Richard N. Collins

Chemical forms of selenium in the metal-resistant bacterium Ralstonia metallidurans CH34 exposed to selenite and selenate.

ABSTRACT Ralstonia metallidurans CH34, a soil bacterium resistant to a variety of metals, is known to reduce selenite to intracellular granules of elemental selenium (Se0). We have studied the kinetics of selenite (SeIV) and selenate (SeVI) accumulation and used X-ray absorption spectroscopy to identify the accumulated form of selenate, as well as possible chemical intermediates during the transformation of these two oxyanions. When introduced during the lag phase, the presence of selenite increased the duration of this phase, as previously observed. Selenite introduction was followed by a period of slow uptake, during which the bacteria contained Se0 and alkyl selenide in equivalent proportions. This suggests that two reactions with similar kinetics take place: an assimilatory pathway leading to alkyl selenide and a slow detoxification pathway leading to Se0. Subsequently, selenite uptake strongly increased (up to 340 mg Se per g of proteins) and Se0 was the predominant transformation product, suggesting an activation of selenite transport and reduction systems after several hours of contact. Exposure to selenate did not induce an increase in the lag phase duration, and the bacteria accumulated approximately 25-fold less Se than when exposed to selenite. SeIV was detected as a transient species in the first 12 h after selenate introduction, Se0 also occurred as a minor species, and the major accumulated form was alkyl selenide. Thus, in the present experimental conditions, selenate mostly follows an assimilatory pathway and the reduction pathway is not activated upon selenate exposure. These results show that R. metallidurans CH34 may be suitable for the remediation of selenite-, but not selenate-, contaminated environments.

Variability of Mn hyperaccumulation in the Australian rainforest tree Gossia bidwillii (Myrtaceae)

This study examines the heterogeneity of the Mn-hyperaccumulative trait in natural stands of the Australian rainforest tree species Gossia bidwillii (Myrtaceae). It is the only known Mn hyperaccumulator from Australia, and has an unusual spatial distribution of Mn in its leaves. G. bidwillii occurs naturally on a range of Mn-containing substrates including ultramafic soils. Leaf samples were collected from individual trees and four small stands, over a longitudinal range of ∼600xa0km. While no variation in the spatial distribution of foliar Mn was detected, considerable variation in Mn concentration was found. G. bidwillii was shown to accumulate Mn when growing on a variety of substrates, and dry weight (DW) foliar Mn concentrations of all trees sampled ranged between 2,740 and 27,470xa0μgxa0g−1. The majority of samples exceeded 10,000xa0μgxa0g−1, the threshold value for Mn hyperaccumulation. The overall frequency distribution of foliar Mn concentration was found to be bimodal, with a small outlier of extreme hyperaccumulators. Highest values were obtained from trees growing on a basaltic krasnozem clay, not ultramafic soil. Soil Mn concentrations were measured, and no relationship was found between foliar Mn concentrations and extractable Mn concentrations in host substrates. Some of the variation in the Mn-hyperaccumulative trait in G. bidwillii throughout its large natural distribution may reflect the unresolved taxonomy of this most widespread species in the genus Gossia. Ability to hyperaccumulate Mn may serve as an additional diagnostic tool for resolving this taxonomy.

Novel pattern of foliar metal distribution in a manganese hyperaccumulator

The primary sequestration of foliar manganese (Mn) in Mn-hyperaccumulating plants can occur in either their photosynthetic or non-photosynthetic tissues, depending on the species. To date, only non-photosynthetic tissues have been found to be the major sinks in other hyperaccumulators. Here, electron (SEM) and proton (PIXE) microprobes were used to generate qualitative energy dispersive (EDS) X-ray maps of leaf cross sections. Two Mn hyperaccumulators, Garcinia amplexicaulis Vieill. (Clusiaceae) and Maytenus fournieri (Panch. and Sebert) Loesn. (Celastraceae), and the Mn accumulator Grevillea exul Lindley (Proteaceae) were studied. PIXE/EDS data obtained here for M. fournieri were in agreement with existing SEM/EDS data showing that the highest localised foliar Mn concentrations were in the epidermal tissues. However, this is the first in situ microprobe investigation of G. amplexicaulis and G. exul. The Mn X-ray maps of G. amplexicaulis revealed a previously undescribed third spatial distribution pattern among Mn-hyperaccumulating species. Manganese was relatively evenly distributed throughout the leaf photosynthetic and non-photosynthetic tissues, while in G. exul it was most highly concentrated in the epidermal cells.

