Lethal critical body residues as measures of Cd,Pb, and Zn bioavailability and toxicity in the earthworm<Emphasis Type="Italic">Eisenia fetida</Emphasis>

Background. Earthworm heavy metal concentrations (critical body residues, CBRs) may be the most relevant measures of heavy metal bioavailability in soils and may be linkable to toxic effects in order to better assess soil ecotoxicity. However, as earthworms possess physiological mechanisms to secrete and/or sequester absorbed metals as toxicologically inactive forms, total earthworm metal concentrations may not relate well with toxicity. Objective  The objectives of this research were to: i) develop LD₅₀s (total earthworm metal concentration associated with 50% mortality) for Cd, Pb, and Zn; ii) evaluate the LD₅₀ for Zn in a lethal Zn-smelter soil; iii) evaluate the lethal mixture toxicity of Cd, Pb, and Zn using earthworm metal concentrations and the toxic unit (TU) approach; and iv) evaluate total and fractionated earthworm concentrations as indicators of sublethal exposure. Methods  Earthworms (Eisenia fetida (Savigny)) were exposed to artificial soils spiked with Cd, Pb, Zn, and a Cd-Pb-Zn equitoxic mixture to estimate lethal CBRs and mixture toxicity. To evaluate the CBR developed for Zn, earthworms were also exposed to Zn-contaminated field soils receiving three different remediation treatments. Earthworm metal concentrations were measured using a procedure devised to isolate toxicologically active metal burdens via separation into cytosolic and pellet fractions. Results and Discussion  Lethal CBRs inducing 50% mortality (LD₅₀, 95% CI) were calculated to be 5.72 (3.54-7.31), 3.33 (2.97-3.69), and 8.19 (4.78-11.6) mmol/kg for Cd, Pb, and Zn, respectively. Zn concentrations of dead earthworms exposed to a lethal remediated Zn-smelter soil were 3-fold above the LD₅₀ for Zn and comparable to earthworm concentrations in lethal Zn-spiked artificial soils, despite a 14-fold difference in total soil Zn concentration between lethal field and artificial soils. An evaluation of the acute mixture toxicity of Cd, Pb, and Zn in artificial soils using the Toxic Unit (TU) approach revealed an LD₅₀ (95% CI) of 0.99 (0.57-1.41) TU, indicating additive toxicity. Conclusions  Total Cd, Pb, and Zn concentrations in earthworms were good indicators of lethal metal exposure, and enabled the calculation at LD₅₀s for lethality. The Zn-LD₅₀ developed in artificial soil was applicable to earthworms exposed to remediated Zn-smelter soil, despite a 14-fold difference in total soil Zn concentrations. Mixture toxicity evaluated using LD₅₀s from each single metal test indicated additive mixture toxicity among Cd, Pb, and Zn. Fractionation of earth worm tissues into cytosolic and pellet digests yielded mixed results for detecting differences in exposure at the sublethal level Recommendation and Outlook  CBRs are useful in describing acute Cd, Pb, and Zn toxicity in earthworms, but linking sublethal exposure to total and/or fractionated residues may be more difficult. More research on detoxification, regulation, and tissue and subcellular partitioning of heavy metals in earthworms and other invertebrates is needed to establish the link between body residue and sublethal exposure and toxicity. Keywords: Bioavailability; Cd; critical body residues; earthworms; metals; Pb; soil; Zn An erratum to this article is available at .     

Uptake and bioaccumulation of heavy elements by two earthworm species from a smelter contaminated area in northern Kosovo

