Acoustic properties of amorphous silica between 1 and 500 mK.

We have made reliable measurements of the sound velocity delta v/v(0) and internal friction Q(-1) in vitreous silica at 1.03, 3.74, and 14.0 kHz between 1 mK and 0.5 K. In contrast with earlier studies that did not span as wide a temperature and frequency range, our measurements of Q(-1) reveal a crossover (as T decreases) only near 10 mK from the T(3) dependence predicted by the standard tunneling model to a T dependence predicted if interactions are accounted for. We find good fits at all frequencies using a single interaction parameter, the prefactor of the interaction-driven relaxation rate, in contrast to earlier claims of a frequency dependent power law. We also show that the discrepancy in the slopes d(delta v/v(0))/d(log(10)T) below and above the sound velocity maximum (1: -1 observed, 1: -2 predicted) can be resolved by assuming a modified distribution of tunneling states.

Evaluation of diagnostic scores with adjustment for covariates.

Diagnostic tests yield measurements on very different types of scales. Quantitative scales may consist of non-negative integers, either unbounded or bounded, with a fixed number of different values, or they may consist of continuous or percentage values. Remembering a different threshold value for each diagnostic variable would be cumbersome, in particular if covariates have to be taken into account. As a convenient way to overcome such problems we propose to compute z-scores for all measurements. They will be adjusted for covariates so that any individual can be judged on any test result on one single scale with an appropriate standard normal quantile as threshold. Two issues need to be addressed: Selection of covariates in the regression model which delivers the adjustment and normality of the residuals. The first will be treated by cross-validation and the latter by applying an appropriate transformation. We apply this methodology to neuropsychological tests and adjust for age, length of education and sex. Normality of residuals is needed on the diagnostically relevant side only. This allows to use parametric transformations, which can be easily implemented, e.g. in database systems. Since we have measurements at baseline and at follow-up we also analyze change values in a similar manner. For ease of interpretation, we transform the resulting z-scores back to the original scale.

Effect of low-level radiation on the low temperature acoustic behavior of a-SiO2.

We report on the mechanical behavior of an a-SiO2 84 kHz torsional oscillator operated between 100> or =T> or =1.0 mK. Below 10 mK we observed well-differentiated transient responses which we attribute to the interaction with low-level background radiation (gamma quanta and cosmic ray micro) and which can be modeled in terms of a change in the spring constant.

Unusual integrase gene expression on the clc genomic island in Pseudomonas sp. strain B13.

An unusual type of gene expression from an integrase promoter was found in cultures of the bacterium Pseudomonas sp. strain B13. The promoter controls expression of the intB13 integrase gene, which is present near the right end of a 105-kb conjugative genomic island (the clc element) encoding catabolism of aromatic compounds. The enzymatic activity of integrase IntB13 is essential for site-specific integration of the clc element into the bacterial host's chromosome. By creating transcription fusions between the intB13 promoter and the gfp gene, we showed that integrase expression in strain B13 was inducible under stationary-phase conditions but, strangely, occurred in only a small proportion of individual bacterial cells rather than equally in the whole population. Integrase expression was significantly stimulated by growing cultures on 3-chlorobenzoate. High cell density, heat shock, osmotic shock, UV irradiation, and treatment with alcohol did not result in measurable integrase expression. The occurrence of the excised form of the clc element and an increase in the rates of clc element transfer in conjugation experiments correlated with the observed induction of the intB13'-gfp fusion in stationary phase and in the presence of 3-chlorobenzoate. This suggested that activation of the intB13 promoter is the first step in stimulation of clc transfer. To our knowledge, this is the first report of a chlorinated compound's stimulating horizontal transfer of the genes encoding its very metabolism.

Characterization of two alternative promoters for integrase expression in the clc genomic island of Pseudomonas sp. strain B13.

