Glycogen-lead relationship in the earthworm Dendrobaena rubida from a heavy metal site

Summary Control individuals contained no lead in the chloragocytes but high -glycogen rosette reserves. Starvation of contaminated earthworms for 4d caused a lead loss and the chloragocytes possessed fewer debris vesicles than those of unstarved worms, suggesting that the debris vesicles may be the route for at least some of the lead loss. No glycogen deposits were observed in the chloragocytes of starved or unstarved earthworms from contaminated soil. Maintenance of contaminated earthworms in potting compost caused lead losses similar to those sustained by starvation, but the chloragocyte cytoplasm possessed -glycogen reserves. Specimens maintained in lead-spiked potting compost showed lead levels similar to those of earthworms taken directly from contaminated soil. No -glycogen accumulations were observed under this enriched regime.Although the possible interference of lead in carbohydrate metabolism is discussed, the results do not wholly support metabolic inhibition by lead. It is hypothesised that lead sequestration is energy-demanding and that in the absence of an energy-rich diet glycogen stores fail to accumulate. In the presence of an organic-rich medium, elevated lead levels preclude glycogen formation, because of the high sequestration-demand, but at lower lead levels -glycogen deposits occur if a high organic diet is available.     

Glycogen-lead relationship in the earthworm Dendrobaena rubida from a heavy metal site

Control individuals contained no lead in the chloragocytes but high alpha-glycogen rosette reserves. Starvation of contaminated earthworms for 4d caused a lead loss and the chlorgocytes possessed fewer debris vesicles than those of unstarved worms, suggesting that the debris vesicles may be the route for at least some of the lead loss. No glycogen deposits were observed in the chloragocytes of starved or unstarved earthworms from contaminated soil. Maintenance of contaminated earthworms in potting compost caused lead losses similar to those sustained by starvation, but the chloragocyte cytoplasm possessed beta-glycogen reserves. Specimens maintained in lead-spiked potting compost showed lead levels similar to those of earthworms taken directly from contaminated soil. No beta-glycogen accumulations were observed under this enriched regime. Although the possible interference of lead in carbohydrate metabolism is discussed, the results do not wholly support metabolic inhibition by lead. It is hypothesised that lead sequestration is energy-demanding and that in the absence of an energy-rich diet glycogen stores fail to accumulate. In the presence of an organic-rich medium, elevated lead levels preclude glycogen formation, because of the high sequestration-demand, but at lower lead levels beta-glycogen deposits occur if a high organic diet is available.     

Fate and effects of cadmium in an experimental marine ecosystem

Two recirculated marine seawater systems (capacities: 150 and 300 l) were used for the study on cadmium accumulation of biological filter sludge, by the musselMytilus edulis and the plaicePleuronectes platessa for a period of up to 300 days. In the contaminated system Cd concentrations varied from 2.5 to 7.0 g/l with a mean of 4.6±1.15 g/l (control: 1.04±0.46 g/l). Accumulation of Cd by the filter sludge from the control system as well as from the contaminated system was significant. Mean concentrations of control-and contaminated sludge over the experimental period were 2.01±0.86 ng Cd/mg dry wt and 9.98±3.55 ng Cd/mg dry wt respectively. Accumulation of Cd byM. edulis both in the control and in the contaminated system was considerable. After 163 days of exposure the whole body burden of mussels rose from 0.3 ng Cd/mg dry wt to around 10 ng Cd/mg dry wt in the controls and to 70 ng Cd/mg dry wt in the contaminated systems. Accumulation of Cd byP. platessa was analysed in backbone, fins, gills, liver, muscle (fillet), otolith and skin (dorsal and ventral) over a period of 280 days. Elevated Cd contents of livers from control specimens were noticed after 200 days and reached 1.75 ng Cd/mg dry wt in liver. There was considerable accumulation in the liver (maximum values: 3.0 ng Cd/mg dry wt) and gills (6.0 ng Cd/mg dry wt) of specimens from the contaminated system, Cd contents of plaice exposed for identical periods of time were very variable. Growth of plaice living in the contaminated system was at times significantly slower than that of the control group. Of the three biological objects tested-filter sludge,M. edulis, P. platessa—the mussel seemed to be the most sensitive indicator of elevated Cd-concentrations, while the reaction of the plaice was slow and less distinct.     

