Field controlled experiments on the physiological responses of maize (Zea mays L.) leaves to low-level air and soil mercury exposures

Thousands of tons of mercury (Hg) are released from anthropogenic and natural sources to the atmosphere in a gaseous elemental form per year, yet little is known regarding the influence of airborne Hg on the physiological activities of plant leaves. In the present study, the effects of low-level air and soil Hg exposures on the gas exchange parameters of maize (Zea mays L.) leaves and their accumulation of Hg, proline, and malondialdehyde (MDA) were examined via field open-top chamber and Hg-enriched soil experiments, respectively. Low-level air Hg exposures (<50 ng m^?3) had little effects on the gas exchange parameters of maize leaves during most of the daytime (p?>?0.05). However, both the net photosynthesis rate and carboxylation efficiency of maize leaves exposed to 50 ng m^?3 air Hg were significantly lower than those exposed to 2 ng m^?3 air Hg in late morning (p?<?0.05). Additionally, the Hg, proline, and MDA concentrations in maize leaves exposed to 20 and 50 ng m^?3 air Hg were significantly higher than those exposed to 2 ng m^?3 air Hg (p?<?0.05). These results indicated that the increase in airborne Hg potentially damaged functional photosynthetic apparatus in plant leaves, inducing free proline accumulation and membrane lipid peroxidation. Due to minor translocation of soil Hg to the leaves, low-level soil Hg exposures (<1,000 ng g^?1) had no significant influences on the gas exchange parameters, or the Hg, proline, and MDA concentrations in maize leaves (p?>?0.05). Compared to soil Hg, airborne Hg easily caused physiological stress to plant leaves. The effects of increasing atmospheric Hg concentration on plant physiology should be of concern.     

Effect of Cuscuta campestris parasitism on the physiological and anatomical changes in untreated and herbicide-treated sugar beet

The effects of field dodder on physiological and anatomical processes in untreated sugar beet plants and the effects of propyzamide on field dodder were examined under controlled conditions. The experiment included the following variants: N—noninfested sugar beet plants (control); I - infested sugar beet plants (untreated), and infested plants treated with propyzamide (1500 g a.i. ha^?1 (T₁) and 2000 g a.i. ha^?1(T₂)). The following parameters were checked: physiological—pigment contents (chlorophyll a, chlorophyll b, total carotenoids); anatomical -leaf parameters: thickness of epidermis, parenchyma and spongy tissue, mesophyll and underside leaf epidermis, and diameter of bundle sheath cells; petiole parameters: diameter of tracheid, petiole hydraulic conductance, xylem surface, phloem cell diameter and phloem area in sugar beet plants. A conventional paraffin wax method was used to prepare the samples for microscopy. Pigment contents were measured spectrophotometrically after methanol extraction. All parameters were measured: prior to herbicide application (0 assessment), then 7, 14, 21, 28 and 35 days after application (DAA). Field dodder was found to affect the pigment contents in untreated sugar beet plants, causing significant reductions. Conversely, reduction in the treated plants decreased 27% to 4% for chlorophyll a, from 21% to 5% for chlorophyll b, and from 28% to 5% for carotenoids (T₁). Also, in treatment T_2, reduction decreased in infested and treated plants from 19% to 2% for chlorophyll a, from 21% to 2% for chlorophyll b, from 23% to 3% for carotenoids and stimulation of 1% and 2% was observed 28 and 35 DAA, respectively. Plants infested (untreated) by field dodder had lower values of most anatomical parameters, compared to noninfested plants. The measured anatomical parameters of sugar beet leaves and petiole had significantly higher values in noninfested plants and plants treated with propyzamide than in untreated plants. Also, the results showed that propyzamide is an adequate herbicide for control of field dodder at the stage of early infestation.     