The clc genomic island is a 105 kb integrative and conjugative element (ICE) in Pseudomonas sp. strain B13, which encodes metabolism of 3-chlorocatechol. The clc island is integrated in a tRNAGly gene, but can excise and form a circular intermediate in which both ends are connected. The integrase gene (intB13) of the clc genomic island is located at the right end, 202 bp from the junction site facing inwards. Fragments upstream of intB13 in the circular form and in the integrated form were fused to a promoterless gfp gene for Green Fluorescent Protein and introduced in monocopy onto the chromosome of strain B13. Quantitative GFP fluorescence measurements in individual cells of the different B13-derivatives revealed that the circular form fragment contained a strong constitutive promoter (Pcirc) driving intB13 expression in all cells. By using primer extension Pcirc could be mapped near the left end of the clc element and Pcirc can therefore only control intB13 expression when left and right ends are connected as in the circular form. Expression from intB13 upstream fragments from the integrated clc element was weaker than that from Pcirc and only occurred in maximally 15% of individual cells in a culture. A promoter (Pint) could be roughly mapped in this region by using reverse-transcription PCR and by successively shortening the fragment from the 5' end. Transposon mutants in cloned left end sequences of the clc element were selected which had lost the activation potential on the Pint promoter and those which resulted in overexpression of GFP from Pint. The DNA sequence of the region of the transposon insertions pointed to a relatively well conserved area among various other genomic islands. The activator mutants mapped in an open reading frame (ORF) encoding a 175 amino acid protein without any significant similarity to functionally characterized proteins in the databases.

Microbial composition and structure of a rotating biological contactor biofilm treating ammonium-rich wastewater without organic carbon.

High nitrogen losses were observed in a rotating biological contactor (RBC) treating ammonium-rich (up to 500 mg NH4(+)-N/L) but organic-carbon-poor leachate from a hazardous waste landfill in Kölliken, Switzerland. The composition and spatial structure of the microbial community in the biofilm on the RBC was analyzed with specific attention for the presence of aerobic ammonium and nitrite oxidizing bacteria and anaerobic ammonium oxidizers. Anaerobic ammonium oxidation (anammox) involves the oxidation of ammonium with nitrite to N2. First the diversity of the biofilm community was determined from sequencing cloned PCR-amplified 16S rDNA fragments. This revealed the presence of a number of very unusual 16S rDNA sequences, but very few sequences related to known ammonium or nitrite oxidizing bacteria. From analysis of biofilm samples by fluorescence in situ hybridization with known phylogenetic probes and by dot-blot hybridization of the same probes to total RNA purified from biofilm samples, the main groups of microorganisms constituting the biofilm were found to be ammonium-oxidizing bacteria from the Nitrosomonas europaea/eutropha group, anaerobic ammonium-oxidizing bacteria of the "Candidatus Kuenenia stuttgartiensis" type, filamentous bacteria from the phylum Bacteroidetes, and nitrite-oxidizing bacteria from the genus Nitrospira. Aerobic and anaerobic ammonium-oxidizing bacteria were present in similar amounts of around 20 to 30% of the biomass, whereas members of the CFB phylum were present at around 7%. Nitrite oxidizing bacteria were only present in relatively low amounts (less than 5% determined with fluorescence in situ hybridization). Data from 16S rRNA dot-blot and in situ hybridization were not in all cases congruent. FISH analysis of thin-sliced and fixed biofilm samples clearly showed that the aerobic nitrifiers were located at the top of the biofilm in an extremely high density and in alternating clusters. Anammox bacteria were exclusively present in the lower half of the biofilm, whereas CFB-type filamentous bacteria were present throughout the biofilm. The structure and composition of these biofilms correlated very nicely with the proposed physiological functional separations in ammonium conversion.

The high-energy proton fluxes in the SAA observed with REM aboard the MIR orbital station.

During two years, from November 1994 to 1996, the particle detector REM measured the highly energetic electron and proton environment at the outside of the MIR orbital station. Using mission averaged data we investigate various aspects of the proton fluxes in the SAA. Comparison with the radiation belt model AP8 reveal important differences.

Corrosion protection by anaerobiosis.

