Alleviation mechanisms of metal(loid) stress in plants by silicon: a review.

Silicon (Si), although not considered as an essential element for plants in general, can ameliorate the phytotoxicity induced by excess metal(loid)s whether non-essential (e.g. Cd, Pb, Cr, Al, As, and Sb) or essential (e.g. Cu, Ni, and Zn). The Si-enhanced resistance allowing plants to cope with this type of abiotic stress has been developed at multiple levels in plants. Restriction of root uptake and immobilization of metal(loid)s in the rhizosphere by Si is probably one of the first defence mechanism. Further, retention of elements in the root apoplasm might enhance the resistance and vigour of plants. At the cellular level, the formation of insoluble complexes between Si and metal(loid)s and their storage within cell walls help plants to decrease available element concentration and restrict symplasmic uptake. Moreover, Si influences the oxidative status of plants by modifying the activity of various antioxidants, improves membrane stability, and acts on gene expression, although its exact role in these processes is still not well understood. This review focuses on all currently known plant-based mechanisms related to Si supply and involved in amelioration of stress caused by excess metal(loid)s.

A new insight on structural and some functional aspects of peri-endodermal thickenings, a specific layer in Noccaea caerulescens roots.

BACKGROUND AND AIMS: Cell walls of the peri-endodermis, a layer adjacent to the endodermis in alpine pennycress (Noccaea caerulescens) roots, form C-shaped peri-endodermal thickenings (PETs). Despite its specific position close to the endodermis, the assumed similarity of PETs to phi thickenings in many other species, and the fact that N. caerulescens is a well-studied heavy-metal-hyperaccumulating plant, the PET as a root trait is still not understood. METHODS: Here, we characterized PET cell walls by histochemical techniques, Raman spectroscopy, immunolabelling and electron microscopy. Moreover, a role of PETs in solute transport was tested and compared with Arabidopsis thaliana plants, which do not form PETs in roots. KEY RESULTS: Cell walls with PETs have a structured relief mainly composed of cellulose and lignin. Suberin, typical of endodermal cells, is missing but pectins are present on the inner surface of the PET. Penetrating dyes are not able to cross PETs either by the apoplasmic or the symplasmic pathway, and a significantly higher content of metals is found in root tissues outside of PETs than in innermost tissues. CONCLUSIONS: Based on their development and chemical composition, PETs are different from the endodermis and closely resemble phi thickenings. Contrarily, the different structure and dye impermeability of PETs, not known in the case of phi thickenings, point to an additional barrier function which makes the peri-endodermis with PETs a unique and rare layer.

Abscisic acid-mediated modifications of radial apoplastic transport pathway play a key role in cadmium uptake in hyperaccumulator Sedum alfredii.

Abscisic acid (ABA) is a key phytohormone underlying plant resistance to toxic metals. However, regulatory effects of ABA on apoplastic transport in roots and consequences for uptake of metal ions are poorly understood. Here, we demonstrate how ABA regulates development of apoplastic barriers in roots of two ecotypes of Sedum alfredii and assess effects on cadmium (Cd) uptake. Under Cd treatment, increased endogenous ABA level was detected in roots of nonhyperaccumulating ecotype (NHE) due to up-regulated expressions of ABA biosynthesis genes (SaABA2, SaNCED), but no change was observed in hyperaccumulating ecotype (HE). Simultaneously, endodermal Casparian strips (CSs) and suberin lamellae (SL) were deposited closer to root tips of NHE compared with HE. Interestingly, the vessel-to-CSs overlap was identified as an ABA-driven anatomical trait. Results of correlation analyses and exogenous applications of ABA/Abamine indicate that ABA regulates development of both types of apoplastic barriers through promoting activities of phenylalanine ammonialyase, peroxidase, and expressions of suberin-related genes (SaCYP86A1, SaGPAT5, and SaKCS20). Using scanning ion-selected electrode technique and PTS tracer confirmed that ABA-promoted deposition of CSs and SL significantly reduced Cd entrance into root stele. Therefore, maintenance of low ABA levels in HE minimized deposition of apoplastic barriers and allowed maximization of Cd uptake via apoplastic pathway.

c-Met and PD-L1 on Circulating Exosomes as Diagnostic and Prognostic Markers for Pancreatic Cancer.

