Enchytraeid Reproduction Test Research Articles

Ecotoxicological laboratory tests with enchytraeids: A reviewThe 7th international symposium on earthworm ecology · Cardiff · Wales · 2002

Summary Worms of the family Enchytraeidae (Oligochaeta, Annelida) are small, inconspicuous animals, which despite their important role in many ecosystems have been virtually neglected as test organisms until about 15 years ago. They belong to the saprophagous soil mesofauna, living mainly in close contact with soil pore water. After it was recognised that they were highly sensitive to anthropogenic stress factors in field studies, a chronic laboratory test using artificial soil, the Enchytraeid Reproduction Test (ERT), was developed for testing of chemicals as well as for soil quality assessment. Having been validated nationally and internationally, the test is currently standardised as a guideline by ASTM, ISO and OECD. In addition, new methods with enchytraeids, such as the use of avoidance behaviour in screening tests and a terrestrial bioaccumulation test are under development. New research could attempt to reduce the high variability in test results as well as examine the influence of natural soil properties on the sensitivity of the worms towards anthropogenic stress factors. In this contribution the most important test methods for the assessment of single chemicals, as well as for potentially contaminated soils are presented. The insights gained so far prove that these animals can play an important role in terrestrial ecotoxicology.

Ecotoxicological laboratory tests with enchytraeids: A review: The 7th international symposium on earthworm ecology · Cardiff · Wales · 2002

Worms of the family Enchytraeidae (Oligochaeta, Annelida) are small, inconspicuous animals, which despite their important role in many ecosystems have been virtually neglected as test organisms until about 15 years ago. They belong to the saprophagous soil mesofauna, living mainly in close contact with soil pore water. After it was recognised that they were highly sensitive to anthropogenic stress factors in field studies, a chronic laboratory test using artificial soil, the Enchytraeid Reproduction Test (ERT), was developed for testing of chemicals as well as for soil quality assessment. Having been validated nationally and internationally, the test is currently standardised as a guideline by ASTM, ISO and OECD. In addition, new methods with enchytraeids, such as the use of avoidance behaviour in screening tests and a terrestrial bioaccumulation test are under development. New research could attempt to reduce the high variability in test results as well as examine the influence of natural soil properties on the sensitivity of the worms towards anthropogenic stress factors. In this contribution the most important test methods for the assessment of single chemicals, as well as for potentially contaminated soils are presented. The insights gained so far prove that these animals can play an important role in terrestrial ecotoxicology.

Statistical results and implications of the enchytraeid reproduction ringtest.

An international ringtest of the enchytraeid reproduction test (ERT) was performed by 29 laboratories, with two substances (carbendazim and 4-nitrophenol) and two test designs (NOEC and ECx). Although many participants had no previous experience in enchytraeid ecotoxicology, the majority of tests were performed accurately and fulfilled the validity criteria, demonstrating method feasibility. However, variability in control reproduction was fairly high and exceeded the proposed validity criterion of a 50% coefficient of variation in 24% of the tests. Variability may be lowered with increasing experience and by using better defined animals, either through synchronized culture or by weighing animals before the test. In all tests, NOECs were higher than the EC10 values, sometimes by more than 1 order of magnitude and usually closer to the EC50. This shows that NOECs, due to their dependence on test design and variability, are an unsatisfactory measure of "no toxicity" for regulatory purposes. Preferably, differences to the control at NOEC concentrations should always be given. To achieve appropriate power and to avoid frequent false negatives, in the ERT, high numbers of replicates would be necessary. We favor using regression approaches with means of replicates whenever possible.

Validating the enchytraeid reproduction test: organisation and results of an international ringtest