As, Cd, Cu, Pb, Sb and Zn concentrations were determined in two earthworm species (Allolobophora rosea and Nicodrilus caliginosus) from a mining and industrial area in northern Kosovo and compared with their contents in the bulk soil and the main soil fractions. Earthworm specimens were collected at fifteen sites located at different distances from a Pb–Zn smelter along a gradient of decreasing contamination. Individuals of A. rosea and N. caliginosus showed similar tissue levels of As, Cd, Cu, Pb, Sb and Zn, suggesting that earthworm species belonging to the same eco-physiological group have a similar propensity to uptake and bioaccumulate heavy elements. Cd, Pb, Sb and Zn concentrations in both earthworm species were positively correlated with the respective total soil contents and generally decreased with distance from the smelter. The bioaccumulation factor (BAF) revealed that Cd and Zn were the only elements bioaccumulated by earthworms. The rank order of BAF values for both species was as follows: Cd > > Zn > > Cu > As = Pb = Sb. The absorption of Cd, Pb, Sb and Zn by earthworms mostly depended on the extractable, reducible and oxidable soil fractions, suggesting that the intestine is likely the most important uptake route. The extractable soil fraction constantly influenced the uptake of these heavy elements, whereas the reducible fraction was important mainly for Pb and Zn. The water soluble fraction had an important role especially for the most mobile heavy elements such as Cd and Zn, suggesting that dermal uptake is not negligible. As a whole, the analytical data indicate that soil fractionation patterns influence the uptake of heavy elements by earthworms, and the extractable fraction is a good predictor of heavy element bioavailability to these invertebrates in soil.     

Effects of heavy metals on the litter consumption by the earthworm Lumbricus rubellus in field soils

Aim of this study was to determine effects of heavy metals on litter consumption by the earthworm Lumbricus rubellus in National Park the “Brabantsche Biesbosch”, the Netherlands. Adult L. rubellus were collected from 12 polluted and from one unpolluted field site. Earthworms collected at the unpolluted site were kept in their native soil and in soil from each of the 12 Biesbosch sites. Earthworms collected in the Biesbosch were kept in their native soils. Non-polluted poplar (Populus sp.) litter was offered as a food source and litter consumption and earthworm biomass were determined after 54 days. Cd, Cu and Zn concentrations were determined in soil, pore water and 0.01 M CaCl₂ extracts of the soil and in earthworms. In spite of low available metal concentrations in the polluted soils, Cd, Cu and Zn concentrations in L. rubellus were increased. The litter consumption rate per biomass was positively related to internal Cd and Zn concentrations of earthworms collected from the Biesbosch and kept in native soil. A possible explanation is an increased demand for energy, needed for the regulation and detoxification of heavy metals. Litter consumption per biomass of earthworms from the reference site and kept in the polluted Biesbosch soils, was not related to any of the determined soil characteristics and metal concentrations.     

The role of earthworms (<Emphasis Type="Italic">Eisenia fetida</Emphasis>) in influencing bioavailability of heavy metals in soils

The effects of earthworm (Eisenia fetida) activity on soil pH, dissolved organic carbon (DOC), microbial populations, fraction distribution and bioavailability of heavy metals (Zn, Cu, Cr, Cd, Co, Ni, and Pb) in five Chinese soils were investigated using pot experiments. A three-step extraction procedure recommended by the European Community Bureau of Reference (BCR; now Standards, Measurements and Testing Programme of the European Community) was used to fractionate the metals in soils into water soluble, exchangeable and carbonate bound (B1), Fe-oxides and Mn-oxides bound (B2) and organic matter and sulfide bound (B3). After the soils were treated with earthworms, the soil pH, water-soluble metal fraction and DOC increased. A significant correlation was obtained between the increased DOC and the increased metals in the water-soluble fraction. The heavy metals in fraction B1 increased after earthworm treatments, while those in fraction B3 decreased. No significant differences were observed for heavy metals in fraction B2. The microbial populations in soil were enumerated with the dilution plate method using several media in the presence of earthworms. The microbial populations increased due to earthworm activity. The biomass of wheat shoots and roots, and the heavy metal concentrations in wheat roots and shoots, were also increased due to the earthworm activity. The present results demonstrated that earthworm activity increases the mobility and bioavailability of heavy metals in soils.     

Uptake and bioaccumulation of heavy elements by two earthworm species from a smelter contaminated area in northern Kosovo