Exosomes are membrane vesicles which offer potential as blood derived biomarkers for malign tumors in clinical practice. Pancreatic cancer is counted among cancer diseases with the highest mortality. The present work seeks to assess whether pancreatic carcinomas release exosomes which express c-Met (proto-oncogene mesenchymal-epithelial transition factor) and PD-L1 (programmed cell death 1 ligand 1), and whether the detection of such expression in serum has diagnostic or prognostic meaning for the affected patients. Exosome isolation was performed on culture media of one benign pancreatic cell line and ten pancreatic carcinoma cell lines as well as on serum samples from 55 patients with pancreatic ductal adenocarcinoma (PDAC), 26 patients with chronic pancreatitis and 10 patients with benign serous cyst adenoma of the pancreas. Exosomes were bound to latex beads and stained with antibodies against c-Met or PD-L1. Analysis of fluorescence intensity was performed by flow cytometry. In terms of c-Met, the mean fluorescence intensity of PDAC-patients was significantly higher than the fluorescence intensity of the comparative patients with benign disease (p < 0.001). A diagnostic test based on c-Met resulted in a sensitivity of 70%, a specificity of 85% and a diagnostic odds ratio of 13:2. The specificity of the test can be further improved by combining it with the established tumor marker carbohydrate antigen 19-9 (CA 19-9). In addition, c-Met-positive patients showed a significantly shorter postoperative survival time (9.5 vs. 21.7 months, p < 0.001). In terms of PD-L1, no significant difference between fluorescence intensity of PDAC-patients and comparative patients was detectable. However, PD-L1-positive PDAC-patients also showed a significantly shorter postoperative survival time (7.8 vs. 17.2 months, p = 0.043). Thus, both markers can be considered as negative prognostic factors.

Formation of a subero-lignified apical deposit in root tip of radish (Raphanus sativus) as a response to copper stress.

Background and Aims: Heavy metals induce changes in root metabolism and physiology, which can lead to a complex remodelling of the root system. The final morphological responses of radish (Raphanus sativus) roots exposed to toxic concentrations of the heavy metal (Cu) include root growth inhibition, differentiation of xylem vessels close to the root tip, enhanced suberin lamellae deposition and enhanced lateral root production. Recently, we have found that such changes in root morphology and anatomy are coupled to the formation of a subero-lignified apical deposit (SLAD) very close to the root tip. Methods: To clarify the details of the formation of a SLAD in the root tip, we conducted experiments with radish roots exposed to a high Cu concentration (60 µm). Histochemical analysis of lignin and suberin as well as analysis of spatial-temporal characteristics of SLAD formation were performed by bright-field, fluorescence and confocal microscopy. Key Results: This unique structure, not longer than 100 µm, consists of modified cell walls of the central cylinder that are encircled by a short cylinder of prematurely suberized endodermal cells. A SLAD starts to form, in both primary and lateral roots, after cessation of root elongation, and it is coupled with xylem differentiation and root branching close to the root apex. We noticed that deposition of phenolic substances into a SLAD, mainly suberin in the endodermis, is spatially separated from suberization or lignification in basally located endodermis. Conclusions: Although the main reason for formation of a SLAD is elusive, we suggest that it is a part of stress-induced responses which relate to decreased root growth or permeability in heavy metal stress.

Root hair abundance impacts cadmium accumulation in Arabidopsis thaliana shoots.

Background and Aims: Root hairs increase the contact area of roots with soil and thereby enhance the capacity for solute uptake. The strict hair/non-hair pattern of Arabidopsis thaliana can change with nutrient deficiency or exposure to toxic elements, which modify root hair density. The effects of root hair density on cadmium (Cd) accumulation in shoots of arabidopsis genotypes with altered root hair development and patterning were studied. Methods: Arabidopsis mutants that are unable to develop root hairs (rhd6-1 and cpc/try) or produce hairy roots (wer/myb23) were compared with the ecotype Columbia (Col-0). Plants were cultivated on nutrient agar for 2 weeks with or without Cd. Cadmium was applied as Cd(NO3)2 at two concentrations, 10 and 100 µm. Shoot biomass, root characteristics (primary root length, lateral root number, lateral root length and root hair density) and Cd concentrations in shoots were assessed. Anatomical features (suberization of the endodermis and development of the xylem) that might influence Cd uptake and translocation were also examined. Key Results: Cadmium inhibited plant growth and reduced root length and the number of lateral roots and root hairs per plant. Suberin lamellae in the root endodermis and xylem differentiation developed closer to the root apex in plants exposed to 100 µm Cd. The latter effect was genotype dependent. Shoot Cd accumulation was correlated with root hair abundance when plants were grown in the presence of 10 µm Cd, but not when grown in the presence of 100 µm Cd, in which treatment the development of suberin lamellae closer to the root tip appeared to restrict Cd accumulation in shoots. Conclusions: Root hair density can have a large effect on Cd accumulation in shoots, suggesting that the symplasmic pathway might play a significant role in the uptake and accumulation of this toxic element.

