Selenium bioavailability and uptake as affected by four different plants in a loamy clay soil with particular attention to mycorrhizae inoculated ryegrass.

The aim of this study was to investigate the influence of plant species, especially of their rhizosphere soil, and of inoculation with an arbuscular mycorrhizal (AM) fungus on the bioavailability of selenium and its transfer in soil-plant systems. A pot experiment was performed with a loamy clay soil and four plant species: maize, lettuce, radish and ryegrass, the last one being inoculated or not with an arbuscular mycorrhizal fungus (Glomus mosseae). Plant biomass and Se concentration in shoots and roots were estimated at harvest. Se bioavailability in rhizosphere and unplanted soil was evaluated using sequential extractions. Plant biomass and selenium uptake varied with plant species. The quantity of rhizosphere soil also differed between plants and was not proportional to plant biomass. The highest plant biomass, Se concentration in plants, and soil to plant transfer factor were obtained with radish. The lowest Se transfer factors were obtained with ryegrass. For the latter, mycorrhizal inoculation did not significantly affect plant growth, but reduced selenium transfer from soil to plant by 30%. In unplanted soil after 65 days aging, more than 90% of added Se was water-extractable. On the contrary, Se concentration in water extracts of rhizosphere soil represented less than 1% and 20% of added Se for ryegrass and maize, respectively. No correlation was found between the water-extractable fraction and Se concentration in plants. The speciation of selenium in the water extracts indicated that selenate was reduced, may be under organic forms, in the rhizosphere soil.

Association constants of Pb2+ with binding sites of fungal biomass using metal-based titrations.

Biosorption is perceived as an alternative method for toxic heavy metal removal/recovery from aqueous effluents. This work focused on derivation of some key quantitative physico-chemical characteristics of a representative biosorbent material required for its further effective exploitation. The newly developed acid-base and metal-based titrator allowed the characterization of the chemisorption active sites of Rhizopus arrhizus biomass and the study of their metal affinity. This experimental approach, combined with an analytical method consisting of transforming the initial data enabled the calculation of the number and capacity of the reactive sites (Qads) and the metal affinity constants (Km) for lead sorption by R. arrhizus biomass. The pKm values for Rhizopus biomass varied between -3 and -6 for sites releasing no protons, -1 and 1 for sites releasing one proton, and > 8 for sites releasing two protons - combined with the Pb precipitation phenomenon. At low temperatures, the active binding site number was lower at lower lead concentrations whereas the precipitation was promoted at higher lead concentration values. Lead adsorption contributed modestly (11%) to its overall uptake and occurred at low lead concentrations onto strong and medium affinity binding sites. Micro-precipitation quickly commenced around active binding sites distinguished by their weak affinity whenever the solution lead concentrations reached 10(-6) or 10(-5) M and represented more than 85% of the total sorbed metal quantity. The work also demonstrated the usefulnes of the methodology reported here for characterizing complex biosorbent materials.

PAH dissipation in a contaminated river sediment under oxic and anoxic conditions.

A batch experiment was conducted to compare PAH degradation in a polluted river sediment under aerobic and anaerobic conditions, and to investigate whether input of fresh organic material (cellulose) could enhance such degradation. All measurements were checked against abiotic control treatments to exclude artifacts of sample preparation and non-biological processes like aging. Three- and four-ring PAHs could be degraded by the indigenous microbial community under aerobic conditions, but anaerobic metabolism based on iron and sulphate reduction was not coupled with PAH degradation of even the simplest 3-ring compounds like phenanthrene. Cellulose addition stimulated both aerobic and anaerobic respiration, but had no effect on PAH dissipation. We conclude that natural attenuation of PAHs in polluted river sediments under anaerobic conditions is exceedingly slow. Dredging and biodegradation on land under aerobic conditions would be required to safely remediate and restore polluted sites.

Distribution and location of polycyclic aromatic hydrocarbons (PAHs) and PAH-degrading bacteria within polluted soil aggregates.

A study was conducted to determine the location and distribution of PAH and PAH-degrading bacteria in different aggregate size fractions of an industrially polluted soil. The estimation of PAH-degrading bacteria using an MPN microplate technique indicated that these bacteria are most numerous in the aggregate size fractions corresponding to fine silt (2-20 microm) and clay (<2 microm) compared to larger fractions or unfractionated soil. PAH concentrations were also highest in the aggregate size fraction corresponding to fine silt. Similar results were found in a spiked soil (incubated for 6 months) with similar carbonated minerals. Transmission electron microscopy observations showed that the autochtonous PAH-degrading bacteria were embedded in the aggregates where PAHs were abundant. In spite of this extensive co-localisation PAH degradation was limited during 6 months incubation. This indicates that factors other than spatial distribution and PAH degrading ability control degradation rates. The fine silt fraction of the industrial soil had an elevated C/N ratio (35) compared to the clay fraction (C/N: 16). Thus the fraction which assumably had the highest specific surface area contained less PAH but similar numbers of PAH-degraders. N thus seem to play an important role in the long term, but as PAH degradation was low in fine size fractions, other sources/factors were probably limiting (easily degradable C, P org, O2 etc.). Based on these findings, soil particle organization and structure of soil aggregates appear to be important for the characterization of a polluted soil (localization and sequestration). Manipulations that modify aggregation in polluted soils could thus potentially influence the accessibility and biodegradability of PAHs.

