Physiological Analysis and Proteome Quantification of Alligator Weed Stems in Response to Potassium Deficiency Stress.

The macronutrient potassium is essential to plant growth, development and stress response. Alligator weed (Alternanthera philoxeroides) has a high tolerance to potassium deficiency (LK) stress. The stem is the primary organ responsible for transporting molecules from the underground root system to the aboveground parts of the plant. However, proteomic changes in response to LK stress are largely unknown in alligator weed stems. In this study, we investigated the physiological and proteomic changes in alligator weed stems under LK stress. First, the chlorophyll and soluble protein content and SOD and POD activity were significantly altered after 15 days of LK treatment. The quantitative proteomic analysis suggested that a total of 296 proteins were differentially abundant proteins (DAPs). The functional annotation analysis revealed that LK stress elicited complex proteomic alterations that were involved in oxidative phosphorylation, plant-pathogen interactions, glycolysis/gluconeogenesis, sugar metabolism, and transport in stems. The subcellular locations analysis suggested 104 proteins showed chloroplastic localization, 81 proteins showed cytoplasmic localization and 40 showed nuclear localization. The protein⁻protein interaction analysis revealed that 56 proteins were involved in the interaction network, including 9 proteins involved in the ribosome network and 9 in the oxidative phosphorylation network. Additionally, the expressed changes of 5 DAPs were similar between the proteomic quantification analysis and the PRM-MS analysis, and the expression levels of eight genes that encode DAPs were further verified using an RT-qPCR analysis. These results provide valuable information on the adaptive mechanisms in alligator weed stems under LK stress and facilitate the development of efficient strategies for genetically engineering potassium-tolerant crops.

Mercury mobility and effects in the salt-marsh plant Halimione portulacoides: Uptake, transport, and toxicity and tolerance mechanisms.

The plant Halimione portulacoides, an abundant species widely distributed in temperate salt-marshes, has been previously assessed as bioindicator and biomonitor of mercury contamination in these ecosystems. The present study aims to assess uptake and distribution of total mercury (THg) and methylmercury (MMHg) within H. portulacoides, potential mercury release by volatilization through leaves, and toxicity and tolerance mechanisms by investigating plant photochemical responses. Stem cuttings of H. portulacoides were collected from a salt-marsh within the Tagus estuary natural protected area, and grown under hydroponic conditions. After root development, plants were exposed to 199HgCl2 and CH3201HgCl, and sampled at specific times (0, 1, 2, 4, 24, 72, 120, 168 (7 days) and 432 h (18 days)). After exposure, roots, stems and leaves were analysed for total 199Hg (T199Hg) and MM201Hg content. Photobiology parameters, namely efficiency and photoprotection capacity, were measured in leaves. Both THg and MMHg were incorporated into the plant root system, stems and leaves, with roots showing much higher levels of both isotope enriched spikes than the other plant tissues. Presence of both mercury isotopes in the stems and leaves and high significant correlations found between roots and stems, and stems and leaves, for both THg and MMHg concentrations, indicate Hg translocation between the roots and above-ground organs. Long-term uptake in stems and leaves, leading to higher Hg content, was more influenced by temperature and radiation than short-term uptake. However, the relatively low levels of both THg and MMHg in the aerial parts of the plant, which were influenced by temperature and radiation, support the possibility of mercury release by stems and leaves, probably via stomata aperture, as a way to eliminate toxic mercury. Regarding photochemical responses, few differences between control and exposed plants were observed, indicating high tolerance of this salt marsh plant to THg and MMHg.

Selenium and silicon reduce cadmium uptake and mitigate cadmium toxicity in Pfaffia glomerata (Spreng.) Pedersen plants by activation antioxidant enzyme system.

