Earthworm Acute Toxicity test (Eisenia foetida) (OECD 207: 1984)

This Standard includes two methods: a contact toxicity test on paper and a toxicity test on artificial soil. The recommended earthworm species is Eisenia foetida, although the norm admits that other species can be used.

The contact test on filter paper is an optional initial screening test used to identify potentially toxic chemicals for earthworms in the soil, which must be complemented by more detailed tests on artificial soil. This test involves exposure of earthworms to the substances evaluated in moistened filter papers placed in vials where a worm is introduced per vial. In this test, a geometric series of at least five or more treatment concentrations should be used with the product under test (expressed in mg/cm2) and at least ten replicates (with one worm per vial) for each treatment, and the respective controls, that is to say a minimum of 60 tests (60 worms). The tests are performed in the dark at 20 ± 2°C and for a period of 48 hours after which the mortality is evaluated. In addition, the standard admits the evaluation of an additional evaluation time of 72 hours.

The test on artificial soil provides toxicity data more representative of the natural exposure of earthworms to chemical products. This test involves keeping earthworms in samples of an accurately defined artificial soil containing the test substance. A geometric series of at least five or more treatment concentrations (expressed in mg/Kg of dry weight) and at least four replicates must be used for each treatment (minimum of 20 tests). A concentration must be used that does not cause mortality and another that causes total mortality. Each replica involves the maintenance of groups of 10 earthworms with artificial earth in glass containers for 14 days in continuous light at 20 ± 2°C (a minimum of 280 earthworms, including positive, negative and solvent controls). Mortality is evaluated 7 and 14 days after application.

After the test, the mortality data will be reported, in addition to any physiological or behavioral changes observed, and the mean lethal concentration (LC50), defined as the concentration of the tested substance that kills 50% of the earthworms during the period of testing.

To carry out the tests, the following information must be provided to the laboratory: test to be evaluated (paper contact or incorporation in artificial soil), concentration range to be evaluated (a minimum of 5 concentrations must be chosen for each of the tests) , nature of the product and its solubility; if it is not soluble in water, the recommended organic solvent should be indicated.