High-throughput procedure for tick surveys of tick-borne encephalitis virus and its application in a national surveillance study in Switzerland

Tick-borne encephalitis (TBE), a viral infection of the central nervous system, is endemic in many Eurasian countries. In Switzerland, TBE risk areas have been characterized by geographic mapping of clinical cases. Since mass vaccination should significantly decrease the number of TBE cases, alternative methods for exposure risk assessment are required. We established a new PCR-based test for the detection of TBE virus (TBEV) in ticks. The protocol involves an automated, high-throughput nucleic acid extraction method (QIAsymphony SP system) and a one-step duplex real-time reverse transcription-PCR (RT-PCR) assay for the detection of European subtype TBEV, including an internal process control. High usability, reproducibility, and equivalent performance for virus concentrations down to 5 x 10(3) viral genome equivalents/microl favor the automated protocol compared to the modified guanidinium thiocyanate-phenol-chloroform extraction procedure. The real-time RT-PCR allows fast, sensitive (limit of detection, 10 RNA copies/microl), and specific (no false-positive test results for other TBEV subtypes, other flaviviruses, or other tick-transmitted pathogens) detection of European subtype TBEV. The new detection method was applied in a national surveillance study, in which 62,343 Ixodes ricinus ticks were screened for the presence of TBE virus. A total of 38 foci of endemicity could be identified, with a mean virus prevalence of 0.46%. The foci do not fully agree with those defined by disease mapping. Therefore, the proposed molecular test procedure constitutes a prerequisite for an appropriate TBE surveillance. Our data are a unique complement of human TBE disease case mapping in Switzerland.

PM(10) remote sensing from geostationary SEVIRI and polar-orbiting MODIS sensors over the complex terrain of the European Alpine region

The subject of this study is to investigate the capability of spaceborne remote sensing data to predict ground concentrations of PM10 over the European Alpine region using satellite derived Aerosol Optical Depth (AOD) from the geostationary Spinning Enhanced Visible and InfraRed Imager (SEVIRI) and the polar-orbiting MODerate resolution Imaging Spectroradiometer (MODIS). The spatial and temporal resolutions of these aerosol products (10 km and 2 measurements per day for MODIS, ∼ 25 km and observation intervals of 15 min for SEVIRI) permit an evaluation of PM estimation from space at different spatial and temporal scales. Different empirical linear relationships between coincident AOD and PM10 observations are evaluated at 13 ground-based PM measurement sites, with the assumption that aerosols are vertically homogeneously distributed below the planetary Boundary Layer Height (BLH). The BLH and Relative Humidity (RH) variability are assessed, as well as their impact on the parameterization. The BLH has a strong influence on the correlation of daily and hourly time series, whilst RH effects are less clear and smaller in magnitude. Despite its lower spatial resolution and AOD accuracy, SEVIRI shows higher correlations than MODIS (rSEV∼ 0.7, rMOD∼ 0.6) with regard to daily averaged PM10. Advantages from MODIS arise only at hourly time scales in mountainous locations but lower correlations were found for both sensors at this time scale (r∼ 0.45). Moreover, the fraction of days in 2008 with at least one satellite observation was 27% for SEVIRI and 17% for MODIS. These results suggest that the frequency of observations plays an important role in PM monitoring, while higher spatial resolution does not generally improve the PM estimation. Ground-based Sun Photometer (SP) measurements are used to validate the satellite-based AOD in the study region and to discuss the impact of aerosols' micro-physical properties in the empirical models. A lower error limit of 30 to 60% in the PM10 assessment from space is estimated in the study area as a result of AOD uncertainties, variability of aerosols properties and the heterogeneity of ground measurement sites. It is concluded that SEVIRI has a similar capacity to map PM as sensors on board polar-orbiting platforms, with the advantage of a higher number of observations. However, the accuracy represents a serious limitation to the applicability of satellites for ground PM mapping, especially in mountainous areas.