Leaf litter leachates have the potential to increase lifespan, body size, and offspring numbers in a clone of Moina macrocopa

Leaf litter processing is one major pathway of the global organic carbon cycle. During this process, a variety of small reactive organic compounds are released and transported to the aquatic environment, and may directly impact aquatic organisms as natural xenobiotics. We hypothesize that different forest stockings produce different leachate qualities, which in turn, stress the aquatic communities and, eventually, separate sensitive from tolerant species. Particularly, leachates from coniferous trees are suspected to have strongly adverse impacts on sensitive species. We exposed individuals of a clone of the model organism, Moina macrocopa, to comparable concentrations (approximately 2 mM) of litter leachates of Norway spruce, Picea abies, Colorado blue spruce, Picea pungens, black poplar, Populus nigra, and sessile oak, Quercus petraea. The animals were fed ad libitum. The following life trait variables were recorded: growth, lifespan, and lifetime offspring. To identify, whether or not exposure to litter leachates provokes an internal oxidative stress in the exposed animals we measured the superoxide anion radical scavenging capacity via photoluminescence. Except of P. abies, exposure to the leachates reduced this antioxidant capacity by approximately 50%. Leachate exposures, except that of Quercus, increased body size and extended lifespan; furthermore, particularly the leachates of both Picea species significantly increased the offspring numbers. This unexpected behavior of exposed Moina may be based on food supplements (e.g., high carbohydrate contents) in the leachates or on yet to be identified regulatory pathways of energy allocation. Overall, our results suggest that the potentially adverse effects of litter leachates can be overruled by either bacterial-growth supporting fractions in the leachates or an internal compensation mechanism in the Moina individuals.     

Effect of crude oil contamination on the chlorophyll content and morpho-anatomy of Cyperus brevifolius (Rottb.) Hassk

Chlorophyll plays a pivotal role in the plant physiology and its productivity. Cultivation of plants in crude oil contaminated soil has a great impact on the synthesis of chlorophyll pigment. Morpho-anatomy of the experimental plant also shows structural deformation in higher concentrations. Keeping this in mind, a laboratory investigation has been carried out to study the effect of crude oil on chlorophyll content and morpho-anatomy of Cyperus brevifolius plant. Fifteen-day-old seedling of the plant was planted in different concentrations of the crude oil mixed soil (i.e., 10,000, 20,000, 30,000, 40,000, and 50,000 ppm). A control setup was also maintained without adding crude oil. Results were recorded after 6 months of plantation. Investigation revealed that there is a great impact of crude oil contamination on chlorophyll content of the leaves of the experimental plant. It also showed that chlorophyll a, chlorophyll b, and total chlorophyll content of leaves grown in different concentrations of crude oil were found to be lower than those of the control plant. Further, results also demonstrated that chlorophyll content was lowest in the treatment that received maximum dose of crude oil. It also showed that chlorophyll content was decreased with increased concentration of crude oil. Results also demonstrated that there was a reduction in plant shoot and root biomass with the increase of crude oil concentration. Results also revealed that the shoot biomass is higher than root biomass. Morphology and anatomy of the experimental plant also show structural deformation in higher concentrations. Accumulation of crude oil on the cuticle of the transverse section of the leaves and shoot forms a thick dark layer. Estimation of the level of pollution in an environment due to oil spill is possible by the in-depth study of the harmful effects of oil on the morphology and anatomy and chlorophyll content of the plants grown in that particular environment.     

Biological effects of ammonia released from a composting plant assessed with lichens

In this study, we investigated whether ammonia emissions from industrial composting of organic waste may influence the surrounding environment, using lichens as bioindicators. To this purpose, samples of N-tolerant and N-sensitive lichens, namely Xanthoria parietina and Evernia prunastri, were transplanted for 1–3 months along transects at increasing distance (0–400 m) from a composting facility in Tuscany, Italy. Atmospheric concentrations of ammonia were measured using passive samplers. The physiological response of lichen transplants was investigated by means of the photosynthetic efficiency (measured as chlorophyll a fluorescence emission), the integrity of cell membranes (measured as electrolyte leakage), and sample viability (measured as enzymatic activity of dehydrogenase). Epiphytic lichen communities were investigated using biodiversity indices. The results showed decreasing concentrations of ammonia, from 48.7 μg/m³ at the composting facility to 2.7 μg/m³ at 400 m. The N-tolerant X. parietina was not affected and some physiological parameters even showed a higher performance, while the N-sensitive E. prunastri showed a reduced performance with increasing atmospheric concentrations approaching the source. A shift from lichen communities composed by meso-acidophilous species (actual condition) to more nitrophilous communities in the near future, approaching the composting facility is suggested. It is concluded that lichens can provide useful data for decision-makers to establish correct science-based environmentally sustainable waste management policies.     

