2 Introduction, Danish Environmental Protection Agency

Mesocosm experiments in the approval procedure for pesticides

2 Introduction

The regulation of pesticide use and protection of non-target species primarily rely on evaluations based on single species tests with organisms belonging to different trophic and taxonomic groups. If specific pesticides are evaluated to constitute a potential hazard to aquatic life, further and extended analysis must be carried out to show that the pesticide does not constitute a risk to the aquatic environment (EU directive 91/414). In line with several other countries Denmark relies on extended risk evaluations based on tests carried out under near-natural conditions (i.e. mesocosms). Several guidelines (e.g. OECD 1996) describe how to carry out mesocosm experiment (experimental design) and what endpoints should be measured. The aim of such guidelines is primarily to define endpoint of regularly concern, which can effectively be addressed only from an appropriate experimental design. Ecological endpoints are those which are directly related to observable changes in the biotic and abiotic components of an aquatic ecosystem. Typically both structural and functional elements are included in the biotic component.

Hypothesis test (i.e. Anova design) is used for investigation of whether the response of a mesocosm unit is different from that of a control unit. Hypothesis tests are used for comparing means and are characterised by having multiple replicates in control and treatment groups. The greater number of replicates, the more accurately is the group variability defined and the greater the power of the test for resolving differences. Hypothesis tests are best for objectively determining if an identified difference between control and treatment groups is statistically significant.

Point estimate tests (i.e. regression design) are designed to evaluate regression relationships and, by using regression equations between pesticide concentration and observed effects, estimate an exposure concentration which will not cause an adverse effect (i.e. No Effect Concentration, NEC) or predict the intensity of an effect at a given exposure level. Regression analysis is used to iteratively fit observed data to theoretical equation. This requires multiple treatments at various concentrations related to a response. The greater number of treatment concentrations along the response gradient, the greater the confidence in the fitted concentration-response line.

Hybrid tests incorporate features of both hypothesis and point estimate tests. Employing both multiple replicates and multiple doses, one can determine if a given treatment level significantly differs from controls and may estimate how different another treatment level will be above or below the given treatment concentration. The dilemma facing an experimenter is, with a limited number of mesocosm units one can reduce the number of replicates to increase the number of exposure concentrations, or as an alternative, reduce the number of dose levels and increase replicates. Fewer replicates will reduce the power to resolve significant effects and fewer dose levels will reduce the confidence in estimating the fit and the NEC.

In this report we have focussed on how to interpret results from mesocosm experiments and subsequently identify "ideal" experimental condition, which satisfy both realism (design of experiments) and regulatory needs, such as consistency of results.