Changes of Ni biogeochemistry in the rhizosphere of the hyperaccumulator Thlaspi goesingense

Processes in the rhizosphere of metal hyperaccumulator species are largely unknown. We investigated root-induced changes of Ni biogeochemistry in the rhizosphere of Thlaspi goesingense Hálácsy in a rhizobox experiment and in related soil chemical and Ni uptake studies. In the rhizobox, a root monolayer was separated from rhizosphere soil by a nylon membrane. Rhizosphere soil was then sliced into 0.5 mm layers and analyzed for changes in soluble (water-extractable, Ni _S) and labile (1 M NH ₄NO ₃-extractable, Ni _L) Ni pools. Ni _L in the rhizosphere was depleted due to excessive uptake in T. goesingense. Ni _S in the rhizosphere increased in contrast to expectations based on the experimental Ni desorption isotherm. Mathematical simulations following the Tinker–Nye–Barber approach overestimated the depletion of the Ni _L and predicted a decrease of Ni _S in the rhizosphere. In a hydroponic experiment, we demonstrated that T. goesingense takes up Ni ²⁺ but excludes metal–organic complexes. The model output was then improved in later versions considering this finding. A sensitivity analysis identified I _max and K _m, derived from the Michaelis–Menten uptake kinetics experiment to be the most sensitive of the model parameters. The model was also sensitive to the accuracy of the estimate of the initial Ni concentration (C _Si) in soil solution. The formation of Ni–DOM complexes in solution could not explain the poor fit as in contrast to previous field experiments, the correlation between soluble Ni and dissolved organic carbon (DOC) was weak. Ion competition of Ni with Ca and Mg could be ruled out as explanation of enhanced Ni solubility in the rhizosphere as the molar ratio of Ni/(Ca + Mg) in solution was not affected. However, a decreased Vanselov coefficient Kv near the root plane indicated (an apparent) lower selectivity of the exchange complex for Ni, possibly due to adsorption of oxalate exuded by T. goesingense roots or associated rhizosphere microbes. This conclusion is supported by field data, showing enhanced oxalate concentrations in the rhizosphere of T. goesingense on the same experimental soil. The implications for phytoextraction and bio-available contaminant stripping (BCS) as well as for future modeling and experimental work are discussed.     

In situ observation of suspended solid aggregates in rivers

Suspended solids are a major pathway of the biogeochemical fate of many contaminants in aquatic systems. Aggregation processes of particles are poorly reported in rivers but they are likely to exist at low flow, mostly because of sticky polysaccharides produced by living organisms. These processes affect suspended particle transport by changing particle sizes and densities and may also limit exchanges of matter between solid and dissolved phases. The main difficulty in floc studies is aggregate fragility, which requires the use of specialized in situ techniques to analyze aggregated suspended solids.We describe two in situ methods for observing suspended particles which do not need heavy field equipment. The first one is based on the filtration of a thin water layer in the natural flow through a membrane which can subsequently be observed by microscope. The second one is based on in situ video snapshots of suspended solids by an endoscope. The video camera linked to the microscope or the endoscope supplies images which are automatically analyzed by image processing to give size distributions.Procedures and validation for both methods are described and results compared with a standard method. The filtration method has been used to trace suspended solids from sewer overflows in the Seine River downstream of Paris. Freshwater flocs are described and a discussion of the fate of aggregates is presented.

---

Résumé Les matières en suspension sont déterminantes dans le cycle biogéochimique de nombreux polluants des systèmes aquatiques. Les phénomènes d'agrégation des particules sont encore mal connus en rivière, mais peuvent se produire dans les fleuves urbains du fait de la lenteur des écoulements et vraisemblablement de l'importance des processus bactériens. Ces phénomènes influencent directement le transport des suspensions en modifiant les tailles et les densités des particules. Ils peuvent également limiter les échanges chimiques entre phase dissoute et particulaire.Le problème de la fragilité des flocs et la complexité des équipements habituellement utilisés in situ nous ont conduit à développer deux méthodes d'observation non destructives des flocs. La première consiste à filtrer une mince lame d'eau directement dans l'écoulement naturel puis à observer les suspensions au microscope. La deuxième méthode est l'observation directe des suspensions dans l'écoulement, grâce à un endoscope. Une caméra vidéo reliée au microscope ou à l'endoscope fournit des images que l'on traite automatiquement (dénombrement et mesure des particules) pour obtenir la granulométrie des agrégats naturels.Nous comparons les résultats des deux méthodes à ceux d'une méthode de référence, puis nous comparons les deux méthodes entre elles. La filtration in situ a été utilisée en Seine à l'aval de gros rejets de temps de pluie de l'agglomération parisienne pour étudier les matières en suspension issues des rejets. La caractérisation des flocs de Seine permet de discuter de leur devenir dans le fleuve.

