Aided phytostabilisation of As- and Cu-contaminated soils using white lupin and combined iron and organic amendments.

An aided phytostabilisation strategy consisting of several composite amendments of iron sulphate and organic materials combined with Lupinus albus L. (white lupin) was evaluated for remediation of an As- and Cu-contaminated soil. Iron sulphate was combined with lime, paper mill sludge (PS), olive mill waste compost (OMWC) or holm oak biochar (BC) and applied to a slightly acidic soil with high concentration of As (∼2200 mg kg-1) and Cu (∼150 mg kg-1). White lupin was grown for 48 days in pots containing amended and non-amended soils and the effect of soil treatments on soluble and extractable trace elements, soil fertility and plant growth and composition was evaluated. The addition of the amendments raised soil pH and reduced soluble As (50-93%) and extractable As and Cu (50-89%). Despite the reduction of As- and Cu-extractable fractions, plant As and Cu uptake was not greatly affected by the amendments. Variations in soil pH and P-Olsen seemed to have influenced As dynamics in the treated soils, although they did not provoke its mobilisation with respect to the non-amended soil. Our results suggest that the freshly formed iron oxides resulting from addition of iron sulphate controlled As dynamics in the treated soils, avoiding its mobilisation due to application of organic materials. The combination of iron sulphate with OMWC and BC is shown as appropriate for aided phytostabilisation of metal(loid)s contaminated soils, as it improved soil fertility and plant nutrition while reduced As and Cu mobility.

Copper microlocalisation and changes in leaf morphology, chloroplast ultrastructure and antioxidative response in white lupin and soybean grown in copper excess.

The microlocalisation of Cu was examined in the leaves of white lupin and soybean grown hydroponically in the presence of 1.6 (control) or 192 µM (excess) Cu, along with its effect on leaf morphology, (ultra)structure and the antioxidative response. The 192 µM dose led to a reduction in the total leaf area and leaf thickness in both species, although more strongly so in white lupin. In the latter species it was also associated with smaller spongy parenchyma cells, and smaller spaces between them, while in the soybean it more strongly reduced the size of the palisade parenchyma and epidermal cells. Energy-dispersive X-ray microanalysis showed that under Cu excess the metal was mainly localised inside the spongy parenchyma cells of the white lupin leaves, and in the lower epidermis cell walls in those of the soybean. Cu excess also promoted ultrastructural chloroplast alterations, reducing the photosynthetic capacity index and the green area of the leaves, especially in the soybean. Despite this, soybean appeared to be more tolerant to Cu excess than white lupin, because soybean displayed (1) lower accumulation of Cu in the leaves, (2) enhanced microlocalisation of Cu in the cell walls and (3) greater levels of induced total -SH content and superoxide dismutase and catalase activities that are expected for better antioxidative responses.

Mitigation of Cu stress by legume-Rhizobium symbiosis in white lupin and soybean plants.

The effect of Bradyrhizobium-legume symbiosis on plant growth, toxicological variables and Cu bioaccumulation was studied in white lupin and soybean plants treated with 1.6, 48, 96 and 192 µM Cu. In both species, those plants grown in the presence of root nodule-forming symbiotic Bradyrhizobium showed less root and shoot growth reduction, plus greater translocation of Cu to the shoot, than those grown without symbiotic Bradyrhizobium. The effective added concentrations of Cu that reduced shoot and root dry weight by 50% (EC50), and the critical toxic concentration that caused a 10% reduction in plant growth (CTC10%), were higher in plants grown with symbiotic Bradyrhizobium, and were in general higher in the roots whether the plants were grown with or without these bacteria. The production of malondialdehyde and total thiols was stimulated by Cu excess in the shoots and roots of white lupin grown with or without symbiotic Bradyrhizobium, but mainly in those without the symbionts. In contrast, in soybean, the increases in malondialdehyde and total thiols associated with rising Cu concentration were a little higher (1.2-5.0 and 1.0-1.6 times respectively) in plants grown with symbiotic Bradyrhizobium than without. Finally, the organ most sensitive to Cu excess was generally the shoot, both in white lupin and soybean grown with or without symbiotic Bradyrhizobium. Further, Bradyrhizobium-legume symbiosis appears to increase the tolerance to Cu excess in both legumes, but mainly in white lupin; plant growth was less reduced and CTC10% and EC50 values increased compared to plants grown without symbiotic Bradyrhizobium. Bradyrhizobium N2 fixation in both legumes would therefore seem to increase the phytoremediation potential of these plants when growing on Cu-contaminated sites.

Efficiency of white lupin in the removal of mercury from contaminated soils: soil and hydroponic experiments.

