Habitat quality, conspecific density, and habitat pre-use affect the dispersal behaviour of two earthworm species, Aporrectodea icterica and Dendrobaena veneta, in a mesocosm experiment

Dispersal capacity is a life-history trait that may have profound consequences for earthworm populations: it influences population dynamics, species persistence and distribution and community structure. It also determines the level of gene flow between populations and affects processes such as local adaptation, speciation and the evolution of life-history traits. It may play a great role in soil functioning by determining the spatial distribution of ecosystem engineers such as earthworms. Dispersal is an evolutionary outcome of the behaviour in response to the ecological constraints of the species. Hence different dispersal behaviour is expected from the different ecological types of earthworms. Nevertheless the dispersal behaviour of earthworms has been little documented.In this work we test a series of basic mechanisms that are fundamental and complementary to understand earthworms dispersal behaviour. We focus on the dispersal triggered by environmental conditions, a fundamental process usually termed “conditional dependent dispersal”. We show experimentally in mesocosms that in one week: 1) earthworm dispersal can be triggered by low habitat quality, either through soil quality or the presence/absence of litter. 2) Earthworms can be subject to positive density dependent dispersal, that is the rate of dispersal increases when density increases; and 3) earthworm dispersal can be reduced by the pre-use of the soil by conspecific individuals that are no longer present.Our results suggest that earthworms may be more mobile than expected from previous estimations, and that they present high capacities of habitat selection. In the light of our findings we elaborate a behavioural scenario of earthworm foraging, and propose several priority working directions.     

The effects of lead shot deposition on soils and crops at a clay pigeon shooting site in northern England

Abstract. The impact of lead shot on soils and crops was examined at a clay pigeon shooting site in northern England. Topsoil cores were collected along a 300 m transect from the shooting range, and the numbers of lead shot pellets per soil core, total and 'plant-available'(0.5 m acetic acid extractable) lead concentrations, organic matter content, pH and cation exchange capacity were determined. The number of oilseed rape plants and their stem diameters were recorded in 1 m² quadrats placed at the soil sampling locations. Total and 'plant-available' lead concentrations in the soil were most but plant numbers per m² and mean stem diameters were least in the area of greatest lead shot deposition. Total lead concentrations in the soil commonly exceeded 5000 mg/kg; these are considerably greater than threshold 'trigger' concentrations proposed by the Department of the Environment, above which soils are considered to be contaminated and warrant further investigation. Concentrations of lead in the oilseed rape plants themselves were also largest in the area of most intense lead shot deposition; in root samples the lead concentration exceeded 400 mg/kg. The management and remediation of contaminated soils at the clay pigeon shooting site are discussed.     

Fluorine contamination of soils and earthworms (Lumbricus spp.) near a site of long-term industrial emission in southern Germany

Summary The F contamination of soils and Lumbricus spp. around a site of long-term industrial emission in southern Germany was examined. Among total, water extractable, and HCl-soluble fractions, the latter most appropriately characterized anthropogenic F accumulation. Based on the HCl-soluble fractions from 88 sampling sites, a contamination map consisting of three zones was established. F accumulation in the calcareous soils of the area was restricted to the top 40–50 cm and can be explained by precipitation as CaF₂. Earthworms (Lumbricus spp.) collected from the different zones reflected the F contamination well in the significant correlations found between total F in earthworms with and without gut and the corresponding soils. The bioaccumulation of F in earthworms is obvious, and may become hazardous for the earthworms themselves and for other animals feeding on contaminated soil and/or its fauna. A significantly higher F value was recorded in the linings of earthworm tubes than in the corresponding soil. F translocation by earthworm burrowing may be a mechanism of subsoil contamination.     