As, Cd, Cu, Pb, Sb and Zn concentrations were determined in two earthworm species (Allolobophora rosea and Nicodrilus caliginosus) from a mining and industrial area in northern Kosovo and compared with their contents in the bulk soil and the main soil fractions. Earthworm specimens were collected at fifteen sites located at different distances from a Pb–Zn smelter along a gradient of decreasing contamination. Individuals of A. rosea and N. caliginosus showed similar tissue levels of As, Cd, Cu, Pb, Sb and Zn, suggesting that earthworm species belonging to the same eco-physiological group have a similar propensity to uptake and bioaccumulate heavy elements. Cd, Pb, Sb and Zn concentrations in both earthworm species were positively correlated with the respective total soil contents and generally decreased with distance from the smelter. The bioaccumulation factor (BAF) revealed that Cd and Zn were the only elements bioaccumulated by earthworms. The rank order of BAF values for both species was as follows: Cd > > Zn > > Cu > As = Pb = Sb. The absorption of Cd, Pb, Sb and Zn by earthworms mostly depended on the extractable, reducible and oxidable soil fractions, suggesting that the intestine is likely the most important uptake route. The extractable soil fraction constantly influenced the uptake of these heavy elements, whereas the reducible fraction was important mainly for Pb and Zn. The water soluble fraction had an important role especially for the most mobile heavy elements such as Cd and Zn, suggesting that dermal uptake is not negligible. As a whole, the analytical data indicate that soil fractionation patterns influence the uptake of heavy elements by earthworms, and the extractable fraction is a good predictor of heavy element bioavailability to these invertebrates in soil.     

Soil properties and metal accumulation by earthworms in the Siena urban area (Italy)

This paper reports the results of a study focused on the metal (Cd, Co, Cr, Cu, Ni, Pb, Sb, U and Zn) distribution in soils and uptake and accumulation by earthworms Nicodrilus caliginosus (Savigny) from urban, peri-urban, green-urban and non-urban zones of Siena municipality (central Italy). The main goal was to define the influence of soil properties and metal soil contents on the uptake of these contaminants by earthworms. Data indicated that Cd, Cu, Pb, Sb and Zn soil contents increased in the following order: non-urban < green-urban < peri-urban < urban soils, suggesting that vehicular traffic affects the distribution of these metals. Pb and Sb were the main soil contaminants and their highest enrichments were found in urban sites where stop-and-go traffic occurs. Concentrations of these traffic-related metals in earthworms showed a distribution pattern similar to that in soil, suggesting that soil contamination influenced the uptake of Cd, Cu, Pb, Sb and Zn by N. caliginosus. There were significant positive correlations between Cd, Pb and Sb earthworm concentrations and their soil contents. The lack of correlation for Cu and Zn could be due to the physiological regulation of these elements by earthworms. Statistical analysis pointed out that the uptake and accumulation of Cd, Cu, Pb, Sb and Zn by earthworms were affected by some soil physicochemical properties such as the organic carbon and carbonate contents that are able to rule the bioavailability of metals in soils.     

Schwermetallanreicherungen in Salzmarschen der südlichen Nordseeküste

Heavy metal accumulations in Salic Fluvisols of the southern North Sea coast The total contents of Cd, Pb, Cu and Zn (HNO₃ bomb digestion) and their EDTA extractable fractions in Salic Fluvisols were investigated. The mean total content of Cd was 0,09 mg/kg, of Pb 44,1 mg/kg, of Cu 11,4 mg/kg and of Zn 105,6 mg/kg. Mostly the highly developed soils showed higher contents than poorly developed ones. The EDTA extractable fractions were comparatively small: they were 59% (Cd), 44% (Pb), 15% (Cu) and 10% (Zn) of the total content. The heavy metal contents of the soils in Elisabeth-Außengroden were higher than of those in Jadebusen. The vertical heavy metal distribution showed a different sedimentation pattern over the last hundred years. Compared with the geological background values Cd was 2- to 3-fold enriched, Pb 7-fold, Cu 3- to 4-fold and Zn 3-fold in soils with high sedimentation rates. Even higher values are likely in soils with smaller sedimentation rates.     

