Riboflavin content in the coelomocytes of contrasting earthworm species is differentially affected by edaphic variables including organic matter and metal content

The riboflavin content in extruded coelomocyte lysates derived from Dendrodrilus rubidus may serve as a sensitive bioindicator of soil metal pollution: the vitamin (B2) content has previously been found to be high in worms from unpolluted soil but low in worms inhabiting Zn/Pb mine soils, aerially deposited Ni-contaminated soil, and in worms experimentally transferred from clean soil to the metalliferous field soils. The aim of the present work was to extend these observations by comparing the number and riboflavin composition of coelomocytes retrieved from three lumbricid species (Allolobophora chlorotica, Dendrobaena veneta, Eisenia andrei) after 4-week exposures to an unpolluted commercial soil, two geochemically contrasting unpolluted field soils, and two different Zn/Pb/Cd-polluted soils from the Bukowno district in South Poland. Whilst eco-physiologically contrasting, these three earthworm species share the trait of possessing relatively high numbers of eleocytes, a category of immune-competent coelomocyte rich in autofluorescent riboflavin. Spectroflurometric analysis indicated that coelomocyte riboflavin content in worms maintained in strongly metalliferous soils or in unpolluted sandy-clay and loamy-sand soils was increased in coleomocytes from epigeic D. veneta and E. andrei species, whilst was decreased in endogeic A. chlorotica. In conclusion, the riboflavin content of earthworm coelomocytes is affected in species-specific ways by edaphic variables, including organic matter and metal pollution.     

Species-specific sensitivity of earthworm coelomocytes to dermal metal (Cd,Cu, Ni,Pb, Zn) exposures: Methodological approach

Earthworm immune-competent cells (coelomocytes) can be adversely affected by soil metal contamination. The aim of the present paper was to perform comparative studies on the coelomocytes of four lumbricid species dermally exposed for 2 days to filter paper soaked with Ni, Cu, Zn, Cd, or Pb chlorides (1 mM). Coelomocytes were expelled by electro-stimulation, counted, and assayed ex vivo by flow cytometry and spectrofluorometry. The coelomocytes of Allolobophora chlorotica and Dendrobaena veneta contain both amoebocytes and riboflavin-storing eleocytes; consequently, they were subjected to assays based on flow cytometric quantification of autofluorescent eleocytes and spectrofluorometric measurement of riboflavin content in coelomocyte lysates. In Al. chlorotica, the number of coelomocytes, the percentage of eleocytes, and the amount of riboflavin were significantly lower in Cu-exposed worms although these cytometric parameters were less affected by Ni, Zn, Cd exposure, and entirely unaffected by Pb exposure. In D. veneta, such cellular effects were also only observed in Cu-exposed worms. The coelomic fluids of Aporrectodea caliginosa and Lumbricus rubellus are constitutionally impoverished of eleocytes; therefore, these species could not be subjected to assays measuring the riboflavin content of these granular cells. Rather, the amoebocytes of Ap. caliginosa and L. rubellus were subjected to flow cytometric measurements of in vitro neutral red uptake (NR). However, the NR uptake assay was technically demanding, requiring a strictly normalized incubation period over all samples to yield useful comparative data. In contrast, the riboflavin content in the coelomocyte lysates of eleocyte-rich species appears to be a robust and convenient immune-function biomarker of environmental stress.     