Degradation of potassium rock by earthworms and responses of bacterial communities in its gut and surrounding substrates after being fed with mineral

Background

Earthworms are an ecosystem''s engineers, contributing to a wide range of nutrient cycling and geochemical processes in the ecosystem. Their activities can increase rates of silicate mineral weathering. Their intestinal microbes usually are thought to be one of the key drivers of mineral degradation mediated by earthworms,but the diversities of the intestinal microorganisms which were relevant with mineral weathering are unclear.

Methodology/Principal Findings

In this report, we show earthworms'' effect on silicate mineral weathering and the responses of bacterial communities in their gut and surrounding substrates after being fed with potassium-bearing rock powder (PBRP). Determination of water-soluble and HNO₃-extractable elements indicated some elements such as Al, Fe and Ca were significantly released from mineral upon the digestion of earthworms. The microbial communities in earthworms'' gut and the surrounding substrates were investigated by amplified ribosomal DNA restriction analysis (ARDRA) and the results showed a higher bacterial diversity in the guts of the earthworms fed with PBRP and the PBRP after being fed to earthworms. UPGMA dendrogram with unweighted UniFrac analysis, considering only taxa that are present, revealed that earthworms'' gut and their surrounding substrate shared similar microbiota. UPGMA dendrogram with weighted UniFrac, considering the relative abundance of microbial lineages, showed the two samples from surrounding substrate and the two samples from earthworms'' gut had similarity in microbial community, respectively.

Conclusions/Significance

Our results indicated earthworms can accelerate degradation of silicate mineral. Earthworms play an important role in ecosystem processe since they not only have some positive effects on soil structure, but also promote nutrient cycling of ecosystem by enhancing the weathering of minerals.     

The accumulation and intracellular compartmentation of cadmium,lead, zinc and calcium in two earthworm species (Dendrobaena rubida and Lumbricus rubellus) living in highly contaminated soil

Summary The earthworm Lumbricus rubellus contained more Ca and Zn, and less Pb and Cd, than Dendrobaena rubida living in the same contaminated disused-mine soil. Differences in the kinetics of Ca turnover may account for some of the inter-specific differences in heavy metal burdens, although the calciferous glands do not seem to be directly involved in heavy metal excretion. A comparison of the present findings with published data indicated that the concentration of soil Ca and the bioavailability of heavy metals, both factors being allied to soil pH, are important exogenous determinants of heavy metal accumulation by different earthworm populations. Electron microprobe X-ray analysis of air-dried smears of chloragogenous tissue showed that the metals were fairly specifically compartmentalized into two distinct organelles in both worms: Ca, Pb and Zn were found (associated with P) in the chloragosomes; Cd was found (with S and probably in stoichiometric association) in a more electron-lucent vesicular component, designated the cadmosome, but which may be identical with the debris vesicles which are characteristic inclusions in conventionally-fixed chloragocytes. The in vivo incorporation of Pb by the chloragosomes of D. rubida was accompanied by the loss of Ca, Zn and P.     

蚯蚓作用下污泥重金属形态变化及其与化学生物学性质变化的关系

城市污泥处理是一项世界性难题,污泥农业利用是其最简单有效的资源化利用方式之一,但污泥中较高的重金属含量限制了其实际推广应用,利用蚯蚓-超富集植物联合修复污泥重金属的方法已引起国内外研究者的关注。以新鲜城市脱水污泥为研究对象,接种赤子爱胜蚓(Eisenia fetida)进行室内培养试验,系统研究蚯蚓作用下污泥重金属形态的变化,及其与污泥氧化还原条件、化学和微生物性质变化的关系,以期为蚯蚓-超富集植物联合修复技术在污泥重金属处理中的应用提供理论依据。结果表明,试验前期蚯蚓在污泥中能正常生长和存活,前20 d总生物量增加了52%。蚯蚓可以显著促进污泥中的Cu、Zn、Cd、Ni等重金属从残渣态和铁锰态等稳定形态向交换态和水溶态等有效形态转化。还可以显著降低污泥中还原性物质的含量,减缓p H值下降速度,降低总有机碳含量,促进铵态氮向硝态氮转化,减少污泥微生物的数量并增加其种群活性。蚯蚓作用下,污泥中重金属的活化程度与还原性物质的含量呈显著负相关,而与微生物种群的活性呈显著正相关(P0.05)。综上所述,蚯蚓可以促进污泥重金属的活化,并改善污泥的肥力条件,为修复植物在污泥中的正常生长和对重金属离子的快速吸收提供有利条件。     

