Spatial distribution, bioaccumulation profiles and risk for consumption of edible bivalves: a comparison among razor clam, Manila clam and cockles in the Venice Lagoon.

Despite their ecological and economical relevance, a lack of data is still occurring about the distribution, abundance, bioaccumulation and risks for consumption of some edible bivalves. The present study has been carried out in the Venice Lagoon taking into account three bivalve species to investigate i) the possible relationship among the biological features-distribution, abundance and bioaccumulation patterns- of razor clams (Solen marginatus Pulteney, 1799), Manila clams (Ruditapes philippinarum Adams and Reeve, 1850) and lagoon cockles (Cerastoderma glaucum Poiret, 1789) and the hydrological features, namely sediment physico-chemical characteristics and contamination; ii) their specific role as bioindicators of inorganic contamination; iii) the possible risks for human health associated with the consumption of these edible bivalves. Results showed that species distribution and abundance, especially for razor clams, was mainly influenced by environmental conditions and sediment granulometric composition, above metal(loid) contamination. The contamination patterns were different among species, as Manila clam generally showed higher bioaccumulation values for most of the metal(loid)s, whilst lagoon cockles preferentially accumulated Ni. Eventually, a serious concern exists for the human consumption, for all species and investigated sites regarding As. This study will raise attention on the effects of bioaccumulation of inorganic pollutants by edible bivalves and risks for consumers' safety, especially concerning razor clams and cockles, for which a critical lack of data on metal(loid) bioaccumulation occurs from the Northern Adriatic Sea.

Functional components of the bacterial CzcCBA efflux system reduce cadmium uptake and accumulation in transgenic tobacco plants.

Cadmium (Cd) is a toxic trace element released into the environment by industrial and agricultural practices, threatening the health of plants and contaminating the food/feed chain. Biotechnology can be used to develop plant varieties with a higher capacity for Cd accumulation (for use in phytoremediation programs) or a lower capacity for Cd accumulation (to reduce Cd levels in food and feed). Here we generated transgenic tobacco plants expressing components of the Pseudomonas putida CzcCBA efflux system. Plants were transformed with combinations of the CzcC, CzcB and CzcA genes, and the impact on Cd mobilization was analysed. Plants expressing PpCzcC showed no differences in Cd accumulation, whereas those expressing PpCzcB or PpCzcA accumulated less Cd in the shoots, but more Cd in the roots. Plants expressing both PpCzcB and PpCzcA accumulated less Cd in the shoots and roots compared to controls, whereas plants expressing all three genes showed a significant reduction in Cd levels only in shoots. These results show that components of the CzcCBA system can be expressed in plants and may be useful for developing plants with a reduced capacity to accumulate Cd in the shoots, potentially reducing the toxicity of food/feed crops cultivated in Cd-contaminated soils.

Assessment of phenolic herbicide toxicity and mode of action by different assays.

A phytotoxicity assay based on seed germination/root elongation has been optimized and used to evaluate the toxic effects of some phenolic herbicides. The method has been improved by investigating the influence of experimental conditions. Lepidium sativum was chosen as the most suitable species, showing high germinability, good repeatability of root length measurements, and low sensitivity to seed pretreatment. DMSO was the most appropriate solvent carrier for less water-soluble compounds. Three dinitrophenols and three hydroxybenzonitriles were tested: dinoterb, DNOC, 2,4-dinitrophenol, chloroxynil, bromoxynil, and ioxynil. Toxicity was also determined using the Vibrio fischeri Microtox® test, and a highly significant correlation was found between EC50 values obtained by the two assays. Dinoterb was the most toxic compound. The toxicity of hydroxybenzonitriles followed the order: ioxynil >bromoxynil >chloroxynil; L. sativum exhibited a slightly higher sensitivity than V. fischeri to these compounds. A QSAR analysis highlighted the importance of hydrophobic, electronic, and hydrogen-bonding interactions, in accordance with a mechanism of toxic action based on protonophoric uncoupling of oxidative phosphorylation. The results suggest that the seed germination/root elongation assay with L. sativum is a valid tool for the assessment of xenobiotic toxicity and can be recommended as part of a test battery.

The symbiosis between Nicotiana tabacum and the endomycorrhizal fungus Funneliformis mosseae increases the plant glutathione level and decreases leaf cadmium and root arsenic contents.