Genetic variation in desiccation tolerance of Dendrobaena octaedra cocoons originating from different climatic regions

The aim of this study was to examine genetic variation in desiccation tolerance in cocoons of the parthenogenetically reproducing earthworm Dendrobaena octaedra by comparing populations originating from different geographic regions (Denmark, Norway and Finland), representing large differences in precipitation and temperature. In one experiment, the tolerance of the three populations to increasing desiccation stress in the range from 100 to 91.6% relative humidity (RH) was examined, aiming to represent ecologically relevant RH values. In a second experiment, the effect of cocoon size on desiccation tolerance was investigated at 92.3% RH in the same three populations. There were highly significant differences in desiccation tolerance between populations, indicating a high genetic differentiation of this trait in D. octaedra. Cocoons from Denmark were much more sensitive (71±14% mortality at 91.6% RH) than cocoons from Norway (21±4% mortality) and Finland (4±5% mortality). Cocoons of worms from Finland and Norway were significantly larger than cocoons produced by worms from Denmark suggesting that cocoons from Denmark lost water at a higher rate when subjected to low humidity. Assuming that slow dehydration is necessary for physiologically based protection mechanisms it may be expected that desiccation tolerance is positively correlated with cocoon size. However, within each of the populations cocoon fresh weight did not have any significant impact on desiccation tolerance. When all populations were pooled there was a significant positive effect of cocoon fresh weight on desiccation tolerance, explaining about 20% of the total variation (linear regression). It seems therefore that genetic variation of desiccation tolerance in D. octaedra cocoons is related to variation in both cocoon size and other, physiologically based tolerance mechanisms.     

A vagrant clone in a peregrine species: Phylogeography, high clonal diversity and geographical distribution in the earthworm Aporrectodea trapezoides (Dugès, 1828)

The peregrine lumbricid Aporrectodea trapezoides is originally from the Palaearctic region but is distributed worldwide. Little is known about its clonal diversity or the existence of a pattern of biogeographical diversification of clones. This study aimed to explore the evolutionary history of A. trapezoides by analysing the mitochondrial (COI and COII) and nuclear (28S rRNA and histone H3) DNA sequences of individuals collected in 11 different countries. High clonal diversity was found for this species, with thirty-seven clones clearly divided into two distinct lineages (I and II). The marked biogeographical boundary between these lineages corresponds to the line separating the Eurosiberian and Mediterranean climates in North Spain. Clone 1 was shared by one-third of the earthworms. While this clone was found in most of the sampled localities, the rest of the clones showed geographically-restricted distributions. This clone may have originated in the Mediterranean area of Central Spain. As it was obviously introduced in Australia and it was found in locations occupied by members of both lineages, we hypothesise that it may have also been introduced in other countries and that it could be a general-purpose genotype able to adapt to a wide range of niches.     

Life cycle and reproductive traits of the earthworm Aporrectodea trapezoides (Dugès, 1828) in laboratory cultures

The parthenogenetic earthworm Aporrectodea trapezoides (Dugès, 1828) is widely distributed all over the world due to European agricultural practices. In order to provide baseline life cycle data, cocoons were obtained from field-collected individuals and their features and viability, incubation period, number of hatchlings and mortality rate were recorded. Singleton and twin earthworms from this first experiment were cultured from hatching during a 490-day period under controlled conditions with biomass, survival, reproductive condition and cocoon production recorded at intervals of 15 days. On average, individuals of isolated-reared A. trapezoides reached maturity at day 153 and body weight at maturity was approximately 1 g. In order to record reproductive traits and differences between field-collected and laboratory-reared individuals, 40 microcosms with an isolated earthworm (20 with field-collected individuals and 20 with laboratory-reared ones) and 40 containing groups of three (20 with each type of individual) were maintained during a complete year under controlled conditions. The amount of soil per individual was the same in both types of microcosm. Both the individuals kept in isolation and those cultured in groups produced cocoons, hence completely proving the obligatory parthenogenetic reproduction in this species without copulation or need of any physical-chemical stimulus. In general, isolated earthworms produced a significantly higher number of cocoons than those in groups of three, and the same was recorded for laboratory-reared earthworms when compared with field-collected ones. This study highlights the importance of knowing the life cycle and reproductive traits of one possible key species in soil management due to its vast distribution and high density in soils, and the species’ highly recommended use in applied studies because of its ease of culture.     