Biofilm-forming bacteria can protect mild (unalloyed) steel from corrosion. Mild steel coupons incubated with Rhodoccocus sp. strain C125 and Pseudomonas putida mt2 in an aerobic phosphate-buffered medium containing benzoate as carbon and energy source, underwent a surface reaction leading to the formation of a corrosion-inhibiting vivianite layer [Fe3(PO4)2]. Electrochemical potential (E) measurements allowed us to follow the buildup of the vivianite cover. The presence of sufficient metabolically active bacteria at the steel surface resulted in an E decrease to -510 mV, the potential of free iron, and a continuous release of ferrous iron. Part of the dissolved iron precipitated as vivianite in a compact layer of two to three microns in thickness. This layer prevented corrosion of mild steel for over two weeks, even in a highly corrosive medium. A concentration of 20 mM phosphate in the medium was found to be a prerequisite for the formation of the vivianite layer.

Measuring mass transfer processes of octane with the help of an alkSalkB::gfp-tagged Escherichia coli.

Diffusion of octane from oily droplets in different microscale settings was measured using Escherichia coli expressing the stable green fluorescent protein (GFP) from the alkB promoter of Pseudomonas oleovorans. GFP fluorescence intensities were determined quantitatively at the single-cell level after 1.0 or 2.5 h incubation and compared with different calibration series using known concentrations of octane. By immobilizing the E. coli sensor cells on the bottom glass plate of a microscope flow chamber, it was possible to monitor the diffusion process for octane in aqueous solution as a function of time and distance from non-aqueous phase droplets of octane alone or oily octane mixtures. When a gas phase was included in the flow chambers, octane transport could be demonstrated from the oily mixtures to the cells through both gas and liquid phase. Assays of non-immobilized sensor cells in microdroplets in the presence or absence of soil particles incubated with octane through the vapour phase revealed a slight reduction in the total amount of induced E. coli cells in the presence of soil. Our results indicate the power of using GFP-marked single-cell biosensors in determining microscale bioavailability of organic pollutants.

The glutathione peroxidase homologous gene from Chlamydomonas reinhardtii is transcriptionally up-regulated by singlet oxygen.

The glutathione peroxidase homologous gene (Gpxh gene) in Chlamydomonas reinhardtii is up-regulated under oxidative stress conditions. The Gpxh gene showed a remarkably strong and fast induction by the singlet oxygen-generating photosensitizers neutral red, methylene blue and rose Bengal. The Gpxh mRNA levels strongly increased, albeit much more slowly, upon exposure to the organic hydroperoxides tert-butyl hydroperoxide (t-BOOH) and cumene hydroperoxide. In contrast, the Gpxh mRNA levels were only weakly induced by exposure to the superoxide-generating compound paraquat and by hydrogen peroxide. A comparison of the Gpxh mRNA levels with those of the heat shock protein HSP70A and the iron superoxide dismutase gene showed qualitative and quantitative differences for the three genes under oxidative stress conditions tested. The Gpxh gene is specifically induced by singlet-oxygen photosensitizers and the relative induction by other compounds is much weaker for Gpxh than for the other genes investigated. Using Gpxh promoter fusions with the arylsulfatase reporter gene, we have shown that the Gpxh was transcriptionally up-regulated by singlet-oxygen photosensitizers. It is also shown that the Gpxh promoter contains a region between 104 and 179 bp upstream of the transcription start that is responsible for the mRNA up-regulation upon exposure to 1O2 but not t-BOOH. Within this region a regulatory sequence homologous to the mammalian cAMP response element (CRE) and activator protein 1 (AP-1) binding site was identified within a 16 bp palindrome.

Factors affecting mass transfer limited biodegradation in saturated porous media.