Exposure of coelomocytes from the earthworm Eisenia hortensis to Cu,Cd, and dimethylbenz[a]anthracene: An in vitro study examining reactive oxygen species production and immune response inhibition

The principal aims of this study were to investigate the in vitro effects of copper (Cu), cadmium (Cd) and the polycyclic aromatic hydrocarbon (PAH) dimethylbenz[a]anthracene (DMBA) on reactive oxygen species (ROS) production and the effect of DMBA on immune function (phagocytosis) in coelomocyte samples isolated from the earthworm Eisenia hortensis. Using a fluorogenic indicator of ROS, dihydrorhodamine 123, we show that earthworm coelomocytes are induced to produce significant increases in ROS when exposed to Cu or Cd at 32–250 μM, an effect diminished by the chelator ethylenediaminetetraacetic acid (EDTA). DMBA also induced significant ROS production in the range of 20–400 μM compared to controls. Testing the effect of DMBA on cellular immune mechanisms, we used a phagocytosis assay to reveal an immunosuppressive effect of DMBA (50–200 μM) on the capacity of hyaline amoebocytes to ingest fluorescent Escherichia coli. These studies illustrate the relative ease and efficiency of these in vitro procedures for testing the toxicological effects of environmental pollutants, including heavy metals and PAHs, using earthworm coelomocytes as indicators of oxidative stress and immune response impairment.     

Allometric estimation of earthworm ash-free dry mass from diameters and lengths of select megascolecid and lumbricid species

We present novel length to ash-free dry mass and preclitellar diameter to ash-free dry mass allometric equations for seven earthworm species from the families Megascolecidae and Lumbricidae, all of which are exotic and most of which are of ecological concern in North America: Eisenia hortensis, Eisenia fetida, Dendrodrilus rubidus, Lumbricus rubellus, Octolasion sp., Amynthas hilgendorfi, and Perionyx excavatus. We also present a length-biomass allometric equation for one enchytraeid, Mesenchytraeus sp. All relationships between length and biomass, and diameter and biomass were statistically significant at the species and family level (P<0.001). The predictive powers of these allometric regressions (as coefficients of determination, r²) were species-specific, and ranged widely from 0.27 to 0.93. Length-biomass regressions provided more predictive power and precision overall than preclitellar diameter-biomass calculations at both the species and the family levels. An ANCOVA followed by orthogonal contrasts determined that, while the slopes of these regressions did not differ significantly between the two earthworm families, significant differences in slopes of length-biomass regressions existed among species within families, indicating the utility of having species-level equations for accurate biomass predictions. With these allometric relationships, we aim to improve the estimation of earthworm biomass in order to facilitate investigations of how exotic-invasive earthworm species impact soil ecosystems.     

Soil amendment with municipal sewage sludge affects the immune system of earthworms Dendrobaena veneta

Dendrobaena veneta is an earthworm species capable of consuming a wide range of organic wastes which may be used as a field indicator of municipal sewage sludge applied to land. The aim of the present 8-week laboratory experiment was to check viability, reproduction and the immune system of D. veneta maintained in soil without food additions (control 0s group) or in soil amended with 25% or 50% municipal sewage sludge (25s and 50s groups, respectively). Reproduction and immunity are important physiological functions whose detailed study can provide information on the effects of pollutants. After the 8-week exposure period, earthworm mortality (2 out 20 individuals) was recorded only in the 50s group. Reproduction was high in the 25s group (44 cocoons and 41 juveniles) whereas reproduction was almost completely inhibited both in the food-deprived control 0s group (1 cocoon, 3 juveniles) and in the 50s group containing a high amount of sludge (2 cocoons). Significantly increased numbers of non-invasively extruded coelomocytes were recorded 3 weeks after the start of the experiment in the 50s group, but they dropped to the food-deprived control level by the end of 8 weeks likely due to exhaustion of the immune system coping with sludge-derived microbes and/or toxins. In contrast, numbers of coelomocytes in the 25s group increased gradually reaching the maximum at the end of the experiments. In conclusion, high amounts of municipal sewage sludge are detrimental to worms, inhibiting reproduction and inflicting mortality. A moderate amount of municipal sewage sludge provides a good source of nutrients for D. veneta, supporting their growth and reproduction for at least 8 weeks. Immunological parameters might serve as useful indicators of earthworm exposure to sewage sludge.     