In this paper the experiences concerning the organisation and results of the enchytraeid reproduction test (ERT) ringtest are summarised (for details see J. Römbke, T. Moser, Organisation and Performance of an International Ringtest for the Validation of the Enchytraeid Reproduction Test, vols. I and II. UBA-Texte 4/99, 1999, 150, 223 pp). • The performance of this ringtest was in line with requirements published by OECD. It was sponsored by the German Federal Environmental Agency (Umweltbundesamt; UBA, Berlin). The UBA was also actively engaged (together with the European Chemicals Bureau, Ispra) in a scientific task force, which consisted of seven scientists experienced in terrestrial ecology and ecotoxicology. • 29 institutions (mainly from universities and contract research laboratories) from 15 countries of Europe and North America participated actively in the ringtest. The co-ordinating laboratory centrally distributed the test chemicals, guidance papers and test organisms to all participants. In addition, several training courses were organised. • Most participants performed two tests with the fungicide Carbendazim and two with 4-nitrophenol. For each chemical, one test was designed according to an EC x approach and the other according to an NOEC approach. Several aspects of the test protocol were modified based on the experiences gained during the ringtest. • A major aspect of the project was the detailed statistical evaluation of the test results (for details see A. Weyers, J. Römbke, T. Moser, T. Ratte, Results of and statistical implications from the enchytraeid reproduction ringtest, 2001, submitted), leading to recommendations for an optimised ecotoxicological test design. A total of 92 tests were performed according to the protocol which is among the highest number ever performed in a ringtest. About 72% met the validity criteria (control mortality <20%, >25 juveniles per 10 adults). • The effects of the two test chemicals on enchytraeid reproduction were in the range expected from data in the literature on oligochaete toxicity. However, statistical evaluation of the data was sometimes difficult as a result of the high variability in the number of juveniles. This variability was caused by several reasons, including hormesis effects or lack of experience of some participants. • Comparison of the data from the NOEC and EC x approaches pointed to a clear advantage in favour of the latter. In most cases EC 10 values were lower than the NOEC values determined in the same test. For details see A. Weyers et al. (loc. cit.). • Reproducibility of the test data and practicability of the ERT ringtest were of the same order of magnitude as other ringtests recently performed. As a result of the ringtest, the ERT draft guideline was significantly improved. The new version is currently being standardised by OECD, ISO and ASTM.

Effects of three chemicals on the survival and reproduction of the oligochaete worm Enchytraeus coronatus in chronic toxicity tests

Using Enchytraeus coronatus as a test organism, we have measured the effects of Derosal (carbendazim as active ingredient), 4-nitrophenol and potassium dichromate on survival (Lethal Concentration: LC50) and on several sublethal variables (Effective Concentrations: EC10, EC50 and the No Observed Effective Concentration: NOEC), in chronic agar tests. Worms were also exposed to carbendazim and potassium dichromate in a soil toxicity test (Enchytraeid Reproduction Test, ERT). Sublethal variables measured in agar tests (number of juveniles and cocoons per adult, number of eggs per cocoon, percent of hatching and adult biomass) were used to facilitate interpretation of the causes of reproductive inhibition of the number of juveniles in the ERT soil toxicity test. Effective concentrations (EC) were estimated using Probit and logistic models. The later provided a better fit for the response of the worms exposed to carbendazim in agar test, in which an increase of the number of juveniles with respect to control was found at low doses. The number of juveniles per adult was the most sensitive variable measured for the three substances in the agar toxicity tests. The reduction in the number of juveniles could be explained mainly by a negative effect on cocoon hatching due to carbendazim (4-week NOEC juveniles/adult = 0.32 mg l—1, 4-week EC10 Reproductive inhibition (RI) = 0.37mg l—1), a result probably related to failure of embryonic development. The effect of 4-nitrophenol on the number of juveniles (4-week NOEC = 0.32mg l—1, 4-week EC10 (RI) = 0.25 mg l—1) was interpreted as a consequence of a reduction in the number of eggs per cocoon, associated with a reduction in parental biomass. Reproductive inhibition (RI) in worms exposed to potassium dichromate (4-week NOEC juveniles/adult = 0.32mg l—1, 4-week EC10 (RI) = 0.40 mg l—1) was explained by a reduction in cocoon production, which apparently was not related to the loss of parental biomass but rather to a failure of worm reproductive physiology. The results of this study provide useful information regarding the effects of carbendazim, 4-nitrophenol and potassium dichromate on worm reproduction biology and contribute to our understanding of the effects of these substances measured in ERT soil toxicity tests.

Enchytraeid Reproduction Test (ERT): Sublethal responses of two Enchytraeus species (Oligochaeta) to toxic chemicals

Enchytraeid Reproduction Test (ERT): Sublethal responses of two Enchytraeus species (Oligochaeta) to toxic chemicals

Comparison of Malathion toxicity using enchytraeid reproduction test and earthworm toxicity test in different soil types

Comparison of Malathion toxicity using enchytraeid reproduction test and earthworm toxicity test in different soil types