Cadmium (Cd) is toxic to plants and animals, making it necessary to develop strategies that seek to reduce its introduction into food chains. Thus, the aim of this study was to investigate whether silicon (Si) and selenium (Se) reduce Cd concentrations in Pfaffia glomerata medicinal plant and attenuate the oxidative stress promoted by this metal. These plants were cultivated in hydroponics under the following treatments: control (nutrient solution), 2.5 µM Se, 2.5 mM Si, 50 µM Cd, 50 µM Cd + 2.5 µM Se, 50 µM Cd + 2.5 mM Si. After 14 days of exposure to treatments, leaves and roots were collected for the determination of dry weight of shoot and roots, Cd concentrations, chlorophyll and carotenoids content, and biochemical parameters (lipid peroxidation and guaiacol peroxidase and superoxide dismutase activities). The data were submitted to analysis of variance and means were compared with Scott-Knott test at 5% error probability. Roots of P. glomerata plants showed a significant reduction on dry weight accumulation when exposed to Cd. However, both Se and Si promoted a significant reduction of deleterious effects of Cd. The Cd concentrations in the tissues were reduced in the presence of Se or Si. Plants treated with Cd together with Se or Si presented higher pigment content than those with only Cd, thus showing a reduction in the negative effects caused by this element. In the treatments in which Se and Si were added in the growth medium together with Cd, an activation of superoxide dismutase and guaiacol peroxidase enzymes was observed in the roots and shoot, which may have contributed to lower lipid peroxidation. Thus, Se and Si reduce Cd concentrations and have potential to ameliorate Cd toxicity in P. glomerata plants, which can be used to increase productivity and quality of medicinal plants.

The mutual restraint effect between the expansion of Alternanthera philoxeroides (Mart.) Griseb and cadmium mobility in aquatic environment.

Alternanthera philoxeroides (Mart.) Griseb is one of the most malignant weeds in its invision habitats. While in the cadmium-contaminated aquatic environment, does A. philoxeroides possess good tolerance and adaptability? To demonstrate the effects of cadmium on A. philoxeroides in the polluted water bodies, a hydroponic stress experiment was conducted over a gradient of Cd concentrations (0, 2.5 and 5mg/l) in triplicate. The seedlings were cultured in a greenhouse and harvested on days 0, 10, 20, 30 and 40, respectively. The results showed the effects of mutual restraint between Cd and A. philoxeroides. The A. philoxeroides seedlings were enriched with large amounts of Cd, and the toxicity of Cd inhibited the rapid growth of A. philoxeroides and induced the rapid degradation of chlorophylls in its tissues. Furthermore, the use of iron plaque effectively immobilized Cd of 1123-2883mg/kg·DW on the root surface, thus it decreased the transferability of Cd in the aquatic environment. Due to its extensive adaptability, good Cd tolerance and the immobilization of Cd predominantly in the roots (the highest Cd concentration enriched was 7588.65±628.90mg/kg·DW in roots). A. philoxeroides effectively restrained the translocation of Cd and partitioned Cd in the roots within water bodies. CAPSULE: The antagonistic effect exists between the invasion of A. philoxeroides and cadmium mobility in aquatic environments.

Higher Novel L-Cys Degradation Activity Results in Lower Organic-S and Biomass in Sarcocornia than the Related Saltwort, Salicornia.

Salicornia and Sarcocornia are almost identical halophytes whose edible succulent shoots hold promise for commercial production in saline water. Enhanced sulfur nutrition may be beneficial to crops naturally grown on high sulfate. However, little is known about sulfate nutrition in halophytes. Here we show that Salicornia europaea (ecotype RN) exhibits a significant increase in biomass and organic-S accumulation in response to supplemental sulfate, whereas Sarcocornia fruticosa (ecotype VM) does not, instead exhibiting increased sulfate accumulation. We investigated the role of two pathways on organic-S and biomass accumulation in Salicornia and Sarcoconia: the sulfate reductive pathway that generates Cys and l-Cys desulfhydrase that degrades Cys to H2S, NH3, and pyruvate. The major function of O-acetyl-Ser-(thiol) lyase (OAS-TL; EC 2.5.1.47) is the formation of l-Cys, but our study shows that the OAS-TL A and OAS-TL B of both halophytes are enzymes that also degrade l-Cys to H2S. This activity was significantly higher in Sarcocornia than in Salicornia, especially upon sulfate supplementation. The activity of the sulfate reductive pathway key enzyme, adenosine 5'-phosphosulfate reductase (APR, EC 1.8.99.2), was significantly higher in Salicornia than in Sarcocornia These results suggest that the low organic-S level in Sarcocornia is the result of high l-Cys degradation rate by OAS-TLs, whereas the greater organic-S and biomass accumulation in Salicornia is the result of higher APR activity and low l-Cys degradation rate, resulting in higher net Cys biosynthesis. These results present an initial road map for halophyte growers to attain better growth rates and nutritional value of Salicornia and Sarcocornia.