Earthworm distribution and abundance along a mineral-peat soil transect

The earthworm fauna along a gradient from undisturbed mineral soil to an artificial acid peat soil up to 25 cm deep and ca. 30 yr old was studied in the Phoenix Park, Dublin. The peat held only 15% of the total numbers of worms in the mineral soil and 9% of the biomass, but 9 out of the 12 species recorded were present in the peat. Aporrectodea caliginosa, A. tuberculata and Allolobophora chlorotica were the most numerous species in the mineral soil but were scarce in the peat where the surfaced-welling, pigmented species L. rubellus, L. castaneus and Satchellius mammalis were most abundant. Low pH and shortage of suitable food were considered to be the main factors limiting earthworm populations in the peat.     

The interactions between soil type and earthworm species determine the properties of earthworm casts

Earthworms are recognized to increase soil porosity, reorganize soil structure, and stimulate soil microflora and nutrient mineralization. The properties of earthworm casts should depend both on earthworm species or ecological group and on soil properties. Interactions between earthworm species and soil types have been suggested, but only poorly demonstrated. In order to better understand those interactions, two hypotheses led our study: (1) Soil type has a greater influence on cast properties than earthworm; (2) Earthworms from different species influence cast properties differently; (3) The intensity and direction of the impact of each earthworm species on cast properties vary with soil properties. Fifteen physical and chemical variables (N–NH₄⁺, N–NO₃⁻, total organic C and N, C/N ratio, CaCO₃, pH, P, K⁺, Mg²⁺, Mn²⁺, Na⁺, CEC, moisture, wettability) were measured in casts of three earthworm species (Lumbricus terrestris, Allolobophora chlorotica and Aporrectodea rosea) produced in three temperate soils. Univariate and multivariate analyses showed that earthworm species and soil types significantly impacted cast properties. pH, N_t, K and Mg contents were interactively altered by both factors. Multivariate analysis showed that a difference of soil type had a major impact on casts properties (62%) compared to the impact of a difference of earthworm species (10%). Cast properties were most impacted by L. terrestris, then by A. chlorotica and last by A. rosea. The response ratio (ratio of the properties of the casts to the properties of the bulk soil) was used to quantify the effect of earthworm species compared to the control soil. It showed a higher response of variables in casts in nutrient-rich soils, especially in casts of L. terrestris. The interactions between earthworm species and soil types on cast properties were discussed with regards to earthworm ecology, properties of the soil, and earthworm modifications of cast microflora.     

Phosphatase activity in earthworm faeces

Wormcasts of Allolobophora caliginosa have been shown to have higher phosphatase activity than occurs in uningested soil, resulting in an increase in inorganic P released by mineralization of organic P. This paper describes similar observations on organic wastes which may be used for vermiculture. Besides A. caliginosa, three species which will feed on dung or other organic substrates were investigated, Eisenia foetida, Dendrobaena veneta and Lumbricus rubellus. When cow dung was used as a substrate, background phosphatase activity was too high for the effects of earthworm activity to be detected and a culture medium was therefore developed from sterilized paper waste sludge with added phytin. Phosphatase activity, assayed by a modified Hoffman method, was higher in the presence of all four earthworm species than in controls. It showed two peaks in relation to pH, at 3–5 and 9–10. The former is attributed to microbial activity and the latter to microbial activity or earthworm alkaline phosphatase. After 1 month, cultures of paper waste sludge with phytin contained about twice as much water soluble P when worms were present than in their absence.     

