Submitted to: Plant and Soil
Publication Type: Peer reviewed journal
Publication Acceptance Date: 2/10/2004
Publication Date: 8/15/2004
Citation: Broadhurst, C.L., Chaney, R.L., Angle, J.S., Erbe, E.F., Maugel, T.K. 2004. Nickel localization and response to increasing ni soil levels in leaves of the ni hyperaccumulator alyssum murale "kotodesh". Plant and Soil Journal. 265(1-2):225-242. Interpretive Summary: The Ni-hyperaccumulator species, Alyssum murale, has begun to be used as a commercial crop for Ni phytoextraction. The plants accumulate up to nearly 3% Ni in their shoots without yield reduction. This remarkable Ni tolerance by A. murale raises the question, where in the plant tissues is the Ni stored?, and which cells accumulate Ni most avidly? These questions were addressed by X-ray fluorescence measurements on frozen sections, and electron microscope examination of the complement fractured tissue. For such examination to be valid, the tissue must remain frozen and no treatments may be used which could cause movement of Ni between cells or tissues. Interesting results were obtained which clarified points addressed by other authors. For example, one author claimed that Ni was stored in leaf trichomes at very high concentration; but the current test found trichomes were very low in Ni but rich in Ca. By growing the plants at varied levels of Ni supply, the cells which store Ni most avidly could be detected; epidermal cells on top and bottom of leaves filled before mesophyll cells. Cell cuticles did not store Ni. And one remarkable finding was that guard cells were very low in Ni when the adjacent epidermal cells were very high in Ni. In the graded series of Ni supply, mesophyll cells accumulated Ni at lower levels than the epidermal cells, but comprise a larger volume of the tissue and at high Ni comprise the major storage area. All evidence supported the view that Ni is mostly stored in vacuoles of leaf cells. Additional testing will be needed to see if differences in Ni storage among A. murale genotypes is correlated with the ability of the genotypes to phytoextract soil Ni.
Technical Abstract: We have developed phytoremediation and phytomining technologies employing Alyssum nickel (Ni) hyperaccumulators to quantitatively extract Ni from soils. Implementation of these technologies requires knowledge of Ni localization patterns for the Alyssum species/ecotypes of interest under realistic growth conditions. We investigated Ni uptake and localization in mature Alyssum murale 'Kotodesh' leaves. Seedlings were grown in potting mix with an increasing series of NiSO4 addition (0, 5, 10, 20, 40, 80 mmol Ni/kg) and in Ni-contaminated soil from metal refining operations. Plants at Ni levels 0, 5, 10, 20 mmol/kg and in refinery soil grew normally. Plants at 40 mmol/kg exhibited the onset of phytotoxicity, and 80 mmol/kg was demonstrably phytotoxic. For each level, cryogenic complement fractures were made from frozen hydrated samples. High-resolution scanning electron microscope (SEM) images were taken of one half. The other half was examined with SEM and semi-quantitative energy dispersive x-ray analysis (SEM-EDX). Our dose-response experiment revealed that Ni localization is more complex and dependent on Ni soil concentration/availability that previously reported. In agreement with previous work, Ni was generally concentrated in the upper and lower epidermis. Specifically, trichome pedicles and epidermal cells associated with trichome attachment were primary Ni compartments, but Ni was not distributed throughout trichomes. Further, increasing Ni levels correlated with decreasing trichome density. Palisade and spongy mesophyll contained relatively high Ni, concentrations (especially mesophyll cells adjacent to the epidermis), and were increasingly important compartments as Ni levels increased. Overall, Ni was not fully excluded from any tissues, although guard cells, substomatal cells, vascular tissue, and trichome rays are probably not preferred Ni compartments in Alyssum murale.