土法炼锌区大气沉降Pb、Zn、Cd及其对土壤质量的影响

Dust emissions from smelters, as a major contributor to heavy metal contamination in soils, could severely influence soil quality. Downwind surface soils within 1.5 km of a zinc smelter, which was active for 10 years but ceased in 2000, in Magu Town, Guizhou Province, China were selected to examine Pb, Zn, and Cd concentrations and their fractionation along a distance gradient from a zinc smelter, and to study the possible effects of Pb, Zn, and Cd accumulation on soil microorganisms by comparing with a reference soil located at a downwind distance of 10 km from the zinc smelter. Soils within 1.5 km of the zinc smelter accumulated high levels of heavy metals Zn (508 mg kg-1), Pb (95.6 mg kg-1), and Cd (5.98 mg kg-1) with low ratios of Zn/Cd (59.1--115) and Pb/Cd (12.4--23.4). Composite pollution indices (CPIs) of surface soils (2.52--15.2) were 3 to 13 times higher than the reference soils. In metal accumulated soils, exchangeable plus carbonate-bound fractions accounted for more than 10% of the total Zn, Pb, and Cd. The saturation degree of metals (SDM) in soils within 1.5 km of the smelter (averaging 1.25) was six times higher than that of the reference soils (0.209). A smaller soil microbial biomass was found more frequently in metal accumulated soils (85.1--438 μg C g-1) than in reference soils (497 μg C g-1), and a negative correlation (P < 0.01) of soil microbial biomass carbon to organic carbon ratio (Cmic/Corg) with SDM was observed. Microbial consumption of carbon sources was more rapid in contaminated soils than in reference soils, and a shift in the substrate utilization pattern was apparent and was negatively correlated with SDM (R = -0.773, P < 0.01). Consequently, dust deposited Pb, Zn, and Cd in soils from zinc smelting were readily mobilized, and weredetrimental to soil quality mainly in respect of microbial biomass.     

Heavy Metal Concentrations,Partitioning, and Storage in Slovak Forest and Arable Soils Along a Deposition Gradient

Along a heavy metal deposition gradient, caused by a Cu smelter, heavy metal concentrations, partitioning, and storage in forest and arable soils were examined. We sampled organic and mineral soil horizons (0—50 cm) at ten pairs of forest and arable sites derived from the same parent material. A-horizons were extracted with a seven-step sequence; O- and subsoil horizons were digested with strong acids (HNO₃/HClO₄). We found high concentrations of Cd (up to 17.38 mg kg^—1 in the O horizons/up to 2.44 mg kg^—1 in the A horizons), Cu (8437/415), Pb (3343/126), and Zn (1482/637) which decreased exponentially with distance from the smelter and with soil depth. The metal concentrations in the organic layers indicate that the average transport distance decreases in the order Cd > Zn > Pb > Cu. With regard to metal partitioning, NH₄NO₃- + NH₄OAc-extractable forms in the A horizons were most affected by the deposition being more pronounced under forest. In the uppermost 50 cm of the four soils nearest to the smelter two to four times higher Cd, Cu, Pb, and Zn storages were found in forest than in arable soils. At greater distance, the higher deposition onto forest soils due to the scavenging effect of the canopy obviously was compensated by stronger leaching.     

Heavy metal levels in earthworms of a forest ecosystem influenced by traffic and air pollution

Soils of a forest ecosystem in Berlin (West) are highly polluted by Pb and less polluted by Cd and Cu. Pb levels in earthworm species depend primarily on soil type and only secondarily on the distance from a highway crossing the forest. The dominating species Lumbricus rubellus and Dendrobaena octaedra show different body burdens especially of Cd. Liming the forest soils decreases the Pb concentrations significantly. Only in D. octaedra has body weight been found to be related to Pb; Pb has been increased from 50 mg kg^–1 in small to 250 mg kg^–1 in large specimens; Cu has been regulated at a more or less constant absolute level (about 0.4 pg per specimen). This species obviously regulates or accumulates the three metals in different ways. Centipedes as predators of earthworms in the sites show markedly less pollution of Pb and Cd (about 2.5 and 0.6 mg kg^–1) than the worms, but higher concentrations of the essential Cu (about 40 mg kg^–1). The use of earthworms as indicators of heavy metal pollution should take into account the biology of the various species, the soil type, the type and amount of organic matter as well as chemical parameters such as pH value or basic anions.     

Oral sealing using glue: a new method to distinguish between intestinal and dermal uptake of metals in earthworms

Earthworms may take up chemicals from soil and pore water, both through their skin (dermal) and by ingestion (oral). It remains unclear, however, what the relative importance of these pathways is. To assess bioavailability of pollutants in soil to earthworms, it is necessary that the contribution of each pathway is known. Lumbricus rubellus were sealed by means of medical histoacryl glue, to block ingestion of soil particles and pore water. For 6 d, these earthworms showed good survival and vitality and no soil ingestion was found. Equal metal uptake was found by sealed and unsealed earthworms exposed to an inert sand matrix continuously flushed with contaminated water. Therefore, pore water uptake via ingestion contributes little to metal accumulation. Uptake rates of Cd, Cu and Pb in sealed and unsealed earthworms exposed to two contaminated field soils were similar. Uptake and elimination kinetics of Zn were significantly lower in sealed earthworms exposed to one of the two field soils. Body concentrations of Cu and Pb could be completely attributed to the dermal route. For internal Cd and Zn concentrations, however, 0-17 and 21-30%, respectively, were derived from ingestion. It is concluded that for metals the dermal route is the uptake route of importance. The sealing method described here may be useful in a variety of earthworm nutrition and contamination-effect studies.     