Over time, anthropogenic activities have led to severe cadmium (Cd) and arsenic (As) pollution in several environments. Plants inhabiting metal(loid)-contaminated areas should be able to sequester and detoxify these toxic elements as soon as they enter roots and leaves. We postulated here that an important role in protecting plants from excessive metal(loid) accumulation and toxicity might be played by arbuscular mycorrhizal (AM) fungi. In fact, human exploitation of plant material derived from Cd- and As-polluted environments may lead to a noxious intake of these toxic elements; in particular, a possible source of Cd and As for humans is given by cigarette and cigar smoke. We investigated the role of AM fungus Funneliformis mosseae (T.H. Nicolson & Gerd.) C. Walker & A. Schüßler in protecting Nicotiana tabacum L. (cv. Petit Havana) from the above-mentioned metal(loid) stress. Our findings proved that the AM symbiosis is effective in increasing the plant tissue content of the antioxidant glutathione (GSH), in influencing the amount of metal(loid)-induced chelators as phytochelatins, and in reducing the Cd and As content in leaves and roots of adult tobacco plants. These results might also prove useful in improving the quality of commercial tobacco, thus reducing the risks to human health due to inhalation of toxic elements contained in smoking products.

Proteomic analysis as a tool for investigating arsenic stress in Pteris vittata roots colonized or not by arbuscular mycorrhizal symbiosis.

Pteris vittata can tolerate very high soil arsenic concentration and rapidly accumulates the metalloid in its fronds. However, its tolerance to arsenic has not been completely explored. Arbuscular mycorrhizal (AM) fungi colonize the root of most terrestrial plants, including ferns. Mycorrhizae are known to affect plant responses in many ways: improving plant nutrition, promoting plant tolerance or resistance to pathogens, drought, salinity and heavy metal stresses. It has been observed that plants growing on arsenic polluted soils are usually mycorrhizal and that AM fungi enhance arsenic tolerance in a number of plant species. The aim of the present work was to study the effects of the AM fungus Glomus mosseae on P. vittata plants treated with arsenic using a proteomic approach. Image analysis showed that 37 spots were differently affected (21 identified). Arsenic treatment affected the expression of 14 spots (12 up-regulated and 2 down-regulated), while in presence of G. mosseae modulated 3 spots (1 up-regulated and 2 down-regulated). G. mosseae, in absence of arsenic, modulated 17 spots (13 up-regulated and 4 down-regulated). Arsenic stress was observed even in an arsenic tolerant plant as P. vittata and a protective effect of AM symbiosis toward arsenic stress was observed.

Design and dynamic simulation of minimal metallo-proteins.

Ab initio in silico design of proteins and enzymes has emerged as a powerful tool to design application-tailored proteins and catalysts for a wide range of applications. Several enzymes exploit the unique features of metal cofactors to achieve catalytic activity otherwise unattainable through the use of only natural amino acid residues. One of the major bottlenecks in ab initio design of novel proteins relies on long-range and epistatic effects that severely limit the possibility of a rational design. Within this framework there is an ongoing effort to reduce protein length and complexity to unlock the full potential of in silico protein design. In this work we specifically address this problem designing and investigating the dynamic features of 10 in silico designed minimal metallo-proteins. In particular, in this paper we investigate whether and to what extent it is possible to design a minimal metallo-enzyme made of only residues involved in metal binding. In this research we address these questions by investigating the ability of 10 different "mini-proteins" with a length shorter than 15 residues. Molecular dynamics studies clearly show that it is possible to design a minimal protein able to bind a metal atom with the correct geometry. It is noteworthy that designed mini-proteins cannot achieve the formation of a canonical hydrophobic core, rather the metal ion provides a "metal core" around which the entire protein is organized. This opens the possibility of designing synthetic enzymes composed of only functional residues organized around a "metal core" which acts as both structural and functional determinat.

Characterization of a former dump site in the Lagoon of Venice contaminated by municipal solid waste incinerator bottom ash, and estimation of possible environmental risk.

Bottom ash from a municipal solid waste incinerator on a former contaminated site, the island of Sacca San Biagio (Lagoon of Venice), was examined in order to evaluate levels of pollutants and their potential mobility and availability. Heavy metal concentrations were determined and the actual contamination of the site was compared with national legislation on polluted sites. The site was mainly contaminated by zinc, copper and lead. Physico-chemical characterization of bottom ash was carried out by SEM (Scanning Electron Microscopy) with micro-analysis by EDS (Energy Dispersive X-ray Spectroscopy) and XRD (X-ray Diffractometry), for information on newly formed minerals. SEM-EDS analysis revealed the presence of particles, compounds and clusters containing heavy metals and, in particular, the presence of barium sulfate, which was assumed to be a site-specific compound. Similarities between bottom ash and atmospheric PM10 collected on the adjacent island of Sacca Fisola were studied and a risk of aerodispersion of the fine fraction of ash was assumed. Lastly, in order to evaluate the potentially available fraction of metals (non-residual fraction) and the directly exchangeable fraction, two single extraction procedures with HCl and citric acid were carried out, respectively. Results showed a relatively low concentration of readily phyto-available metals, as well as the high concentrations found for some heavy metals (Cu, Pb, Zn) in the potentially mobilizable fraction.

Distribution of arsenic compounds in Mytilus galloprovincialis of the Venice lagoon (Italy).