Effects of cadmium on the photosynthetic activity in mature and young leaves of soybean plants

Cadmium (Cd) is a widely spread pollutant and can be easily taken up by crop from soil, resulting in a serious health issue for humans. The objective of this study was to comparatively investigate the photosynthetic activity, chlorophyll a fluorescence, chlorophyll contents, and spectral reflectance in mature and young leaves of soybean plants after being treated with different concentrations of Cd for 10 days. The photosynthetic rate, chlorophyll contents, actual photochemical efficiency of PSII, and photochemical quenching in the young leaves decreased more significantly with increasing concentrations of Cd in the nutrient solution, compared with those in the mature leaves, though the young leaves had less Cd concentrations. Thus, there was more excessive excited energy produced in the young leaves than that in the mature leaves. In the young leaves, due to more excessive excited energy, more reactive oxygen species may be generated, which further damaged the photosynthetic apparatus. It was supported by the fact that the decrease of reflectance in near-infrared wavelengths of the young leaves was more noticeable than that of the mature leaves. In addition, the chlorophyll a fluorescence transients of the young leaves was significantly different from that in the mature leaves, indicating that the electron transport of young leaves were inhibited much more severely than that of the mature leaves. These observations imply that the responses of photosynthetic activity of soybean leaves to Cd stress depend on their growth stage, and the Cd-induced inhibition of photosynthetic activity might be attributed to the decrease in chlorophyll contents and the decrease in mesophyll CO₂ assimilation ability cause by the Cd, which further decreased the consumption of ATP and NADPH, leading to accumulation of NADPH on the acceptor sides of the PSI, and then feedback inhibited electron transport in chloroplasts.     

Physiological effects of tetracycline antibiotic pollutants on non-target aquatic Microcystis aeruginosa

This study aimed to evaluate the aquatic toxicity of three typical tetracycline antibiotics, including tetracycline, oxytetracycline, and chlortetracycline, on the cyanobacterium Microcystis aeruginosa. The cell density, chlorophyll a content, protein content, and enzymatic antioxidant activities were determined. The results showed that the cell growth was significantly inhibited by the three compounds at a low concentration. The chlorophyll a and protein content decreased significantly after exposure to 0.05 mg L^?1 of each compound for 9 d. When exposed to 0.2–1 mg L^?1 of tetracycline, the superoxide dismutase (SOD) activity increased, but peroxidase (POD) and catalase (CAT) activities decreased. In contrast, when exposed to oxytetracycline and chlortetracycline at different concentrations ranging from 0.2 to 1 mg L^?1 and from 0.01 to 0.05 mg L^?1, the SOD activity decreased, but the POD and CAT activities increased. These findings indicate that tetracycline antibiotics influence cell growth and protein synthesis, and they also induce oxidative stress in M. aeruginosa at environmentally similar concentrations. Thus, this study may provide further insights into the toxic effects of tetracycline antibiotics and the controlled use of antibiotics.     

The linear accumulation of atmospheric mercury by vegetable and grass leaves: Potential biomonitors for atmospheric mercury pollution