The effect of feeding behavior on Hg accumulation in the ecophysiologically different earthworms Lumbricus terrestris and Octolaseon cyaneum: A microcosm experiment

 A soil microcosm experiment was performed to assess the uptake of Hg from various Hg-spiked food sources (soil, leaf litter and root litter of Trifolium alexandrinum) by two earthworm species, Lumbricus terrestris (anecic) and Octolaseon cyaneum (endogeic). Treatments were applied in which one of the three food sources was Hg spiked and the other two were not. Additional treatments in which all or none of the food sources were Hg spiked were used as controls. Uptake of Hg from soil into tissues of both earthworm species was significantly higher than uptake of Hg from leaf litter or root litter, indicating that soil may be the most important pool for the uptake of Hg into earthworms. In addition, the anecic L. terrestris significantly accumulated Hg from all Hg-spiked food sources (leaf litter, root litter and soil), whereas the endogeic O. cyaneum took up Hg mainly from soil particles. Interestingly, there was no further increase in Hg in L. terrestris when all food sources were Hg spiked compared to the single Hg-spiked sources. This may be attributed to the relatively high Hg content in the soil, which may have influenced the feeding behavior of the earthworms, although their biomass did not significantly decline. We suggest that, in addition to the physiological differences, feeding behavior may also play a role in the contrasting uptake of Hg by the two earthworm species.     

Experimental evidence for the role of earthworms in compacted soil regeneration based on field observations and results from a semi-field experiment

In laboratory experiments, earthworms are often observed to burrow through compacted soil layers, leading to the general assumption that these animals play a significant role in regenerating compacted soils in agricultural plots. To demonstrate this role under field conditions, the abundance of earthworm macropores inside compacted zones was estimated on plots under reduced (RT) or conventional tillage (CT). Then, different types of compacted zones typically found in CT (plough pan and compacted clods) and RT plots (compacted volume under wheel tracks) were experimentally simulated in wooden boxes, buried in the field and inoculated with different earthworm species. After 6 weeks of incubation, the number of macropores inside the compacted zones was examined. Field observations showed that approximately 10% and 30% of the compacted zones were colonised by at least one macropore in CT and RT plots, respectively. A significantly greater number of anecics was found in RT plots, but we could not conclude that this ecological type of earthworm plays a more major role in the regeneration process in these plots since there were fewer compacted zones and these covered a smaller area in CT. The semi-field experiment provided evidence that earthworm-mediated regeneration of compacted zones is possible and its nature varies between ecological types of earthworm. Lumbricus terrestris, which makes individual burrows that are vertical and deep, was the main species to cross through the plough pan. The other three earthworm species (Aporrectodea giardi, A. caliginosa and A. rosea) did burrow inside the other types of compacted zones (“wheel tracks” and “compacted clods”). In every case, however, macropore density was far greater in non-compacted zones, illustrating that avoidance of compacted soil by earthworms is important and should be taken into account when extrapolating results from laboratory studies.     

Genetic structure among earthworms (Lumbricus terrestris L.) from different sampling sites in western Germany based on random amplified polymorphic DNA

Earthworms are being used as bio-indicators to assess terrestrial pollution. However, it is often not known whether their populations possess a uniform genetic structure, which would allow comparison of residues or biological properties of earthworms from different sampling locations. In order to investigate this point, random amplified polymorphic DNA (RAPD) variation was surveyed in earthworms (Lumbricus terrestris) from five different sampling sites in Germany. Forty oligonucleotide RAPD primers (10 base pairs in length) were screened, three of which produced high polymorphic band patterns. A total of 61 DNA fragments were detected in 90 individuals of L. terrestris from five sampling sites with 49 (80.3%) RAPD markers being polymorphic. The genetic similarities within (band sharing rates between 0.756 and 0.795) and among the L. terrestris populations (0.635) were similar even at widely separate locations. Inter-population variation in the RAPD pattern for all five earthworm populations accounted for 37.9% of the total variation, while intra-population variation for three adjacent Saarland populations accounted for only 18.0% of the total variation. Principal component analysis (PCA) and the genetic distances of the populations confirm these results. Twenty-four percent of the genetic distance is caused by geographical isolation as shown by a test for isolation by distance. These results show that L. terrestris fulfils the genetic qualifications for a bio-indicator particularly at closely located sampling sites. However, the results also suggest that earthworm studies of widely separated locations should include genetic characterisation of the earthworm samples.     