     

In situ phosphorus release experiments in the Warnow River (Mecklenburg,northern Germany)

Results are presented of in situ benthic phosphorus release experiments in an undercut bank of an impounded river. Due to high sedimentation of phytoplankton biomass high oxygen consumption rates between 259.4 and 947.0 mg O₂ m^–2 d^–1 developed, leading to almost anaerobic conditions and phosphorus releases between 175.2 and 236.3 mgP m^–2 d^–1 over a period of 18 days.In a second series of experiments the water column overlying the sediment was aerated, resulting in much lower P release rates (1.1 to 32.9 mgP m^–2 d^–1) over a period of 30 days. The influence of pH and nitrate was studied by adjusting pH and adding NO₃ ^– to the overlying water. Increasing pH positively affected P release rates and enhanced NO₃ ^– levels led to an increase of benthic P release, too.     

The effects of root-induced pH changes on the depletion of inorganic and organic phosphorus in the rhizosphere

A new method allowing control of rhizosphere pH and mineral nutrition was applied to study depletion of various organic and inorganic phosphorus fractions extractable sequentially with 0.5M KHCO₃ (pH 8.5), 0.1M NaOH and residual P extractable with 6M H₂SO₄ from the rhizosphere soil.Soil pH was affected about 2 mm from the root mat. Depletion zones of inorganic P (KHCO₃-P_i) extractable with 0.5M KHCO₃ extended up to about 4 mm but the depletion zones of all other P fractions were about 1 mm only. The root-induced decrease of soil pH from 6.7 to 5.5 increased the depletion of total P from all fractions by 20% and depletion of KHCO₃-P_i and residual P by 34% and 43%, respectively. Depletion of organic P (KHCO₃-P_o) extractable with 0.5M KHCO₃ was not affected by a change in rhizosphere pH. With constant or increased pH, depletion of inorganic P (NaOH-P_i) was 17% and organic P (NaOH-P_o) was 22% higher than with decreased pH. Only 54–60% of total P withdrawn from all fractions was from KHCO₃-P_i. Substantial amounts of KHCO₃-P_o and NaOH-P_o were mineralized and withdrawn from the rhizosphere within 1 mm from the root mat, as 11–15% of total P withdrawn originated from the organic P fractions. A remaining 11–16% was derived from NaOH-P_i, and 15–18% from residual P fractions likely to be rather immobile. Thus, 40–46% of the P withdrawn near the root mat of rape originated from non-mobile P fractions normally not included in 0.5M NaHCO₃ extraction used to obtain an index of plant-available soil P.     

In situ localized amplification and contact replication of many individual DNA molecules

We describe a method to clone and amplify DNA by performing the polymerase chain reaction (PCR) in a thin polyacrylamide film poured on a glass microscope slide. The polyacrylamide matrix retards the diffusion of the linear DNA molecules so that the amplification products remain localized near their respective templates. At the end of the reaction, a number of PCR colonies, or 'polonies', have formed, each one grown from a single template molecule. As many as 5 million clones can be amplified in parallel on a single slide. If an Acrydite modification is included at the 5' end of one of the primers, the amplified DNA will be covalently attached to the polyacrylamide matrix, allowing further enzymatic manipulations to be performed on all clones simultaneously. We describe techniques to make replicas of these polony slides, and high throughput sequencing protocols for this technology. Other applications are also discussed.     

In situ identification of protein structural changes in prion-infected tissue

Transmissible spongiform encephalopathies (TSE) are fatal neurodegenerative disorders characterized by the conversion of the normal prion protein (PrP(C)) into aggregates of its pathological conformer (PrP(Sc)). The mechanism behind this structural conversion is unclear. We report the identification of disease-related protein structural differences directly within the tissue environment. Utilizing a synchrotron infrared (IR) light source, IR images of protein structure were obtained at a subcellular resolution, revealing regions of decreased alpha-helical content and elevated beta-sheet structure in and around infected neurons in the 263 K scrapie hamster model. PrP(Sc) immunostaining of the same tissue demonstrated that the elevated beta-sheet regions correspond to regions where the misfolded structure of PrP(Sc) is located. No evidence of these structural changes was observed in normal neurons.     

