The earthworm species Eisenia fetida and Eisenia andrei are commonly used in ecotoxicological standard tests. In the present study, we compared the sensitivity of E. fetida with that of two soil-dwelling earthworm species (Aporrectodea caliginosa and Lumbricus terrestris) in order to evaluate the capacity of E. fetida to predict effects of the insecticide imidacloprid. Responses were compared using two endpoints, a biochemical (changes in heat shock protein level (hsp70)) and a behavioural (avoidance behaviour).

Purpose The use of only one or a few species—representing an entire taxon—in ecotoxicological standard tests poses risk of underestimating the impact of toxicants on the environment. In earthworm ecotoxicity tests, the species Eisenia fetida or Eisenia andrei are commonly used, and there is evidence that these species respond relatively insensitive towards environmental pollution. With the present study, we wanted to evaluate the risk of underestimating effects of the insecticide imidacloprid in soil organisms by comparing E. fetida with two other earthworm species ( Aporrectodea caliginosa and Lumbricus terrestris ) regarding their sensitivities towards soil contaminated with this widely used insecticide. Materials and methods In laboratory experiments, the specimens were individually exposed to various concentrations of the pesticide (0.2, 0.66, 2 and 4 mg kg −1 dry weight (DW)) for 1, 7 and 14 days. Afterwards, histopathological changes in the midgut, chloragogenous tissue and skin, as well as body mass changes, were assessed. Results and discussion While significant changes in body mass in E. fetida and A. caliginosa occurred after exposure to imidacloprid concentrations as low as 0.2 (7 days) and 0.66 mg kg −1 DW (14 days), significant body mass changes in L. terrestris observed to 2 and 4 mg kg −1 DW, for 7 and 14 days of exposure, respectively. The histopathological examinations revealed that significant cellular changes already occurred after 24 h exposure to the lowest test concentrations in all species, but the degree of detrimental effects as well as species-specific differences were dependent on the monitor tissue. In general, E. fetida seemed to be more sensitive than L. terrestris concerning cellular alterations, but the hierarchy in species-specific differences was less obvious than for body mass change. Conclusions Even if E. fetida proved to be the most sensitive species in this study, general differences in sensitivity make evident that always a range of species—being representatives of an animal taxon—in ecotoxicological tests should be tested in order to avoid underestimations of effects. In the case of testing only one species, an increase of safety factors should be considered. Since effects already occurred at environmentally relevant concentrations, the use of imidacloprid in agriculture might be of great concern.

Adverse effects of agrochemicals on earthworms' burrowing behaviour can have crucial impacts on the entire ecosystem. In the present study, we have therefore assessed short- and long-term effects on burrowing behaviour in the earthworm species Aporrectodea caliginosa and Lumbricus terrestris after exposure to a range of imidacloprid concentrations (0.2-4 mg kg(-1) dry weight (DW)) for different exposure times (1, 7, 14 d). 2D-terraria were used for the examination of post-exposure short-term effects (24-96 h), while post-exposure long-term effects were assessed by means of X-ray burrow reconstruction in three dimensional soil cores (6 weeks). For the latter each core was incubated with two specimens of L. terrestris and four of A. calignosa. Short-term effects on the burrowing behaviour (2D) of A. caliginosa were already detected at the lowest test concentration (0.2 mg kg(-1) DW), whereas such effects in L. terrestris were not observed until exposure to concentrations 10 times higher (2 mg kg(-1) DW). For both species tested in the 2D-terraria, "total burrow length after 24 h" and "maximal burrow depth after 24 h" were the most sensitive endpoints. 3D reconstructions of the burrow systems made by both earthworm species in the repacked soil cores revealed a significant linear decrease in burrow volume with increasing imidacloprid concentration. Since many of the observed effects occurred at imidacloprid concentrations relevant to natural conditions and since reduced activities of earthworms in soils can have crucial impacts on the ecosystem level, our results are of environmental concern.

Earthworms play key roles in soils and sub-lethal effects of environmental toxicants on these organisms should be taken seriously, since they might have detrimental effects on higher ecological levels. In laboratory experiments we have assessed sub-lethal effects (body mass change and cast production) of imidacloprid on two earthworm species commonly found in different agricultural soils (Lumbricus terrestris and Aporrectodea caliginosa). After 7 days of exposure in contaminated soil, a significant loss of body mass was found in both species exposed to imidacloprid concentrations as low as 0.66 mg kg(-1) dry soil. These losses ranged from 18.3 to 39% for A. caliginosa and from 7.4 to 32.4% for L. terrestris, respectively. Changes in cast production, a new biomarker previously validated using L. terrestris, was assessed by soil sieving using the recommended mesh size (5.6 mm) for L. terrestris and three different mesh sizes for A. caliginosa (5.6, 4 and 3.15 mm). The 4 mm mesh size proved to be the most suitable sieve size for A. caliginosa. Cast production increased by 26.2% in A. caliginosa and by 28.1% in L. terrestris at the lowest imidacloprid concentration tested (0.2 mg kg(-1) dry soil), but significantly decreased at higher concentrations (equal to and above 0.66 mg kg(-1) dry soil) in both earthworm species after the 7 days exposure experiment. These decreases in cast production ranged from 44.5 to 96.9% in A. caliginosa and from 42.4 to 95.7% in L. terrestris. The EC(50) for cast production were 0.84 (L. terrestris) and 0.76 mg kg(-1) dry soil (A. caliginosa), respectively. The detected sub-lethal effects were found close to the predicted environmental concentration (PEC) of imidacloprid, which is in the range of 0.33-0.66 mg kg(-1) dry soil. The biomarkers used in the present study, body mass change and changes in cast production, may be of ecological relevance and have shown high sensitivity for imidacloprid exposure of earthworms. The measurement of changes in cast production should be considered for inclusion in current standard tests.