Moderate irrigation intervals facilitate establishment of two desert shrubs in the Taklimakan Desert Highway Shelterbelt in China.

BACKGROUND: Water influences various physiological and ecological processes of plants in different ecosystems, especially in desert ecosystems. The purpose of this study is to investigate the response of physiological and morphological acclimation of two shrubs Haloxylon ammodendron and Calligonum mongolicunl to variations in irrigation intervals. METHODOLOGY/PRINCIPAL FINDINGS: The irrigation frequency was set as 1-, 2-, 4-, 8- and 12-week intervals respectively from March to October during 2012-2014 to investigate the response of physiological and morphological acclimation of two desert shrubs Haloxylon ammodendron and Calligonum mongolicunl to variations in the irrigation system. The irrigation interval significantly affected the individual-scale carbon acquisition and biomass allocation pattern of both species. Under good water conditions (1- and 2-week intervals), carbon assimilation was significantly higher than other treatments; while, under water shortage conditions (8- and 12-week intervals), there was much defoliation; and under moderate irrigation intervals (4 weeks), the assimilative organs grew gently with almost no defoliation occurring. CONCLUSION/SIGNIFICANCE: Both studied species maintained similar ecophysiologically adaptive strategies, while C. mongolicunl was more sensitive to drought stress because of its shallow root system and preferential belowground allocation of resources. A moderate irrigation interval of 4 weeks was a suitable pattern for both plants since it not only saved water but also met the water demands of the plants.

Physiological Integration Affects Expansion of an Amphibious Clonal Plant from Terrestrial to Cu-Polluted Aquatic Environments.

The effects of physiological integration on clonal plants growing in aquatic and terrestrial habitats have been extensively studied, but little is known about the role in the extension of amphibious clonal plants in the heterogeneous aquatic-terrestrial ecotones, especially when the water environments are polluted by heavy metals. Ramets of the amphibious clonal herb Alternanthera philoxeroides were rooted in unpolluted soil and polluted water at three concentrations of Cu. The extension of populations from unpolluted terrestrial to polluted aqueous environments mainly relied on stem elongation rather than production of new ramets. The absorbed Cu in the ramets growing in polluted water could be spread horizontally to other ramets in unpolluted soil via physiological integration and redistributed in different organs. The performances of ramets in both terrestrial and aquatic habitats were negatively correlated with Cu intensities in different organs of plants. It is concluded that physiological integration might lessen the fitness of connected ramets in heterogeneously polluted environments. The mechanical strength of the stems decreased with increasing Cu levels, especially in polluted water. We suggest that, except for direct toxicity to growth and expansion, heavy metal pollution might also increase the mechanical risk in breaking failure of plants.

Effects of Cu Pollution on the Expansion of an Amphibious Clonal Herb in Aquatic-Terrestrial Ecotones.

Physiological integration can enhance the performance of clonal plants in aquatic and terrestrial heterogeneous habitats and associated ecotones. Similar to nutrients, pollutants may be transported among connected ramets via physiological integration. Few studies have examined the expansion of amphibious clonal plants from terrestrial to aquatic environments, particularly when the local water supply is polluted with heavy metals. A greenhouse experiment was conducted using the amphibious plant Alternanthera philoxeroides to determine whether Cu can spread among clonal plants and examine the corresponding effects of this pollution on the expansion of clonal plants in aquatic-terrestrial ecotones. Ramets from the same clonal fragments were rooted in unpolluted soil and polluted water at five different levels. The responses of the ramets in terrestrial and aquatic habitats were quantified via traits associated with growth, morphology and Cu accumulation. The results indicated that ramets in soil and water significantly differed in nearly all of these traits. The expansion of populations from terrestrial to polluted aquatic habitats was facilitated by stem elongation rather than new ramet production. The accumulated Cu in polluted ramets can be horizontally transported to other ramets in soil via connected stolons. In terms of clonal growth patterns, variations in Cu pollution intensity were negatively correlated with variations in the morphological and growth traits of ramets in polluted aquatic habitats and unpolluted soil. We concluded that Cu ions are distributed among the clones and accumulated in different ramet tissues in heterogeneous habitats. Therefore, we suggest that Cu pollution of aquatic-terrestrial ecotones, especially at high levels, can affect the growth and expansion of the whole clones because Cu ions are shared between integrated ramets.