茶叶皂角用于提高大豆油-水体系洗脱修复焦化厂土壤中多环芳烃、Cd、Ni的可行性研究

Mixed contaminated brownfield sites have brought serious risks to human health and environmental safety. With the purpose of removing polycyclic aromatic hydrocarbons(PAHs) and heavy metals from a coking plant site, an innovative technology for ex-situ washing was developed in the present work. The combination of 15.0 mL L~(-1) soybean oil and 7.5 g L~(-1) tea saponin proved an effective method to extract co-pollutants from soil. After two consecutive washing cycles, the efficiency rates of removal for 3-, 4-,5(+6)-ring, and total PAHs, Cd, and Ni were approximately 98.2%, 96.4%, 92.3%, 96.3%, 94.1%, and 89.4%, respectively. Meanwhile,as evaluated by Tenax extraction method and metal stability indices, the residual PAHs and heavy metals after consecutive washing mainly existed in the form with extremely low bioaccessibility in the soil. Thus, in the soil after two washing cycles, there appeared limited environmental transfer risk of co-pollutants. Moreover, a subsequent precipitation method with alkaline solution and PAHdegrading strain Sphingobium sp. PHE9 inoculation effectively removed 84.6%–100% of Cd, 82.5%–91.7% of Ni, and 92.6%–98.4% of PAHs from the first and second washing solvents. The recovered solvents also exhibited a high recycling effectiveness. Therefore, the combined cleanup strategy proposed in this study proved environmentally friendly, which also played a major role in risk assessment and management in mixed polluted sites.     

Lead shot and teal (Anas crecca) in the Camargue, Southern France: Effects of embedded and ingested pellets on survival

The negative consequences of lead shot ingestion have been demonstrated in captive birds, and the prevalence of embedded shot has been measured in wild waterfowl several times. However, the long-term consequences of these two indirect outcomes of hunting on waterfowl survival have merely been investigated. Using data from about 40,000 X-rayed teals (Anas crecca), this study attempts to evaluate these effects. We used multivariate logistic regressions to model the probability of carrying shot in the flesh or in the gizzard while testing for various explanatory variables such as sex, age, time or morphological measures like mass and flattened wing length. Because of technical issues, we used a non-parametric sign test rather than a complete capture-recapture analysis to evaluate the effect of shot on teal survival. This test was applied to the differences in time between ringing and recovery for lead categories (no lead shot, ?1 shot in the flesh but none in the gizzard, ?1 shot in the gizzard but none in the flesh) compared two by two. We show that, overall, males are more likely to carry embedded shot than females whereas the latter are more likely to carry lead pellets in the gizzard. Similarly, adults are more likely to carry shot in the flesh whereas first year individuals are more likely to have pellets in the gizzard. Embedded shot tend to accumulate in the ducks body over time, i.e. with the number of encounters with hunters, with no significant effect on survival. Conversely, the probability of carrying shot in the gizzard decreases over time, because lead-poisoning quickly leads the individual to death. Several possible biological interpretations are put forward to explain those results. This study demonstrates that negative impacts of ingested lead on teal survival arise after only one single pellet is ingested, advocating the ban of lead ammunition.     

Microbial utilisation of two proteins adsorbed to a vertisol clay fraction: toxin from Bacillus thuringiensis subsp. tenebrionis and bovine serum albumin

Clay minerals have been shown to reduce the extent and rate of biodegradation of several compounds. Here, we investigated the ability of soil clays to protect proteins from biodegradation: the insecticidal protein from Bacillus thuringiensis subsp. tenebrionis (Btt toxin) and Bovine Serum Albumin (BSA). The two proteins adsorbed in large amounts (up to 0.24 g BSA g⁻¹ clay and 0.74 g Btt toxin g⁻¹ clay) and irreversibly to smectite clay particles from a vertisol. We measured the growth of a soil inoculum in the presence of each of proteins as the sole source of carbon. When clay was present in the medium, microbial growth was directly proportional to the amount of free protein (i.e. nonadsorbed). Hence, the two proteins were unavailable when adsorbed to clay. The clay had little influence on the ability of microorganisms to hydrolyse a soluble substrate. The inhibitory effect of clays on utilisation of BSA and Btt toxin was interpreted as being the result of the adsorption of the proteins to clay, which rendered the proteins unavailable for microbial utilisation.     