In situ observation of coral recruitment using fluorescence census techniques

A novel method for the observation of recruitment of fluorescent taxa was developed combining fluorescence census techniques with conventional microscopic examination. The new technique was used to observe coral recruitment on natural limestone plates over a period of 4 months on Meras reef, North Sulawesi, Indonesia. During this period, fluorescence photos were taken of each plate on a weekly basis. This allowed for the detailed observation of fluorescent coral recruitment in-situ. After specific time periods, the plates were sampled and the detected recruits were categorized to family level using their skeletal structure. The fluorescence census technique detected 97.6% of all coral recruits bigger than one millimetre in diameter. The diameter measurements of the recruit's skeletons were used to estimate growth rates at family level. Using the photo series method, the time of settlement of fluorescent recruits on the plates was determined. The results showed roughly linear diameter growth of recruits in the first two months. Acroporidae-polyps were 1.14 mm in diameter when they settled and grew at rate of approximately 0.18 mm in diameter per week. Pocilloporidae-polyps settled at approximately 1.14 mm and showed a growth rate of approximately 0.23 mm in diameter per week. Recruits of three families were observed in detail over time on the artificial plates using this method. While recruitment was fairly continuous, an increase in pocilloporid recruits was observed during the month of September. Similar increases were observed for acroporid recruits during the middle of November and the second quarter of December. This method identifies the limits of detection for the fluorescence census technique used and represents a useful method for the temporal fine scale observation of recruitment in situ.     

Measurement of microbial biomass phosphorus in rhizosphere soil

³²P-labelled monocalcium phosphate solution was supplied by point injection to the root system of wheat plants grown in soil cores in a controlled environment. There was no detectable incorporation of³²P into organic P fractions in the soil remaining after roots were removed, confirming field observations. The techniques used to measure organic P (including biomass P) could detect an incorporation of³²P into soil microbial biomass equivalent to 0.3 μgP.g^?1 soil, compared to a total soil biomass P content estimated to be ca. 6.5 μgP.g^?1 soil. The limited incorporation of the added P into microbial biomass in the root-free soil may be due partly to a limited diffusion of³²P into the non-rhizosphere soil and partly to the removal of³²P-labelled microbial biomass adhering to or in very close association with the root surface. it is proposed that in studies of soil nutrient status, total soil biomass P (roots + soil flora + microfauna) should be measured, rather than attempting an estimate of microbial P. A sequential extraction procedure using a single soil sample, where a biocide is added to the extracting solution, is proposed as an alternative to the conventional procedure for measuring soil biomass P where two soil samples, one treated with a biocide, are extracted simultaneously.     

In situ observation of mobility and anchoring of PKCbetaI in plasma membrane

We employed fluorescence correlation spectroscopy (FCS) to analyze the characteristics of biomolecules in living cells. Protein kinase C (PKC) changes its subcellular localization from cytosol to the plasma membrane by its ligand. Using FCS, we found PKCbetaI labeled with enhanced green fluorescent protein freely diffusing in cytosol. Upon 12-O-tetradecanoylphorbol-13-acetate activation, a large part of PKCbetaI is anchored in the plasma membrane but some PKCbetaI still moves freely near the plasma membrane. These results indicate that a diffusion-driven transport mechanism is appropriate for the molecular mechanism of the PKCbetaI localization change.     

In situ observation of the generation of isothiocyanates from sinigrin in horseradish and wasabi

Sulfur K-edge X-ray absorption spectroscopy has been used to determine the chemical identity of the sulfur-containing species in horseradish (Armoracia lapthifolia) and wasabi (Wasabia japonica) in situ, before and after cell disruption. The major sulfur-containing species in the intact root is sinigrin (1-thio-beta-D-glucopyranose 1-N-(sulfoxy)-3-buteneimidate) and related congeners. Disrupting the cells by applying local pressure allowed the conversion of the sulfur moieties in sinigrin to isothiocyanates and sulfate in approximately equimolar amounts. In contrast to previous suggestions, no detectable thiocyanates were formed, but an unusual thio intermediate may have been identified for the first time.     

The biogeochemistry of phosphorus after the first century of soil development on Rakata Island, Krakatau, Indonesia

This study examined the accumulation of organic carbon (C) and fractions ofsoil phosphorus (P) in soils developing in volcanic ash deposited in the1883 eruption of Krakatau. Organic C has accumulated at rates of 45 to 127g/m²/yr during 110 years of soil development, resulting inprofiles with as much as 14 kgC/m². Most soil P is found inthe HCl-extractable forms, representing apatite. A loss of HCl-extractableP from the surface horizons is associated with a marked accumulation ofNaOH-extractable organic P bound to Al. A bioassay with hill rice suggeststhat P is limiting to plant growth in these soils, perhaps as a result ofthe rapid accumulation of P in organic forms.     

