Potential of Castanea sativa for biomonitoring As, Hg, Pb, and Tl: A focus on their distribution in plant tissues from a former mining district.

Bioavailability of potentially toxic elements (PTEs) from the Earth's crust in the soil, e.g., As, Hg, Tl, and Pb, can pose a potential environmental and health risk because of human activities, especially related to mining extraction. The biomonitoring allows to detect PTE contamination through their measurement in living organisms as trees. However, the choice of which plant species and tissue to analyse is a key point to be evaluated in relation to PTE absorption and translocation. The aim of this work was to assess the As, Hg, Tl, and Pb distribution in Castanea sativa Mill. plant tissues, given its importance for both biomass and food production. The study identified two sites in the Alpi Apuane (Italy), with similar environmental conditions (e.g., elevation, exposure, forest type, and tree species) but different soil PTE levels. The topsoil was characterized, and the PTE fractions with different bioavailability were measured. The PTE concentrations were also analysed in chestnut plant tissues (leaves, bark, wood, nuts, and shells) in parallel with and evaluation of plant health status through the determination of micro and macronutrient concentrations and the leaf C and N isotope composition (δ13C or δ15N). Chestnut trees showed a good health status highlighting its suitability for Tl, As, Hg, and Pb biomonitoring, displaying a tissue-specific PTE allocation. Thallium and Hg were detected in all plant tissues at similar concentrations, As was found in leaves, wood, and nuts while Pb only in the bark. The δ15N negatively correlated with leaf Mn and Tl concentrations, suggesting possible changes in N source and/or plant metabolism due to the high contamination level and acid soil pH. Thallium in La Culla site trees was associated with its presence in the carbonate rocks but not in the topsoil, highlighting the potentiality of chestnut in providing valuable information for geochemical surveying.

Early Physiological, Cytological and Antioxidative Responses of the Edible Halophyte Chenopodium quinoa Exposed to Salt Stress.

Quinoa (Chenopodium quinoa Willd.) is a plant of South American origin recently valorized for its nutritional and nutraceutical properties in human diet. Quinoa is cultivated in many parts of the world, with a selection of varieties with good adaptability to extreme climatic conditions and salt stress. The variety Red Faro, native to southern Chile but harvested in Tunisia, was considered for salt stress resistance, considering its seed germination and 10-day seedling growth at increasing doses of NaCl (0, 100, 200 and 300 mM). Seedlings were spectrophotometrically analyzed for antioxidant secondary metabolites (polyphenols, flavonoids, flavonols and anthocyanins), antioxidant capacity (ORAC, oxygen radical absorbance capacity, DPPH*, 2,2-diphenyl-1-pic-rylhydrazyl), antioxidant enzyme activity (superoxide dismutase (SOD), guaiacol peroxidase (GPX), ascorbate peroxidase (APX) and catalase (CAT)) and mineral nutrient content in root and shoot tissues. Cytogenetic analysis of root tip was performed to check for meristematic activity and the possible presence of chromosomal abnormalities induced by salt stress. The results indicated a general increase in antioxidant molecules and antioxidant enzymes NaCl dose-dependent, no effects on seed germination but negative effects on seedling growth, and little effect on root meristems mitotic activity. These results indicated that stress conditions can induce an increase in biologically active molecules that could be used for nutraceutical purposes.

Synchrotron Radiation Spectroscopy and Transmission Electron Microscopy Techniques to Evaluate TiO2 NPs Incorporation, Speciation, and Impact on Root Cells Ultrastructure of Pisum sativum L. Plants.

Biosolids (Bs) for use in agriculture are an important way for introducing and transferring TiO2 nanoparticles (NPs) to plants and food chain. Roots of Pisum sativum L. plants grown in Bs-amended soils spiked with TiO2 800 mg/kg as rutile NPs, anatase NPs, mixture of both NPs and submicron particles (SMPs) were investigated by Transmission Electron Microscopy (TEM), synchrotron radiation based micro X-ray Fluorescence and micro X-ray Absorption Near-Edge Structure (µXRF/µXANES) and Inductively Coupled Plasma Optical Emission Spectrometry (ICP-OES). TEM analysis showed damages in cells ultrastructure of all treated samples, although a more evident effect was observed with single anatase or rutile NPs treatments. Micro-XRF and TEM evidenced the presence of nano and SMPs mainly in the cortex cells near the rhizodermis. Micro-XRF/micro-XANES analysis revealed anatase, rutile, and ilmenite as the main TiO2 polymorphs in the original soil and Bs, and the preferential anatase uptake by the roots. For all treatments Ti concentration in the roots increased by 38-56%, however plants translocation factor (TF) increased mostly with NPs treatment (261-315%) and less with SMPs (about 85%), with respect to control. In addition, all samples showed a limited transfer of TiO2 to the shoots (very low TF value). These findings evidenced a potential toxicity of TiO2 NPs present in Bs and accumulating in soil, suggesting the necessity of appropriate regulations for the occurrence of NPs in Bs used in agriculture.

