Mechanisms of Uptake and Translocation of Thallium in Brassica Vegetables: An X-ray Fluorescence Microspectroscopic Investigation.

Most nonoccupational human exposure to thallium (Tl) occurs via consumption of contaminated food crops. Brassica cultivars are common crops that can accumulate more than 500 µg Tl g-1. Knowledge of Tl uptake and translocation mechanisms in Brassica cultivars is fundamental to developing methods to inhibit Tl uptake or conversely for potential use in phytoremediation of polluted soils. Brassica cultivars (25 in total) were subjected to Tl dosing to screen for Tl accumulation. Seven high Tl-accumulating varieties were selected for follow-up Tl dosing experiments. The highest Tl accumulating Brassica cultivars were analyzed by synchrotron-based micro-X-ray fluorescence to investigate the Tl distribution and synchrotron-based X-ray absorption near-edge structure spectroscopy (XANES) to unravel Tl chemical speciation. The cultivars exhibited different Tl tolerance and accumulation patterns with some reaching up to 8300 µg Tl g-1. The translocation factors for all the cultivars were >1 with Brassica oleracea var. acephala (kale) having the highest translocation factor of 167. In this cultivar, Tl is preferentially localized in the venules toward the apex and along the foliar margins and in minute hot spots in the leaf blade. This study revealed through scanning electron microscopy and X-ray fluorescence analysis that highly Tl-enriched crystals occur in the stoma openings of the leaves. The finding is further validated by XANES spectra that show that Tl(I) dominates in the aqueous as well as in the solid form. The high accumulation of Tl in these Brassica crops has important implications for food safety and results of this study help to understand the mechanisms of Tl uptake and translocation in these crops.

Chemical transformations of arsenic in the rhizosphere-root interface of Pityrogramma calomelanos and Pteris vittata.

Pityrogramma calomelanos and Pteris vittata are cosmopolitan fern species that are the strongest known arsenic (As) hyperaccumulators, with potential to be used in the remediation of arsenic-contaminated mine tailings. However, it is currently unknown what chemical processes lead to uptake of As in the roots. This information is critical to identify As-contaminated soils that can be phytoremediated, or to improve the phytoremediation process. Therefore, this study identified the in situ distribution of As in the root interface leading to uptake in P. calomelanos and P. vittata, using a combination of synchrotron micro-X-ray fluorescence spectroscopy and X-ray absorption near-edge structure imaging to reveal chemical transformations of arsenic in the rhizosphere-root interface of these ferns. The dominant form of As in soils was As(V), even in As(III)-dosed soils, and the major form in P. calomelanos roots was As(III), while it was As(V) in P. vittata roots. Arsenic was cycled from roots growing in As-rich soil to roots growing in control soil. This study combined novel analytical approaches to elucidate the As cycling in the rhizosphere and roots enabling insights for further application in phytotechnologies to remediated As-polluted soils.

Comprehensive insights in thallium ecophysiology in the hyperaccumulator Biscutella laevigata.

Biscutella laevigata is the strongest known thallium (Tl) hyperaccumulator plant species. However, little is known about the ecophysiological processes leading to root uptake and translocation of Tl in this species, and the interactions between Tl and its chemical analogue potassium (K). Biscutella laevigata was subjected to hydroponics experimentation in which it was exposed to Tl and K, and it was investigated in a rhizobox experiment. Laboratory-based micro-X-ray fluorescence spectroscopy (µ-XRF) was used to reveal the Tl distribution in the roots and leaves, while synchrotron-based µ-XRF was utilised to reveal elemental distribution in the seed. The results show that in the seed Tl was mainly localised in the endosperm and cotyledons. In mature plants, Tl was highest in the intermediate leaves (16,100 µg g-1), while it was one order of magnitude lower in the stem and roots. Potassium did not inhibit or enhance Tl uptake in B.laevigata. At the organ level, Tl was localised in the blade and margins of the leaves. Roots foraged for Tl and cycled Tl across roots growing in the control soils. Biscutella laevigata has ostensibly evolved specialised mechanisms to tolerate high Tl concentrations in its shoots. The lack of interactions and competition between Tl and K suggests that it is unlikely that Tl is taken up via K channels, but high affinity Tl transporters remain to be identified in this species. Thallium is not only highly toxic but also a valuable metal and Tl phytoextraction using B. laevigata should be explored.