茶树根际土壤磷的解吸特性

为了探明茶树根际土壤磷的释放过程与供应机制,采用外源磷吸附培养方法,研究了不同母质、不同种植年限茶树根际土壤磷的解吸过程与特性,并用最小二乘法进行最优函数拟合.结果表明:茶树根际土壤与非根际土壤磷的解吸过程有明显的差异.茶树根际土壤的磷解吸能力极显著高于非根际土壤;与非根际土壤相比,根际土壤的平均有效磷含量、平均解吸率和平均β值(单位吸附量中的解吸量)分别高出5.49 mg.kg-1、1.7%和24.4%.不同成土母质发育的茶树根际土壤磷解吸能力为花岗岩风化物>第四纪红色粘土>板页岩风化物.随着种植年限的延长,3种母质茶树根际土壤的有效磷与磷解吸能力均有不同程度地提高.     

Assessing changes in the genetic diversity of potato gene banks. 2. In situ vs ex situ

An important question in the conservation of potato germ plasm is whether germ plasm in the gene bank, although stable, still represents the in situ populations from which it was collected, sometimes many decades ago. The answer would direct objective decisions regarding the value of re-collections and in situ preservation. The present study was undertaken as a project of the Association of Potato Inter-gene-bank Collaborators (APIC). It measured genetic differentiation between potato germ plasm maintained in the US gene bank for many years and current in situ populations re-collected from the same original sites in the wild. Solanum jamesii and Solanum ?fendleri from the United States were used as representatives of potato germ plasm. Re-collections were carried out in 1992 at the same locations at which gene bank-conserved accessions had been collected in 1958 and 1978. RAPD markers revealed significant genetic differences between gene bank-conserved and re-collected in?situ populations for all seven comparisons of S. jamesii (diploid outcrosser), and 12 of 16 comparisons within S. ?fendleri (tetraploid inbreeder). The average genetic similarities were 65.2% for S. ?jamesii and 80.4% for S. ?fendleri. Possible explanations and consequences of these unexpectedly large differences are discussed.     

Impacts of environmental changes on the biogeochemistry of aquatic humic substances

Humic substances (HS) are the main constituent of the organic carbon pool in stained aquatic ecosystems. HS absorb visible and ultraviolet (UV) light, have acid-base properties and metal and nutrient binding abilities. Based on these characteristics, UV irradiation, pH and the trophic status of aquatic ecosystems will influence the impact of HS on element cycling in surface waters. With climatic change and environmental pollution, UV irradiance, acidification and eutrophication may increase further. In this paper impacts of UV irradiation, pH and eutrophication on the structure, properties and biodegradation of aquatic HS are discussed.     

Microbial biogeochemistry and phosphorus limitation in cryoconite holes on glaciers across the Taylor Valley,McMurdo Dry Valleys,Antarctica

Biogeochemistry - Zooplankton communities are important components of aquatic food webs in part because they recycle nutrients and carbon. Lacking a complete understanding of the composition of...     

In situ filtration responses of Daphnia galeata to changes in food quality

In the stoichiometric study of phyto–zooplankton interactions,a controversy exists about how Daphnia species regulate theirfeeding rate when submitted to low-quality food (i.e. high dietarycarbon:phosphorus [C:P] ratio). In this study, we gathered dataover 3 years on in situ clearance rates of a Daphnia galeatapopulation, by conducting grazing experiments from April 1998to October 2000 in the Esch-sur-Sûre reservoir (Grand-Duchyof Luxembourg). Observed clearance rates (2.5–13.5 mLindividual^–1 day^–1, mean 7.0 mL individual^–1day^–1) were correlated with population and environmentalvariables. Mean body size of Daphnia individuals was the bestpredictor of clearance rate (r² = 0.639), followed by watertemperature (r² = 0.262) and P concentration in the seston (r²     

In situ observation of spherical DNA assembly in water and the controlled release of bound dyes

Three strands of 30-mer oligodeoxyribonucleotides (ODNs) were designed to form three-way junctions that possess self-complementary sticky ends. The morphology of self-assembled ODNs in water was observed in situ by confocal laser scanning fluorescence microscopy. The three-way junctions self-assembled into spherical assemblies, in accordance with transmission and scanning electron microscopy. The size of nucleospheres was in the range of several tens of nanometers to micrometers, which varied depending on the concentration of ODNs and added salts. Fluorescence images of spherical ODN assemblies suggested that the nucleospheres possess multiwalled structures. The fluorescence of sodium 1-anilinonaphthalene-8-sulfonate in the presence of nucleospheres revealed that the interior of nucleospheres possesses polarity corresponding to that between methanol and ethanol. A dye-inclusion experiment showed that cationic and even anionic dyes were adsorbed to the interior of the nucleospheres. The dye-included nucleospheres released dyes by thermal dissociation or digestion of the constituent ODNs.