Transformation and fixation of Zn in two polluted soils by changes of pH and organic ligands

Although the movement of Zn from the solid to the solution phase of soils is a significant process preceding plant uptake, the quantity of metal that can be solubilised through rhizosphere processes is also extremely important. Therefore, the consequences of plant-derived organic ligands, and changes in pH on the isotopically exchangeable quantity of Zn (E value) were examined in a polluted acid and calcareous soil. Variations in pH were facilitated through the use of dilute NaOH or HNO3 solutions. The organic ligands studied included 0.25–5 mM concentrations of sodium tartrate, the free acid and sodium salt of citrate, histidine, and deoxymugineic acid. As expected, the organic ligands and a reduction in pH increased the solution concentration of Zn in these soils. Furthermore, through the application of isotopic dilution techniques, it was determined that some of these organic ligands could significantly increase the quantity of isotopically exchangeable Zn. However, with the exception of the 5 mM sodium citrate treatment in the calcareous soil, pH was the overriding parameter that effected changes in the E value. Reducing the pH by approximately 2 units increased the E value by 39 and 80%, respectively, in the acid and calcareous soil. Conversely, evidence for Zn fixation (a decrease of the E value) was observed in the acid soil when pH was increased. The exudation of organic ligands and variations of pH induced by plants have the potential to significantly vary the quantity of phytoavailable Zn in these 2 polluted soils.

Seleno-l-Methionine Is the Predominant Organic Form of Selenium in Cupriavidus metallidurans CH34 Exposed to Selenite or Selenate

ABSTRACT The accumulated organic form of selenium previously detected by X-ray absorption near-edge structure (XANES) analyses in Cupriavidus metallidurans CH34 exposed to selenite or selenate was identified as seleno-l-methionine by coupling high-performance liquid chromatography to inductively coupled plasma-mass spectrometry.

Resistance, accumulation and transformation of selenium by the cyanobacterium Synechocystis sp. PCC 6803 after exposure to inorganic SeVI or SeIV

Summary Our purpose was to investigate the ability of Synechocystis sp . PCC 6803, a photosynthetic prokaryote isolated from fresh water, to resist, incorporate and reduce the oxidized forms of selenium including selenite and selenate, the major selenium species present in aquatic systems. Selenium speciation and the chemical intermediates during selenium transformation were determined by X-ray absorption near edge structure (XANES) spectroscopy. The possible internalisation pathways involving selenium and the metabolic fate of selenate and selenite were examined. Selenate metabolism seemed to proceed via the sulfate reduction pathway resulting in the formation of the R-Se-H, R-Se-R and R-Se-Se-R species. The transformation of selenate to toxic amino acids may explain the high sensitivity of Synechocystis to selenate. Several mechanisms of selenium reduction seem to compete during selenite assimilation. A specific mechanism may transform internalised selenite into selenide and, subsequently induce the biosynthesis of selenoproteins. A non-specific mechanism may interfere with thiols, such as glutathione in the cell cytoplasm, or with proteins in the periplasm of the bacteria, notably thioredoxins. Several hypotheses concerning the complex transformation of selenium in Synechocystis could therefore be proposed.

Influence of thermodynamic database on the modelisation of americium(III) speciation in a simulated biological medium

Summary An intercomparison exercise was led in order to study the impact of thermodynamic database (TDB) on the speciation of americium(III) in biochemical media. A first exercice was led, considering only the americium(III) species in solution. Even with different data sets, all the exercises are in relatively good agreement, concluding to successive complexation of the radioelement by phosphate (pH range from 0 to 3), citrate (pH range from 3 to 8) and hydroxo-carbonate (pH range from 8 to 14). Nevertheless, the speciation of americium differs for each pH range from one work to an other. These results are still observed when the speciation calculation includes the formation of solids. Nevertheless, some workers did not integrate in their data set the formation constant of Am(PO4). This solid is known to be very insoluble in typical biological media. All the calculations where Am(PO4) was not in the TDB presented a speciation where americium is solvated (mostly under citrate forms). When the formation of Am(PO4) is considered, the actinide is quantitatively present under this solid species. This work is a representative example of the impact of the thermodynamic data used in speciation exercises. Some exercices, using uncompleted TDB had wrong results and led to the conclusion of the necessity in the use of expertised and extended TDB.