Samples of Mytilus galloprovincialis collected in different sites of the Venice lagoon (Italy) were investigated for total arsenic concentrations by ICP-AES and for single arsenic species by HPLC-ICP-MS. For this purpose, an analytical procedure for the sensitive and efficient speciation of the arsenic species As(III), As(V), monomethylarsonic acid (MMA), dimethylarsinic acid (DMA), arsenobetaine (AB), arsenocholine (AC), and four arsenosugars was optimised. The total arsenic and the single arsenic species were determined in both the hepatopancreas (digestive gland) and the remaining soft tissues in order to verify the different arsenic accumulation in the body parts of mussels. Arsenic compounds were extracted from the mussels with a methanol/water mixture; the extracts were evaporated to dryness, redissolved in water, and chromatographed in an anion-exchange column, a Hamilton PRP-X100. Only small quantities or traces of inorganic arsenic were detected in the mussels. The majority of arsenic compounds detected in the extracts were organic species, with a predominance of arsenobetaine and of an arsenosugar. In addition, a greater arsenic accumulation in the digestive glands of mussels was observed.

Effects of copper on Sambucus nigra L. seedlings studied by electron paramagnetic resonance and atomic absorption spectroscopies.

We present in this work preliminary results on the translocation of copper, manganese and iron from soil to leaves of Sambucus nigra L. seedlings in the presence of increasing copper concentrations in the growth medium. The use of electron spin resonance spectroscopy and the comparison with atomic absorption spectroscopy is not presently common in the study of metal absorption in plants. This approach gave promising results both on the detection of some metal ions, as well as of radical species related to the presence of Cu2+ in plant tissues. Copper was found to act synergistically in manganese uptake.

Speciation of heavy metals in sediments of the lagoon of Venice collected in the industrial area.

In this study a procedure of sequential extraction of metals from sediments was applied to samples from the lagoon of Venice. Compared to the measurement of total concentrations, the knowledge of metal partitioning among sediment phases can help in estimating the risk posed by metal polluted sediments and give information on contamination sources and on the factors that influence metal enrichment in sediments. The results permitted to estimate for each metal the percentage of the total content that could be potentially remobilized from sediments, owing to changes in environmental conditions. The immediately available fraction represented in general a low proportion of the total content. An important proportion of most metals (except for Zn, Cd and Cu) was associated to residual phases and therefore was not expected to be released under the normal environmental conditions. The large amounts of metals bound to Fe-Mn oxides/hydroxides confirmed the strong scavenging efficiency of this phase. The carbonate phase was the preferential associative form for Pb, Cd and Zn, whereas Cu was bound in prevalence to the organic matter/sulphide phase.

Toxicity of tributyltin and triphenyltin to early life-stages of Paracentrotus lividus (Echinodermata: Echinoidea).

Sperm cell and embryo toxicity tests using the Mediterranean sea urchin Paracentrotus lividus were performed to assess the toxicity of tributyltin chloride, bis(tributyltin)oxide, triphenyltin acetate, and triphenyltin hydroxide. Toxicity values (mean effective concentration [EC50]) ranged from 2.97 to 18.5 microg/L for sperm cells and from 1.11 to 2.62 microg/L for embryos. For sperm cells, the toxicity of the two tributyl compounds was significantly greater than that of two triphenyl compounds; for embryos, the triphenyl compounds appeared to be more toxic. Study of embryotoxic effects highlighted closely concentration-dependent damages, the most sensitive stages corresponding to the crucial phases of differentiation (gastrula and prisma). Both EC50 and no-observed-effect concentration values for the four organotin compounds are similar to those reported in the literature for early life stages of other marine organisms.

Substituted aniline interaction with submitochondrial particles and quantitative structure-activity relationships.

The toxic effects of eighteen substituted anilines were determined by means of a short-term in vitro assay, using submitochondrial particles (SMP) as biosensors. The assay allows for the quantification of the effects of toxicants that act specifically on mitochondrial respiratory functions, like uncouplers and inhibitors, or non-specifically, by disturbing the structure and functioning of the inner mitochondrial membrane. The obtained EC(50) values range from 72.5 to 1910 micromol/l. The type and position of the substituents are of fundamental importance in determining the toxic potency. In general, the presence of electron-withdrawing substituents produces higher toxic effects, whereas electron-donating groups seem to reduce the toxicity. Quantitative structure-activity relationships (QSAR) showed that toxicity values were correlated with the Hammett sigma constant and with hydrogen bonding capacity descriptors, such as E(LUMO), E(HOMO) and Q(+). The results indicate that toxicity increases with increasing the hydrogen bonding donor capacity of the NH(2) group and support the hypothesis of a mechanism of action based on hydrogen bonding formation between the amino group of anilines and polar groups at the membrane/water interface. Such an interaction would cause a derangement of the membrane structure and, as a consequence, a disturbance of its functioning.