Non-target effects of the microbial control agents Pseudomonas fluorescens DR54 and Clonostachys rosea IK726 in soils cropped with barley followed by sugar beet: a greenhouse assessment

Non-target effects of a bacterial (Pseudomonas fluorescens DR54) and a fungal (Clonostachys rosea IK726) microbial control agent (MCA), on the indigenous microbiota in bulk soil and rhizosphere of barley, and subsequent a sugar beet crop, were studied in a greenhouse experiment. MCAs were introduced by seed and soil inoculation. Bulk and rhizosphere soils were sampled regularly during the growth of barley and sugar beet. The soils were assayed for the fate of MCAs and various features of the indigenous soil microbiota. At the end of the experiment (193 d), DR54 and IK726 had declined by a factor of 10⁶ and 20, respectively, and DR54 showed a short-lasting growth increase in the sugar beet rhizosphere. In general, the non-target effects were small and transient. IK726 seemed to have general stimulating effects on soil enzyme activity and the soil microbiota, and resulted in a significant increase in plant dry weight. The plant growth-promoting effect of DR54 was less pronounced and the DR54 displaced indigenous pseudomonads. DR54 stimulated growth of protozoans with a tolerance for the anti-fungal compound viscosinamide produced by DR54. Treatment with the fungicide Fungazil had no effects on plant growth or soil microorganisms. Phospholipid fatty acid (PLFA) analysis detected the perturbations of the soil microbial community structure in the MCA treatments as well as the return to non- perturbed conditions reflecting the decline of inoculant populations. The PLFA technique appears to be suitable for in situ monitoring of MCA non-target effects on the soil microbiota, but should be combined with assays for MCA survival and soil enzyme activity.     

A phylogenetic microarray targeting 16S rRNA genes from the bacterial division Acidobacteria reveals a lineage-specific distribution in a soil clay fraction

We designed an oligonucleotide microarray using probe sequences based upon a phylogenetic analysis of 16S rRNA genes recovered from members of the bacterial division Acidobacteria. A total of 42,194 oligonucleotide probes targeting members of the Acidobacteria division at multiple phylogenetic levels were included on a high-density microarray. Positive control hybridizations revealed a linear relationship between hybridization signal and template concentration, and a substantial decrease in non-specific hybridization was achieved through the addition of 2.5 M betaine to the hybridization buffer. A mean hybridization signal value was calculated for each Acidobacteria lineage, with the resultant lineage-specific hybridization data revealing strong predictive value for the positive control hybridizations. The Acidobacteria phylochip was then used to evaluate Acidobacteria rRNA genes from a Wisconsin soil and within a soil clay fraction. The Acidobacteria hybridization profile revealed the predominance of Acidobacteria subdivisions four and six, and also suggested a decrease in the abundance of subdivision six relative to subdivision four in the soil clay fraction. The change in relative abundance of these subdivisions in a soil clay fraction was supported by data from quantitative PCR. These results support the utility of a phylogenetic microarray in revealing changes in microbial population-level distributions in a complex soil microbial assemblage.     

Methane emissions from stems of Fraxinus mandshurica var. japonica trees in a floodplain forest

We measured methane (CH₄) emissions from the stem surfaces of mature Fraxinus mandshurica var. japonica trees in a floodplain forest. Flux measurements were conducted almost monthly from May to October 2005, and positive CH₄ fluxes were detected throughout the study period, including the leafless season. The mean CH₄ flux was 176 and 97 μg CH₄ m⁻² h⁻¹ at the lower (15 cm above the ground) and upper (70 cm above the ground) stem positions, respectively. The CH₄ concentration was lower in soil gas than in ambient air to a depth of at least 40 cm. One possible source of CH₄ emitted from the stems might be the dissolved CH₄ in groundwater; maximum concentrations were 10,000 times higher than atmospheric CH₄ concentrations. Our results suggest that CH₄ transport from the submerged soil layer to the atmosphere may occur through internal air spaces in tree bodies.     