This study examined the ability of the white lupin to remove mercury (Hg) from a hydroponic system (Hg concentrations 0, 1.25, 2.5, 5 and 10 micromol/L) and from soil in pots and lysimeters (total Hg concentration (19.2 +/- 1.9) mg/kg availability 0.07%, and (28.9 +/- 0.4) mg/kg availability 0.09%, respectively), and investigated the accumulation and distribution of Hg in different parts of the plant. White lupin roots efficiently took up Hg, but its translocation to the harvestable parts of the plant was low. The Hg concentration in the seeds posed no risk to human health according to the recommendations of the World Health Organization, but the shoots should not be used as fodder for livestock, at least when unmixed with other fodder crops. The accumulation of Hg in the hydroponically-grown plants was linear over the concentration range tested. The amount of Hg retained in the roots, relative to the shoots, was almost constant irrespective of Hg dose (90%). In the soil experiments, Hg accumulation increased with exposure time and was the greater in the lysimeter than in the pot experiments. Although Hg removal was the greater in the hydroponic system, revealing the potential of the white lupin to extract Hg, bioaccumulation was the greatest in the lysimeter-grown plants; the latter system more likely reflects the true behaviour of white lupin in the field when Hg availability is a factor that limits Hg removal. The present results suggest that the white lupin could be used in long-term soil reclamation strategies that include the goal of profitable land use in Hg-polluted areas.

Subcellular compartmentalisation of cadmium in white lupins determined by energy-dispersive X-ray microanalysis.

The microlocalisation of cadmium (Cd) at the tissue-cellular level in Lupinus albus L. cv. Multolupa was determined by energy-dispersive X-ray microanalysis (EDXMA). Experimental plants were grown on Cd-treated (0 and 150 microM) perlite for 35 days. In leaves, Cd was found inside cells (cytoplasm or vacuoles), especially in the vascular bundle cells. Cd-induced damage of the chloroplast structure was also detected. EDXMA of the roots showed the cell wall to be the main area of Cd binding at the cellular level; only a small amount of Cd was found in the vacuoles. At the tissue level, a decreasing Cd gradient was seen from the outer to the inner root cortical parenchyma. Cd and S were found co-localised in the vascular cylinder.

Cadmium in white lupin nodules: impact on nitrogen and carbon metabolism.

The aims of this work were to investigate the microlocalisation of cadmium (Cd) in Lupinus albus L. cv. Multolupa nodules, and to determine its effects on carbon and nitrogen metabolism. Nodulated white lupin plants were grown in a growth chamber with or without Cd (150 µM). Energy-dispersive X-ray microanalysis showed the walls of the outer nodule cortex cells to be the main area of Cd retention, helping to reduce the harmful effect Cd might have on the amount of N(2) fixed by the bacteroids. Sucrose synthase activity declined by 33% in the nodules of the Cd-treated plants, and smaller reductions were recorded in glutamine synthetase, aspartate aminotransferase, alkaline invertase and NADP-dependent isocitrate dehydrogenase activities. The Cd treatment also sharply reduced nodule concentrations of malate, succinate and citrate, while that of starch doubled, but that of sucrose experienced no significant change. In summary, the present results show that white lupins accumulate significant amounts of Cd in their root nodules. However, the activity of some enzymes involved in ammonium assimilation did decline, promoting a reduction in the plant N content. The downregulation of sucrose synthase limits the availability of carbon to the bacteroids, which might interfere with their respiration. Carbon metabolism therefore plays a primary role in the impaired function of the white lupin root nodule caused by Cd, while N metabolism appears to have a more secondary involvement.

Interaction and accumulation of manganese and cadmium in the manganese accumulator Lupinus albus.

The effects of the interaction between Mn and Cd on the growth of the white lupin (Lupinus albus), uptake of these metals, their accumulation, and effects on heavy metal stress indicators were studied under glasshouse conditions. Plants were grown with and without Mn and/or Cd for 4 weeks. The absence of Mn and Cd led to lipid peroxidation-induced loss of flavonoids and anthocyanins in the roots, reduced the size of the plant canopy, and led to the appearance of proteoid roots. Sensitivity to Cd in white lupin was enhanced by a low Mn supply, despite lower Cd uptake and accumulation (leaf Mn:Cd concentration ratio <3), as evidenced by increased lipid peroxidation in the leaves and strong inhibition of growth. However, when the Mn supply was adequate, the plants showed few symptoms of Cd toxicity, even though Cd uptake and accumulation increased. A Mn:Cd ratio of up to 20 was enough to minimize Cd stress in the leaf, reflecting the plants' relative tolerance to Cd under such conditions. Irrespective of the Mn supply, the increase in antioxidant compounds observed in the roots of Cd-treated plants might act as a protective mechanism by minimizing the oxidative stress caused by Cd exposure. In summary, high leaf Mn concentrations seem to render white lupins more tolerant to Cd stress.