Validation of Web Accessibility

Tato práce se zabývá přístupností webových stránek a možnostmi její validace. Základní částí je analýza současných metodik používaných ve světě - se zaměřením na USA, Evropu, a hlavně Českou republiku. Na ni navazuje přehled nejznámějších nástrojů pro kontrolu přístupnosti, z jejichž vlastností vychází specifikace, návrh a implementace vlastní aplikace, která je výsledkem této práce.This thesis deals with the accessibility of websites and its validation capabilities. The essential part is the analysis of current methodologies used in the world - with a focus on U.S., Europe, and especially the Czech Republic. It is followed by an overview of the best known tools for checking accessibility of their properties based specification, design and implementation of custom applications, which is the result of this work.

A Non-Linear Finite Element Model for the LHC Main Dipole Coil Cross-Section

The production of the dipole magnets for the Large Hadron Collider is at its final stage. Nevertheless, some mechanical instabilities are still observed for which no clear explanation has been found yet. A FE modelization of the dipole cold mass cross-section had already been developed at CERN, mainly for magnetic analysis, taking into account conductor blocks and a frictionless behavior. This paper describes a new ANSYS® model of the dipole coil cross-section, featuring individual turns inside conductor blocks, and implementing friction and the mechanical non-linear behavior of insulated cables. Preliminary results, comparison with measurements performed in industry and ongoing developments are discussed

A Finite Element Model for Mechanical Analysis of LHC Main Dipole Magnet Coils

After years of studies and observations, the mechanical stability of the LHC main dipole magnets still remains an open issue. The robustness of these magnets has already been asserted and their reliability in operation is not far from being proven. However, anomalous mechanical behaviors sometimes observed are not yet completely understood. A finite element model, which has been recently developed at CERN, aims at providing an instrument for better explaining these anomalies. Cable modeling and contact between elements, friction and mechanical hysteresis are the key features of this model. The simulation of the hysteresis experienced by the coil during collaring, presented here, is the starting point for the representation of the whole life cycle of the dipole coil

Molekularbiologische und biochemische Untersuchungen zur Biosynthese von Clorobiocin in Streptomyces roseochromogenes DS 12.976