Salt tolerance of Beta macrocarpa is associated with efficient osmotic adjustment and increased apoplastic water content.

The chenopod Beta macrocarpa Guss (wild Swiss chard) is known for its salt tolerance, but the mechanisms involved are still debated. In order to elucidate the processes involved, we grew wild Swiss chard exposed to three salinity levels (0, 100 and 200 mm NaCl) for 45 days, and determined several physiological parameters at the end of this time. All plants survived despite reductions in growth, photosynthesis and stomatal conductance in plants exposed to salinity (100 and 200 mm NaCl). As expected, the negative effects of salinity were more pronounced at 200 mm than at 100 mm NaCl: (i) leaf apoplastic water content was maintained or increased despite a significant reduction in leaf water potential, revealing the halophytic character of B. macrocarpa; (ii) osmotic adjustment occurred, which presumably enhanced the driving force for water extraction from soil, and avoided toxic build up of Na(+) and Cl(-) in the mesophyll apoplast of leaves. Osmotic adjustment mainly occurred through accumulation of inorganic ions and to a lesser extent soluble sugars; proline was not implicated in osmotic adjustment. Overall, two important mechanisms of salt tolerance in B. macrocarpa were identified: osmotic and apoplastic water adjustment.

Sensitivity of growth and biomass allocation patterns to increasing nitrogen: a comparison between ephemerals and annuals in the Gurbantunggut Desert, north-western China.

BACKGROUND AND AIMS: Biomass accumulation and allocation patterns are critical to quantifying ecosystem dynamics. However, these patterns differ among species, and they can change in response to nutrient availability even among genetically related individuals. In order to understand this complexity further, this study examined three ephemeral species (with very short vegetative growth periods) and three annual species (with significantly longer vegetative growth periods) in the Gurbantunggut Desert, north-western China, to determine their responses to different nitrogen (N) supplements under natural conditions. METHODS: Nitrogen was added to the soil at rates of 0, 0.5, 1.0, 3.0, 6.0 and 24.0 g N m(-2) year(-1). Plants were sampled at various intervals to measure relative growth rate and shoot and root dry mass. KEY RESULTS: Compared with annuals, ephemerals grew more rapidly, increased shoot and root biomass with increasing N application rates and significantly decreased root/shoot ratios. Nevertheless, changes in the biomass allocation of some species (i.e. Erodium oxyrrhynchum) in response to the N treatment were largely a consequence of changes in overall plant size, which was inconsistent with an optimal partitioning model. An isometric log shoot vs. log root scaling relationship for the final biomass harvest was observed for each species and all annuals, while pooled data of three ephemerals showed an allometric scaling relationship. CONCLUSIONS: These results indicate that ephemerals and annuals differ observably in their biomass allocation patterns in response to soil N supplements, although an isometric log shoot vs. log root scaling relationship was maintained across all species. These findings highlight that different life history strategies behave differently in response to N application even when interspecific scaling relationships remain nearly isometric.

Effect of Hg, As and Pb on biomass production, photosynthetic rate, nutrients uptake and phytochelatin induction in Pfaffia glomerata.

Plantlets of Pfaffia glomerata (Spreng.) were exposed for 28 days to three different metal/metalloid (Hg, Pb and As) with different levels (Hg 1; As 25, 50, 100 and Pb 100 and 400 µM) to analyze the possible phytochelatin initiation and affects on growth and photosynthetic pigments vis-à-vis metal accumulation potential of plants. The plantlets showed significant Hg, As and Pb accumulation in roots (150, 1267.67 and 2129 µg g(-1) DW respectively); however, a low root to shoot metal translocation was observed. It was interesting to note that all tested macronutrient (Mg, K, Ca) was higher in shoots and just opposite in case of micronutrients (Cu, Fe, Zn), was recorded highest in roots. The growth of plantlets (analyzed in terms of length and dry weight) was negatively affected by various metal treatments. In addition, the level of photosynthetic pigments alters significantly in response to all metal/metalloid treatment. In response to all tested metal/metalloids in plants only As induced phytochelatins (PC2, PC3 and PC4) in roots, and in shoots, GSH was observed in all tested metal/metalloids. In conclusion, P. glomerata plantlets could not cooperatively induce phytochelatins under any of Hg and Pb levels.