Adsorption behavior of redox-active suppressor additives: Combined electrochemical and STM studies

The redox chemistry and the related surface phase behavior of Safranine (SAF) and Janus Green B (JGB) have been studied by means of cyclic voltammetry in combination with in situ Scanning Tunneling Microscopy using HOPG (Highly Oriented Pyrolytic Graphite) and single crystalline Cu(1 0 0) as model substrates, both revealing different widths of the accessible potential windows. JGB and SAF serve as prototypical heterocyclic suppressor/leveler additives that are used for the metallization of 3D-TSVs (3D Through Silicon Vias) following a classical "leveling" concept. SAF can be considered as the reductive decomposition product of JGB that is formed at the copper/electrolyte interface upon electroplating. Both additives reveal a pronounced pH-dependent redox-chemistry with redox-transitions lying close to or even beyond the anodic limit of the copper potential window. Affected by these redox-processes are in particular the aromatic cores of those heterocycles that can be (quasi)reversibly reduced by a two electron transfer process within the potential window of copper. Therefore we identify the reduced form of those dyes as the active components for the suppressing/leveling effect in copper plating. STM data clearly shows a dye surface phase behavior that is crucially determined by its potential-dependent redox-chemistry. This will be exemplarily discussed for the SAF dye. On chloride-modified Cu(1 0 0) mono-reduced SAF forms a structurally well-defined monolayer of cationic stacking polymers. However, this coupled anion/cation layer reveals only minor suppressing capabilities with respect to the copper dissolution and deposition processes. Complete reduction of the aromatic heterocycle finally leads to the 3D precipitation of hydrophobic reaction products. 3D clusters of this SAF precipitate are discussed as the active structural motif for the suppressing effect of these dyes. (C) 2011 Elsevier Ltd. All rights reserved.

Sensitivity of pelagic calcification to ocean acidification

Ocean acidification might reduce the ability of calcifying plankton to produce and maintain their shells of calcite, or of aragonite, the more soluble form of CaCO3. In addition to possibly large biological impacts, reduced CaCO3 production corresponds to a negative feedback on atmospheric CO2. In order to explore the sensitivity of the ocean carbon cycle to increasing concentrations of atmospheric CO2, we use the new biogeochemical Bern3D/PISCES model. The model reproduces the large scale distributions of biogeochemical tracers. With a range of sensitivity studies, we explore the effect of (i) using different parameterizations of CaCO3 production fitted to available laboratory and field experiments, of (ii) letting calcite and aragonite be produced by auto- and heterotrophic plankton groups, and of (iii) using carbon emissions from the range of the most recent IPCC Representative Concentration Pathways (RCP). Under a high-emission scenario, the CaCO3 production of all the model versions decreases from ~1 Pg C yr−1 to between 0.36 and 0.82 Pg C yr−1 by the year 2100. The changes in CaCO3 production and dissolution resulting from ocean acidification provide only a small feedback on atmospheric CO2 of −1 to −11 ppm by the year 2100, despite the wide range of parameterizations, model versions and scenarios included in our study. A potential upper limit of the CO2-calcification/dissolution feedback of −30 ppm by the year 2100 is computed by setting calcification to zero after 2000 in a high 21st century emission scenario. The similarity of feedback estimates yielded by the model version with calcite produced by nanophytoplankton and the one with calcite, respectively aragonite produced by mesozooplankton suggests that expending biogeochemical models to calcifying zooplankton might not be needed to simulate biogeochemical impacts on the marine carbonate cycle. The changes in saturation state confirm previous studies indicating that future anthropogenic CO2 emissions may lead to irreversible changes in ΩA for several centuries. Furthermore, due to the long-term changes in the deep ocean, the ratio of open water CaCO3 dissolution to production stabilizes by the year 2500 at a value that is 30–50% higher than at pre-industrial times when carbon emissions are set to zero after 2100.