Analytical determination of the microbial utilization and transformation of humic acids extracted from municipal refuse.

Humic substances are usually the refractory part of natural organic matter, and in a landfill they can retain inorganic and organic micropollutants. This study has investigated analytically whether humic acids (HA) extracted by use of alkali from either fresh municipal refuse or from refuse disposed of in a landfill for up to 12 months can resist microbial degradation under aerobic conditions. When added as a supplementary nutrient source, up to 63.6% of HA was utilized and this percentage was enhanced to a mean value of 88.5% when different HA preparations were used as the sole source of carbon. In cultures of a soil microbial community containing the same preparations as sole sources of nitrogen, HA was usually completely utilized. The remaining HA re-isolated from some microbial cultures were highly depleted in carbon and, simultaneously, the nitrogen content was enhanced. The FTIR spectra were indicative of strong participation of aliphatic structural units in the refuse-related HA preparations. Because of the microbial activity, different carbonaceous substances were primarily removed from the HA structure, and an increase in nitrogenous molecular groups became apparent. The structural transformations brought about by soil microorganisms "in vitro" corresponded to those occurring naturally in HA obtained from refuse aged for 12 months in a landfill.

A multidisciplinary approach to assess history, environmental risks, and remediation feasability of soils contaminated by metallurgical activities. Part A: chemical and physical properties of metals and leaching ability.

Three soils contaminated by heavy metals (HMs) and aliphatic hydrocarbons originating from different industrial sources were characterized in a multidisciplinary study combining chemical, physical (Part A), and mineralogical (Part B) approaches to define history, environmental risks, and remediation feasibility. These were an agricultural soil located nearby a Zn/Pb smelter and two soils from a steel metallurgical (siderurgy) waste land. High Pb and Zn contents were reported for all, and high Cr, Cd, and aliphatic hydrocarbons were present in different combinations in two out of three soils. Carbonate matrixes more strongly cemented fine particles in the agricultural soil than in both siderurgical ones, but buffering capacities were stronger in the latter. After the disruption of these cements by ultrasounds, HM were concentrated in the <50-microm fraction of the agricultural and of one siderurgical soil. Sequential extractions indicated for all the soils a preferential association of HM with carbonates, Fe-Mn oxides, organic matter, and sulfides. The stirring of one siderurgical soil with water resulted in an immediate leaching of hexavalent chromium (0.3 mg x L(-1)) involving potential ecotoxicological risks.

A multidisciplinary approach to assess history, environmental risks, and remediation feasability of soils contaminated by metallurgical activities. Part B: direct metal speciation in the solid phase.

Three soils contaminated by industrial smelting activities previously characterized using chemical and physical approaches (Part A, Venditti et al. [2000] Arch Environ Contam Toxicol [in press]) were further investigated through a complementary mineralogical approach. X-ray diffraction allowed identification of the main mineral components: carbonates (calcite, siderite) and iron oxides (magnetite, hematite, goethite) prevailed in both siderurgical soils, and siliceous compounds were more abundant in the agricultural soil. Scanning electron microscopy coupled with energy dispersive x-ray microanalysis showed that carbonates and oxides contained low amounts of heavy metals (HMs), but the potential solubility of such species under acidic conditions associated to their abundance may result in a high mobility of associated contaminants. Microanalysis also revealed peculiar minor mineral forms (metallic nodules, alloys, ore residues) that were highly informative about the history of soil contamination. Small metal-rich particles were observed (brass, lead and zinc oxides). In such compounds, HMs showed high chemical stability and were consequently poorly mobile and toxic, but also poorly removable through chemical leaching. A risk of mechanical dispersion was suggested for these particles, particularly after the chemical and/or mechanical disruption of carbonate cements. These features allowed completion and confirmation of the conclusions achieved after chemical and physical investigations, providing accurate information about the history, environmental risks and remediation feasability of industrial soil contamination.

Microbial utilization and transformation of humic acid-like substances extracted from a mixture of municipal refuse and sewage sludge disposed of in a landfill.