The apoplasmic pathway via the root apex and lateral roots contributes to Cd hyperaccumulation in the hyperaccumulator Sedum alfredii.

Although the significance of apoplasmic barriers in roots with regards to the uptake of toxic elements is generally known, the contribution of apoplasmic bypasses (ABs) to cadmium (Cd) hyperaccumulation is little understood. Here, we employed a combination of stable isotopic tracer techniques, an ABs tracer, hydraulic measurements, suberin lamellae staining, metabolic inhibitors, and antitranspirants to investigate and quantify the impact of the ABs on translocation of Cd to the xylem in roots of a hyperaccumulating (H) ecotype and a non-hyperaccumulating (NH) ecotype of Sedum alfredii. In the H ecotype, the Cd content in the xylem sap was proportional to hydrostatic pressure, which was attributed to pressure-driven flow via the ABs. The contribution of the ABs to Cd transportation to the xylem was dependent on the Cd concentration applied to the H ecotype (up to 37% at the highest concentration used). Cd-treated H ecotype roots showed significantly higher hydraulic conductance compared with the NH ecotype (76 vs 52 × 10­8 m s­1MPa­1), which is in accordance with less extensive suberization due to reduced expression of suberin-related genes. The main entry sites of apoplasmically transported Cd were localized in the root apexes and lateral roots of the H ecotype, where suberin lamellae were not well developed. These findings highlight the significance of the apoplasmic bypass in Cd hyperaccumulation in hyperaccumulating ecotypes of S. alfredii.

Formation of silica aggregates in sorghum root endodermis is predetermined by cell wall architecture and development.

Background and Aims: Deposition of silica in plant cell walls improves their mechanical properties and helps plants to withstand various stress conditions. Its mechanism is still not understood and silica-cell wall interactions are elusive. The objective of this study was to investigate the effect of silica deposition on the development and structure of sorghum root endodermis and to identify the cell wall components involved in silicification. Methods: Sorghum bicolor seedlings were grown hydroponically with (Si+) or without (Si-) silicon supplementation. Primary roots were used to investigate the transcription of silicon transporters by quantitative RT-PCR. Silica aggregation was induced also under in vitro conditions in detached root segments. The development and architecture of endodermal cell walls were analysed by histochemistry, microscopy and Raman spectroscopy. Water retention capability was compared between silicified and non-silicified roots. Raman spectroscopy analyses of isolated silica aggregates were also carried out. Key Results: Active uptake of silicic acid is provided at the root apex, where silicon transporters Lsi1 and Lsi2 are expressed. The locations of silica aggregation are established during the development of tertiary endodermal cell walls, even in the absence of silicon. Silica aggregation takes place in non-lignified spots in the endodermal cell walls, which progressively accumulate silicic acid, and its condensation initiates at arabinoxylan-ferulic acid complexes. Silicification does not support root water retention capability; however, it decreases root growth inhibition imposed by desiccation. Conclusion: A model is proposed in which the formation of silica aggregates in sorghum roots is predetermined by a modified cell wall architecture and takes place as governed by endodermal development. The interaction with silica is provided by arabinoxylan-ferulic acid complexes and interferes with further deposition of lignin. Due to contrasting hydrophobicity, silicification and lignification do not represent functionally equivalent modifications of plant cell walls.

Asymmetrical development of root endodermis and exodermis in reaction to abiotic stresses.