Quantification of nitrogen excretion rates for three lumbricid earthworms using 15N

 Nitrogen excretion rates of ¹⁵N-labeled earthworms and contributions of ¹⁵N excretion products to organic (dissolved organic N) and inorganic (NH₄-N, NO₃-N) soil N pools were determined at 10  °C and 18  °C under laboratory conditions. Juvenile and adult Lumbricus terrestris L., pre-clitellate and adult Aporrectodea tuberculata (Eisen), and adult Lumbricus rubellus (Hoffmeister) were labeled with ¹⁵N by providing earthworms with ¹⁵N-labeled organic substrates for 5–6 weeks. The quantity of ¹⁵N excreted in unlabeled soil was measured after 48 h, and daily N excretion rates were calculated. N excretion rates ranged from 274.4 to 744 μg N g^–1 earthworm fresh weight day^–1, with a daily turnover of 0.3–0.9% of earthworm tissue N. The N excretion rates of juvenile L. terrestris were significantly lower than adult L. terrestris, and there was no difference in the N excretion rates of pre-clitellate and adult A. tuberculata. Extractable N pools, particularly NH₄-N, were greater in soils incubated with earthworms for 48 h than soils incubated without earthworms. Between 13 and 40% of excreted ¹⁵N was found in the ¹⁵N-mineral N (NH₄-N+NO₃-N) pool, and 13–23% was in the ¹⁵N-DON pool. Other fates of excreted ¹⁵N may have been incorporation in microbial biomass, chemical or physical protection in non-extractable N forms, or gaseous N losses. Earthworm excretion rates were combined with earthworm biomass measurements to estimate N flux from earthworm populations through excretion. Annual earthworm excretion was estimated at 41.5 kg N ha^–1 in an inorganically-fertilized corn agroecosystem, and was equivalent to 22% of crop N uptake. Our results suggest that the earthworms could contribute significantly to N cycling in corn agroecosystems through excretion processes. Received: 12 April 1999     

Metal soil pollution differentially affects both the behaviour and exposure of <Emphasis Type="Italic">A. caliginosa</Emphasis> and <Emphasis Type="Italic">L. terrestris</Emphasis>: a mesocosm study

The effects on two earthworm species of a gradient of metal contamination in soil collected close to a 50-year-old lead recycling factory were investigated in mesocosms filled with soil sampled at three distances from the factory (10, 30 and 60 m). After 5 weeks of exposure, earthworm litter consumption and weight change were measured. Burrow systems were analysed using X-ray tomography, and water infiltration was measured. No significant differences in earthworm weight or activity were observed between mesocosms filled with soil from 30 and 60 m. In contrast, both earthworm species significantly lost weight and burrowed less in the soil sampled at 10 m. In the cores filled with the soil collected at 10-m distance, Aporrectodea caliginosa avoided the highly contaminated first layer (0–5 cm) and burrowed deeper whereas Lumbricus terrestris burrowed relatively more in this layer. We assume that these different reactions are associated with their ecological types. Epi-anecic earthworms forage litter at the soil surface, whereas endogeic earthworms are geophagous and thus are able to forage deeper. This was further corroborated by the bioaccumulation factors measured for each species: for L. terrestris, BAF values for Pb and Cd only decreased slightly in the 10-m soil correlating with their overall reduced activity. However, BAF values for A. caliginosa were 20-fold lower compared to those observed in soil from 30 and 60 m. These modifications in burrowing behaviour in the 10-m mesocosms resulted in a significant and marked decrease in water infiltration rates but only for L. terrestris.     

