Herman Clijsters

Reclamation of a bare industrial area contaminated by non-ferrous metals: in situ metal immobilization and revegetation.

To reduce the environmental impact of a 135-ha bare industrial area with a highly phytotoxic soil contaminated by non-ferrous metals, a rehabilitation strategy was developed, aimed at the restoration of a vegetation cover. Two different techniques to overcome the high phytotoxicity of the soil were first evaluated on a laboratory scale: reduction of soil phytotoxicity by the addition of a powerful metal immobilizing substance and use of metal-tolerant plants. Since a combination of both approaches proved most promising, this strategy was subsequently utilized in a 3-ha field experiment on the most contaminated location of the industrial area. After soil treatment and sowing of seeds of metal-tolerant grasses, a complete and healthy vegetation cover was quickly established, even at the sites where metal concentrations were extremely high. The reduction of soil phytotoxicity by addition of beringite was immediate and was confirmed 15 months after the treatment. Thirty months after the reclamation activities, the vegetation is still healthy and vegetative and generative plant propagation is abundant.

Induction of enzyme capacity in plants as a result of heavy metal toxicity: Dose-response relations in Phaseolus vulgaris L., treated with zinc and cadmium

Toxic doses of zinc and cadmium inhibit shoot growth but increase the capacity of several leaf enzymes in dwarf beans (Phaseolus vulgaris L.). Both effects were studied as a function of the metal concentration applied to the plant. There was a linear relationship between the metal content of the primary leaf and the nutrient solution. When leaf metal content exceeded a toxic threshold value, shoot growth became inhibited and an increase in capacity of the following enzymes was measured in the leaf: glucose-6-phosphate dehydrogenase, glutamate dehydrogenase, isocitrate dehydrogenase, malic enzyme, glutamate-oxaloacetate transminase, peroxidase. The threshold values were similar for growth inhibition as well as for enzyme capacity induction. Both effects were strongly correlated to each other, especially under conditions of toxic zinc treatment. Measurement of enzyme capacity might therefore provide a useful criterion for the evaluation of the phytotoxicity of soils, contaminated by zinc and/or cadmium.

Antioxidative responses of wheat treated with realistic concentration of cadmium

Abstract Two cvs. of wheat differently sensitive to many stress factors (cv. Ofanto less sensitive than cv. Adamello) were grown in a controlled environment with cadmium near threshold concentrations supplying the metal at equal-effect concentrations. Cd excess determined in both cvs. a reduction in water and turgor potential but a maintenance of relative water content. Cv Ofanto showed a higher capacity of Cd exclusion from roots but a higher translocation to shoots in comparison with cv. Adamello. Notwithstanding the higher metal concentration in leaves of cv. Ofanto, K + leakage was more pronounced in Adamello suggesting that mechanisms of Cd detoxification and tolerance such as vacuolar compartmentalisation were activated in the first one. In Adamello plants, ethylene rose at the lowest metal concentration and the activation in roots of the antioxidative enzymes catalase, ascorbate peroxidase and guaiacol peroxidase came into play whereas in Ofanto ethylene and catalase did not change. Following cadmium treatment, superoxide dismutase activity was reduced or remained at the control value in roots and in leaves. For both cultivars ascorbate peroxidase, syringaldazine peroxidase and guaiacol peroxidase activities were always higher in roots than in leaves. These activities were induced by Cd in Ofanto leaves, whereas in Adamello leaves they remained at control levels or increased somewhat at the highest metal concentration. Cadmium changed the peroxidase isozyme pattern in both cultivars. Cv. Ofanto showed, as for other stress such as drought, salinity, nickel and copper, a co-tolerance towards Cd. Analogies in the response to other metals such as copper could be found in activation of catalase at the lower metal concentration in cv. Adamello and in the induction of ascorbate peroxidase in leaves of cv. Ofanto.

Evaluation of metal mobility, plant availability and immobilization by chemical agents in a limed-silty soil.