Background and Aims In the present study, we show that development of endodermis and exodermis is sensitively regulated by water accessibility. As cadmium (Cd) is known to induce xeromorphic effects in plants, maize roots were exposed also to Cd to understand the developmental process of suberin lamella deposition in response to a local Cd source. Methods In a first experiment, maize roots were cultivated in vitro and unilaterally exposed to water-containing medium from one side and to air from the other. In a second experiment, the roots were placed between two agar medium layers with a strip of Cd-containing medium attached locally and unilaterally to the root surface. Key Results The development of suberin lamella (the second stage of exodermal and endodermal development) started asymmetrically, preferentially closer to the root tip on the side exposed to the air. In the root contact with Cd in a spatially limited area exposed to one side of the root, suberin lamella was preferentially developed in the contact region and additionally along the whole length of the root basipetally from the contact area. However, the development was unilateral and asymmetrical, facing the treated side. The same pattern occurred irrespective of the distance of Cd application from the root apex. Conclusions These developmental characteristics indicate a sensitive response of root endodermis and exodermis in the protection of vascular tissues against abiotic stresses.

Cadmium translocation by contractile roots differs from that in regular, non-contractile roots.

BACKGROUND AND AIMS: Contractile roots are known and studied mainly in connection with the process of shrinkage of their basal parts, which acts to pull the shoot of the plant deeper into the ground. Previous studies have shown that the specific structure of these roots results in more intensive water uptake at the base, which is in contrast to regular root types. The purpose of this study was to find out whether the basal parts of contractile roots are also more active in translocation of cadmium to the shoot. METHODS: Plants of the South African ornamental species Tritonia gladiolaris were cultivated in vitro for 2 months, at which point they possessed well-developed contractile roots. They were then transferred to Petri dishes with horizontally separated compartments of agar containing 50 µmol Cd(NO3)2 in the region of the root base or the root apex. Seedlings of 4-d-old maize (Zea mays) plants, which do not possess contractile roots, were also transferred to similar Petri dishes. The concentrations of Cd in the leaves of the plants were compared after 10 d of cultivation. Anatomical analyses of Tritonia roots were performed using appropriately stained freehand cross-sections. KEY RESULTS: The process of contraction required specific anatomical adaptation of the root base in Tritonia, with less lignified and less suberized tissues in comparison with the subapical part of the root. These unusual developmental characteristics were accompanied by more intensive translocation of Cd ions from the basal part of contractile roots to the leaves than from the apical-subapical root parts. The opposite effects were seen in the non-contractile roots of maize, with higher uptake and transport by the apical parts of the root and lower uptake and transport by the basal part. CONCLUSIONS: The specific characteristics of contractile roots may have a significant impact on the uptake of ions, including toxic metals from the soil surface layers. This may be important for plant nutrition, for example in the uptake of nutrients from upper soil layers, which are richer in humus in otherwise nutrient-poor soils, and also has implications for the uptake of surface-soil pollutants.

Silicon alleviates cadmium toxicity by enhanced photosynthetic rate and modified bundle sheath's cell chloroplasts ultrastructure in maize.

Silicon was shown to alleviate the negative effects of various biotic and abiotic stresses on plant growth. Although the positive role of Si on toxic and heavy metal Cd has been already described, the mechanisms have been explained only partially and still remain unclear. In the present study we investigated the effect of Si on photosynthetic-related processes in maize exposed to two different levels of Cd via measurements of net photosynthetic rate (AN), chlorophyll a fluorescence and pigment analysis, as well as studies of leaf tissue anatomy and cell ultrastructure using bright-field and transmission electron microscopy. We found that Si actively alleviated the toxic syndromes of Cd by increasing AN, effective photochemical quantum yield of photosystem II (ϕPSII) and content of assimilation pigments, although did not decrease the concentration of Cd in leaf tissues. Cadmium did not affect the leaf anatomy and ultrastructure of leaf mesophyll's cell chloroplasts; however, Cd negatively affected thylakoid formation in chloroplasts of bundle sheath cells, and this was alleviated by Si. Improved thylakoid formation in bundle sheath's cell chloroplasts may contribute to Si-induced enhancement of photosynthesis and related increase in biomass production in C4 plant maize.

New method for visualization of silica phytoliths in Sorghum bicolor roots by fluorescence microscopy revealed silicate concentration-dependent phytolith formation.