Comparative stability studies of poly(2-methyl-2-oxazoline) and poly(ethylene glycol) brush coatings

Non-fouling surfaces that resist non-specific adsorption of proteins, bacteria, and higher organisms are of particular interest in diverse applications ranging from marine coatings to diagnostic devices and biomedical implants. Poly(ethylene glycol) (PEG) is the most frequently used polymer to impart surfaces with such non-fouling properties. Nevertheless, limitations in PEG stability have stimulated research on alternative polymers that are potentially more stable than PEG. Among them, we previously investigated poly(2-methyl-2-oxazoline) (PMOXA), a peptidomimetic polymer, and found that PMOXA shows excellent anti-fouling properties. Here, we compare the stability of films self-assembled from graft copolymers exposing a dense brush layer of PEG and PMOXA side chains, respectively, in physiological and oxidative media. Before media exposure both film types prevented the adsorption of full serum proteins to below the detection limit of optical waveguide in situ measurements. Before and after media exposure for up to 2 weeks, the total film thickness, chemical composition, and total adsorbed mass of the films were quantified using variable angle spectroscopic ellipsometry (VASE), X-ray photoelectron spectroscopy (XPS), and optical waveguide lightmode spectroscopy (OWLS), respectively. We found (i) that PMOXA graft copolymer films were significantly more stable than PEG graft copolymer films and kept their protein-repellent properties under all investigated conditions and (ii) that film degradation was due to side chain degradation rather than due to copolymer desorption.

Combined use of transcranial magnetic stimulation and metal electrode implants: a theoretical assessment of safety considerations

This paper provides a theoretical assessment of the safety considerations encountered in the simultaneous use of transcranial magnetic stimulation (TMS) and neurological interventions involving implanted metallic electrodes, such as electrocorticography. Metal implants are subject to magnetic forces due to fast alternating magnetic fields produced by the TMS coil. The question of whether the mechanical movement of the implants leads to irreversible damage of brain tissue is addressed by an electromagnetic simulation which quantifies the magnitude of imposed magnetic forces. The assessment is followed by a careful mechanical analysis determining the maximum tolerable force which does not cause irreversible tissue damage. Results of this investigation provide useful information on the range of TMS stimulator output powers which can be safely used in patients having metallic implants. It is shown that conventional TMS applications can be considered safe when applied on patients with typical electrode implants as the induced stress in the brain tissue remains well below the limit of tissue damage.

Performance Tuning Non-Uniform Sampling for Sensitivity Enhancement of Signal-Limited Biological NMR

Non-uniform sampling (NUS) has been established as a route to obtaining true sensitivity enhancements when recording indirect dimensions of decaying signals in the same total experimental time as traditional uniform incrementation of the indirect evolution period. Theory and experiments have shown that NUS can yield up to two-fold improvements in the intrinsic signal-to-noise ratio (SNR) of each dimension, while even conservative protocols can yield 20-40 % improvements in the intrinsic SNR of NMR data. Applications of biological NMR that can benefit from these improvements are emerging, and in this work we develop some practical aspects of applying NUS nD-NMR to studies that approach the traditional detection limit of nD-NMR spectroscopy. Conditions for obtaining high NUS sensitivity enhancements are considered here in the context of enabling H-1,N-15-HSQC experiments on natural abundance protein samples and H-1,C-13-HMBC experiments on a challenging natural product. Through systematic studies we arrive at more precise guidelines to contrast sensitivity enhancements with reduced line shape constraints, and report an alternative sampling density based on a quarter-wave sinusoidal distribution that returns the highest fidelity we have seen to date in line shapes obtained by maximum entropy processing of non-uniformly sampled data.