Effects of heavy metals on the structure and functioning of detritivore communities in a contaminated floodplain area

The aim of this study was to determine the effects of heavy metal pollution on the structure and functioning of detritivore soil communities that consist of isopods, millipedes and earthworms, in 15 heavily polluted flood plain soils, located in the delta area of the rivers Rhine and Meuse, in the Netherlands. The 15 study sites represent a gradient in Zn, Cu and Cd concentrations. The structural attributes of the detritivore community, which were assessed, were the species richness and densities in the field sites. The functioning of the detritivore community was studied by determining organic matter decomposition using litter bags and feeding activity with the bait-lamina method. Concentrations of Cd, Cu and Zn were measured in soil, pore water and 0.01 M CaCl₂ extracts of the soil, in adult earthworms and plant leaves. Results show that metal pollution is not a dominating factor determining the species richness and densities of the selected detritivore groups, although the biomass of the earthworm Lumbricus rubellus was positively and significantly correlated to Zn concentrations in pore water and 0.01 M CaCl₂ extracts. Litter decomposition was significantly and positively correlated to detritivore biomass and 0.01 M CaCl₂ extractable Cd concentrations in soil and negatively to pH-CaCl₂, although the range of pH values was very small. It can be concluded that in spite of high metal levels in the soil, bioavailable concentrations are too low to result in clear negative effects on the structure and functioning of detritivores in the Biesbosch, the Netherlands.     

Heavy‐metal mobilization and uptake by mycorrhizal and nonmycorrhizal willows (Salix × dasyclados)

Ectomycorrhizal fungi have been shown to affect metal transfer from the soil to the host plant, but the use of these fungi for increased phytoextraction of heavy metals has been scarcely investigated. Therefore, a two‐factorial pot experiment was conducted with Salix × dasyclados and (1) two contaminated soils with different concentrations of NH₄NO₃‐extractable metals and (2) two strains of the ectomycorrhizal fungus Paxillus involutus (one strain originating from a noncontaminated site—Pax1, and another from a contaminated site—Pax2). The inoculation with Pax2 increased the phytoavailability of Cd in the soils. Inoculation with both fungal strains increased the stem and root biomass, but had no effect on metal concentrations in the stems. Decreased Cd and increased Cu concentrations were observed in the roots of inoculated willows. The inoculation with P. involutus increased Cd (up to 22%), Zn (up to 48%), and Cu content in the stems. Decreased Pb content (Cu and Pb content were always <1 mg per plant) occurred in the stems from plants at the soil with the higher concentration of NH₄NO₃‐extractable metals. Contrary to this, in the soil with lower concentrations of NH₄NO₃‐extractable metals, the inoculation had no significant effects on the total uptake of Zn and Cu and even caused decreased Cd (Pax2) and Pb (Pax1) contents in the stems. Strain Pax2 had higher colonization densities, but the plants had lower mycorrhizal dependencies in the contaminated soils than after inoculation with the strain Pax1. Generally, metal extractability in the soils substantially affected the mycorrhizal dependency and heavy‐metal uptake of the willows. We concluded, that the inoculation with P. involutus offers an opportunity to particularly increase the phytoextraction of Zn, but the metal extractability and fungal strain effects have to be tested.     

Heavy metal accumulation by two earthworm species and its relationship to total and DTPA-extractable metals in soils