Genotoxicity of the food additive E171, titanium dioxide, in the plants Lens culinaris L. and Allium cepa L.

E171 (titanium dioxide, TiO2), an authorized foods and beverage additive, is also used in food packaging and in pharmaceutical and cosmetic preparations. E171 is considered to be an inert and non-digestible material, not storable in animal tissues, but the possible presence of TiO2 nanoparticles (NP) may present a risk to human health and the environment. We determined the presence of 15% TiO2 NP in a commercial E171 food additive product, by electron microscopy. The biological effects of E171 were assessed in Lens culinaris and Allium cepa for the following endpoints: percentage of germination, root elongation, mitotic index, presence of chromosomal abnormalities, and micronuclei. The results indicated low phytotoxicity but dose-dependent genotoxicity. We also observed internalization of TiO2 NP and ultrastructural alterations in the root systems.

TiO2 nanoparticles in a biosolid-amended soil and their implication in soil nutrients, microorganisms and Pisum sativum nutrition.

The wide use of nanoparticles (NPs), gives concern about their possible negative implications in the environment and living organisms. In particular, titanium dioxide (TiO2) NPs are accumulated in biosolids (Bs) coming from wastewater treatment plants, which in turn are used as farm soil amendments and are becoming an important way of NPs entrance in the terrestrial ecosystems. In this study, to simulate a low and cumulative load of TiO2 NPs, 80 and 800 mg TiO2per Kg of soil were spiked in the Bs prior to its addition to soil. The effects of different crystal phases of TiO2 NPs (pure anatase and pure rutile or their mixture) and their non-coated bulk counterparts (larger particles) on the availability of mineral nutrients and on the status of the bacterial communities together with the nutritional status of Pisum sativum L. plants were evaluated. Results showed the reduction, to different extents, on the availability of important soil mineral nutrients (e.g. Mn 65%, Fe 20%, P 27%, averagely), in some cases size- (e.g. P) and dose-dependent. Bacterial biodiversity was also affected by the presence of high TiO2 dose in soil. The mineral nutrition of pea plants was also altered, showing the main reduction in Mn (80% in the roots and 50% in the shoots), K, Zn, P (respectively, 80, 40, and 35% in the roots), and an increase of N in the shoots, with possible consequences on the quality of the crop. The present study gives new integrated data on the effects of TiO2 NPs in the soil-plant system, on the soil health and on the nutritional quality of crops, rising new implications for future policies and human health.

An integrated approach to highlight biological responses of Pisum sativum root to nano-TiO2 exposure in a biosolid-amended agricultural soil.

This study focused on crop plant response to a simultaneous exposure to biosolid and TiO2 at micro- and nano-scale, being biosolid one of the major sink of TiO2 nanoparticles released into the soil environment. We settled an experimental design as much as possible realistic, at microcosm scale, using the crop Pisum sativum. This experimental design supported the hypotheses that the presence of biosolid in the farming soil might influence plant growth and metabolism and that, after TiO2 spiking, the different dimension and crystal forms of TiO2 might be otherwise bioavailable and differently interacting with the plant system. To test these hypotheses, we have considered different aspects of the response elicited by TiO2 and biosolid at cellular and organism level, focusing on the root system, with an integrative approach. In our experimental conditions, the presence of biosolid disturbed plant growth of P. sativum, causing cellular damages at root level, probably through mechanisms not only oxidative stress-dependent but also involving altered signalling processes. These disturbances could depend on non-humified compounds and/or on the presence of toxic elements and of nanoparticles in the biosolid-amended soil. The addition of TiO2 particles in the sludge-amended soil, further altered plant growth and induced oxidative and ultrastructural damages. Although non typical dose-effect response was detected, the most responsiveness treatments were found for the anatase crystal form, alone or mixed with rutile. Based on ultrastructural observations, we could hypothesise that the toxicity level of TiO2 nanoparticles may depend on the cell ability to isolate nanoparticles in subcellular compartments, avoiding their interaction with organelles and/or metabolic processes. The results of the present work suggest reflections on the promising practice of soil amendments and on the use of nanomaterials and their safety for food plants and living organisms.