Determination of free and total valproic acid in human plasma by capillary electrophoresis with contactless conductivity detection

A new approach for the determination of free and total valproic acid in small samples of 140 μL human plasma based on capillary electrophoresis with contactless conductivity detection is proposed. A dispersive liquid-liquid microextraction technique was employed in order to remove biological matrices prior to instrumental analysis. The free valproic acid was determined by isolating free valproic acid from protein-bound valproic acid by ultrafiltration under centrifugation of 100 μL sample. The filtrate was acidified to turn valproic acid into its protonated neutral form and then extracted. The determination of total valproic acid was carried out by acidifying 40 μL untreated plasma to release the protein-bound valproic acid prior to extraction. A solution consisting of 10 mM histidine, 10 mM 3-(N-morpholino)propanesulfonic acid and 10 μM hexadecyltrimethylammonium bromide of pH 6.5 was used as background electrolyte for the electrophoretic separation. The method showed good linearity in the range of 0.4-300 μg/mL with a correlation coefficient of 0.9996. The limit of detection was 0.08 μg/mL, and the reproducibility of the peak area was excellent (RSD=0.7-3.5%, n=3, for the concentration range from 1 to 150 μg/mL). The results for the free and total valproic acid concentration in human plasma were found to be comparable to those obtained with a standard immunoassay. The corresponding correlation coefficients were 0.9847 for free and 0.9521 for total valproic acid.

Ocular penetration of caspofungin in a rabbit uveitis model

BACKGROUND: Little is known about the ocular penetration of echinocandin antifungals. We studied the ocular distribution of systemically administered caspofungin in a rabbit uveitis model. METHODS: Caspofungin (1 mg/kg per day) was given intravenously to rabbits as a single dose or as repeated daily doses on 7 days starting 24 h after induction of unilateral uveitis by intravitreal endotoxin injection. Caspofungin concentrations were determined by high-performance liquid chromatography in the cornea, aqueous humor, vitreous humor, and serum 4, 8, 16, and 24 h after administration of a single dose and 24 h after the last of seven doses. RESULTS: The mean caspofungin concentration in the aqueous of the inflamed eye 4 and 8 h after single-dose administration was 1.30 +/- 0.39 mug/ml and 1.12 +/- 0.34 mug/ml, respectively. Drug concentrations decreased to 0.24 +/- 0.09 mug/ml at 16 h and 0.26 +/- 0.14 mug/ml at 24 h. In the vitreous of inflamed eyes drug levels were undetectable at all time points. No drug was found in the aqueous of inflamed eyes 24 h after the last of seven repeated doses, and the vitreous only contained trace amounts. In the corneas of inflamed eyes concentrations reached 1.64 +/- 0.48 mug/g at 4 h, peaked at 2.16 +/- 1.14 mug/g at 8 h, and declined to 1.87 +/- 0.52 mug/g and 1.49 +/- 0.48 mug/g at 16 and 24 h, respectively. After repeated dosing, corneal concentrations of caspofungin were 0.8 and 1.0 mug/g and below the limit of detection in two of four animals. In non-inflamed eyes no drug was detectable in the aqueous and vitreous humor, and the corneas at any time point. CONCLUSIONS: In our model, caspofungin reached therapeutically relevant levels in the aqueous and cornea but not in the vitreous humor of inflamed eyes. Intraocular drug deposition was critically dependent on a disrupted blood-eye barrier. These findings suggest a limited role for caspofungin in the treatment of fungal endophthalmitis.

An unusual patient with hypercalciuria, recurrent nephrolithiasis, hypomagnesemia and G227R mutation of Paracellin-1. An unusual patient with hypercalciuria and hypomagnesemia unresponsive to thiazide diuretics