Metal-contaminated soils in the vicinity of industrial sites become of ever-increasing concern. Diagnostic criteria and ecological technologies for soil remediation should be calibrated for various soil conditions; actually, our knowledge of calcareous soil is poor. Silty soils near smelters at Evin (Pas de Calais, France) have been contaminated by non-ferrous metal fallout and regularly limed using foams. Therefore, the mobility, bioavailability, and potential phytotoxicity of Cd, Pb and Zn, were investigated using single soil extractions (i.e. water, 0.1 n Ca(NO(3))(2), and EDTA pH 7), and vegetation experiments, in parallel with a biological test based on (iso)-enzymes in leaves and roots, before and following soil treatment with chemical agents, i.e. Thomas basic slags (TBS), hydrous manganese oxide (HMO), steel shots (ST) and beringite. No visible toxicity symptoms developed on the above-ground parts of ryegrass, tobacco and bean plants grown in potted soil under controlled environmental conditions. Cd, Zn and Pb uptake resulted in high concentrations in the above-ground plant parts, but the enzyme capacities in leaves and roots, and the peroxidase pattern indicated that these metal concentrations were not phytotoxic for beans as test plants. The addition of chemical agents to the soil did not increase biomass production, but treatment with either HMO, ST or beringite markedly decreased the mobility of Cd, Zn and Pb. These agents were proven to be effective in mitigating the Cd uptake by plants. HMO and ST decreased either Pb or Zn uptake by ryegrass. TBS was effective in lowering Pb uptake by the same species. Beringite decreased Cd uptake by beans. If fallout could be restricted, the metal content of food crops in this area should be lowered by soil treatment. However, the differences in Cd uptake between plant species were not suppressed, regardless of the type of agents applied to the soil.

Peroxidases in roots and primary leaves of Phaseolus vulgaris Copper and Zinc Phytotoxicity: a comparison

Summary The contribution of peroxidases was studied in roots and primary leaves of bean seedlings as a part of the defence mechanism against copper and zinc imposed oxidative stress. Based on the metal content of the different plant organs, copper was mainly «stored» in the roots, whereas zinc was readily transported to the upper ground parts. In the roots copper had an increasing effect on the capacities of guaiacol and syringaldazine peroxidases. A possible role for lignification in establishing an apoplastic transport barrier in roots is hypothesised. Zinc, on the other hand, had a limited impact on the root metabolism. Future studies should focus on other protection mechanisms in this plant organ. In the primary leaves, both elements had a pronounced (increasing) effect on the peroxidase activities. The data obtained in this study show that peroxidases contribute to the antioxidative defence of the primary leaves of 10 days old bean seedlings after application of both metals and that an interorgan signal transduction pathway is involved. It is pointed out that additional experimental approaches are necessary before the protection mechanism against different metals can be fully compared.

Selected bioavailability assays to test the efficacy of amendment-induced immobilization of lead in soils

Lead immobilization in 10 soils contaminated with Pb from different origin was examined using lime (CaCO3), a mix of cyclonic ash and steelshots (CA+ST), and a North Carolina phosphate rock. The immobilization efficacy of the three amendments was evaluated using single (CaCl2solution) and sequential (BCR method) chemical extractions in tandem with a microbiological Pb biosensor (BIOMET), a Pb phytotoxicity test, Pb plant uptake, and a Physiologically Based Extraction Test (PBET) mimicking Pb bioavailability in the human gastro-intestinal tract. The results demonstrated the necessity of using a diverse suite of bioavailability methodology when in situ metal immobilization is assessed. Sequential (BCR) extractions and PBET analysis indicated that PR was the most effective amendment. PR however, proved ineffective in totally preventing Pb phytotoxicity and Pb uptake on all soils tested. On the contrary, CA+ST and lime decreased BIOMET Pb, Pb phytotoxicity, and Pb uptake to a far greater extent than did PR. BIOMET detectable Pb and Pb uptake, however, were strongly related to Pb in soluble or exchangeable soil fractions (i.e., CaCl2 extractable). By combining these fractions with the acid-extractable Pb, accomplished by using acetic acid extractant, the recently developed BCR sequential extraction scheme appeared to have lost some valuable information on judging Pb bioavailability. The data show that different amendments do not behave consistently across different soils with different sources of contamination. Different indices for measuring Pb bioavailability are also not necessarily consistent within particular soil and amendment combinations.