Aminocoumarin antibiotics, such as novobiocin, clorobiocin and coumermycin A1, are produced by various Streptomyces strains and are very potent against gram-positive pathogenic bacteria including methicillin-resistant Staphylococcus strains. Bacterial DNA gyrase is the target of the aminocoumarin antibiotics. Until recently, novobiocin (Albamycin®, Pharmacia-Upjohn) was licensed in the United States for the treatment of infections with gram-positive bacteria and has been shown to enhance the cytotoxic activities of the anti-tumor drugs etoposide and teniposide. So far, the therapeutic use of aminocoumarin antibiotics is limited due to their low solubility in water, toxicity in eukaryotes and poor penetration in gram-negative bacteria. Combinatorial biosynthesis may offer a chance to develop novel aminocoumarins with improved properties. The biosynthetic gene clusters of novobiocin and coumermycin A1 were already sequenced in our laboratory. The first task of my thesis was to identify the biosynthetic gene cluster of the third "classical" aminocoumarin antibiotic, clorobiocin. The cluster was cloned by screening a cosmid library of Streptomyces roseochromogenes DS 12.976 with two heterologous probes from the novobiocin biosynthetic gene cluster. Sequence analysis revealed 29 open reading frames with striking similarity to the biosynthetic gene clusters of novobiocin and coumermycin A1. A comparison of the gene clusters of clorobiocin, novobiocin and coumermycin A1 showed that the structural differences between the three antibiotics were remarkably well reflected by differences in the organization of the biosynthetic gene clusters. The second part of my thesis was to elucidate the biosynthesis of 3-dimethylallyl-4-hydroxybenzoate moiety (Ring A) of clorobiocin and novobiocin by biochemical and molecular biological studies. Comparison of the three aminocoumarin clusters allowed us to identify three genes in the novobiocin and clorobiocin clusters for which no homologues existed in the coumermycin cluster. We speculated that these genes might be involved in the biosynthesis of Ring A (which is absent in coumermycin A1). These genes were: a) cloR and novR, which showed sequence similarity to putative aldolases; b) cloF and novF, which showed sequence similarities to dehydrogenases; c) cloQ and novQ, which did not show sequence similarities to known genes in the database. A biosynthetic pathway for clorobiocin was proposed in which activated ß-hydroxytyrosine was a common intermediate in the formation of the aminocoumarin ring (Ring B) and Ring A (Fig. 7, p. 40). However, analysis of S. roseochromogenes mutants (cloI- mutant, cloQ- mutant and cloR- mutant) revealed that Ring A and Ring B are formed by two distinct and independent pathways and that cloQ and cloR are essential for the formation of Ring A. CloQ was expressed in E. coli, purified and identified as an aromatic prenyltransferase. It is a soluble, monomeric, 35 kDa protein. 4-Hydroxyphenylpyruvate (4HPP) and dimethylallyl diphosphate (DMAPP) were identified as the substrates of this enzyme, with Km values determined as 25 and 35 µM, respectively. CloQ was found to be dissimilar from most prenyltransferases described so far and may indicate the existence of a new class of prenyltransferases. CloR was expressed in E. coli, purified and identified as a bifunctional non-heme iron oxygenase. It is a soluble, tetrameric protein. CloR converts 3-dimethylallyl-4-hydroxyphenylpyruvate via 3-dimethylallyl-4-hydroxymandelic acid (3DMA-4HMA) to Ring A. Therefore it catalyzes two consecutive oxidative decarboxylation steps. 18O2 labelling experiments showed that two oxygen atoms are incorporated into the intermediate 3DMA-4HMA in the first reaction step, but only one further oxygen is incorporated into the final product Ring A during the second reaction step. CloR does not show sequence similarity to known oxygenases. It apparently presents a novel member of the diverse family of the non-heme iron (II) and a-keto dependent oxygenases, with 3DMA-4HPP functioning both as a-ketoacid and as hydroxylation substrate. The reaction catalyzed by CloR represents a new pathway to benzoic acids in nature. In the third part of my thesis, a novO- mutant was created from the novobiocin producer S. spheroides, using a new inactivation method: the PCR targeting system. This mutant produced a derivative of novobiocin lacking the methyl group on the aminocoumarin ring.This provided functional proof for the role of novO in novobiocin biosynthesis. The mutant will be used for further experiments in combinatorial