Culture and horticulture: Protecting soil quality in urban gardening.

Urban cultivation for food production is of growing importance. The quality of urban soil can be improved by tillage and the incorporation of organic matter, or can be degraded by chemical treatments. Urban gardeners have a role in this process, through the selection of various cultivation techniques. Our study focuses on an allotment area in the town of Pisa (Italy), which since 1995 has been run as a municipal vegetable garden by the residents. We analysed the soil and compared the data with those collected five years previously, to verify the possible changes in soil properties and fertility. We also interviewed the gardeners regarding their backgrounds, motivations and cultivation practices. We looked for possible changes in the soil quality attributable to the cultivation techniques. We found that the allotment holders influenced the soil quality through the cultivation techniques. Organic carbon, electrical conductivity and the content of copper increased unevenly in relation to the gardeners' cultivation practices. At the same time the study highlights that the urban gardeners were not completely aware of how to protect and enhance the fertility and the quality of urban soil. We believe that town councils should be responsible for providing correct information to the allotment holders and thus prevent the possible misuse of urban soil to grow food, as this can affect everyone's health.

Overcoming limitation of "recalcitrant areas" to phytoextraction process: The synergistic effects of exogenous cytokinins and nitrogen treatments.

The aim of the present work was to test the efficiency of the phytoextraction process involving the use of exogenous phytohormone (cytokinins, CKs) and fertilizer (nitrogen, N) treatments in phytotechnologies to address risk management in "recalcitrant areas". The CKs and N treatments, alone or combined (CKs + N) in a Modulated Application (MA), were tested on the crop plant Helianthus annuus, common to Mediterranean area, fast growing and with high biomass production. Plants were grown on boron (B) contaminated sediments (collected from a geothermal area located in Tuscany (Italy). Plant growth, B uptake, together with plant stress parameters were investigated. Boron is easily taken up and translocated by some crop plants, but the high phytotoxicity can dramatically impact the plant growth and consequently the applicability and efficiency of the phytoextraction process. As indicators of plant stress, oxidative balance and photosynthetic parameters were investigated to give a deeper insight of phytotoxic mechanisms. Results showed that while each treatment (CKs and N alone) had significantly positive effects on plant health, the MA treatment provided a synergistic effect on morphological parameters and biomass production as a whole. After MA treatment, plants showed antioxidant activity comparable to that of the control (unpolluted sediments) and showed an increase of net photosynthesis. Moreover, our data showed very high values of B uptake and translocation (about 800 mg kg-1 in shoots), without any alteration triggered by the treatments (CKs and N alone or combined in MA). B phytoextraction resulted increased about fivefold with the MA treatments, while each treatment alone increased only two or three folds when treated with either CKs or N. The MA treatment is not "contaminant specific", so it could be applied in other "recalcitrant areas" where different types of contaminations occur, in order to overcome limitations of plant growth.

Evidence for the natural origins of anomalously high chromium levels in soils of the Cecina Valley (Italy).

The problem of high levels of chromium is one of the most important issues in soils of the Mediterranean area, in particular those deriving from ophiolitic parent materials. Very often the chromium concentration is greater than the threshold values of legislation on soil pollution and the knowledge of the origin of contamination (natural or anthropogenic) is important to formulate risk characterization. This study evaluated the soils from three coastal areas of the Cecina Valley (Tuscany, Italy) to understand the origin of chromium in the soils, where high levels of hexavalent chromium were found in well and spring waters of the areas. The main soil characteristics and the correlations among the values of chromium and nickel were determined. Chromium speciation was evaluated by synchrotron radiation X-ray absorption spectroscopy. The results showed the presence of only trivalent chromium in soil and a positive linear correlation between chromium and nickel (e.g. r = 0.76 for the Marina di Bibbona-Bolgheri area), corroborating the hypothesis of a geogenic origin of contamination. This hypothesis was also supported by the low CRI index for the soils with high total Cr content, indicating a higher presence of refractory minerals in the Marina di Bibbona-Bolgheri area than Cecina and Collemezzano areas. The refractory material found in soils was attributed to the presence of ophiolite outcrops in the surroundings and their sedimentary remnants. The weathering of ultramafic-derived constituents and their regional-scale transport are believed to be responsible for the enrichment of chromium and nickel in the investigated soils.