One question in the use of plants as biomonitors for atmospheric mercury (Hg) is to confirm the linear relationships of Hg concentrations between air and leaves. To explore the origin of Hg in the vegetable and grass leaves, open top chambers (OTCs) experiment was conducted to study the relationships of Hg concentrations between air and leaves of lettuce (Lactuca sativa L.), radish (Raphanus sativus L.), alfalfa (Medicago sativa L.) and ryegrass (Lolium perenne L.). The influence of Hg in soil on Hg accumulation in leaves was studied simultaneously by soil Hg-enriched experiment. Hg concentrations in grass and vegetable leaves and roots were measured in both experiments. Results from OTCs experiment showed that Hg concentrations in leaves of the four species were significantly positively correlated with those in air during the growth time (p?<?0.05), while results from soil Hg-enriched experiment indicated that soil-borne Hg had significant influence on Hg accumulation in the roots of each plant (p?<?0.05), and some influence on vegetable leaves (p?<?0.05), but no significant influence on Hg accumulation in grass leaves (p?>?0.05). Thus, Hg in grass leaves is mainly originated from the atmosphere, and grass leaves are more suitable as potential biomonitors for atmospheric Hg pollution. The effect detection limits (EDLs) for the leaves of alfalfa and ryegrass were 15.1 and 22.2 ng g^–1, respectively, and the biological detection limit (BDL) for alfalfa and ryegrass was 3.4 ng m^–3.     

Assessment of silver nanoparticle-induced physiological and molecular changes in Arabidopsis thaliana

In this study, the effect of silver nanoparticles and silver ions on Arabidopsis thaliana was investigated at physiological and molecular levels. The seedlings were grown in sublethal concentrations of silver nanoparticles and silver ions (0.2, 0.5, and 1 mg/L) in 1/4 Hoagland’s medium for 14 days under submerged hydroponic conditions. Significantly higher reduction in the total chlorophyll and increase in anthocyanin content were observed after exposure to 0.5 and 1 mg/L silver nanoparticles as compared to similar concentrations of silver ions. Lipid peroxidation increased significantly after exposure to 0.2, 0.5, and 1 mg/L of silver nanoparticles and 0.5 and 1 mg/L of silver ions. Qualitative analysis with dichloro-dihydro-fluorescein diacetate and rhodamine 123 fluorescence showed a dose-dependent increase in reactive oxygen species production and changes in mitochondrial membrane potential in the roots of seedlings exposed to different concentrations of silver nanoparticles. Real-time PCR analysis showed significant upregulation in the expression of sulfur assimilation, glutathione biosynthesis, glutathione S-transferase, and glutathione reductase genes upon exposure to silver nanoparticles as compared with silver ions. Overall, based on the physiological and molecular level responses, it was observed that exposure to silver nanoparticles exerted more toxic response than silver ions in A. thaliana.     

Evaluation of growth and biochemical indicators of Salvinia natans exposed to zinc oxide nanoparticles and zinc accumulation in plants

The adverse effects of zinc oxide nanoparticles (ZnO NPs) with an average diameter of 25 nm on the aquatic plant Salvinia natans (L.) All. were determined. Growth, superoxide dismutase (SOD) activity, catalase (CAT) activity, peroxidase activity, and chlorophyll content of the plants were measured after 7 days of exposure to different concentrations of ZnO NPs (1 to 50 mg L^?1). The particle distribution in the culture medium (without plants) during the first 24 h was determined using a Nanotrac 250 particle analyzer. We also investigated the zinc accumulation in leaves and roots of the plant after 7 days of exposure. Exposure to 50 mg L^?1 ZnO NPs significantly increased SOD and CAT activities (P?<?0.05) and significantly depressed photosynthetic pigments (P?<?0.05). However, plant growth was not significantly affected (P?>?0.05). NPs completely precipitated at the bottom of the container at 8 h except for the portions of dissolution and aggregation on the roots. ZnO NPs at a concentration of 50 mg L^?1 can adversely affect S. natans, and their stress is affected by their aggregation and dissolution.     

Polycyclic aromatic hydrocarbon contamination in an urban area assessed by Quercus ilex leaves and soil