Soil- and surfactant-enhanced reductive dechlorination of carbon tetrachloride in the presence of Shewanella putrefaciens 200

Abstract Sorption of organic contaminants to soils has been shown to limit bioavailability and biodegradation in some systems. Use of surfactants has been proposed to reverse this effect. In this study, the effects of a high organic carbon content soil and a nonionic surfactant (Triton X-100) on the reductive dechlorination of carbon tetrachloride (CCl 4 ) were examined in anaerobic systems containing Shewanella putrefaciens . Although more than 70% of the added CCl 4 was sorbed to the soil phase in these systems, the reductive dechlorination of CCl 4 was not diminished. Rather, rates of CCl 4 dechlorination in systems containing soil were enhanced relative to systems containing non-sorptive sand slurries. This enhancement was also observed in sterile soil slurries to which a chemical reductant, dithiothreitol was added. It appears that the organic soil used in these experiments contains some catalytic factor capable of transforming CCl 4 in the presence of an appropriate chemical or microbial reductant. The addition of Triton X-100 to sand and soil slurries containing S. putrefaciens resulted in increased CCl 4 degradation in both systems. The effect of Triton could not be explained by: (i) surfactant induced changes in the distribution of CCl 4 , (i.e. decreased sorption) or the rate of CCl 4 desorption; (ii) a direct reaction between Triton and CCl 4 ; or (iii) increased cell numbers resulting from use of the surfactant as a substrate. Rather, it appears that Triton X-100 addition resulted in lysis of bacterial cells, a release of biochemical reductant, and enhanced reductive transformation of CCl 4 . These results provide insights to guide the development of more effective direct or indirect bioremediation strategies.

Potential phytoavailability of anthropogenic cobalt in soils as measured by isotope dilution techniques

Isotope dilution is a useful technique to determine the potential phytoavailability of an element in soil. This method involves equilibrating an isotope with soil and then sampling the labile metal pool by analysis of the soil solution (E value) or plants growing in the soil (L value). The work reported here was conducted to evaluate the distribution coefficient (Kd), and the potential phytoavailability (E value) of cobalt (Co) in eight soils subjected to the atmospheric deposition of anthropogenic Co. Multiple regression analyses demonstrated that the K(d) of isotopically exchangeable Co in these soils was best modelled with two parameters: soil pH and organic carbon (OC) content (log Kd=0.85(pH)+1.1(logOC)-5.0, R2=0.94, p<0.01). Cobalt E values ranged from 1.5 to 37% of total soil Co concentrations. No evidence was obtained to suggest that Co(III), if present, was isotopically exchangeable in these soils and it was concluded that the Co E values consisted solely of Co(II). Cobalt L values, measured with Triticum aestivum L. (46 days), of two of these soils (varying in soil pH, e.g. 5.0 and 7.2) were statistically (p<0.05) different to E values. However, when changes of bulk soil pH on Co E values were considered, the two values were statistically (p<0.05) similar indicating that processes affecting soil pH during plant growth can alter isotopically exchangeable concentrations of Co.

Assessment of uranium and selenium speciation in human and bacterial biological models to probe changes in their structural environment

Abstract This study illustrates the potential of physicochemical techniques to speciate uranium (U) and selenium (Se) in biological samples. Speciation, defined here as the study of structural environment, of both toxic elements, was characterized at several levels in biological media and directly in human cells or bacteria once the metal(loid)s were internalized. External speciation that is extracellular speciation in culture media was predicted by thermodynamic equilibrium computer modelling using the JChess software and validated by spectroscopic measurements (XANES and EXAFS). Internal speciation that is intracellular speciation in eukaryotic and prokaryotic cells was studied in vitro with a soil bacterium Cupriavidus metallidurans CH34 and ROS 17/2.8 osteoblasts, human cells responsible for bone formation. XANES, EXAFS, HPLC-ICP-MS and SDS-PAGE coupled to particle induced X-ray emission (PIXE) permitted the identification and quantification of complexes formed with organic or inorganic molecules and/or larger proteins.