Microbial degradation rates in the subsurface are not only limited by the physiological capacity of the organisms, but also by inefficient supply of nutrients to the microbes. Although mass transfer limitation of biodegradation in the subsurface has been postulated for years, experimental evidence is still scarce. In the column experiments described here, diffusive transport of 4-nitroanisole from the bulk solution to cells of Rhodococcus opacus strain AS2 immobilized on glass beads or sand appeared to be responsible for the slow transformation rates observed. Assuming steady state, we applied a coupled transformation/transport equation to these data (Best equation) and apparent bead-related mass transfer coefficients were found to increase in proportion to the surface area covered with bacteria. This implies that mass transfer coefficients for individual cells remained constant. In an idealized oligotrophic environment where cells are only loosely clustered and do not shield each other, we would therefore expect biodegradation rates to be independent from the longitudinal distribution of the total biomass along a given flow path. Moreover, apparent mass transfer coefficients increased with the grain size of the column fillings, but did not change upon varying the flow rate. With a limiting external transport step, overall transformation fluxes do not become saturated at concentrations as low as predicted for Michaelis-Menten-type kinetics. Mass transfer limitation thus offers a justification for the common assumption that biodegradation rates in the subsurface follow first order kinetics in a wide concentration range.

Enrichment and characterization of an anammox bacterium from a rotating biological contactor treating ammonium-rich leachate.

Anaerobic ammonium oxidation with nitrite to N2 (anammox) is a recently discovered microbial reaction with interesting potential for nitrogen removal from wastewater. We enriched an anammox culture from a rotating disk contactor (near Kölliken, Switzerland) that was used to treat ammonium-rich leachate with low organic carbon content. This enrichment led to a relative population size of 88% anammox bacteria. The microorganism carrying out the anammox reaction was identified by analysis of the 16S rDNA sequence and by fluorescence in situ hybridization (FISH) with 16S-rRNA-targeting probes. The percentage sequence identity between the 16S rDNA sequences of the Kölliken anammox organism and the archetype anammox strain Candidatus Brocadia anammoxidans was 90.9%, but between 98.5 and 98.9% with Candidatus Kuenenia stuttgartiensis, an organism identified in biofilms by molecular methods. The Kölliken culture catalyzed the anaerobic oxidation of ammonium with nitrite in a manner seemingly identical to that of Candidatus B. anammoxidans, but exhibited higher tolerance to phosphate (up to 20 mM) and to nitrite (up to 13 mM) and was active at lower cell densities. Anammox activity was observed only between pH 6.5 and 9, with an optimum at pH 8 and a temperature optimum at 37 degrees C. Hydroxylamine and hydrazine, which are intermediates of the anammox reaction of Candidatus B. anammoxidans, were utilized by the Kölliken organisms, and approximately 15% of the nitrite utilized during autotrophic growth was converted to nitrate. Electron microscopy showed a protein-rich region in the center of the cells surrounded by a doughnut-shaped region containing ribosomes and DNA. This doughnut-shape region was observed with FISH as having a higher fluorescence intensity. Similar to Candidatus B. anammoxidans, the Kölliken anammox organism typically formed homogenous clusters containing up to several hundred cells within an extracellular matrix.

Chemical properties of catechols and their molecular modes of toxic action in cells, from microorganisms to mammals.

Catechols can undergo a variety of chemical reactions. In this review, we particularly focus on complex formations and the redox chemistry of catechols, which play an inportant role in the toxicity of catechols. In the presence of heavy metals, such as iron or copper, stable complexes can be formed. In the presence of oxidizing agents, catechols can be oxidized to semiquinone radicals and in a next step to o-benzoquinones. Heavy metals may catalyse redox reactions in which catechols are involved. Further chemical properties like the acidity constant and the lipophilicity of different catechols are shortly described as well. As a consequence of the chemical properties and the chemical reactions of catechols, many different reactions can occur with biomolecules such as DNA, proteins and membranes, ultimately leading to non-repairable damage. Reactions with nucleic acids such as adduct formation and strand breaks are discussed among others. Interactions with proteins causing protein and enzyme inactivation are described. The membrane-catechol interactions discussed here are lipid peroxidation and uncoupling. The deleterious effect of the interactions between catechols and the different biomolecules is discussed in the context of the observed toxicities, caused by catechols.