Fat but slim: Criteria of seed attractiveness for earthworms

Earthworms were shown to significantly affect seeds and seedlings survival via their ingestion and digestion for nutritive purposes. Such selective feeding of earthworms on plant seeds is likely to favour certain plant species and to affect seed bank composition, plant recruitment and plant community structure. Relationships between earthworms and seeds, particularly seed traits that determine attractiveness of seeds for earthworms, are yet to be determined. In this study, the influence of six seed traits was tested on the ingestion, digestion and germination of seeds by two earthworm species (Lumbricus terrestris, anecic and Satchellius mammalis, epigeic). The seed traits tested were their length, width, weight, shape, oil content and the presence of trichomes on their surface. Each earthworm species was introduced into a microcosm with eleven seed species from a chalk grassland that represented those different traits. Ingested, digested and germinated seeds were counted after voiding the guts of the earthworms. Univariate and multivariate analyses showed that seed length, width, weight and seed oil content could significantly affect the ingestion of seeds for both earthworm species. Seed width and seed oil content were the two traits that influenced the digestion of seeds the most, but only for L. terrestris. We also found that seed ingestion was earthworm species-specific but we found no correlation between earthworm traits and number of ingested or digested seeds. Few seeds germinated from L. terrestris casts and no seeds germinated from S. mammalis casts. Implications in terms of plant evolution strategies are further discussed.     

Flooding responses of three earthworm species,Allolobophora chlorotica,Aporrectodea caliginosa and Lumbricus rubellus,in a laboratory-controlled environment

To get a better understanding of earthworm’ responses towards flooding, three laboratory experiments were performed with the species Allolobophora chlorotica, Aporrectodea caliginosa and Lumbricus rubellus.Flooding response was determined in a pot experiment, in which the earthworms were incubated for 42 days in flooded or non-flooded soil, with or without heavy metal pollution. To determine moisture preference, earthworms were incubated for 9 days in aquaria with five compartments, containing soil with a moisture gradient (%, w/w), ranging from 35%, 45% (field capacity), 55%, 65% (saturated) to 65%+ (saturated and an extra water layer). Effects on earthworm health were studied by incubating earthworms of each species for 42 days in soil with the same range of moisture contents and determining the dry/wet weight ratio and dry weight gain as an indication of earthworm health.A. chlorotica was tolerant to water, although the worms tended to escape from flooded soil. Their health was significantly lower in the flooded soils (P<0.05). A. caliginosa showed little response to flooding. This species, however, was affected by the heavy metal pollution in the pot experiment. L. rubellus was sensitive towards flooding, with escape and avoidance behaviour being the main mechanism of survival. This species was able to survive when forced to stay in flooded soil for at least 42 days, but this significantly reduced its health (P<0.05).These results suggest that earthworms are able to survive in inundated soils, but there are large differences between species in response to flooding conditions.     

Movement response of Collembola to the excreta of two earthworm species: Importance of ammonium content and nitrogen forms

Several studies reported variable effects of earthworms on microarthropod density and variety. The present study tests the attraction of seven collembolan species belonging to four families, to the excreta of two earthworm species belonging to two families and two ecological categories, Aporrectodea giardi and Hormogaster elisae. Our objectives were (1) to better understand the impact of earthworms on the composition and density of Collembola communities, and (2) to dissect mechanisms involved in the attraction. Experiments were performed in Petri dishes containing two half-disks of filter paper, one with earthworm excreta, i.e. casts or a mix of mucus and urine, and the other with natural soil aggregates or water, respectively. Collembola were introduced half-way between the two half-disks and their number was counted on each half-disk and compared over 140 min. The content of ammonium in casts and mucus-urine of both earthworm species was analyzed to determine whether it altered the responses of Collembola faced with different types of earthworm excreta. The behaviour of Collembola varied strongly among the seven collembolan species, and with type of excreta and earthworm species. Six collembolan species were attracted to the mucus and urine of at least one earthworm species. The mucus-urine mixture of A. giardi, with low ammonium content, was generally more attractive than that of H. elisae, which was even repulsive in some cases, probably because of high levels of ammonium. The attraction to casts of the two earthworm species was less frequent and more variable. Folsomia candida was neither attracted to the casts nor to the mucus and urine of any earthworm species. Therefore, (1) earthworm species with different ecology, and different nitrogen excretion pathway impact differently the behaviour of collembolan species belonging to the same family or arising from the same habitat, and (2) variations in the sensitivity to ammonium among collembolan species partially explain the variable response of Collembola to earthworm excreta.     