The redox status of plant cells (AsA and GSH) is sensitive to zinc imposed oxidative stress in roots and primary leaves of Phaseolus vulgaris

Abstract The ascorbate-glutathione cycle has been shown to be of great importance in multiple stress reactions. In the present study, alterations in the antioxidant pools combined with the evolution of the capacities of enzymes involved in defence against oxidative stress was investigated in roots and primary leaves of Phaseolus vulgaris L. after a 50-μM Zn application to roots. A difference has to be made between both plant organs. In roots, an overall oxidation of ascorbate occurred immediately after the start of the zinc treatment. This was accompanied by a decrease in ascorbate peroxidase (APOD; EC 1.11.1.11) and glutathione reductase (GR; EC 1.6.4.2) capacities, the latter resulting in an increase of glutathione disulphide (GSSG). In addition, in the primary leaves ascorbate was strongly affected by zinc toxicity and an immediate increase in the total amount could be observed. Whereas enhancement of the capacities constituting the ascorbate-glutathione pathway was observed, it only limited the effects of oxidative stress in primary leaves. At the end of the experiment, a strong increase in ascorbate oxidation was noticed suggesting that an excess of zinc can cause oxidative stress.

Physiological Responses to Heavy Metals in Higher Plants; Defence against Oxidative Stress

Depending on the physiological process investigated heavy metal phytotoxicity can be either inhibitory or stimulatory. Photosynthesis and its partial light and dark reactions are inhibited; the activity of various enzymes, located in several cell compartiments, is increased. These enzymes are mostly induced since metals affect the transcription activity. They appear to be related to the plant defence against oxidative stress caused by metal phytotoxicity. Careful examination of the time course of this induction reveals differences in response between the metals applied

The Effect of Recombinant Heavy Metal-Resistant Endophytic Bacteria on Heavy Metal Uptake by Their Host Plant

ABSTRACT The ncc-nre nickel resistance system of Ralstonia metallidurans 31A was efficiently expressed in Burkholderia cepacia L.S.2.4 and Herbaspirillum seropedicae LMG2284. The heterologous expression of ncc-nre encoded nickel resistance was accompanied by nickel removal from the culture medium. B. cepacia L.S.2.4:: ncc-nre and H. seropedicae LMG2284::ncc-nre were able to remove 35 and 15% nickel, respectively. The capacity to remove nickel through sequestration or bio-precipitation processes and consequently lowering the free nickel concentration could offer interesting benefits for these endophytic bacteria and their host plants. Once inoculated in their host plant, they could possibly alter the nickel speciation and therefore decrease the free ions and thus toxic concentration for the plant metabolism. Lupinus luteus L, when grown on a nickel enriched substrate and inoculated with B. cepacia L.S.2.4::ncc-nre, showed a significant increase (30%) of nickel concentration in the roots, whereas the nickel concentration in the shoots remained comparable with that of the control plants. The inoculation of Lolium perenne (cv Atlas) with the nickel resistance derivative of H. seropedicae LMG2284::ncc-nre resulted in a significant decrease of the nickel concentration in the roots (11%) as well as in the shoots (14%). As this phenomenon was also observed in the Lolium perenne plants inoculated with the wild-type strain LMG2284, the nickel resistance characteristics probably are not responsible for the altered nickel uptake observed.

Inhibition of Photosynthesis in Phaseolus vulgaris by Treatment with Toxic Concentration of Zinc: Effect on Ribulose-1,5-bisphosphate Carboxylase/ Oxygenase

Summary Dwarf beans (Phaseolus vulgaris L.cv Limburgse Vroege) were cultivated on a medium containing a toxic non-lethal Zn2+ concentration.The ribulose-1,5-bisphosphate carboxylase/oxygenase capacity of these Zn2+ treated plants was compared with the capacity of control plants, grown under standard nutrient conditions.The carboxylase capacity, measured on crude leaf extracts, was significantly inhibited by Zn2+ treatment.As the oxygenase capacity was not affected, the carboxylase/oxygenase capacity ratio decreased significantly.This lower ratio was restored up to the control value by in vitro inactivation of the enzyme, followed by reactivation in the presence of Mg2+.This suggests a partial substitution in vivo of Zn2+ for Mg2+ in the ternary enzyme-CO2metal2+ complex.Further evidence for this substitution was provided by the extremely low Mg/Zn concentration ratio found in the chloroplast, as well as by the higher KM(CO2) of the enzyme.