Root responses to localised soil arsenic enrichment in the fern Pityrogramma calomelanos var. austroamericana grown in rhizoboxes.

The terrestrial fern Pityrogramma calomelanos, a cosmopolitan tropical species, is one of the strongest known arsenic (As) hyperaccumulator plants. This study aimed to determine whether P. calomelanos preferentially forages for arsenite (As3+) or arsenate (As5+) in As-contaminated soils, and whether a positive root response to As enhances accumulation in P. calomelanos. Therefore, an experiment using rhizoboxes divided in two halves were constructed with a control soil (C) and As3+ or As5+ dosed soil at either 50 and 100 µg g-1 As. Micro-X-ray Fluorescence elemental mapping (µXRF) was employed to analyze the distribution of As in roots and fronds, and Scanning Electron Microscopy with Energy Dispersive Spectroscopy (SEM-EDS) was used to determine As distribution in the reproductive tissues of P. calomelanos. The results showed that Pityrogramma roots do not specifically forage for As-contaminated soil; the area based on pixel counts was similar across all the treatments with no statistical differences. However, frond biomass was slightly higher in the treatments C ǀ As3+ and C ǀ As5+, and the highest accumulation of As in fronds was in the As5+ ǀ As3+ (100 µg g-1) treatment, with 3418 and 2370 µg g-1 in old and young fronds respectively. Arsenic cycling across the roots was observed by the µXRF mapping; in C ǀ As5+ (100) the As was higher and evenly distributed in both sections, whilst in C ǀ As3+ (50), the As was higher in the As3+ side. The µXRF mapping showed a broader As distribution in older fronds, where As was highest in the rachis and extended into the pinnule through the midrib. Pityrogramma calomelanos does not specifically root forage for As-enriched zones in the soil and grows healthily without signs of toxicity at lower (50 µg g-1) and higher (100 µg g-1) concentrations of As3+ and As5+ in the soil.

Quantification of spatial metal accumulation patterns in Noccaea caerulescens by X-ray fluorescence image processing for genetic studies.

BACKGROUND: Hyperaccumulation of trace elements is a rare trait among plants which is being investigated to advance our understanding of the regulation of metal accumulation and applications in phytotechnologies. Noccaea caerulescens (Brassicaceae) is an intensively studied hyperaccumulator model plant capable of attaining extremely high tissue concentrations of zinc and nickel with substantial genetic variation at the population-level. Micro-X-ray Fluorescence spectroscopy (µXRF) mapping is a sensitive high-resolution technique to obtain information of the spatial distribution of the plant metallome in hydrated samples. We used laboratory-based µXRF to characterize a collection of 86 genetically diverse Noccaea caerulescens accessions from across Europe. We developed an image-processing method to segment different plant substructures in the µXRF images. We introduced the concentration quotient (CQ) to quantify spatial patterns of metal accumulation and linked that to genetic variation. RESULTS: Image processing resulted in automated segmentation of µXRF plant images into petiole, leaf margin, leaf interveinal and leaf vasculature substructures. The harmonic means of recall and precision (F1 score) were 0.79, 0.80, 0.67, and 0.68, respectively. Spatial metal accumulation as determined by CQ is highly heritable in Noccaea caerulescens for all substructures, with broad-sense heritability (H2) ranging from 76 to 92%, and correlates only weakly with other heritable traits. Insertion of noise into the image segmentation algorithm barely decreases heritability scores of CQ for the segmented substructures, illustrating the robustness of the trait and the quantification method. Very low heritability was found for CQ if randomly generated substructures were compared, validating the approach. CONCLUSIONS: A strategy for segmenting µXRF images of Noccaea caerulescens is proposed and the concentration quotient is developed to provide a quantitative measure of metal accumulation pattern, which can be used to determine genetic variation for such pattern. The metric is robust to segmentation error and provides reliable H2 estimates. This strategy provides an avenue for quantifying XRF data for analysis of the genetics of metal distribution patterns in plants and the subsequent discovery of new genes that regulate metal homeostasis and sequestration in plants.