive work on respiratory toxicity testing can be found here.

One-Year Dog Toxicity Test

Many countries used to require both a three-month (sub-chronic) study and one-year (chronic) study in dogs as part of the data requirements for registering pesticide active ingredients. In these tests, beagles are exposed to pesticides orally or via inhalation. Because the two studies have been required for more than 30 years, there is an extensive database of results. Beginning in the late 1990s, numerous scientific articles1-10 have been published comparing the results of the two studies and supporting the conclusion that a one-year test in addition to the three-month test is of little value and that its elimination would not compromise human safety or protection.

In the US, following urging from Science Consortium member PETA US and other groups, the EPA conducted its own comparison of data from the two tests and, after reaching the same conclusion as previous investigators, removed the one-year study from its requirements in 2007.11 The EU passed legislation in March 2013 also eliminating the requirement for the one-year dog study. Consortium scientists subsequently began contacting regulatory agencies in other parts of the world, including Canada, Australia, Japan, South Korea, India, and Brazil, urging them to follow suit.

Following discussions between the Science Consortium and Health Canada staff, Canada eliminated its requirement in March 2016. Japan and South Korea eliminated their requirements in April 2018, and Brazil eliminated theirs in July 2019. Australia, China, and India have indicated that they can waive or do not require the one-year test.

Advancing the Use of Non-Animal Methods

On 17 March 2016, the EPA OPP announced its commitment to transition towards the use of non-animal testing approaches that will enhance the quality of risk assessment and ensure better protection of human health and the environment. This announcement was followed in September 2019 with a commitment to replace the use of mammals in pesticide toxicity tests by 2035. In June 2020, the EPA issued a work plan to prioritise agency efforts towards meeting this goal and created a metrics webpage to help gauge progress and identify priorities for future work. The EPA OPP has taken numerous steps to reach its goal, including the following:

  • It issued guidance for waiving acute dermal toxicity testing of pesticide formulations (2016) and active ingredients (2020).
  • It instituted a policy accepting the use of in vitro defined approaches for skin sensitisation testing of single chemicals (2018).
  • It instituted a policy to reduce the number of test concentrations – and therefore fish – used in fish bioconcentration tests (2020).
  • It conducted a risk assessment for the re-registration of six isothiazolinones using in vitro tests and an artificial neural network-based defined approach (2020).
  • It is investigating the use of the GHS additivity formula, which uses toxicity data from the individual ingredients in formulations to estimate the toxicity of end-products without having to conduct additional testing (report expected in 2021).

The Science Consortium participates in a workgroup of pesticide industry members, animal welfare representatives, and other stakeholders convened by the EPA OPP that discusses current efforts to reduce the number of animals used in the six-pack of acute toxicity testing. In addition, a Science Consortium scientist participates in discussions of the Pesticide Program Dialog Committee (PPDC) and has been involved in its 21st Century Toxicology Workgroup. The PPDC is a federal advisory committee for the EPA OPP with members representing industry, growers, and the animal protection, environmental, and farmworker communities. It provides the EPA OPP with feedback on various regulatory, policy, and programme implementation issues related to pesticides.

The Science Consortium organises training opportunities to familiarise regulatory staff with non-animal test methods. These sessions are led by in silico and in vitro method experts, such as the Institute for In Vitro Sciences and the Laboratory of Mathematical Chemistry. The Science Consortium co-organises a webinar series on the use of new approach methods in risk assessment with the agency.

In the EU, the Science Consortium participates in member state competent authority meetings for biocides, in which data requirements are discussed and updated. The Science Consortium works with the European Commission, member state authorities, animal protection organisations, and industry representatives on a number of far-reaching regulatory initiatives, such as the Chemicals Strategy for Sustainability, to replace tests on animals with robust non-animal approaches.

Similarly, in India, the Science Consortium works with the Central Insecticide Board and Registration Committee and industry members, providing scientific recommendations on the implementation of effective, non-animal methods.

  • References

    [1]Gerbracht U, Spielmann H. The use of dogs as second species in regulatory testing of pesticides. I. Interspecies comparison. Arch Toxicol. 1998;72(6):319–329.

    [2]Spielmann H, Gerbracht U. The use of dogs as second species in regulatory testing of pesticides. Part II: Subacute, subchronic and chronic studies in the dog. Arch Toxicol. 2001;75(1):1–21.

    [3]Baetcke KP, Phang W, Dellarco V. A comparison of the results of studies on pesticides from 12- or 24-month dog studies with dog studies of shorter duration. Health Effects Division, Office of Pesticide Programs. U.S. Environmental Protection Agency. Available at Published 2005.

    [4]Box RJ, Spielmann H. Use of the dog as non-rodent species in the safety testing schedule associated with the registration of crop and plant protection products (pesticides): Present status. Arch Toxicol. 2005;79(11):615–626.

    [5]Doe JE, Boobis AR, Blacker A, et al. A tiered approach to systemic toxicity testing for agricultural chemical safety assessment. Crit Rev Toxicol. 2006;36(1):37–68.

    [6]U.S. Environmental Protection Agency. Length of dog toxicity study(ies) that is appropriate for chronic RfD determinations of pesticide chemicals. Health Effects Division, Office of Pesticide Programs. Available at (Docket # EPA-HQ-OPP-2004-0387-0179). Published 2006.

    [7]van Ravenzwaay B. Initiatives to decrease redundancy in animal testing of pesticides. ALTEX. 2010;27(3):112-114.

    [8]Dellarco VL, Rowland J, May B. A retrospective analysis of toxicity studies in dogs and impact on the chronic reference dose for conventional pesticide chemicals. Crit Rev Toxicol. 2010;40(1):16–23.

    [9]Kobel W, Fegert I, Billington R, et al. A 1-year toxicity study in dogs is no longer a scientifically justifiable core data requirement for the safety assessment of pesticides. Crit Rev Toxicol. 2010;40(1):1–15.

    [10]Kobel W, Fegert I, Billington R, et al. Relevance of the 1-year dog study in assessing human health risks for registration of pesticides. An update to include pesticides registered in Japan. Crit Rev Toxicol. 2014;44(10):842–848.

    [11]US Environmental Protection Agency. Pesticides: data requirements for conventional chemicals, technical amendments, and data requirements for biochemical and microbial pesticides; final rules. Federal Register Notice 72(207):60934-60988. Available at Published 2007.

For a list of our publications, see here.