Ecophysiological adaptations of two halophytes to salt stress: photosynthesis, PS II photochemistry and anti-oxidant feedback--implications for resilience in climate change.

Halimione portulacoides and Sarcocornia fruticosa commonly exhibit a reddish coloration especially in high evaporation periods, due to betacyanin production in response to stress. Although sharing the same area in salt marshes, they present different strategies to overcome salinity stress. While S. fruticosa present a dilution strategy, increasing succulence, H. portulacoides appears to have developed an ionic compartmentalization strategy. Nevertheless, there's still a decrease in the photosynthetic activity in different extents. While in S. fruticosa, the impairment of photosynthetic activity is due to a decrease in the flow from the electron transport chain to the quinone pool; in H. portulacoides the process is affected far more early, with high amounts of energy dissipated at the PSII light harvesting centers. This photosynthetic impairment leads to energy accumulation and consequently to the production of reactive oxygen species (ROS). SOD was particularly active in stressed individuals, although this increment is rather more significant in S. fruticosa than in H. portulacoides suggesting that H. portulacoides may have a maximum salt concentration at which can sustain cellular balance between ROS production and scavenging. These different ecophysiological responses have great importance while evaluating the impacts climate change driven increase of sediment salinity on halophyte physiology and on the marsh community and ecosystem services.

Effect of vulculic acid produced by Nimbya alternantherae on the photosynthetic apparatus of Alternanthera philoxeroides.

The effect of the toxin vulculic acid produced by Nimbya alternantherae, on the photosynthetic apparatus of Alternanthera philoxeroides, was investigated via the photochemical activity and SDS-PAGE of protein on thylakoid membranes, fast chlorophyll a fluorescence transient measurements and the JIP-test. The electron transport rate of photosystem II (PSII), non-cyclic photophosphorylation activity, as well as the activity of chloroplast ATPase and Rubisco reduced significantly after vulculic acid treatment. Vulculic acid affected the O-J-I-P fluorescence induction kinetics, showing an increase of the parameters FV/FO, VK and VJ and a decrease of FO, FM, PIABS, φPo, ψEo, φEo, φRo, δRo and PItotal. In addition, it significantly decreased the amounts of major photosystem I (PSI) and PSII proteins. It is concluded that vulculic acid is a photosynthetic inhibitor with multiple action sites. The main targets are the light harvesting complex (LHC) and the oxygen evolving complex (OEC) on the PSII donor side. Vulculic acid blocks electron transport beyond QA and on the PSI acceptor side by digesting major PSI and PSII proteins.

[Subcellular distribution and phytotoxicity of cadmium in Alternanthera philoxeroides leaves].

A hydroponic experiment was conducted to study the subcellular distribution of Cd and mineral elements in Alternanthera philoxeroides leaves and the leaves anti-oxidative capacity and chlorophyll, soluble protein, and phytochelatins (PCs) contents under the stress of different concentration (0, 0.05, 0.1, 0.2, 0.4 mmol x L(-1)) Cd. With the increasing concentration of Cd in culture medium, the Cd content in all subcellular components of A. philoxeroides leaves increased significantly, and mainly distributed in cell wall, followed by in soluble fractions, and in chloroplast and mitochondria. When the Cd concentration in the medium exceeded 0.2 mmol x L(-1), the Cd was more allocated in soluble fractions than in cell wall. Cd stress resulted in an obvious imbalance of mineral elements uptake. With the increasing concentration of Cd, the Ca content in all subcellular components, especially in cell wall, increased significantly, whereas the P and K contents in cell wall and soluble fractions as well as the Mg and Fe contents in chloroplast decreased. In the meantime, the soluble protein and chlorophyll contents showed a decreasing trend, the glutathione and ascorbic acid contents decreased after an initial increase, the total anti-oxidative capacity (T-AOC) increased progressively, and the PCs accumulated in large quantity. These results suggested that A. philoxeroides had definite resistance to the water body Cd, and there was a dosage-effect relationship between the Cd enrichment in the subcellular components of A. philoxeroides leaves and the phytotoxicity of Cd. The imbalance of the mineral elements in subcellular components and the decrease of soluble protein and chlorophyll contents in chloroplast indicated the obvious phytotoxicity of Cd, while the massive accumulation of Ca in cell wall and the increased levels of PCs and T-AOC suggested the stronger resistance of A. philoxeroides to Cd stress. There was a definite correlation between the PCs production by A. philoxeroides and the toxicity of Cd, suggesting that the PCs could be considered as a sensitive biomarker for estimating the Cd phytotoxicity.