The purpose of the research was to establish whether humic acid-like substances (HA) related to municipal refuse disposed of in a landfill can resist microbial degradation and if they contribute, in that way, to long-term stabilization of landfill refuse. Using a mixture of 0.1 M Na(4)P(2)O(7) + 0.1 M NaOH, we extracted HA from municipal refuse mixed with sewage sludge and disposed of for up to 12 months, in a 40-m(3) model landfill. In laboratory experiments under aerobic conditions, up to 50% of HA was utilized as a supplementary source of nutrients by an assemblage of soil microorganisms in only 21 days. The microbial utilization was enhanced to over 80%, and up to 98%, respectively, if HA served as the sole source of carbon or nitrogen. Remaining HA which could be re-isolated from microbial cultures were lower in carbon (<12%) and nitrogen (<2.3%). Spectroscopic analysis (UV, Vis, FTIR) indicated losses, especially in aliphatic structural units, and a relative enhancement in aromatic structures. It was postulated that for their high degree of degradability, HA indigenous to that anthropogenic environment would not play an important role in the long-term stabilization of landfill refuse.

Copper speciation and microbial activity in long-term contaminated soils.

Most soil quality guidelines do not distinguish among the various forms of metals in soils; insoluble, nonreactive, and nonbioavailable forms are deemed as hazardous as highly soluble, reactive, and toxic forms. The objective of this study was to better understand the long-term effects of copper on microorganisms in relation to its chemical speciation in the soil environment. Carbon mineralization processes and the global structure of different microbial communities (fungi, eubacteria, actinomycetes) are still affected after more than 50 years of copper contamination in 20 soils sampled from two different agricultural sites. The microbial respiration lag period (LP) preceding the beginning of mineralization process increases with the level of soil copper contamination and is not significantly affected by other environmental factors such as soil pH and soil organic matter (SOM) content. The total copper concentration showed the best correlation with the LP when each site is considered separately. However, when considering the whole set of data, soil solution free Cu2+ activity (pCu2+) is the best predictor of Cu toxicity determined by LP (quite likely because pCu2+ integrates the soil physicochemical variability). The maximum mineralization rate (MMR), even if well correlated with the pCu2+, appears not to be a good biomonitor of copper contamination in soils since it is highly sensitive to soil characteristics such as SOM content. This study emphasizes the importance of the physicochemical properties of the environment on soil heavy metal toxicity and on soil toxicological measurements. These properties must be characterized in soil toxicological studies with respect to (1) their interactions with heavy metals, and (2) their direct impact on the selected biological test. The measurement of pCu2+ to characterize the level of soil contamination and of lag period as a bioindicator of metal effects in the soil are recognized as useful tools for the evaluation of the biological quality of soils.

Enzyme-linked immunofiltration assay To estimate attachment of thiobacilli to pyrite

An enzyme-linked immunofiltration assay (ELIFA) has been developed in order to estimate directly and specifically Thiobacillus ferrooxidans attachment on sulfide minerals. This method derives from the enzyme-linked immunosorbent assay but is performed on filtration membranes which allow the retention of mineral particles for a subsequent immunoenzymatic reaction in microtiter plates. The polyclonal antiserum used in this study was raised against T. ferrooxidans DSM 583 and recognized cell surface antigens present on bacteria belonging to the genus Thiobacillus. This antiserum and the ELIFA allowed the direct quantification of attached bacteria with high sensitivity (10(4) bacteria were detected per well of the microtiter plate). The mean value of bacterial attachment has been estimated to be about 10(5) bacteria mg-1 of pyrite at a particle size of 56 to 65 &mgr;m. The geometric coverage ratio of pyrite by T. ferrooxidans ranged from 0.25 to 2.25%. This suggests an attachment of T. ferrooxidans on the pyrite surface to well-defined limited sites with specific electrochemical or surface properties. ELIFA was shown to be compatible with the measurement of variable levels of adhesion. Therefore, this method may be used to establish adhesion isotherms of T. ferrooxidans on various sulfide minerals exhibiting different physicochemical properties in order to understand the mechanisms of bacterial interaction with mineral surfaces.

Cyanide Degradation under Alkaline Conditions by a Strain of Fusarium solani Isolated from Contaminated Soils.