Methanosarcina lacustris sp. nov., a new psychrotolerant methanogenic archaeon from anoxic lake sediments.

A new psychrotolerant methanogenic archaeon strain ZS was isolated from anoxic lake sediments (Switzerland). The cells of the organism were non-motile cocci, 1.5-3.5 microm in diameter. The cells aggregated and formed pseudoparenchyma. The cell wall was Gram-positive. The organism utilized methanol, mono-, di-, trimethylamine and H2/CO2 with methane production. The temperature range for growth was 1-35 degrees C with an optimum at 25 degrees C. The DNA G+C content of the organism was 43.4. mol%. Analysis of the 16S rRNA gene sequence showed that strain ZS was phylogenetically closely related to members of the genus Methanosarcina, but clearly differed from all described species of this genus (95.6-97.6% of sequence similarity). The level of DNA-DNA hybridization of strain ZS with Methanosarcina barkeri and Methanosarcina mazei was 15 and 31%, respectively. Based on the results of physiological and phylogenetic studies strain ZS can be assigned to a new species of the genus Methanasarcina. The name Methanosarcina lacustris sp. nov. is proposed. The type strain is ZS (= DSM 13486T, VKM B-2268).

Proton Irradiation Facility and space radiation monitoring at the Paul Scherrer Institute.

The Proton Irradiation Facility (PIF) has been designed and constructed, in cooperation between Paul Scherrer Institute (PSI) and European Space Agency (ESA), for terrestrial proton testing of components and materials for spacecraft. Emphasis has been given to generating realistic proton spectra encountered by space-flights at any potential orbit. The facility, designed in a user-friendly manner, can be readily adapted to the individual requirements of experimenters. It is available for general use serving also in testing of radiation monitors and for proton experiments in different scientific disciplines. The Radiation Environment Monitor REM has been developed for measurements of the spacecraft radiation conditions. Two instruments were launched into space, one into a Geo-stationary Transfer Orbit on board of the STRV-1b satellite and one into a Low Earth Orbit on the Russian MIR station. The next generation of monitors (SREMs--Standard REMs) is currently under development in partnership of ESA, PSI and Contraves-Space. They will operate both as minimum intrusive monitors, which provide radiation housekeeping data and alert the spacecraft when the radiation level crosses allowed limits and as small scientific devices measuring particle spectra and fluxes. Future missions as e.g. INTEGRAL, STRV-1c and PROBA will be equipped with new SREMs.

[Effect of microorganisms and seasonal factors on the isotope composition of organic carbon from Black Sea suspensions]. / Vliianie mikroorganizmov i sezonnykh faktorov na izotopnyi sostav organicheskogo ugleroda vzvesi chernogo moria.

The isotopic composition of particulate organic carbon (POC) from the Black Sea deep-water zone was studied during a Russian-Swiss expedition in May 1998. POC from the upper part of the hydrogen sulfide zone (the C-layer) was found to be considerably enriched with the 12C isotope, as compared to the POC of the oxycline and anaerobic zone. In the C-layer waters, the concurrent presence of dissolved oxygen and hydrogen sulfide and an increased rate of dark CO2 fixation were recorded, suggesting that the change in the POC isotopic composition occurs at the expense of newly formed isotopically light organic matter of the biomass of autotrophic bacteria involved in the sulfur cycle. In the anaerobic waters below the C-layer, the organic matter of the biomass of autotrophs is consumed by the community of heterotrophic microorganisms; this results in weighting of the POC isotopic composition. Analysis of the data obtained and data available in the literature allows an inference to be made about the considerable seasonable variability of the POC delta 13C value, which depends on the ratio of terrigenic and planktonogenic components in the particulate organic matter.

Bacterial phosphating of mild (unalloyed) steel.