A transit peptide-like sorting signal at the C terminus directs the Bienertia sinuspersici preprotein receptor Toc159 to the chloroplast outer membrane.

Although Toc159 is known to be one of the key GTPase receptors for selective recognition of chloroplast preproteins, the mechanism for its targeting to the chloroplast surface remains unclear. To compare the targeting of these GTPase receptors, we identified two Toc159 isoforms and a Toc34 from Bienertia sinuspersici, a single-cell C4 species with dimorphic chloroplasts in individual chlorenchyma cells. Fluorescent protein tagging and immunogold studies revealed that the localization patterns of Toc159 were distinctive from those of Toc34, suggesting different targeting pathways. Bioinformatics analyses indicated that the C-terminal tails (CTs) of Toc159 possess physicochemical and structural properties of chloroplast transit peptides (cTPs). These results were further confirmed by fluorescent protein tagging, which showed the targeting of CT fusion proteins to the chloroplast surface. The CT of Bs Toc159 in reverse orientation functioned as a cleavable cTP that guided the fluorescent protein to the stroma. Moreover, a Bs Toc34 mutant protein was retargeted to the chloroplast envelope using the CTs of Toc159 or reverse sequences of other cTPs, suggesting their conserved functions. Together, our data show that the C terminus and the central GTPase domain represent a novel dual domain-mediated sorting mechanism that might account for the partitioning of Toc159 between the cytosol and the chloroplast envelope for preprotein recognition.

Tolerance and accumulation of copper in the salt-marsh shrub Halimione portulacoides.

The present study evaluated the tolerance and accumulation potential in the salt-marsh shrub Halimione portulacoides under moderate and high external Cu levels. A greenhouse experiment was conducted in order to investigate the effects of a range of external Cu concentrations (0 to 60 mmol l(-1)) on growth and photosynthetic performance by measuring gas exchange, chlorophyll fluorescence parameters and photosynthetic pigments. We also determined total copper, nitrogen, phosphorus and sulfur concentrations in the plant tissues. H. portulacoides survived with external Cu concentrations of up to 35 mmol Cu l(-1), although the excess of metal resulted in a biomass reduction of 48%. The effects of Cu on growth were linked to a drastic reduction in net photosynthesis. However, H. portulacoides tolerated Cu levels of up to 15 mmol Cu l(-1) without suffering adverse physiological effects. Our results indicate that this species could play an important role in the restoration of Cu-contaminated soils.

Compartmentalization and ultrastructural alterations induced by chromium in aquatic macrophytes.

The aim of the present study was to identify the sites of accumulation of Cr in the species of macrophytes that are abundant in the Cachoeira river, namely, Alternanthera philoxeroides, Borreria scabiosoides, Polygonum ferrugineum and Eichhornia crassipes. Plants were grown in nutritive solution supplemented with 0.25 and 50 mg l(-1) of CrCl(3)·6H(2)O. Samples of plant tissues were digested with HNO(3)/HCl in a closed-vessel microwave system and the concentrations of Cr determined using inductively-coupled plasma mass spectrometry (ICP-MS). The ultrastructure of root, stem and leaf tissue was examined using transmission electron microscopy (TEM) and secondary ion mass spectrometry (SIMS) in order to determine the sites of accumulation of Cr and to detect possible alterations in cell organelles induced by the presence of the metal. Chromium accumulated principally in the roots of the four macrophytes (8.6-30 mg kg(-1) dw), with much lower concentrations present in the stems and leaves (3.8-8.6 and 0.01-9.0 mg kg(-1) dw, respectively). Within root tissue, Cr was present mainly in the vacuoles of parenchyma cells and cell walls of xylem and parenchyma. Alterations in the shape of the chloroplasts and nuclei were detected in A. philoxeroides and B. scabiosoides, suggesting a possible application of these aquatic plants as biomarkers from Cr contamination.