We investigated the PAH contamination of Naples urban area, densely populated and with high traffic flow, by analyses of environmental matrices: soil and Quercus ilex leaves. Being some PAHs demonstrated to have hazardous effects on human health, the accumulation of carcinogenic and toxic PAHs (expressed as B(a)Peq) was evaluated in the leaves and soil. The main sources of the PAHs were discriminated by the diagnostic ratios in the two matrices. The urban area appeared heavily contaminated by PAHs, showing in soil and leaves total PAH concentrations also fivefold higher than those from the remote area. The soil mainly accumulated heavy PAHs, whereas leaves the lightest ones. Median values of carcinogenic PAH concentrations were higher in soil (440 ng g^?1 d.w.) and leaves (340 ng g^?1 d.w.) from the urban than the remote area (60 and 70 ng g^?1 d.w., respectively, for soil and leaves). Also, median B(a)Peq concentrations were higher both in soil and leaves from the urban (137 and 63 ng g^?1 d.w., respectively) than those from the remote area (19 and 49 ng g^?1 d.w., respectively). Different from the soils, the diagnostic ratios found for the leaves discerned PAH sources in the remote and urban areas, highlighting a great contribution of vehicular traffic emission as main PAH source in the urban area.     

Feeding preferences of two detritivores related to size and metal content of leaves: the crustaceans Atyaephyra desmarestii (Millet) and Echinogammarus meridionalis (Pinkster)

The equilibrium of the structure and functioning of freshwater ecosystems is dependent of detritivores that link all the other functional groups. The preference for feeding leaves with different diameters (particle size) and leaves with metal contamination (several concentrations of the essential metals copper and zinc) were determined for two detritivores, the decapod Atyaephyra desmarestii and the amphipod Echinogammarus meridionalis. Several no-choice and multi-choice assays were done to determinate which leaf diameter the amphipod and the decapod species would eat when they had or not had alternatives available and include a set of dual-choice assays with contaminated and uncontaminated foods. No significant preference was shown by either species relative to the diameter of leaves, either on no-choice or multi-choice assays. The presence of essential metals on food did not had any influence on the feeding choice of these organisms over the concentration range studied. Both showed no preference on ingesting food spiked with these essential metals, except E. meridionalis which preferred ingesting leaves with 2.19 μg.l^?1 of copper instead of uncontaminated leaves. For further works, despite no preference for leaves with a certain diameter, the leaves with 0.70 cm (0.385 cm²of area) and with 0.50 cm (1.767 cm² of area) should be used for A. desmarestii and E. meridionalis, respectively. Furthermore, to maintain E. meridionalis, the diet should include some percentage of copper in order to accomplish metabolic needs.     

Physiological characteristics of <Emphasis Type="Italic">Plantago major</Emphasis> under SO<Subscript>2</Subscript> exposure as affected by foliar iron spray

Sulfur dioxide (SO₂) is considered as a main air pollutant in industrialized areas that can damage vegetation. In the present study, we investigated how exposure to SO₂ and foliar application of iron (Fe) would affect certain physiological characteristics of Plantago major. The plant seedlings exposed or unexposed to SO₂ (3900 μg m^?3) were non-supplemented or supplemented with Fe (3 g L^?1) as foliar spray. Plants were exposed to SO₂ for 6 weeks in 100 × 70 × 70 cm chambers. Fumigation of plants with SO₂ was performed for 3 h daily for 3 days per week (alternate day). Lower leaf Fe concentration in the plants exposed to SO₂ at no added Fe treatment was accompanied with incidence of chlorosis symptoms and reduced chlorophyll concentration. No visible chlorotic symptoms were observed on the SO₂-exposed plants supplied with Fe that accumulated higher Fe in their leaves. Both at with and without added Fe treatments, catalase (CAT) and peroxidase (POD) activity was higher in the plants fumigated with SO₂ in comparison with those non-fumigated with SO₂. Foliar application of Fe was also effective in increasing activity of antioxidant enzymes CAT and POD. Exposure to SO₂ led to reduced cellulose but enhanced lignin content of plant leaf cell wall. The results obtained showed that foliar application of Fe was effective in reducing the effects of exposure to SO₂ on cell wall composition. In contrast to SO₂, application of Fe increased cellulose while decreased lignin content of the leaf cell wall. This might be due to reduced oxidative stress induced by SO₂ in plants supplied with Fe compared with those unsupplied with Fe.     