Effects of cropping and tree density on earthworm community composition and densities in central Cameroon

Earthworms can have positive effects upon crop growth in the tropics. If soils are to be managed sustainably, then more attention should be paid to the effects of cultivation and cropping practices upon earthworms. When forest vegetation is cleared, slashed, burned and land is tilled and cultivated, earthworm abundance, diversity and activity are reduced. Conversely, retaining trees in agroecosystems may maintain earthworm populations during the cropping phase.Here, we assessed the impact on earthworm species diversity and densities of crop cultivation in the understorey of timber plantations thinned to two tree densities and compared these with uncropped, undisturbed timber plantation controls. The plots were reassessed after two and a half years of fallow to see whether populations had recovered. The experiment was in central Cameroon.Seventeen earthworm species were recorded from Eudrilidae subfamilies Eudrilinae and Pareudrilinae, Ocnerodrilidae and Acanthodrilidae, most of which were endemics. This included two new species from two new genera from the sub-family Pareudrilinae, one new species from one new genus of Ocnerodrilidae, two new species of Dichogaster and one new species of Legonodrilus. Ten species were epigeic, six were endogeic and one was anecic.Generally, earthworm densities were lower in cropped plots than in the undisturbed plantation control. The most abundant species was a Legonodrilus sp. nov. with average densities of 49 individuals m⁻² in the crop phase and 80 ind. m⁻² in the fallow phase. By the fallow phase, densities in the low tree density (120 ind. m⁻²) were higher than in the high density (40 ind. m⁻²). The densities of the epigeic Acanthodrilidae were significantly reduced to 7 ind. m⁻² in the cropped plots compared with 42 ind. m⁻² in the control plots. The effects of cropping were thus species-specific and more work is required to identify which of these endemics are the ecosystem engineers in the system.     

Earthworm populations in a northern U.S. Cornbelt soil are not affected by long-term cultivation of Bt maize expressing Cry1Ab and Cry3Bb1 proteins

Earthworms, which play a key role in biogeochemical processes in soil ecosystems, could be negatively affected by the cultivation of transgenic Bt crops. Studies to date have found few effects of Bt maize on earthworm species. If adverse effects occur, they are likely to be chronic or sub-lethal and expressed over large spatial and temporal scales. Our objective in the present study was to investigate potential effects on earthworm populations in soil cultivated with Bt maize in a large multiple-year field study. We surveyed the earthworm populations in 0.16-ha experimental field plots of two varieties of Cry1Ab Bt maize, one variety of Cry3Bb1 Bt maize, and three non-transgenic control varieties cultivated for four years. Four earthworm species were found in our sample: Aporrectodea caliginosa, Aporrectodea trapezoides, Aporrectodea tuberculata (collectively, the A. caliginosa species complex), and Lumbricus terrestris. We found no significant differences in the biomass of juveniles and adults for all four species between Bt and non-Bt maize varieties. From this and previous studies, we conclude that the effects of Cry1Ab and Cry3Bb1 Bt maize on the A. caliginosa species complex and L. terrestris are small. Nonetheless, general conclusions about the effects of Bt maize on earthworm populations are not warranted due to the small number of species tested. In future laboratory studies, earthworm species should be selected according to their association with a Bt crop and the impact of that species to valued soil ecosystem processes.     

Using FAME profiles for the characterization of animal wastes and vermicomposts

This study investigated the possibility of fingerprinting different organic wastes (cow, pig and horse manure) and the vermicomposts produced by different earthworm species (Eisenia andrei, Eudrilus eugeniae and Lumbricus rubellus) analyzing the profiles of fatty acid methyl esters (FAMEs). We found clear differences between their microbial communities, demonstrating the power and sensitivity of the total FAME analysis. In addition, qualitative and quantitative analyses of specific biomarkers permitted to determine differences between samples and to evaluate the effect of earthworms in the decomposition of organic matter. Fatty acid profiles were largely determined by the different vermicomposting earthworm species. Fatty acid 18:2ω6 increased significantly in horse manure vermicomposted by L. rubellus and in cow manure vermicomposted by the three earthworm species, whereas it decreased significantly in pig manure vermicomposted by L. rubellus and E. eugeniae. Fatty acid 20:4ω6 increased significantly in all vermicomposts obtained with the three earthworm species.     