Arbuscular mycorrhizal contribution to heavy metal uptake by maize (Zea mays L.) in pot culture with contaminated soil

In two pot-culture experiments with maize in a silty loam (P2 soil) contaminated by atmospheric deposition from a metal smelter, root colonization with indigenous or introduced arbuscular mycorrhizal (AM) fungi and their influence on plant metal uptake (Cd, Zn, Cu, Pb, Mn) were investigated. Soil was -irradiated for the nonmycorrhizal control. In experiment 1, nonirradiated soil provided the mycorrhizal treatment, whereas in experiment 2 the irradiated soil was inoculated with spores of a fungal culture from P2 soil or a laboratory reference culture, Glomus mosseae. Light intensity was considerably higher in experiment 2 and resulted in a fourfold higher shoot and tenfold higher root biomass. Under the conditions of experiment 1, biomass was significantly higher and Cd, Cu, Zn and Mn concentrations significantly lower in the mycorrhizal plants than in the nonmycorrhizal plants, suggesting a protection against metal toxicity. In contrast, in experiment 2, biomass did not differ between treatments and only Cu root concentration was decreased with G. mosseae-inoculated plants, whereas Cu shoot concentration was significantly increased with the indigenous P2 fungal culture. The latter achieved a significantly higher root colonization than G. mosseae (31.7 and 19.1%, respectively) suggesting its higher metal tolerance. Zn shoot concentration was higher in both mycorrhizal treatments and Pb concentrations, particularly in the roots, also tended to increase with mycorrhizal colonization. Cd concentrations were not altered between treatments. Cu and Zn, but not Pb and Cd root-shoot translocation increased with mycorrhizal colonization. The results show that the influence of AM on plant metal uptake depends on plant growth conditions, on the fungal partner and on the metal, and cannot be generalized. It is suggested that metal-tolerant mycorrhizal inoculants might be considered for soil reclamation, since under adverse conditions AM may be more important for plant metal resistance. Under the optimized conditions of normal agricultural practice, however, AM colonization even may increase plant metal absorption from polluted soils.     

Effects of lead on bioaccumulation patterns and the ecophysiological response inMonochoria korsakowi

Phytoextraction is a feasible method for eliminating heavy metals from contaminated soil. However, only a few plant species can hyperaccumulate toxic metals. We examined bioaccumulation patterns and the ecophysiological responses (i.e., photosynthetic pigments, total antioxidative capacity, and variations in antioxidant enzymes) in plants ofMonochoria korsakowi that were exposed to various concentrations of lead. Contents increased significantly when more lead was applied, with most of this metal being found in the roots. Biomass for each organ type decreased with higher lead concentrations. Chlorophyll a contents declined in proportion to lead amounts, whereas those of chlorophyllb were not significantly diminished. Carotenoid contents were not influenced by such treatment. Total antioxidant capacity in the leaf blades of treated plants increased greatly with higher lead concentrations. We conclude thatM. korsakowi could be used as a potential phytoremediator of heavy metals in contaminated soils.     

A field test of a sediment bioassay with the oligochaete worm Tubifex tubifex (Müller, 1774)

A sediment toxicity test using reproduction in the tubificid oligochaete Tubifex tubifex was tested in 11 geographic locations in the Laurentian Great Lakes where sediments have previously been documented as being contaminated. Samples were taken from both potentially contaminated and clean sites in each of the 11 areas. Using test endpoints such as survival and growth, results from 72 samples were classified using cluster and ordination techniques. Six response groups were identified and four groups of toxic sites determined. Toxic effects ranged from mortality of adult worms to reduced reproduction. Using correlation and discriminant analysis there was good evidence that high concentrations of metals such as copper and lead were responsible for the observed effects at some sites.     