Altered physiology, cell structure, and gene expression of Theobroma cacao seedlings subjected to Cu toxicity

Seedlings of Theobroma cacao CCN 51 genotype were grown under greenhouse conditions and exposed to increasing concentrations of Cu (0.005, 1, 2, 4, 8, 16, and 32 mg Cu L^?1) in nutrient solution. When doses were equal or higher than 8 mg Cu L^?1, after 24 h of treatment application, leaf gas exchange was highly affected and changes in chloroplasts thylakoids of leaf mesophyll cells and plasmolysis of cells from the root cortical region were observed. In addition, cell membranes of roots and leaves were damaged. In leaves, 96 h after treatments started, increases in the percentage of electrolyte leakage through membranes were observed with increases of Cu in the nutrient solution. Moreover, there was an increase in the concentration of thiobarbituric acid-reactive substances in roots due to lipid peroxidation of membranes. Chemical analysis showed that increases in Cu concentrations in vegetative organs of T. cacao increased with the increase of the metal in the nutrient solution, but there was a greater accumulation of Cu in roots than in shoots. The excess of Cu interfered in the levels of Mn, Zn, Fe, Mg, K, and Ca in different organs of T. cacao. Analysis of gene expression via RTq-PCR showed increased levels of MT2b, SODCyt, and PER-1 expression in roots and of MT2b, PSBA, PSBO, SODCyt, and SODChI in leaves. Hence, it was concluded that Cu in nutrient solution at doses equal or above 8 mg L^?1 significantly affected leaf gas exchange, cell ultrastructure, and transport of mineral nutrients in seedlings of this T. cacao genotype.     

Effects of copper sulfate on growth and physiological responses of Limoniastrum monopetalum

A glasshouse study of the coastal shrub Limoniastrum monopetalum was carried out to evaluate its tolerance and capacity to accumulate copper. We investigate the effects of Cu from 0 to 60 mmol l^?1 on the growth, photosynthetic apparatus, and nutrient uptake of L. monopetalum, by measuring gas exchange, chlorophyll fluorescence parameters, photosynthetic pigments, and total copper, nitrogen, phosphorus, sulfur, calcium, and magnesium content in the plant tissues. Although L. monopetalum did not survive at 60 mmol l^?1 Cu, the species demonstrated a high tolerance to Cu-induced stress, since all plants survived external Cu concentrations of up to 35 mmol l^?1 and displayed similar growth in the Cu-enriched medium as in the control treatment of up to the external level of 15 mmol Cu l^?1 (1,000 mg Cu l^?1). The reduced growth registered in plants exposed to 35 mmol Cu l^?1 can be attributed to reduced photosynthetic carbon assimilation associated with the adverse effect of the metal on the photochemical apparatus and a reduction in the absorption of essential nutrients. Copper tolerance was associated with the capacity of the plant to accumulate the metal in its roots and effectively prevent its translocation to photosynthetic tissues. L. monopetalum has the characteristics of a Cu-excluder plant and could be used in the revegetation of Cu-contaminated soils.     

Cadmium tolerance in six poplar species

Selection of poplar species with greater Cd tolerance and exploiting the physiological mechanisms involved in Cd tolerance are crucial for application of these species to phyto-remediation. The aim of this study is to investigate variation in Cd tolerance among the six poplar species and its underlying physiological mechanisms. Cuttings of six Populus species were cultivated for 10 weeks before exposure to either 0 or 200 μM CdSO₄ for 20 days. Gas exchange in mature leaves was determined by a portable photosynthesis system. Cd concentrations in tissues were analyzed by a flame atomic absorbance spectrometry. Subsequently, Cd amount per plant, bio-concentration factor (BCF) and translocation factor (T _f) were calculated. Nonenzymatic compounds and activities of antioxidative enzymes in tissues were analyzed spectrophotometrically. Cd exposure caused decline in photosynthesis in four poplar species including Populus cathayana (zhonghua 1). Among the six species, P. cathayana (zhonghua 1) displayed the highest Cd concentrations in tissues, the largest Cd amount in aerial parts, the highest BCF in aerial parts and T _f under Cd exposure. Under Cd stress, increases in total soluble sugars in roots but decreases in starch in roots, wood, and leaves of P. cathayana (zhonghua 1) were found. Induced O ₂ ^?? and H₂O₂ production in roots and leaves, and increases in free proline, soluble phenolics, and activities of antioxidative enzymes were observed in P. cathayana (zhonghua 1). Based on results of this pot experiment, it is concluded that P. cathayana (zhonghua 1) is superior to other five species for Cd phyto-remediation, and its well-coordinated physiological changes under Cd exposure confer the great Cd tolerance of this species.     