Silica phytoliths are microscopic structures of amorphous hydrated silica (SiO2 · nH2O) formed by specialized plant cells. Besides their biological roles, physical, chemical, and structural properties of biogenic silica offer a wide spectrum of applications in many fields of industry and technology. Therefore, processes involved in their formation recently become a very interesting topic to study. However, optical transparency and microscopic sizes of silica phytoliths do not allow their visualization and localization by classical light microscopy methods. Their observation thus requires phytolith isolation, technically difficult or lengthy sample preparation procedures, or a work with toxic chemicals. In this paper we are proposing a novel method for visualization of silica phytoliths in Sorghum bicolor root endodermal cells by fluorescence microscopy using alkali mounting solution (pH 12). This method offers an easy and quick preparation of the samples and high contrast imaging. Based on our results we can assume that the proposed fluorescent method for silica phytolith investigation allows observation of multiple samples in relatively short time period and thus might be applicable also for high-throughput screenings. Using this method we found out that after a 3-day cultivation of sorghum plants the minimal needed concentration of sodium silicate, limiting the formation of silica phytoliths in the root endodermis, was 25 µmol dm(-3). The positive correlation of sodium silicate concentration in the substrate with the phytolith diameter was also observed.

Cadmium uptake, localization and stress-induced morphogenic response in the fern Pteris vittata.

Cadmium uptake, tissue localization and structural changes induced at cellular level are essential to understand Cd tolerance in plants. In this study we have exposed plants of Pteris vittata to different concentrations of CdCl2 (0, 30, 60, 100 µM) to evaluate the tolerance of the fern to cadmium. Cadmium content determination and its histochemical localization showed that P. vittata not only takes up, but also transports and accumulates cadmium in the aboveground tissues, delocalizing it mainly in the less bioactive tissues of the frond, the trichomes and the scales. Cadmium tolerance in P. vittata was strictly related to morphogenic response induced by the metal itself in the root system. Adaptive response regarded changes of the root apex size, the developmental pattern of root hairs, the differentiation of xylem elements and endodermal suberin lamellae. All the considered parameters suggest that, in our experimental conditions, 60 µM of Cd may represent the highest concentration that P. vittata can tolerate; indeed this Cd level even improves the absorbance features of the root and allows good transport and accumulation of the metal in the fronds. The results of this study can provide useful information for phytoremediation strategies of soils contaminated by Cd, exploiting the established ability of P. vittata to transport, delocalize in the aboveground biomass and accumulate polluting metals.

Silicon nanoparticles vs trace elements toxicity: Modus operandi and its omics bases.

Phytotoxicity of trace elements (commonly misunderstood as 'heavy metals') includes impairment of functional groups of enzymes, photo-assembly, redox homeostasis, and nutrient status in higher plants. Silicon nanoparticles (SiNPs) can ameliorate trace element toxicity. We discuss SiNPs response against several essential (such as Cu, Ni, Mn, Mo, and Zn) and non-essential (including Cd, Pb, Hg, Al, Cr, Sb, Se, and As) trace elements. SiNPs hinder root uptake and transport of trace elements as the first line of defence. SiNPs charge plant antioxidant defence against trace elements-induced oxidative stress. The enrolment of SiNPs in gene expressions was also noticed on many occasions. These genes are associated with several anatomical and physiological phenomena, such as cell wall composition, photosynthesis, and metal uptake and transport. On this note, we dedicate the later sections of this review to support an enhanced understanding of SiNPs influence on the metabolomic, proteomic, and genomic profile of plants under trace elements toxicity.

Mechanisms of Arbuscular Mycorrhizal Fungi Increasing Silicon Uptake by Rice.

Arbuscular mycorrhizal fungi (AMF) influence silicon (Si) uptake by plants, but the mechanisms remain unclear. This study investigated the mechanisms of AMF-mediated Si uptake by rice, a model Si-accumulating plant, and explored the tripartite interactions among AMF, Si, and phosphorus (P). AMF inoculation increased shoot Si content by 97% when supplied with silicic acid and by 29% with calcium silicate and upregulated expression of Si transporters Lsi1 and Lsi2 in roots. Supplying Si only to AMF hyphae increased the root Si content by 113%, indicating direct Si uptake by hyphae. Mechanisms of AMF-induced Si uptake were elucidated: 1) direct Si uptake by hyphae, 2) increased silicate dissolution, and 3) upregulation of Si transporters. Silicon application also increased AMF colonization by 28%, and the absence of interactions was observed on P uptake. Altogether, AMF support Si acquisition and Si fosters AMF colonization in rice, whereas the P uptake depends more on AMF than on Si.