The concentrations of Zn, Cd, Pb and Cu in earthworm tissues were compared with the total and DTPA-extractable contents of these heavy metals in contaminated soils. Samples were taken from a pasture polluted by waste from a metallurgic industry over 70 y ago. Three individuals of Aporrectodea caliginosa and Lumbricus rubellus and soil samples were collected at six points along a gradient of increasing pollution. Total metal contents of earthworms, soil, and metals extracted by DTPA from the soil were measured. Total heavy metal contents of the soils ranged from 165.7 to 1231.7 mg Zn kg⁻¹, 2.7 to 5.2 mg Cd kg⁻¹, 45.8 to 465.5 mg Pb kg⁻¹ and 30.0 to 107.5 mg Cu kg⁻¹. Their correlations with metals extracted by DTPA were highly significant. Contents of the metals in earthworm tissues were higher in A. caliginosa than in L. rubellus, with values ranging from 556 to 3381 mg Zn kg⁻¹, 11.6 to 102.9 mg Cd kg⁻¹, 1.9 to 182.8 mg Pb kg⁻¹ and 17.9 to 35.9 mg Cu kg⁻¹ in A. caliginosa, and from 667.9 to 2645 mg Zn kg⁻¹, 7.7 to 26.3 mg Cd kg⁻¹, 0.5 to 37.9 mg Pb kg⁻¹ and 16.0 to 37.6 mg Cu kg⁻¹ in L. rubellus, respectively. Correlations between body loads in earthworms with either total or DTPA-extractable contents of soil metals were significant, except for Cd in L. rubellus and Cu in A. caliginosa. Considering its simple analytical procedure, DTPA-extractable fraction may be preferable to total metal content as a predictor of bio-concentrations of heavy metals in earthworms. Biota-to-Soil Accumulation Factor (BSAF) of these four metals are Cd>Zn>Cu>Pb, with range of mean values between: Cd (6.18-17.02), Zn (1.95-7.91), Cu (0.27-0.89) and Pb (0.08-0.38) in A. caliginosa, and Cd (3.64-6.34), Zn (1.5-6.35), Cu (0.29-0.87) and Pb (0.04-0.13) in L. rubellus. The BSAF of Ca, Fe and Mn are Ca>Mn>Fe, with mean values of: Ca (0.46-1.31), Mn (0.041-0.111), Fe (0.017-0.07) in A. caliginosa and Ca (0.98-2.13), Mn (0.14-0.23), Fe (0.019-0.048) in L. rubellus, respectively. Results of principal component analysis showed that the two earthworm species differ in the pattern of metal bioaccumulation which is related to their ecological roles in contaminated soils.     

Enhanced Lead Uptake by an Association of Plant and Earthworm Bioaugmented with Bacteria

Lead (Pb) is recorded as the second most hazardous pollutant of the environment. Previous cases of Pb bioremediation has been reported using single biosystem, but very few reports are available in biological approaches using multi-biosystems to achieve an enhanced bioremoval of Pb. The present study evaluated the capacity of a unique association of Pennisetum purpureum, a hyperaccumulator plant, and Lumbricus terrestris (earthworm) bioaugmented with a Pb-resistant bacterium, obtained from an industrially contaminated site and identified as isolate VITMVCJ1 Klebsiella variicola, to bioremediate Pb. The Pb-resistant gene was amplified in the bacterial isolate VITMVCJ1. The study was conducted for 60 d. Results verified that the bioaugmentation process enhanced 1) root and shoot length of the plants, 2) chlorophyll content of the plants, and 3) biofilm-producing ability of the microbes from the rhizosphere region of the plants. The total phenolic and flavonoid contents were found to be lower in the plants in the bioaugmented setup. The study also observed a reduction in the toxic effects of Pb on earthworm and plant. The earthworm was used to assess the Pb-induced stress syndrome after exposure to sublethal concentrations of Pb in the soil. A reduction in the content of malondialdehyde, a potential biomarker, on exposure to Pb demonstrated the role of the inoculum to alleviate heavy metal-induced stress in earthworms. All three symbionts accumulated Pb; Pb was accumulated mainly in the root of the plant, and poorly in the shoot of the plant and body mass of the earthworm. The bioaugmentation system exhibited stable and excellent uptake of Pb from the contaminated soils. The results of the present study suggest the positive effect of the synergistic association of the plant and earthworm with appropriate microbes for the bioremoval of Pb.     

Nitrification and denitrification activities of zinc-treated soils worked by the earthworm<Emphasis Type="Italic"> Pheretima</Emphasis> sp.

Little information is available on the roles of earthworms in N transformation and biological activities in heavy metal-contaminated soils. The purpose of the present paper is to examine the effects of earthworm on nitrifying and denitrifying enzyme activities of Zn-treated soils. ZnCl₂ solution was added to soils at rates ranging from 0 to 400 mg Zn kg^–1 and was allowed to fractionate and stabilize for 60 days. The Zn-tolerant earthworm Pheretima sp., which had been previously screened, was then introduced into Zn-treated soils and allowed to work for 40 days. Original soils without the addition of Zn and introduction of earthworms but incubated under the same conditions were used as controls. Zn-treated soils without the introduction of earthworms were also used as controls. Results showed that earthworm activity increased both nitrifying and denitrifying enzyme activities of the soils. These beneficial effects of earthworms were particularly evident in the treatments where Zn had an adverse effect. Zn added at a rate 200 mg Zn kg^–1 generally had a stimulating effect on the enzyme activities measured. However, Zn added at a rate >200 mg Zn kg^–1 had an adverse effect on nitrifying and denitrifying enzyme activities, which could be alleviated or even completely eliminated by earthworm activities.     