Nitrogen fertilizer improves boron phytoextraction by Brassica juncea grown in contaminated sediments and alleviates plant stress.

In this study we evaluated the effect of different fertilizer treatments on Brassica plants grown on boron-contaminated sediments. Experiments were conducted in the laboratory and on the lysimeter scale. At laboratory scale (microcosm), five different fertilizers were tested for a 35-d period. On the lysimeter scale, nitrogen fertilization was tested at three different doses and plants were allowed to grow until the end of the vegetative phase (70 d). Results showed that nitrogen application had effectively increased plant biomass production, while B uptake was not affected. Total B phytoextracted increased three-fold when the highest nitrogen dose was applied. Phytotoxicity on Brassica was evaluated by biochemical parameters. In plants grown in unfertilized B-contaminated sediments, the activity of antioxidant enzymes superoxide dismutase (SOD), ascorbate peroxidase (APX) and pyrogallol peroxidase (PPX) increased, whereas catalase (CAT) decreased with respect to control plants. Addition of N progressively mitigated the alteration of enzymatic activity, thus suggesting that N can aid in alleviating B-induced oxidative stress. SOD activity was restored to control levels just at the lowest N treatment, whereas the CAT inhibition was partially restored only at the highest one. N application also lowered the B-induced increase in APX and PPX activities. Increased glutathione reductase activity indicated the need to restore the oxidative balance of glutathione. Data also suggest a role of glutathione and phytochelatins in B defense mechanisms. Results suggest that the nitrogen fertilizer was effective in improving B phytoextraction by increasing Brassica biomass and by alleviating B-induced oxidative stress.

The effects of exogenous plant growth regulators in the phytoextraction of heavy metals.

The term "assisted phytoextraction" usually refers to the process of applying a chemical additive to contaminated soil in order to increase the metal uptake by crop plants. In this study three commercially available plant growth regulators (PGRs) based on cytokinins (CKs) were used to boost the assisted phytoextraction of Pb and Zn in contaminated soil collected from a former manufactured gas-plant site. The effects of EDTA treatment in soil and PGR treatment in leaves of Helianthus annuus were investigated in terms of dry weight biomass, Pb and Zn accumulation in the upper parts of the plants, Pb and Zn phytoextraction efficiency and transpiration rate. Metal solubility in soil and its subsequent accumulation in shoots were markedly enhanced by EDTA. The combined effects of EDTA and cytokine resulted in an increase in the Pb and Zn phytoextraction efficiency (up to 890% and 330%, respectively, compared to untreated plants) and up to a 50% increase in foliar transpiration. Our results indicate that exogenous PGRs based on CKs can positively assist the phytoextraction increasing the biomass production, the metal accumulation in shoots and the plant transpiration. The observed increase in biomass could be related to its action in stimulation of cell division and shoot initiation. On the other hand, the increase in metal accumulation in upper parts of plant could be related to both the role of PGRs in the enhancement of plant resistance to stress (as toxic metals) and the increase in transpiration rate, i.e. flux of water-soluble soil components and contaminants by the regulation of stomatal opening.

Combined application of Triton X-100 and Sinorhizobium sp. Pb002 inoculum for the improvement of lead phytoextraction by Brassica juncea in EDTA amended soil.

The process of EDTA-assisted lead phytoextraction from the Bovisa (Milan, Italy) brownfield soil was optimized in microcosms vegetated with Brassica juncea. An autochthonous plant growth-promoting rhizobacterium (PGPR), Sinorhizobium sp. Pb002, was isolated from the rhizosphere of B. juncea grown on the Pb-contaminated soil in presence of 2 mM EDTA. The strain was augmented (10(8) CFU g(-1) soil) in vegetated microcosms to stimulate B. juncea biomass production and, hence, its phytoextraction potential. Triton X-100 was also added to microcosms at 5 and 10 times the critical micelle concentration (cmc) to increase the permeability of root barriers to the EDTA-Pb complexes. Triton X-100 amendment determined an increase in Pb concentration within plant tissues. However it contextually exerted a phytotoxic effect. Sinorhizobium sp. Pb002 augmentation was crucial to plant survival in presence of both bioavailable lead and Triton X-100. The combination of the two treatments produced up to 56% increase in the efficiency of lead phytoextraction by B. juncea. The effects of these treatments on the structure of the soil bacterial community were evaluated by 16S rDNA denaturing gradient gel electrophoresis (DGGE).