Ozone exposure induces the activation of leaf senescence-related processes and morphological and growth changes in seedlings of Mediterranean tree species

Four Mediterranean tree taxa, Quercus ilex subsp. ilex, Quercus ilex subsp. ballota, Olea europaea cv. vulgaris and Ceratonia siliqua, were exposed to different ozone (O(3)) concentrations in open top chambers (OTCs) during 2 years. Three treatments were applied: charcoal-filtered air (CF), non-filtered air (NF) and non-filtered air plus 40 ppb(v) of O(3) (NF +). The photochemical maximal efficiency, Fv/Fm, decreased in NF + plants during the second year of exposure, especially during the most stressful Mediterranean seasons (winter and summer). An increase of delta(13)C was found in three of the four studied species during the first year of exposure. This finding was only maintained in C. siliqua during the second year. Decreases in the chlorophyll content were detected during the first year of fumigations in all the species studied, but not during the second year. The NF + treatment induced changes in foliar anatomical characteristics, especially in leaf mass per area (LMA) and spongy parenchyma thickness, which increased in some species. A reduction in N content and an increase in delta(15)N were found in all species during the second year when exposed in the NF + OTCs, suggesting a change in their retranslocation pattern linked to an acceleration of leaf senescence, as also indicated by the above mentioned biochemical and anatomical foliar changes. The two Q. ilex subspecies were the most sensitive species since the changes in N concentration, delta(15)N, chlorophyll, leaf area, LMA and biomass occurred at ambient O(3) concentrations. However, C. siliqua was the most responsive species (29% biomass reduction) when exposed to the NF + treatment, followed by the two Q. ilex subspecies (14-20%) and O. europaea (no significant reduction). Ozone resistance of the latter species was linked to some plant traits such as chlorophyll concentrations, or spongy parenchyma thickness.     

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 HNO₃?CHClO₄ (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 ??g 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.     

Experimental exposure of juvenile savannah monitors (Varanus exanthematicus) to an environmentally relevant mixture of three contaminants: effects and accumulation in tissues

Using varanids as indicators of pollution in African continental wetlands was previously proposed. The present study aimed at understanding experimentally how monitors absorb and accumulate pollutants and how they are affected. The relevance of non-destructive sampling was also evaluated. Savannah monitors (Varanus exanthematicus) were orally exposed during 6 months to a mixture of lead, 4,4′-dichlorodiphenyltrichloroethane (4,4′-DDT) and chlorpyrifos-ethyl (CPF) or to the vehicle only. Proportionally to their mass, exposed monitors received the same dose: 20 then 10 mg lead?kg^?1, 2 then 0.5 mg CPF?kg^?1 and 4 mg 4,4′-DDT?kg^?1. Individuals surviving contamination were euthanized after 4 or 6 months of experiment. Tissues were analysed for lead by atomic absorption spectrophotometry and for DDT and CPF by gas chromatography. Exposed monitors absorbed all three pollutants but only lead (essentially in bone, tail tips and phalanxes) and 4,4′-DDT plus its main metabolites (essentially in fat and liver) accumulated. CPF killed ten individuals. Clear correlations occurred between the total quantity of lead or 4,4′-DDT administered and concentrations in tissues. Tail tips and skin samples are recommended non-destructive indicators for lead and organochlorine pesticides contamination, respectively. This work confirms that monitors can be used as relevant indicators of environmental pollution by lead and organochlorine pesticides. Although varanids withstand heavy lead and DDT contamination, our results suggest that CPF can be lethal at very low doses to the herpetofauna and emphasize the importance of considering all taxa in impact assessment studies, including reptiles.