Several cyanide-tolerant microorganisms have been selected from alkaline wastes and soils contaminated with cyanide. Among them, a fungus identified as Fusarium solani IHEM 8026 shows a good potential for cyanide biodegradation under alkaline conditions (pH 9.2 to 10.7). Results of K(sup14)CN biodegradation studies show that fungal metabolism seems to proceed by a two-step hydrolytic mechanism: (i) the first reaction involves the conversion of cyanide to formamide by a cyanide-hydrolyzing enzyme, cyanide hydratase (EC 4.2.1.66); and (ii) the second reaction consists of the conversion of formamide to formate, which is associated with fungal growth. No growth occurred during the first step of cyanide degradation, suggesting that cyanide is toxic to some degree even in cyanide-degrading microorganisms, such as F. solani. The presence of organic nutrients in the medium has a major influence on the occurrence of the second step. Addition of small amounts of yeast extract led to fungal growth, whereas no growth was observed in media containing cyanide as the sole source of carbon and nitrogen. The simple hydrolytic detoxification pathway identified in the present study could be used for the treatment of many industrial alkaline effluents and wastes containing free cyanide without a prior acidification step, thus limiting the risk of cyanhydric acid volatilization; this should be of great interest from an environmental and health point of view.

Occurrences at mineral-bacteria interface during oxidation of arsenopyrite by Thiobacillus ferrooxidans.

The combination of an improved bacterial desorption method, scanning electron microscopy (SEM), diffuse reflectance and transmission infrared Fourier transform spectroscopy, and a desorption-leaching device like high-pressure liquid chromatography (HPLC) was used to analyze bacterial populations (adhering and free bacteria) and surface-oxidized phases (ferric arsenates and elemental sulfur) during the arsenopyrite biooxidation by Thiobacillus ferrooxidans. The bacterial distribution, the physicochemical composition of the leachate, the evolution of corrosion patterns, and the nature and amount of the surface-oxidized chemical species characterized different behavior for each step of arsenopyrite bioleaching. The first step is characterized by a slow but strong adhesion of bacteria to mineral surfaces, the appearance of a surface phase of elemental sulfur, the weak solubilization of Fe(II), As(III), and As(V), and the presence of the first corrosion patterns, which follow the fragility zones and the crystallographic orientation of mineral grains. After this short step, growth of the unattached bacteria begins, while ferrous ions in solution are oxidized by them. Ferric ions produced by the bacteria can oxidize the sulfide directly and are regenerated by Fe(II) bacterial oxidation. At this time, a bioleaching cycle takes place and a coarse surface phase of ferric arsenate (FeAsO(4) . xH(2)O where x approximately 2) and deep ovoid pores appear. At the end of the bioleaching cycle, the high concentration of Fe(III) and As(V) in solution promotes the precipitation of a second phase of amorphous ferric arsenate (FeAsO(4) . xH(2)O where x approximately 4) in the leachate. Then the biooxidation process ceases: The bacteria adhering to the mineral sufaces are coated by the ferric arsenates and the concentration of Fe(III) on the leachate is found to have decreased greatly. Both oxidation mechanisms (direct and indirect oxidation) have been stopped. (c) 1995 John Wiley & Sons, Inc.

Quantification of the intragranular porosity formed in bioleaching of pyrite by Thiobacillus ferroxidans.

During the bacterial oxidation of a pure pyrite by Thiobacillus ferrooxidans, a great number of corrosion tunnels appear that are easily revealed by scanning electron microscopy observations. This involves an increase in the surface area without significant granulometric reduction of mineral grains. Thus, the evaluation of intragranular porosity, determined by elution front analysis, allows one to estimate accurately the fraction of oxidized sulphide, because of the development of deep holes (propagating pore mechanism). After 60 days of bioleaching, the intragranular porosity represents about 34% of the initial sulphide volume, which corresponds to 25 km of tunnels (2 microm i.d.) per gram of pyrite. On other hand, the granulometric reduction ( approximately 7%) is responsible for a 23% decrease of the initial sulphide volume. The elution front analysis appears as a nondestructive method for measuring the intragranular porosity of the bioleached pyrite.

Corrosion and Electrochemical Oxidation of a Pyrite by Thiobacillus ferrooxidans.

The oxidation of a pure pyrite by Thiobacillus ferrooxidans is not really a constant phenomenon; it must be considered to be more like a succession of different steps which need characterization. Electrochemical studies using a combination of a platinum electrode and a specific pyrite electrode (packed-ground-pyrite electrode) revealed four steps in the bioleaching process. Each step can be identified by the electrochemical behavior (redox potentials) of pyrite, which in turn can be related to chemical (leachate content), bacterial (growth), and physical (corrosion patterns) parameters of the leaching process. A comparison of the oxidation rates of iron and sulfur indicated the nonstoichiometric bacterial oxidation of a pure pyrite in which superficial phenomena, aqueous oxidation, and deep crystal dissolution are successively involved.

[Chemoembolization of hepatic metastases. Preliminary study]. / Chimio-embolisation des métastases hépatiques. Etude préliminaire.

Twenty-one patients with hepatic metastases were treated by chemoembolization over a 2-year period, the technique consisting of selective injection of a mixture of Spongel powder and Adriamycin plus 5-FU into the hepatic artery. Indications and results are discussed as a function of clinical, biologic and radiologic criteria.