Reduced seed production, inbreeding, and pollen shortage in a small population of a threatened tree, Magnolia stellata

We investigated how genetic factors and pollen shortage affect seed production in a small population of Magnolia stellata, a threatened insect-pollinated tree. We used microsatellite and manual pollination techniques, and compared a small population to a large one. Compared to the large population, the small population showed low allelic variation and had an F_IS significantly greater than 0 in adults, indicating that genetic deterioration, including genetic drift and inbreeding, may have occurred in adults. Manual self-pollination lowered seed production relative to manual cross-pollination to a different extent between populations: δ (the magnitude of inbreeding depression due to self-fertilization) was lower in the small population than in the large population. However, under natural pollination, the estimated embryo mortality rates after ovules self-fertilized were similar between the two populations because the primary selfing rate was higher in the small population. The ovule mortality rate due to pollen shortage and that due to factors independent of pollen shortage and selfing were both approximately 10% higher in the small population, suggesting that pollen transfer may have decreased and genetic deterioration effects may have increased in that population. These factors reduced seed production in the small population (female reproductive success = 0.3%) compared to the large population (2.6%). Our results suggest that seed production in the small population of M. stellata is strictly limited by elevated pollen shortage, selfing, and genetic deterioration in adults, which accelerate the risk of extinction.     

大麦过氧化物体膜上的抗坏血酸过氧化物酶拟南芥的耐盐机制

Ascorbate peroxidases （APX）, localized in the cytosol, peroxisome, mitochondria, and chloroplasts of plant cells, catalyze the reduction of H2O2 to water by using ascorbic acid as the specific electron donor. To determine the role of peroxisomal type ascorbate peroxidase （pAPX）, an antioxidant enzyme, in protection against salt-induced oxidative stress, transgenic Arabidopsis thaliana plant carrying a pAPX gene （HvAPX1） from barley （Hordeum vulgate L.） was analyzed. The transgenic line pAPX3 was found to be more tolerant to salt stress than the wild type. Irrespective of salt stress, there were no significant differences in Na^＋, K^＋, Ca^2＋, and Mg^2＋ contents and the ratio of K^＋ to Na^＋ between pAPX3 and the wild type. Clearly, the salt tolerance in pAPX3 was not due to the maintenance and reestablishment of cellular ion homeostasis. However, the degree of H2O2 and lipid peroxidation （measured as the levels of malondialdehyde） accumulation under salt stress was higher in the wild type than in pAPX3. The mechanism of salt tolerance in transgenic pAPX3 can thus be explained by reduction of oxidative stress injury. Under all conditions tested, activities of superoxide, glutathione reductase, and catalase were not significantly different between pAPX3 and the wild type. In contrast, the activity of APX was significantly higher in the transgenic plant than in wild type under salt stress. These results suggested that in higher plants, HvAPX1 played an important role in salt tolerance and was a candidate gene for developing salttolerant crop plants.     

Glycosidation of apigenin results in a loss of its activity on different growth parameters of arbuscular mycorrhizal fungi from the genus Glomus and Gigaspora

The effect of different concentrations (0.5, 2 and 8 μM) of apigenin and its glycosidated form 5,7,4′-hydroxy flavone glycoside on arbuscular mycorrhizal (AM) fungal spore germination, hyphal growth, hyphal branching, the formation of entry points and root colonization of Gigaspora. rosea, Gi. margarita, Glomus mosseae and G. intraradices was tested. The lowest apigenin concentration (0.5 μM) nearly doubled hyphal branching, the formation of entry points and root colonization of all four tested fungi, whereas higher concentrations (2 and 8 μM) nearly doubled the hyphal growth of Gi. margarita, G. mosseae and G. intraradices. In none of the treatments with the apigenin-glycoside any effect on AM fungi could be observed. Our data show that apigenin exhibits an AM fungal genus and even species activity and we provide strong evidence that glycosidation results in a loss of its activity towards AM fungi.