A 19-year-old female patient with hypercalciuria and recurrent nephrolithiasis/urinary tract infection unresponsive to thiazide type diuretics is presented. The patient first experienced nephrolithiasis at the age of 4 years. Afterwards, recurrent passages of stones and urinary tract infection occurred. On diagnostic evaluation at the age of 19 years, she also had hypocitraturia and hypomagnesemia. Her serum calcium concentrations were near the lower limit of normal (8.5-8.8 mg/dl; normal range: 8.5-10.5), her serum magnesium concentrations were 1.15-1.24 mg/dl (normal range: 1.4-2.5) and urinary calcium excretion was 900 mg/24 h. PTH concentrations were increased (110-156 pg/ml; normal range: 10-65). We tried to treat the patient with hydrochlorothiazide at a dose of 50 mg/day. During treatment with thiazide diuretics, PTH concentration remained high and the patient had recurrent urinary tract infections and passages of stones. Serum magnesium concentration did not normalize even under the parenteral magnesium infusion. Her mother had a history of nephrolithiasis 20 years ago. Severe hypomagnesemia in association with hypercalciuria/urinary stones is reported as a rare autosomal recessive disorder caused by impaired reabsorption of magnesium and calcium in the thick assending limp of Henle's loop. Recent studies showed that mutations in the CLDN16 gene encoding paracellin-1 cause the disorder. In exon 4, a homozygous nucleotide exchange (G679C) was identified for the patient. This results in a point mutation at position Glycine227, which is replaced by an Arginine residue (G227R). The mother was heterozygous for this mutation. G227 is located in the fourth transmembrane domain and is highly conserved in the claudin gene family. This case indicates the pathogenetic role of paracellin-1 mutation in familial hypomagnesemia with hypercalciuria and nephrocalcinosis and further underlines the risk of stone formation in heterozygous mutation carriers.

Generic detection of coronaviruses and differentiation at the prototype strain level by reverse transcription-PCR and nonfluorescent low-density microarray

A nonfluorescent low-cost, low-density oligonucleotide array was designed for detecting the whole coronavirus genus after reverse transcription (RT)-PCR. The limit of detection was 15.7 copies/reaction. The clinical detection limit in patients with severe acute respiratory syndrome was 100 copies/sample. In 39 children suffering from coronavirus 229E, NL63, OC43, or HKU1, the sensitivity was equal to that of individual real-time RT-PCRs.

Integration of external and internal dosimetry in Switzerland

Individual monitoring regulations in Switzerland are based on the ICRP60 recommendations. The annual limit of 20 mSv for the effective dose applies to the sum of external and internal radiation. External radiation is monitored monthly or quarterly with TLD, DIS or CR-39 dosemeters by 10 approved external dosimetry services and reported as H(p)(10) and H(p)(0.07). Internal monitoring is done in two steps. At the workplace, simple screening measurements are done frequently in order to recognise a possible incorporation. If a nuclide dependent activity threshold is exceeded then one of the seven approved dosimetry services for internal radiation does an incorporation measurement to assess the committed effective dose E(50). The dosimetry services report all the measured or assessed dose values to the employer and to the National Dose Registry. The employer records the annually accumulated dose values into the individual dose certificate of the occupationally exposed person, both the external dose H(p)(10) and the internal dose E(50) as well as the total effective dose E = H(p)(10)+E(50). Based on the national dose registry an annual report on the dosimetry in Switzerland is published which contains the statistics for the total effective dose, as well as separate statistics for external and internal exposure.

Impact of changing definitions for myocardial infarction: a report from the AMIS registry

BACKGROUND: To assess the impact of the new definitions of myocardial infarction, we retrospectively analyzed 9190 patients from 63 hospitals with reported peak troponin values included between 2001 and 2007 in the Swiss AMIS (Acute Myocardial Infarction in Switzerland) Plus registry. METHODS: Patients were classified as belonging to the "classic" myocardial infarction group (peak total CK or CK-MB above the upper limit of normal, or troponin T [TnT] >0.1 microg/L or troponin I [TnI] >0.1-0.8 microg/L [depending on the assay]) or "new" myocardial infarction group (TnT >0.01 microg/L or TnI >0.01-0.07 microg/L). RESULTS: There were 489 patients in the "new" group who were similar to the 8701 "classic" patients in terms of age, sex, and prevalence of both diabetes and renal failure, but more frequently had a history of prior coronary artery disease, hypertension, and hyperlipidemia. At admission, they less frequently had ST elevation on their electrocardiogram, were more frequently in Killip class I, and received less primary percutaneous coronary intervention. Hospital mortality was 3.5% in the "new" and 6.7% in the "classic" myocardial infarction group (P=.004). In a subset of patients with a longer follow-up, mortality at 3 and 12 months was 1% and 5.6%, respectively, for "new" and 1.6% and 4%, respectively, for "classic" myocardial infarction (NS). CONCLUSIONS: Patients with minimal elevation of serum troponin have smaller infarctions, less aggressive treatment, fewer early complications, and a better early prognosis than patients with higher serum biomarker levels. After discharge, however, their prognosis currently appears no different from that of patients with a "classic" myocardial infarction event.