biosynthesis.Aminocoumarin-Antibiotika wie Novobiocin, Clorobiocin und Coumermycin A1 werden von verschiedenen Streptomycetenstämmen produziert und besitzen große Wirksamkeit gegenüber grampositiven pathogenen Bakterien, einschließlich Methicillin-resistenter Staphylococcusstämme. Das Zielmolekül der Aminocoumarin-Antibiotika ist die bakterielle DNA-Gyrase. Bis vor kurzen wurde Novobiocin (Albamycin®, Pharmacia-Upjohn) in den Vereinigten Staaten zur Behandlung von Infektionen mit grampositiven Bakterien eingesetzt. Es konnte gezeigt werden, dass es die cytotoxische Aktivität von Cytostatika wie Etoposid und Teniposid verstärkt. Aufgrund ihrer schlechten Wasserlöslichkeit, ihrer Toxizität für Eukaryoten und ihrer schlechten Aufnahme in gramnegative Bakterien ist der therapeutische Einsatz der Aminocoumarin-Antibiotika bislang stark eingeschränkt. Kombinatorische Biosynthese könnte eine Möglichkeit zur Entwicklung neuer Aminocoumarine mit verbesserten Eigenschaften darstellen. Voraussetzung dafür ist die genaue Kenntnis der Biosynthese der betreffenden Antibiotika. Die Biosynthesegencluster von Novobiocin und Coumermycin A1 waren bereits in unserem Labor sequenziert worden. Die erste Aufgabe meiner Doktorarbeit war die Identifizierung des Biosynthesegenclusters des dritten "klassischen" Aminocoumarin-Antibiotikums Clorobiocin. Das Cluster wurde durch Screening einer Cosmidbank von Streptomyces roseochromogenes DS 12.976 mit zwei heterologen Sonden des Novobiocin-Biosynthesegenclusters kloniert. Vergleichende Sequenzanalyse ergab 29 offene Leserahmen mit deutlicher Übereinstimmung zu den Biosynthesegenclustern von Novobiocin und Coumermycin A1. Ein Vergleich der Gencluster von Clorobiocin, Novobiocin und Coumermycin A1 zeigte, dass sich die strukturellen Unterschiede zwischen den drei Antibiotika erstaunlich genau in der Organisation der Cluster widerspiegeln. Der zweite Teil meiner Arbeit bestand in der Aufklärung der Biosynthese der 3-Dimethylallyl-4-hydroxybenzoat Gruppe (Ring A) von Clorobiocin und Novobiocin, sowohl durch biochemische als auch durch molekularbiologische Untersuchungen. Der Vergleich der drei Aminocoumarin-Cluster ermöglichte uns die Identifizierung von drei Gene, die sowohl im Novobiocin- als auch im Clorobiocin-Cluster vorkommen, für die aber keine Homologe im Coumermycin A1-Cluster existieren. Wir vermuteten, dass diese Gene an der Biosynthese von Ring A beteiligt sein könnten, da dieser in Coumermycin A1 nicht vorhanden ist. Diese Gene sind: a) cloR und novR, die Sequenzähnlichkeit zu putativen Aldolasen zeigten; b) cloF und novF, die Sequenzähnlichkeit zu Dehydrogenasen zeigten; und c) cloQ und novQ, welche keinerlei Sequenzähnlichkeit zu bekannten Genen in den Datenbanken aufwiesen. Es wurde ein Weg für die Biosynthese von Clorobiocin vorgeschlagen demzufolge aktiviertes ß-Hydroxytyrosin als gemeinsames Zwischenprodukt bei der Bildung des Aminocoumarinrings (Ring B) und des Ring A fungiert (Fig. 7, p. 40). Es konnte jedoch durch die Herstellung und Analyse von S. roseochromogenes Mutanten (cloI-, cloQ-, und cloR-) gezeigt werden, dass Ring A und Ring B auf zwei verschiedenen, unabhängigen Wegen gebildet werden. CloQ wurde in E. coli exprimiert und gereinigt und als eine aromatische Prenyltransferase identifiziert. CloQ ist ein lösliches, monomeres, 35 kD Protein. 4-Hydroxyphenylpyruvat (4HPP) und Dimethylallyldiphosphat (DMAPP) wurden als Substrate dieses Enzyms identifiziert und die Km-Werte mit 25 µM bzw. 35 µM bestimmt. Wir konnten zeigen, dass CloQ sich von den meisten bisher bekannten Prenyltransferasen unterscheidet und möglicherweise einer neuen Klasse von Prenyltransferasen angehört. CloR wurde in E. coli exprimiert und gereinigt und als eine bifunktionale non-Häm Eisen-Oxygenase identifiziert. Es ist ein lösliches, tetrameres Protein. CloR setzt 3-Dimethylallyl-4-hydroxyphenylpyruvat über 3-Dimethylallyl-4-hydroxymandelsäure zu Ring A um. Dies erfolgt in zwei aufeinanderfolgenden Decarboxylierungsschritten. 18O2-Markierungsexperimente zeigten, dass im ersten Reaktionsschritt zwei Sauerstoffatome in das 3DMA-4HMA Zwischenprodukt eingebaut werden, wohingegen aber im zweiten Schritt nur ein weiteres Sauerstoffatom in das Ring A Endprodukt inkorporiert wird. CloR zeigt keine Sequenzähnlichkeit zu bekannten Oxygenasen. Anscheinend stellt es ein neues Mitglied der divergenten Familie der non-Häm Eisen (II) und a-Ketosäure-abhängigen Oxygenasen dar, bei dem 3DMA-4HPP sowohl als a-Ketosäure als auch als Hydroxylierungssubstrat fungiert. Die von CloR katalysierte Reaktion stellt einen neuen Weg zur Biosynthese von Benzoesäuren dar