Mild (unalloyed) steel electrodes were incubated in phosphate-buffered cultures of aerobic, biofilm-forming Rhodococcus sp. strain C125 and Pseudomonas putida mt2. A resulting surface reaction leading to the formation of a corrosion-inhibiting vivianite layer was accompanied by a characteristic electrochemical potential (E) curve. First, E increased slightly due to the interaction of phosphate with the iron oxides covering the steel surface. Subsequently, E decreased rapidly and after 1 day reached -510 mV, the potential of free iron, indicating the removal of the iron oxides. At this point, only scattered patches of bacteria covered the surface. A surface reaction, in which iron was released and vivianite precipitated, started. E remained at -510 mV for about 2 days, during which the vivianite layer grew steadily. Thereafter, E increased markedly to the initial value, and the release of iron stopped. Changes in E and formation of vivianite were results of bacterial activity, with oxygen consumption by the biofilm being the driving force. These findings indicate that biofilms may protect steel surfaces and might be used as an alternative method to combat corrosion.

DNA degradation by the mixture of copper and catechol is caused by DNA-copper-hydroperoxo complexes, probably DNA-Cu(I)OOH.

Free hydroxyl radicals (free (.)OH), singlet oxygen ((1)O(2)), or (. )OH produced by DNA-copper-hydroperoxo complexes are possible DNA-damaging reactive oxygen species (ROS) in the reaction system containing copper, catechol, and DNA. para-Chlorobenzoic acid (pCBA) degradation studies revealed that CuCl(2) mixed with catechol produced free (.)OH. In the presence of DNA, however, inhibition of the pCBA degradation suggested that another ROS is responsible for the DNA degradation. Of a series of ROS scavengers investigated, only KI, NaN(3), and Na-formate-all of the salts tested-strongly inhibited the DNA degradation, suggesting that the ionic strength rather than the reactivity of the individual scavengers could be responsible for the observed inhibition. The ionic strength effect was confirmed by increasing the concentration of phosphate buffer, which is a poor (.)OH scavenger, and was interpreted as the result of destabilization of DNA-copper-hydroperoxo complexes. Piperidine-labile site patterns in DNA degraded by copper and catechol showed that the mixture of Cu(II) and catechol degrades DNA via the intermediate formation of a DNA-copper-hydroperoxo complex. Replacement of guanine by 7-deazaguanine did not retard the DNA degradation, suggesting that the DNA-copper-hydroperoxo complexes do not bind to the guanine N-7 as proposed in the literature.

Characterization of a second tfd gene cluster for chlorophenol and chlorocatechol metabolism on plasmid pJP4 in Ralstonia eutropha JMP134(pJP4).

Within the 5.9-kb DNA region between the tfdR and tfdK genes on the 2,4-dichlorophenoxyacetic acid (2,4-D) catabolic plasmid pJP4 from Ralstonia eutropha JMP134, we identified five open reading frames (ORFs) with significant homology to the genes for chlorocatechol and chlorophenol metabolism (tfdCDEF and tfdB) already present elsewhere on pJP4. The five ORFs were organized and assigned as follows: tfdD(II)C(II)E(II)F(II) and tfdB(II) (in short, the tfd(II) cluster), by analogy to tfdCDEF and tfdB (the tfd(I) cluster). Primer extension analysis of mRNA isolated from 2,4-D-grown R. eutropha JMP134 identified a single transcription start site in front of the first gene of the cluster, tfdD(II), suggesting an operon-like organization for the tfd(II) genes. By expressing each ORF in Escherichia coli, we confirmed that tfdD(II) coded for a chloromuconate cycloisomerase, tfdC(II) coded for a chlorocatechol 1, 2-dioxygenase, tfdE(II) coded for a dienelactone hydrolase, tfdF(II) coded for a maleylacetate reductase, and tfdB(II) coded for a chlorophenol hydroxylase. Dot blot hybridizations of mRNA isolated from R. eutropha JMP134 showed that both tfd(I) and tfd(II) genes are transcribed upon induction with 2,4-D. Thus, the functions encoded by the tfd(II) genes seem to be redundant with respect to those of the tfd(I) cluster. One reason why the tfd(II) genes do not disappear from plasmid pJP4 might be the necessity for keeping the regulatory genes for the 2,4-D pathway expression tfdR and tfdS.