Isolation,purification and partial characterization of chloragocytes from the earthworm species Lumbricus terrestris

Chloragocytes were isolated from the earthworm species Lumbricus terrestris. After mechanical dissociation and sedimentation through Percoll, a highly purified fraction of viable chloragocytes was obtained. The isolated chloragocytes accumulated the vital dye neutral red and reduced the tetrazolium dye MTT, thereby indicating cellular integrity. Time of flight flow cytometric analyses revealed a main population of large and highly granulated cells in the 30-33 m size range. Hydrolase measurements showed that -D-N-acetyl-glucosaminidase and acid phosphatase exhibited the highest activities (146.6 and 24.9 mU/mg of protein, respectively), possibly indicating a major role for these 2 hydrolases in the physiological function of chloragocytes. In contrast, other acid hydrolases such as -D-galactosidase and -D-glucuronidase had specific activities of respectively 26 and 182 times lower than the glucosaminidase. The specific activity of the membrane-bound alkaline phosphatase was comparable to that of its acid counterpart (18.9 vs. 24.9 mU/mg of protein, respectively) and this level of activity may show an important trans-membrane activity in chloragocytes. The cytoplasmic and mitochondrial enzyme isocitrate dehydrogenase had a level of activity comparable to that of the exclusively cytoplasmic enzyme lactate dehydrogenase (6.6 vs. 8.1 mIU/mg of protein, respectively). When L. terrestris chloragocyte homogenates were separated on Percoll, results showed that hydrolases and dehydrogenases were mainly associated with the lighter materials that remained above the Percoll layer. Nonetheless, the detection of significant proportions (15-25%) of the total recovered activity of acid phosphatase and-galactosidase in the enriched chloragosome fraction supports the notion that some chloragosomes may be 'lysosome-like' organelles.     

Influence of <Emphasis Type="Italic">Lumbricus terrestris</Emphasis> earthworms on the structure of the yeast community of forest litter

The taxonomic structure of yeast communities was studied in forest litter and soil, as well as in substrates transformed by the activity of Lumbricus terrestris earthworms (leaves in heaps, the gut contents, and coproliths). The activity of L. terrestris has a weak effect on the total yeast abundance but results in substantial changes in the community taxonomic composition. The share of ascomycetous yeasts is significantly higher in the substrates associated with the activity of earthworms. The teleomorphic ascomycetes Williopsis saturnus were isolated from the gut contents. The effect of earthworms on the composition of the yeast community in the process of forest litter destruction is more pronounced than seasonal changes.     

The performance of vetivers (Chrysopogon zizaniodes and Chrysopogon nemoralis) on heavy metals phytoremediation: laboratory investigation

Abstract

Phytoremediation with vetiver was investigated in relation to heavy metal contaminated soil in Thailand. The work compared the performance of two species of vetiver named Songkhla 3 (Chrysopogon zizaniodes) and Prachuap Khiri Khan (Chrysopogon nemoralis) in absorbing lead, zinc, and cadmium in contaminated soils. Toxicity Characteristic Leaching Procedure (TCLP), and Allium tests were conducted to determine toxicity of treated soil. Ethylenediaminetetraacetic acid (EDTA) was also used to increase heavy metals concentration in solution in soil, which led to an increase in translocation and bioaccumulation factors. In general, results showed that concentration of heavy metals decreased in soil and increased in both the shoots and roots of vetivers during a 4-month treatment period. TCLP results indicated that the concentration of zinc and cadmium in contaminated soil was reduced over treatment time, and significantly increased after EDTA was applied. To confirm vetiver performance in phytoremediation, Allium testing showed that remained heavy metals in treated soils had no effect on nucleus aberration. Songkhla 3 and Prachuap Khiri Khan showed similar trends in their ability to remediate lead, zinc, and cadmium from contaminated soil. Both species could accumulate higher concentrations of heavy metals in their shoots and roots over time, and with EDTA application.     