The salt influence on aprotic electrolytes

Tato práce se zabývá kapalnými aprotickými elektrolyty s rozpouštědly na bázi směsí sulfolanu s dalším rozpouštědlem, ve spojení se sodnými a lithnými solemi. U těchto elektrolytů jsou vyhodnocovány jejích vlastnosti a to vzhledem k jejich použití v lithno – iontových akumulátorech z pohledu bezpečnosti a elektrických vlastností. Pro bezpečnostní rizika je měřen bod vzplanuti a z elektrických vlastností je měřena měrná elektrická vodivost a u vybraných vzorků stanovena kapacita dvojvrstvy.This thesis deals with the liquid aprotic electrolytes with solvent on the basis of sulfolane blends with other solvent, in combination with the sodium and lithium salts. In these electrolytes are evaluated their properties due to their use in lithium - ion batteries in terms of safety and electrical properties. For safety risk is measured flashpoint and for electrical properties is measured electrical conductivity for selected samples are intended capacities of double layers.

Substance use and psychiatric symptoms in subjects referred to the Drug Addiction Services by the Prefecture

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Prenatal Insecticide Exposures and Birth Weight and Length among an Urban Minority Cohort

We reported previously that insecticide exposures were widespread among minority women in New York City during pregnancy and that levels of the organophosphate chlorpyrifos in umbilical cord plasma were inversely associated with birth weight and length. Here we expand analyses to include additional insecticides (the organophosphate diazinon and the carbamate propoxur), a larger sample size (n = 314 mother–newborn pairs), and insecticide measurements in maternal personal air during pregnancy as well as in umbilical cord plasma at delivery. Controlling for potential confounders, we found no association between maternal personal air insecticide levels and birth weight, length, or head circumference. For each log unit increase in cord plasma chlorpyrifos levels, birth weight decreased by 42.6 g [95% confidence interval (CI), −81.8 to −3.8, p = 0.03] and birth length decreased by 0.24 cm (95% CI, −0.47 to −0.01, p = 0.04). Combined measures of (ln)cord plasma chlorpyrifos and diazinon (adjusted for relative potency) were also inversely associated with birth weight and length (p < 0.05). Birth weight averaged 186.3 g less (95% CI, −375.2 to −45.5) among newborns with the highest compared with lowest 26% of exposure levels (p = 0.01). Further, the associations between birth weight and length and cord plasma chlorpyrifos and diazinon were highly significant (p ≤ 0.007) among newborns born before the 2000–2001 U.S. Environmental Protection Agency’s regulatory actions to phase out residential use of these insecticides. Among newborns born after January 2001, exposure levels were substantially lower, and no association with fetal growth was apparent (p > 0.8). The propoxur metabolite 2-isopropoxyphenol in cord plasma was inversely associated with birth length, a finding of borderline significance (p = 0.05) after controlling for chlorpyrifos and diazinon. Results indicate that prenatal chlorpyrifos exposures have impaired fetal growth among this minority cohort and that diazinon exposures may have contributed to the effects. Findings support recent regulatory action to phase out residential uses of the insecticides

An off-momentum beam loss feedback controller and graphical user interface for the LHC

During LHC operation, a campaign to validate the configuration of the LHC collimation system is conducted every few months. This is performed by means of loss maps, where specific beam losses are voluntarily generated with the resulting loss patterns compared to expectations. The LHC collimators have to protect the machine from both betatron and off-momentum losses. In order to validate the off-momentum protection, beam losses are generated by shifting the RF frequency using a low intensity beam. This is a delicate process that, in the past, often led to the beam being dumped due to excessive losses. To avoid this, a feed-back system based on the100 Hz data stream from the LHC Beam Loss system has been implemented. When given a target RF frequency, the feedback system approaches this frequency in steps while monitoring the losses until the selected loss pattern conditions are reached, so avoiding the excessive losses that lead to a beam dump. This paper will describe the LHC off-momentum beam loss feedback system and the results achieved.peer-reviewe