Nutrient content of earthworm casts influenced by different mulch types

The aim of this study was to investigate the effects of different mulch types on the quality and quantity of casts of different lumbricid earthworm species. Nutrient contents (organic carbon C_org, total nitrogen N_t and inorganic phosphate Pi), and acid phosphatase activity (SPA) of earthworm casts and soil aggregates were measured and then compared. This short time experiment was conducted in the laboratory, simulating field conditions of mulch management in temperate agroecosystems. In microcosms, the two common field species Lumbricus terrestris and Octolasion cyaneum were inoculated. Barley, lupin, maize, or sugarbeet were used as mulch according to amounts usually applied in the field. Nutrient contents and phosphatase activity in worm casts were generally enhanced compared to the soil. Nutrient values were higher in the casts of the detritivorous L. terrestris than those of the geophageous O. cyaneum. Conversely, the phosphatase activity was increased in the casts of O. cyaneum compared to L. terrestris in case of the barley and sugar-beet treatments. Cast production was related to the earthworms' change of biomass (ΔB) as well as, in the case of L. terrestris, to nitrogen content of the mulch. Due to their chemical compounds, the mulch types influence both the food selection of the worms and the acceptance by microorganisms.     

Species richness and zoogeographic affinities of earthworms in Cyprus

A new survey taken in the East Mediterranean island of Cyprus showed an increase in the number of earthworm species (from the six recorded earlier to 16). These species belong to two earthworm families: Lumbricidae (15 species) and Acanthodrilidae (one species). The majority of these species are autochthonous (57–64%) and five to seven species (36–44%) were introduced. The autochthonous species are (i) endemic of the Levant (Dendrobaena semitica), (ii) distributed in Anatolia and the Caucasus (D. pentheri), (iii) distributed from Europe up to Anatolia (Allolobophora nematogena, D. pantaleonis, Proctodrilus tuberculatus) (iv) distributed from Europe up to the Levant (Murchieona minuscula), (v) distributed in the Transcaucasian region and in the East Mediterranean (Helodrilus patriarchalis), or (vi) widely distributed along the Mediterranean basin (D. byblica, Eiseniella neapolitana, and Octodrilus complanatus). The data obtained indicate that the Cypriote earthworm fauna is significantly richer in species than previously thought and that it has more zoogeographic affinities with the Anatolian fauna than with those of Levant.     

Occurrence and distribution of earthworms in agricultural landscapes across Europe with regard to testing for responses to plant protection products

Purpose

Within the regulatory framework for authorisation of plant protection products (PPPs) (EU Directive 91/414/1991/EEC replaced by Regulation (EC) 1107/2009), higher tier risk assessments and earthworm field tests are conducted in different countries across Europe. This paper describes dominant earthworm species for regulatory and biogeographical regions in agricultural landscapes across Europe and examines regional differences in earthworm communities and densities and their respective response to a toxic reference.

Materials and methods

For the assessment of earthworm abundance and species distribution, data of untreated control plots from 30 earthworm field studies were analysed; each conducted according to the ISO 11268–3 (1999) guideline by European Crop Protection Association member companies in the context of registration of PPPs. For the evaluation of the response to PPPs under different regional and climatic conditions, the effect on earthworm abundance was assessed by comparing plots treated with toxic references with untreated control plots. Additionally, a comparative literature review was included providing an overview of earthworm species composition and densities in agricultural crops from 14 European countries.