Use of earthworms Eisenia andrei on the bioremediation of contaminated area in north of Tunisia and microbial soil enzymes as bioindicator of change on heavy metals speciation

Purpose

A better understanding is required of the potential of soil biota in controlling the availability and mobility of heavy metals and ascertaining their toxicity. The objectives of this work are to assess, first, the modification of heavy metal speciation induced by earthworms Eisenia andrei and, second, the consequence of this metal speciation change on soil enzyme activities as an easy bioindicator of stress.

Materials and methods

The experiment was conducted on six sites from Jebel Ressas Mines, which are characterized by a gradient heavy metal contamination (Pb, Zn, and Cd). Earthworms E. andrei were introduced in these six soils for 60 days. We had performed heavy metal speciation both in the presence and absence of worms. Modifications of activities of seven enzymes implicated in C, N, and P biochemical cycles were used as a bioindicator of metal stress. We had used the co-inertia statistical method to evaluate the correlation between change in heavy metal speciation induced by earthworms and the enzyme activities in soils.

Results and discussion

Our results suggested that earthworms modified the heavy metal dynamic and speciation. They decrease the amount of metal associated with the most available fraction, such as exchangeable one, and increase the amount of metal bound to the more stable fraction, like Mn and Fe oxide ones. On the same hand, enzyme activities increased in majority of the soils, following earthworm activity, but this effect is dependent on the amount of soil contamination. Moreover, the co-inertia results denote that change in heavy metal speciation significantly influences the soil enzyme activities in Jebel Ressas soils, especially β-glucosidase, urease, deshydrogenase, and fluorescein diacetate hydrolysis (FDA), and can be considered as bioindicators of metal toxicity and biological quality in the contaminated area.

Conclusions

By reducing the availability of heavy metals, the earthworms are useful in the bioremediation of heavy metal contaminated soils. Soil enzymes β- glucosidase, urease, deshydrogenase, and FDA can be used to assess the changes in metal speciation and can let us, therefore, predict if the soils are bioremediated.

     

Carbon and trace element fluxes in the pore water of an urban soil following greenwaste compost, woody and biochar amendments, inoculated with the earthworm Lumbricus terrestris

The benefits of adding composted organic materials to soils to enhance carbon storage could be countered by the mobilisation of some harmful pollutants commonly found in frequently degraded urban soils. Therefore non-composted materials could be a safer option. In the present study, carbon and trace element fluxes in soil pore water were studied in response to the surface mulch addition and the incorporation into an urban soil of greenwaste compost versus two non-composted amendments; a woody oversize material and biochar following inoculation with the vertical burrowing earthworm Lumbricus terrestris. The aim was to establish (i) to what extent the non-composted amendments impacted on mobility of soluble trace elements in the soil, compared to the composted amendment, and (ii) if/how this was regulated by earthworm activity.Both composted and non-composted amendments enhanced dissolved organic carbon (DOC) in soil pore water to ∼100-300 mg l⁻¹ in the upper depth of the soil profile above which they were applied as a mulch and similarly within the mesocosms in which they were mixed. Dissolved organic carbon, dissolved nitrogen (DTN) and trace metals, especially Cu and Pb, where enhanced to the greatest extent by greenwaste compost, because of strong co-mobilisation of metals by DOC. Biochar enhanced As and Cu mobility in the field profile and, additionally Pb in the mesocosms, with no effect on Cd. The woody, oversize amendment neither greatly increased DOC nor As, Cu, Pb or Zn mobility although, unlike the other amendments, earthworms increased DOC and Cd mobility when soils were amended with this material.This study concludes that non-composted amendments had a lower impact on DOC and thus trace element co-mobility than the composted greenwaste in this urban soil, whilst the general influence of earthworms was to reduce DOC and hence associated trace element mobility. In wider environmental terms the addition of non-composted materials to some urban soils, versus composted greenwaste could reduce the risk of mobilising potentially harmful elements, whilst usefully improving soil quality.