Zinc alleviates mercury-induced oxidative stress in Pfaffia glomerata (Spreng.) Pedersen.

The possible role of zinc (Zn) to reverse the oxidative stress caused by mercury (Hg) was investigated in Pfaffia glomerata plantlets. Thirty-day-old acclimatized plantlets of P. glomerata were exposed to four treatments: control, 50 µM Zn, 50 µM Hg and 50 µM Zn + 50 µM Hg for 9 days. In Zn + Hg treatment, shoot and root Hg concentrations were 59 and 24% smaller than that plants exposed to 50 µM Hg added alone. An increase in the Zn concentration in the shoot of plants exposed to Zn + Hg occurred, although in the roots Zn concentration was not altered, when compared to the control. Fresh and dry weights, as well as the activity of δ-aminolevulinic acid dehydratase (δ-ALA-D) in Hg-treated plants were significantly reduced. Percentage survival, fresh and dry weights and δ-ALA-D activity of plants treated by 50 µM Zn + 50 µM Hg were greater than of that treated by Hg alone. Moreover, Zn treatment reduced the lipid peroxidation caused by Hg, being this effect related to increased root superoxide dismutase activity, and shoot catalase and ascorbate peroxidase activities. In conclusion, the presence of Zn in the substrate caused a significant reduction in the oxidative stress induced by Hg.

Lead induced responses of Pfaffia glomerata, an economically important Brazilian medicinal plant, under in vitro culture conditions.

Plantlets of Pfaffia glomerata (Spreng.) were exposed in vitro for 30 days to five lead levels (0-400 µM) to analyze the effects on growth and oxidative stress and responses of various antioxidants vis-à-vis lead accumulation. The plantlets showed significant lead accumulation in roots (1,532 µg g(-1) DW) with a low root to shoot lead translocation (ca. 3.6%). The growth of plantlets was negatively affected by various lead treatments, although the level of photosynthetic pigments did not alter significantly in response to any lead treatment. However, plantlets suffered from oxidative stress as suggested by the significant increase in malondialdehyde levels in root (8.48 µmol g(-1) FW) and shoot (3.20 µmol g(-1) FW) tissues with increasing lead treatments. In response to the imposed toxicity, increases in the activities of catalase in root (4.14 ∆E min(-1) mg(-1) protein) and shoot (3.46 ∆E min(-1) mg(-1) protein) and superoxide dismutase in root (345.32 units mg(-1) protein) and shoot (75.26 units mg(-1) protein), respectively, were observed, while the levels of non-protein thiols and ascorbic acid were not affected significantly in either roots or shoots.

Zinc compartmentation in Halimione portulacoides (L.) Aellen and some effects on leaf ultrastructure.

INTRODUCTION: The halophyte Halimione portulacoides collected in a polluted area of the river Sado estuary (Portugal) and obtained from hydroponic cultures was used to evaluate the compartmentation of Zn and its preferential binding sites. In parallel, we tried to assess if the minimum available Zn concentration found in marsh soil induces changes at the ultrastructural level. MATERIALS AND METHODS: A sequential extraction method was used to study the Zn compartmentation within the cell. Both dried plant samples and extracts/residues from compartmentation studies were digested by HNO3­HClO4 (4:1) until dryness and analyzed by atomic absorption spectrophotometry. Segments of young leaves, previously exposed to Zn were fixed in glutaraldehyde and osmium tetroxide. Ultrathin sections were stained and examined by transmission electron microscopy at 80 kV. RESULTS AND DISCUSSION: Proteins and carbohydrates of the cell walls constitute preferential binding sites of Zn, containing between 25% and 33% and between 30% and 40% of the total, respectively. Hydroponic plants accumulate Zn in their leaves up to (194 µg g−1) without visible damage or changes in the protein and chlorophyll concentrations, compared with the controls. Chlorenchyma chloroplasts of Zn-treated plants exhibited an unusual number of starch grains, which can be seen as an alert mechanism. CONCLUSIONS AND PERSPECTIVES: Although so far the levels of Zn in the leaves within the studied area have not reached high values, monitoring them remains a priority. Also, issues related with starch synthesis and organic ligands must be evaluated. The understanding of the predictable behavior of this halophyte is our main goal, and the results here presented can contribute to this achievement.