Results and discussion

The assessment of earthworm field studies from six different European countries revealed that common earthworm species of anecic and endogeic ecological groups are present at most field sites. Dominant species groups of endogeic and anecic earthworms can be defined that are abundant in all assessed countries. These are the endogeic species Aporrectodea caliginosa, Aporrectodea rosea and Allolobophora chlorotica, and the anecic species Lumbricus terrestris (Northern and Central Europe) and Lumbricus friendi (Southern Europe). Taking into account the high variability in total earthworm abundances, it can be concluded that the variability within regions was larger than the variability between regions.

Conclusions

Analysis of the earthworm community and data of toxic references lead to the conclusion that testing in different zones is not considered necessary.     

Impact of five different tillage systems on soil organic carbon content and the density,biomass, and community composition of earthworms after a ten year period

To assess the impact of different types of soil tillage on the density, biomass, and community composition of earthworms, a long-term field study was performed in which soils were tilled in different ways for ten years. This study included five different types of tillage: (i) plough, (ii) grubber, (iii) disc harrow, (iv) mulch sowing, and (v) direct sowing. At the end of the experiment the earthworm density, biomass, and community composition, and the SOC (soil organic carbon) content were determined. The results show that density, biomass, and community composition of earthworm populations varied in relation to the type of soil tillage used. The density of anecic earthworm species decreased when soils were managed by conventional ploughing, relative to reduced tillage practices, whereas conversely the density of endogeic species increased. Additionally, the varying types of soil tillage influenced the abundance and biomass of different earthworm species in different ways. The density of Aporrectodea caliginosa was positively influenced by ploughing, whereas Aporrectodea longa, Lumbricus castaneus, and Satchellius mammalis showed a positive relationship to the grubber and Allolobophora chlorotica to direct sowing. We attribute these changes to modifications in the vertical distribution of SOC and varying potentials for mechanical damage of earthworms by tillage. A decrease in tillage intensity modified the vertical SOC distribution in the topsoil and consequently revealed positive effects on earthworm biodiversity, thus sustaining soil functioning.     

Potential of the Silpho Moor experimental birch plots as a habitat for Lumbricus terrestris

It is argued that the podzol of Silpho Moor could be converted under birch to a typical brown earth only if it could sustain an earthworm population with a Lumbricus terrestris biomass of not less than 100 g m^?2. No worms of this species were found under experimental birch plots 30 yr old. The presence of podzolizing species in the ground vegetation; the low pH of the raw humus layer; the low Ca and N supply in the soil, litter and rainfall; and the impeded soil drainage lead to the conclusion that colonization by L. terrestris is unlikely.     

Novel microsatellite loci for the compost earthworm Eisenia fetida: A genetic comparison of three North American vermiculture stocks

Earthworms are important components of soil ecosystems worldwide, and have been used extensively as indicator species in ecotoxicology studies. Our understanding of mating systems, population structure, and genetic diversity in earthworms is limited by the current lack of available genetic tools. To address this gap, we developed 16 novel microsatellite markers for the compost earthworm Eisenia fetida, one of the most widely studied earthworm species. We tested the new markers in 3 putative populations of worms from commercial vermiculture operations in Canada and the U.S.A. All 16 loci were variable in at least one population, with the number of alleles per locus ranging from 3 to 10, observed heterozygosity from 0.000 to 0.783, and polymorphic information content from 0.032 to 0.587. One group had significantly reduced heterozygosity compared to the other 2, but overall there were only minor genetic differences among the tested suppliers, suggesting a possible bottleneck for this species in North America. The microsatellite loci we describe here will be extremely useful tools for future field and laboratory studies of E. fetida. Body size and condition in adult worms varied significantly by supplier, and breeding success and cocoon production were significantly higher in breeding pairs from one supplier compared to the other two. In the absence of evidence for genetic divergence among populations, our results suggest that the early rearing environment may be an important factor affecting adult breeding performance. We caution that the source of worms may affect the outcome of laboratory experiments involving growth and reproduction.