## 4.3.4. Selection of test organisms - Eco animals

Author: Michiel Kraak

Reviewers: Kees van Gestel, Jörg Römbke

Learning objectives:

You should be able to

- name the requirements for suitable laboratory ecotoxicity test organisms.
- list the most commonly used standard test organisms per environmental compartment.
- argue the need for more than one test species and the need for non-standard test organisms.

Key words: Test organism, standardized laboratory ecotoxicity tests, environmental compartment, habitat, different trophic levels

Introduction

Standardized laboratory ecotoxicity tests require constant test conditions, standardized endpoints (see section on Endpoints) and good performance in control treatments. Actually, in reliable, reproducible and easy to perform toxicity tests, the test compound should be the only variable. This sets high demands on the choice of the test organisms.

For a proper risk assessment, it is crucial that test species are representative of the community or ecosystem to be protected. Criteria for selection of organisms to be used in toxicity tests have been summarized by Van Gestel et al. (1997). They include: 1. Practical arguments, including feasibility, cost-effectiveness and rapidity of the test, 2. Acceptability and standardisation of the tests, including the generation of reproducible results, and 3. Ecological significance, including sensitivity, biological validity etc. The most practical requirement is that the test organism should be easy to culture and maintain, but equally important is that the test species should be sensitive towards different stressors. These two main requirements are, however, frequently conflicting. Species that are easy to culture are often less sensitive, simply because they are mostly generalists, while sensitive species are often specialists, making it much harder to culture them. For scientific and societal support of the choice of the test organisms, preferably they should be both ecologically and economically relevant or serve as flagship species, but again, these are opposite requirements. Economically relevant species, like crops and cattle, hardly play any role in natural ecosystems, while ecologically highly relevant species have no obvious economic value. This is reflected by the research efforts on these species, since much more is known about economically relevant species than about ecologically relevant species.

There is no species that is most sensitive to all pollutants. Which species is most sensitive depends on the mode of action and possibly also other properties of the chemical, the exposure route, its availability and the properties of the organism (e.g., presence of specific targets, physiology, etc.). It is therefore important to always test a number of species, with different life traits, functions, and positions in the food web. According to Van Gestel et al. (1997) such a battery of test species should be:

1. Representative of the ecosystem to protect, so including organisms having different life-histories, representing different functional groups, different taxonomic groups and different routes of exposure;

2. Representative of responses relevant for the protection of populations and communities; and

3. Uniform, so all tests in a battery should be applicable to the same test media and applying to the same test conditions, e.g. the same range of pH values.

Representation of environmental compartments

Each environmental compartment, water, air, soil and sediment, requires its specific set of test organisms. The most commonly applied test organisms are daphnids (Daphnia magna) for water, chironomids (Chironomus riparius) for sediments and earthworms (Eisenia fetida) for soil. For air, in the field of inhalation toxicology, humans and rodents are actually the most studied organism. In ecotoxicology, air testing is mostly restricted to plants, concerning studies on toxic gasses. Besides the most commonly applied organisms, there is a long list of other standard test organisms for which test protocols are available (Table 1; OECD site).

Table 1. Non-exhaustive list of standard ecotoxicity test species.

Environmental compartment(s) | Organism group | Test species
Water | Plant | Myriophyllum spicatum
Water | Plant | Lemna
Water | Algae | Species of choice
Water | Cyanobacteria | Species of choice
Water | Fish | Danio rerio
Water | Fish | Oryzias latipes
Water | Amphibian | Xenopus laevis
Water | Insect | Chironomus riparius
Water | Crustacean | Daphnia magna
Water | Snail | Lymnaea stagnalis
Water | Snail | Potamopyrgus antipodarum
Water-sediment | Plant | Myriophyllum spicatum
Water-sediment | Insect | Chironomus riparius
Water-sediment | Oligochaete worm | Lumbriculus variegatus
Sediment | Anaerobic bacteria | Sewage sludge
Soil | Plant | Species of choice
Soil | Oligochaete worm | Eisenia fetida or E. andrei
Soil | Oligochaete worm | Enchytraeus albidus or E. crypticus
Soil | Collembolan | Folsomia candida or F. fimetaria
Soil | Mite | Hypoaspis (Geolaelaps) aculeifer
Soil | Microorganisms | Natural microbial community
Dung | Insect | Scathophaga stercoraria
Dung | Insect | Musca autumnalis
Air-soil | Plant | Species of choice
Terrestrial | Bird | Species of choice
Terrestrial | Insect | Apis mellifera
Terrestrial | Insect | Bombus terrestris/B. impatiens
Terrestrial | Insect | Aphidius rhopalosiphi
Terrestrial | Mite | Typhlodromus pyri

Non-standard test organisms

The use of standard test organisms in standard ecotoxicity tests performed according to internationally accepted protocols strongly reduces the uncertainties in ecotoxicity testing. Yet, there are good reasons for deviating from these protocols. The species in Table 1 are listed according to their corresponding environmental compartment, but ignores differences between ecosystems and habitats. Soils may differ extensively in composition, depending on e.g. the sand, clay or silt content, and properties, e.g. pH and water content, each harbouring different species. Likewise, stagnant and current water have few species in common. This implies that based on ecological arguments there may be good reasons to select non-standard test organisms. Effects of compounds in streams can be better estimated with riverine insects rather than with the stagnant water inhabiting daphnids, while the compost worm Eisenia fetida is not necessarily the most appropriate species for sandy soils. The list of non-standard test organisms is of course endless, but if the methods are well documented in the open literature, there are no limitations to employ these alternative species. They do involve, however, experimental challenges, since non-standard test organisms may be hard to culture and to maintain under laboratory conditions and no protocols are available for the ecotoxicity test. Thus increasing the ecological relevance of ecotoxicity tests also increases the logistical and experimental constraints (see chapter 6 on Risk assessment).

Increasing the number of test species

The vast majority of toxicity tests is performed with a single test species, resulting in large margins of uncertainty concerning the hazardousness of compounds. To reduce these uncertainties and to increase ecological relevance it is advised to incorporate more test species belonging to different trophic levels, for water e.g. algae, daphnids and fish. For deriving environmental quality standards from Species Sensitivity Distributions (see section on SSDs) toxicity data is required for minimal eight species belonging to different taxonomical groups. This obviously causes tension between the scientific requirements and the available financial resources.

References

OECD site. https://www.oecd-ilibrary.org/environment/oecd-guidelines-for-the-testing-of-chemicals-section-2-effects-on-biotic-systems_20745761.

Van Gestel, C.A.M., Léon, C.D., Van Straalen, N.M. (1997). Evaluation of soil fauna ecotoxicity tests regarding their use in risk assessment. In: Tarradellas, J., Bitton, G., Rossel, D. (Eds). Soil Ecotoxicology. CRC Press, Inc., Boca Raton: 291-317.