A comparative study of gut microbiota profiles of earthworms fed in three different substrates

The gut microbiome of earthworms has a complex interdependence with the host. When the soil minerals pass through earthworm’s gut, they may affect the gut microbiota. To gain insight into the response of gut microbiota to the passed minerals, we fed earthworm (Eisenia fetida) on nutrient-poor soil and ore powder, and used high throughput sequencing to characterize the earthworm intestinal microbial community to find evidence for a core bacterial community of the E. fetida. The results showed that earthworms’ gut maintained a core microbiome that appeared in all samples. These core microbiota may play a significant role in a species’ environmental interactions. The composition of intestinal microbiomes varied with substrates. The earthworm guts from two nutrient-poor substrates had similar microbial communities and they were different from nutrient-rich substrate. Proteobacteria and Bacteroidetes were more abundant in the gut of earthworms kept on a nutrient-poor substrate such as ore powder or mineral soil than in the gut of earthworms kept in organic-rich compost soil; some of these microorganisms may help earthworms survive in nutrient-poor substrates.     

Subcellular localization of polluting metals in roadside earthworms exposed to traffic exhaust gases.

The aim was to make a subcellular localization of metals in tissue from lumbricid earthworms exposed to environmental pollution. Scanning transmission electron microscopy in combination with energy dispersive X-ray microanalysis of tissue fixed in glutaraldehyde only, and with no electron staining, was used. Zinc was registered in the metachromatic mucous granules of the epidermal cells, and zinc, iron and lead in the chloragosomes of the chloragocytes suggesting that metal may be excreted together with the slime or stored in chloragosomes. Relatively few metal nuclear inclusions were encountered probably due to the fact that some metal leaks out during the preparation process. A comparison is made with a chemical analysis of cellular fractions (Talberg, 1977).     

Effects of heavy metals on growth and ultrastructure ofChara vulgaris

Summary The toxicity of some heavy metals to the common macrophytic freshwater algaChara vulgaris was studied under laboratory conditions. For experiments, apical tips of algae containing two internodes were cultivated for fourteen days in the presence of various concentrations of cadmium, mercury or lead (as triethyl lead or lead nitrate). Fifty percent growth inhibition occurred with concentrations of 8.5×10^–8 M (9.5 ppb) cadmium, 7.5×10^–7M (150ppb) mercury, 1.6×10^–6 M (330ppb) organic lead or 4× 10^–5 M (8000 ppb) inorganic lead. Sublethal concentrations of these metals caused alterations in the fine structure of internodal cells which turned out to be at least partly metal-specific or in the case of lead, the effects depended on whether the lead was ionic or organically bound. Cadmium and inorganic lead induced disorders of cell wall microfibrils which resulted in local wall protuberances. Mercury affected the chloroplasts which mostly showed considerably increased grana stacks. In addition, mercury caused a dilation of the endoplasmic reticulum and of the mitochondrial tubuli. Organic lead damaged the membrane system of chloroplasts; sheet- or tubule-like thylakoids were disarranged and showed whorl-like structures. At higher concentrations of organic lead, tubular invaginations of the plasmalemma (charasomes) disappeared. The fine structure of nuclei was not altered by any of the metals.     

Algae sequester heavy metals via synthesis of phytochelatin complexes

A Cd-binding complex was isolated from Chlorella fusca and has been shown to be composed of phytochelating peptides, (-Glu-Cys) _n -Gly, n=2–5. Members of six of the ten classes of Phycophyta revealed phytochelatin synthesis after exposure to cadmium ions. Phytochelatin was also induced by ions of lead, zinc, silver, copper and mercury. These experiments uneqiovocally demonstrated that algae sequester heavy metals by an identical mechanism as higher plants, namely via complexation to phytochelatins.     

Effects of heavy metal stress on free amino acids in the haemolymph and proteins in haemolymph and total body tissue of Lymantria dispar larvae parasitized by Glyptapanteles liparidis