There is currently a lack of ecotoxicity tests adapted to earthworm species of higher ecological relevance and whose endpoints could be directly related to their ecological role in the soil. We propose a new and relatively simple ecotoxicity test based on the estimation of cast production (CP) by Lumbricus terrestris under laboratory conditions. CP was found to be linearly correlated to earthworm biomass and to be greatly influenced by soil water content. Azinphos-methyl had no effect on CP at all the concentrations tested. Significant decreases were observed at the normal application rate for other pesticides with (imidacloprid, carbaryl, methomyl) or without (ethyl-parathion and chlorpyrifos-ethyl) a clear concentration-effect response. For the highest concentration tested, reduction in CP varied between 35 and 67%. CP is straightforward and rapidly measured and ecologically meaningful. We thus believe it to be of great use as an endpoint in ecotoxicity testing.

The physiological and biochemical adaptation mechanisms of heat-tolerant snail species are poorly known. We investigated cellular adaptations to heat stress in four heat-exposed snail populations (two of Xeropicta derbentina, one each of Cernuella virgata and Theba pisana). Snails were exposed to different temperature regimes (25, 33, 38, 40, 43, 45, 48, 50 and 528C) in the laboratory for 8 h. After exposure, immobility rates were determined and a qualitative and semi-quantitative histopathological analysis of the digestive gland (calcium and digestive cells) and mucocytes in the foot was conducted. In addition, the coloration of the shells was densitometrically determined. The shells of T. pisana were significantly darker than the shells of the other populations. The immobility rates and histopathological examinations showed that X. derbentina (both populations) was slightly better adapted to heat than C. virgata, and much better adapted than T. pisana. The observed differences in heat tolerance seemed to be connected to the ability for rapid and extreme proliferation of calcium cells in the digestive gland.

On the basis of studies with laboratory strains of Drosophila and Arabidopsis, it has been hypothesized that potential buffers to the expression of phenotypic morphological variation, such as Hsp90 and possibly Hsp70, represent important components of Waddington's widget, which may confer capacitive evolution. As studies on field populations of living organisms to test this hypothesis are lacking, we tested whether a heat response strategy involving high stress protein levels is associated with low morphological variation and vice versa, using four natural populations of Mediterranean pulmonate snails. In response to 8 hr of elevated temperatures, a population of Xeropicta derbentina with uniform shell pigmentation pattern showed remarkably high Hsp70 but low Hsp90 levels. In contrast, a highly variable population of Cernuella virgata kept both Hsp90 and Hsp70 levels low when held at diverse though environmentally relevant temperatures. Two other populations (Theba pisana and another X. derbentina population) with intermediate variation in shell pigmentation pattern were also intermediate in inducing Hsp70, though Hsp90 was maintained at a low level. The observed correlation of stress protein levels and coloration pattern variation provide the first indirect evidence for an association of stress proteins with Waddington's widget under natural conditions.

On the one hand, snow-packing protects tress from low-temperature related injuries, when light, but on the other hand it can also cause severe damage to trees and forests when excessively heavy and can bring about great economic losses in forestry and disturbances in the ecosystems. (Jalkanen, R. and Konopka, B., 1998) The most common form of damage is stem breakage, but tress can also be bent, uprooted and a consequential damage through insect or fungal attacks is more likely to happen. The severity of snow damage is related to tree characteristics, so that slightly tapering stems, asymmetric crowns and rigid horizontal branching are all associated with higher risk. (Nykänen et al., 1997) In Northern Finland, where winds & storms are fed by moisture from the Baltic Sea, tree breakage under extreme snow loading is considered to be the major limiting factor at timberline. (Marchand, P. 1987) As in the case of Riisitunturi snow damage is found from 300 m a.s.l. upwards and about 30% of the trees carry evidence of snow damage. (Eurola, S. and Huttunen, A., 1984) In the future, assuming the global change in climate, both mean temperature and precipitation could rise in northern latitudes and the risk of snow damage could increase compared to the present day, because of an increased frequency of snowfalls at temperatures of around zero. (Nykänen et al., 1997)