Ultrasound-guided paravertebral puncture and placement of catheters in human cadavers: an imaging study

BACKGROUND: During paravertebral block, the anterolateral limit of the paravertebral space, which consists of the pleura, should preferably not be perforated. Also it is possible that, during the block, the constant superior costotransverse ligament can be missed in the loss-of-resistance technique. We therefore aimed to develop a new technique for an ultrasound-guided puncture of the paravertebral space. METHODS: We performed 20 punctures and catheter placements in 10 human cadavers. A sonographic view showing the pleura and the superior costotransverse ligament was obtained with a slightly oblique scan using a curved array transducer. After inline approach, injection of 10 ml normal saline confirmed the correct position of the needle tip, distended the space, and enabled catheter insertion. The spread of contrast dye injected through the catheters was assessed by CT scans. RESULTS: The superior costotransverse ligament and the paravertebral space were easy to identify. The needle tip reached the paravertebral space without problems under visualization. In contrast, the introduction of the catheter was difficult. The CT scan revealed a correct paravertebral spread of contrast in 11 cases. Out of the remaining, one catheter was found in the pleural space, in six cases there was an epidural, and in two cases there was a prevertebral spread of contrast dye. CONCLUSIONS: We successfully developed a technique for an accurate ultrasound-guided puncture of the paravertebral space. We also showed that when a catheter is introduced through the needle with the tip lying in the paravertebral space, there is a high probability of catheter misplacement into the epidural, mediastinal, or pleural spaces.

PCR and real-time PCR primers developed for detection and identification of Bifidobacterium thermophilum in faeces

BACKGROUND: Culture-independent methods based on the 16S ribosomal RNA molecule are nowadays widely used for assessment of the composition of the intestinal microbiota, in relation to host health or probiotic efficacy. Because Bifidobacterium thermophilum was only recently isolated from human faeces until now, no specific real-time PCR (qPCR) assay has been developed for detection of this species as component of the bifidobacterial community of the human intestinal flora. RESULTS: Design of specific primers and probe was achieved based on comparison of 108 published bifidobacterial 16S rDNA sequences with the recently published sequence of the human faecal isolate B. thermophilum RBL67. Specificity of the primer was tested in silico by similarity search against the sequence database and confirmed experimentally by PCR amplification on 17 Bifidobacterium strains, representing 12 different species, and two Lactobacillus strains. The qPCR assay developed was linear for B. thermophilum RBL67 DNA quantities ranging from 0.02 ng/microl to 200 ng/microl and showed a detection limit of 10(5) cells per gram faeces. The application of this new qPCR assay allowed to detect the presence of B. thermophilum in one sample from a 6-month old breast-fed baby among 17 human faecal samples tested. Additionally, the specific qPCR primers in combination with selective plating experiments led to the isolation of F9K9, a faecal isolate from a 4-month old breast-fed baby. The 16S rDNA sequence of this isolate is 99.93% similar to that of B. thermophilum RBL67 and confirmed the applicability of the new qPCR assay in faecal samples. CONCLUSION: A new B. thermophilum-specific qPCR assay was developed based on species-specific target nucleotides in the 16S rDNA. It can be used to further characterize the composition of the bifidobacterial community in the human gastrointestinal tract. Until recently, B. thermophilum was considered as a species of animal origin, but here we confirm with the application of this new PCR assay the presence of B. thermophilum strains in the human gut.