Heavy metal contamination of the forest pest insect Lymantria dispar (L.) (Lepidoptera; Lymantriidae), the gypsy moth, can alter its haemolymph composition, as has already been shown for carbohydrates and lipids in recent studies. L. dispar larvae are frequently parasitized by Glyptapanteles liparidis (Bouché) (Hymenoptera; Braconidae) larvae, which can—to some extent—regulate the population size of the pest insect. The parasitoids feed on the haemolymph of L. dispar larvae; hence, a different haemolymph composition of the host alters the trophic situation of the parasitoids. The aim of the present study was to investigate whether metal contamination also affects the concentrations of free amino acids in L. dispar haemolymph, and protein concentrations in their haemolymph and tissue. L. dispar larvae were parasitized on the first day of the second instar and then reared on diets contaminated with Cd, Pb, Cu or Zn at two concentrations each. Haemolymph and total body tissue of the larvae (fourth instar/third day) were analyzed. The concentrations of the free amino acids were elevated in five out of the eight contamination groups (Cd6, Pb4, Cu6, Cu10, Zn60), whereas haemolymph protein concentrations were significantly reduced in all contaminated individuals. The haemolymph protein concentration was 18 mg/ml in the control group and decreased to less than 10 mg/ml due to cadmium and zinc contamination at both concentrations and in the low copper contamination group. In contrast, total body proteins (136 g/mg dry weight in the control group) were elevated due to heavy metal stress. Analyses of haemolymph protein concentrations during the fourth instar demonstrated an increase of the proteins from day one to day four (followed by a decrease on the fifth day) in the control group and the cadmium contamination group. A steady increase of proteins from the first to the fifth day in the copper and zinc contaminated larvae indicated a retarded development in these groups. Thus, the present study along with other recent studies demonstrated, that heavy metal stress changes the concentrations of all main haemolymph compounds of L. dispar larvae.     

Development of a technology for commercial phytoextraction of nickel: economic and technical considerations

In recent R&D work, we have made progress in developing a commercial technology using hyperaccumulator plant species to phytoextract nickel (Ni) from contaminated and/or Ni-rich soils. An on-going program is being carried out to develop a genetically improved phytoextraction plant that combines favorable agronomic and Ni accumulation characteristics. Genetically diverse Ni hyperaccumulator species and ecotypes of Alyssum were collected and then evaluated in both greenhouse and field using serpentine and Ni-refinery contaminated soils. Large genetic variation was found in those studies. Mean shoot Ni concentrations in field-grown plants ranged from 4200 to 20400 mg kg^–1. We have been studying several soil management practices that may affect the efficiency of Ni phytoextraction. Soil pH is an important factor affecting absorption of metals by plants. An unexpected result of both greenhouse and field experiments was that Ni uptake by two Alyssum species was reduced at lower soil pH and increased at higher soil pH. At higher pH, plant yield was improved also. In soil fertility management studies, we found that N application significantly increased plant biomass, but did not affect plant shoot Ni concentration. These findings indicate that soil management will be important for commercial phytoextraction. A number of field trials have been carried out to study planting methods, population density, weed control practices, harvest schedule and methods, pollination control, and seed processing. Such crop management studies have improved phytoextraction efficiency and provide a tool for farmers to conduct commercial production. We have done some work to develop efficient and cost-effective methods of Ni recovery. Recovery of energy by biomass burning or pyrolysis could help make phytoextraction more cost-effective. The progress made in our recent studies will enable us to apply this technology commercially in the near future.     

Seasonal variation in the abundance, biomass and biodiversity of earthworms in soils contaminated with metal emissions from a primary smelting works

1. Earthworms (Annelida: Oligochaeta) were sampled on four occasions (spring, summer, autumn and winter) at 14 sites along two transects from a primary lead/ zinc/cadmium smelting works at Avonmouth, UK.
2. Total abundance and biomass of earthworms decreased with proximity to the smelter. No worms were collected from the two sites closest to the factory (<0·6 km) and catches were significantly lower than controls at a further five sites (<3 km).
3. Seasonal composition of sampled communities differed only for summer with lower numbers of individuals and species collected at all sites. Reduced catches in the summer sample is a response to drought.
4. Species richness was lowest at sites close to the factory. For example, worms such as Aporrectodea caliginosa (Savigny) and Allolobophora chlorotica (Savigny) that were dominant at relatively clean sites further from the smelter are absent from the most contaminated soils.
5. Reduced species richness resulted in lower Shannon–Weiner diversity and higher Berger–Parker dominance. Multivariate cluster analysis for spring, summer and winter indicated that sites could be split into three groups based upon relative species composition. In autumn, two clusters were identified.
6. The absence of sensitive species from sites close to the smelting works supports the inclusion of earthworms as a key group in a terrestrial prediction and classification scheme for quantifying the effects of pollutants on soil biodiversity. However, sampling should be carried out in spring or autumn to obtain an accurate picture of community structure.