Bioavailability and toxicity of arsenic in maize (Zea mays L.) grown in contaminated soils.

The growth of maize (Zea mays L.), hybrid Valentina, was compared in two types of contaminated soil substrates (Ochre and Heap) with different arsenic (As) concentration originating from an old mining area in Slovakia. Although the total As concentration in Heap soil was 2.6 times lower than in Ochre soil (90 and 237 mg kg⁻¹, respectively), plants grown in Heap soil accumulated more As in their tissues (shoot and root As concentration being 4 and 5.5 times higher, respectively) and were markedly smaller, which produced significantly less biomass and flowered later in comparison with Ochre soil grown plants.

Silicon nanoparticles (SiNPs) restore photosynthesis and essential oil content by upgrading enzymatic antioxidant metabolism in lemongrass (Cymbopogon flexuosus) under salt stress.

Lemongrass (Cymbopogon flexuosus) has great relevance considering the substantial commercial potential of its essential oil. Nevertheless, the increasing soil salinity poses an imminent threat to lemongrass cultivation given its moderate salt-sensitivity. For this, we used silicon nanoparticles (SiNPs) to stimulate salt tolerance in lemongrass considering SiNPs special relevance to stress settings. Five foliar sprays of SiNPs 150 mg L-1 were applied weekly to NaCl 160 and 240 mM-stressed plants. The data indicated that SiNPs minimised oxidative stress markers (lipid peroxidation, H2O2 content) while triggering a general activation of growth, photosynthetic performance, enzymatic antioxidant system including superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD), and osmolyte proline (PRO). SiNPs amplified stomatal conductance and photosynthetic CO2 assimilation rate by about 24% and 21% in NaCl 160 mM-stressed plants. Associated benefits contributed to pronounced plant phenotype over their stressed counterparts, as we found. Foliar SiNPs sprays assuaged plant height by 30% and 64%, dry weight by 31% and 59%, and leaf area by 31% and 50% under NaCl 160 and 240 mM concentrations, respectively. SiNPs relieved enzymatic antioxidants (SOD, CAT, POD) and osmolyte (PRO) in lemongrass plants stressed with NaCl 160 mM (9%, 11%, 9%, and 12%, respectively) and NaCl 240 mM (13%, 18%, 15%, and 23%, respectively). The same treatment supported the oil biosynthesis improving essential oil content by 22% and 44% during 160 and 240 mM salt stress, respectively. We found SiNPs can completely overcome NaCl 160 mM stress while significantly palliating NaCl 240 mM stress. Thus, we propose that SiNPs can be a useful biotechnological tool to palliate salinity stress in lemongrass and related crops.

Copper and zinc accumulation, distribution, and tolerance in Pistia stratiotes L.; revealing the role of root caps.

Pollution by potentially toxic trace metals, such as copper or zinc, is global. Both Cu and Zn are essential microelements, which in higher concentrations become toxic. The aquatic plant Pistia stratiotes(L. has great potential for phytoremediation. Also, it has an unusually large and easily detachable root cap, which makes it a suitable model for studying the potential role of the root cap in metal uptake. Plant response to environmentally relevant concentrations of Cu (0.1, 0.3, and 1 µM) and Zn (0.3, 1, and 3 µM) was investigated with the aim of studying their interaction and distribution at the root tissue level as well as revealing their tolerance mechanisms. Changes in the root anatomy and plant ionome were determined using light and fluorescence microscopy, ICP-MS, and µXRF imaging. Alterations in photosynthetic activity caused by Cu or Zn excesses were monitored by direct imaging of fast chlorophyll fluorescence kinetics (OJIP). Fe and Mn were preferentially localized in the root cap, while Ca, Cu, Ni, and Zn were mainly in the root tip regardless of the Cu/Zn treatment. Translocation of Cu and Zn to the leaves increased with higher doses, however the translocation factor was the lowest in the highest treatments. Measurements of photosynthetic parameters showed a higher susceptibility of electron transport flux from QA to QB under increasing Cu than Zn supply. This, along with our findings regarding the root anatomy and the differences in Ca accumulation and distribution, led to the conclusion that P. stratiotes is more effective for Zn remediation than Cu.