Le typage bactérien est une mesure de contrôle essentielle pour lutter contre la diffusion des bactéries multirésistantes, mais les techniques actuelles sont longues et coûteuses. L'objectif de cette thèse était d'évaluer la capacité de la technique MALDI-TOF MS à typer les 3 principales espèces bactériennes responsables d'infections nosocomiales - Escherichia coli, Staphylococcus aureus et Pseudomonas aeruginosa. L'analyse par MALDI-TOF MS permet d'identifier les souches de E. coli appartenant au phylogroupe B2, souches les plus virulentes parmi les souches extra-intestinales, et au STI 31, clone très impliqué dans la dissémination des P-lactamases à spectre étendu. Chez S. aureus, cette technique reconnait les souches appartenant au CC398, impliquées dans une proportion importante d'infections graves chez des personnes fragiles. La technique MALDI-TOF MS identifie également cinq clones de P. aeruginosa à haut risque épidémique - STI 11, STI 75, ST235, ST253, ST395. Si nous avons confirmé des pics caractéristiques du phylogroupe B2 décrit également par d'autres auteurs, les pics biomarqueurs identifiant E. coli ST131 ou des clones épidémiques de S. aureus varient suivant les études. Différents paramètres peuvent influencer les résultats de typage par MALDI-TOF MS et doivent donc être standardisés. La technique MALDI-TOF MS permet d'identifier certains clones épidémiques. En gardant à l'esprit que le choix d'une méthode de typage doit être fait en fonction des objectifs mais aussi des performances des différents systèmes disponibles, la technique MALDI-TOF MS pourrait se positionner comme un outil de typage de première ligne. / Bacterial typing is crucial to tackle the spread of bacterial pathogens but current methods are time-consuming and costly. The objective of this study was to evaluate the ability of MALDI-TOF MS to type the 3 main bacterial species re.sponsible for nosocomial infections - Escherichia coti, Staphylococcus aureus and Pseudomonas aeruginosa. Analysis of MALDI-TOF mass spectra allow the identification (i) of E. coti isolates belonging to phylogroup B2, that fosters the most virulent extra-intestinal strains, and (ii) of E. coli STl 31, a clone involved in the dissemination of extended-spectrum β-lactamases. MALDI-TOF MS also recognizes S. aureus isolates belonging to CC398, a pathogen responsible for invasive infections in fragile patients and associated with a higher 30-day mortality. Furthermore the MALDI-TOF MS based typing method identifies the major high risk epidemic clones of P. aeruginosa STl 11, STl 75, ST235, ST253, and ST395. We confirmed phylogroup B2 specific peaks also described by other authors. However the identification of E. coli STl 31 or epidemic clones of S. aureus is based on biomarkers that differs between studies. Different parameters can influence the MALDI-TOF MS typing results and must therefore be standardized. MALDI-TOF-based typing methods allow the detection of epidemic pathogens. Keeping in mind that the choice of a method for routine bacterial typing should be made according to the objectives but also the performance of the different systems available, the MALDI-TOF MS technique could become a first-line typing method.

This diploma thesis deals with Bisphenol A, especially with its impact on the aquatic ecosystem. Information about its properties, production and current use are summarized here. Its harmful impact on the environment, especially on the aquatic ecosystem and the human body is characterized. Also the methods of its degradation within the aquatic environment have been described. A summary of the options of a determination of Bisphenol A in water samples is incorporated and the method of gas chromatography with mass spectrometry (GC/TOF MS) and comprehensive two-dimensional gas chromatography with mass spectrometry (GCxGC/TOF-MS) is compared in the experimental part. Analytical determination precedes the isolation of the analyte from the water samples by solid phase extraction (SPE) using SupelcleanTM ENVITM - 18 and derivatization using the silylation reagent, N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA). The result of this work is the analysis of a series of real samples from wastewater treatment plants Brno Modřice and Luhačovice by a two-dimensional gas chromatography with mass spectrometry (GCxGC/TOF-MS).

Over the last decades organic light-emitting diodes (OLEDs) and organic solar cells (OSCs) have gained considerable attention as efficient, flexible, lightweight, and potentially low-cost technology for lighting and display applications or as a renewable energy source, respectively. However, achieving long-term stability remains challenging. Revealing and understanding aging processes is therefore of great interest. This work presents fundamental investigations to understand and circumvent organic device degradation. In the first part, single materials used in organic devices were investigated. By tailoring an attenuated total reflection infrared (ATR-IR) spectrometer to the specific needs and subsequent measurements, it is shown that the tris(8-hydroxyquinoline)aluminum (Alq3) molecule, a well known fluorescent green emitter, degrades during air exposure by the formation of carbonyl groups. By using a laser desorption/ionization time of flight mass spectrometer (LDI-TOF-MS) it was shown that a,w-bis-(dicyanovinylen)-sexithiophen (DCV6T-Bu4), a well known small-molecule material which is used as part of the active layer, reacts with oxygen during ultraviolet (UV) irradiation. By using climate boxes and a sun simulator the impact of dry and humid air as well as sunlight on C60, a widely-used acceptor molecule in organic solar cells, was investigated. The breaking of the C60 cage to C58 and C56 and the further reaction of these components with oxygen as well as the dimerization of C58 and C56 molecules were found. The degradation products such as C58O increase with air exposure time but they are independent of the humidity level of the ambient air as well as sunlight irradiation. Subsequent annealing leads to a decrease of the C58O concentration. Many efficient n-dopants are prone to degradation in air, due to the low ionization potentials, thereby limiting the processing conditions. It was found that the air exposure of the highly efficient n-dopant tetrakis(1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidinato)ditungsten(II) (W2(hpp)4) leads to oxidation reactions of the molecule to [W(hpp)2 + O] and other degradation products. The decay constant of W2(hpp)4 and the matching mean growth time of the [W(hpp)2 + O] degradation as well as a second very quick degradation of the dopant could be determined. The two decay constants can be explained by the assumption that W2(hpp)4 molecules, which are involved in the charge transfer, do degrade slower due to the fact that the charge transfer leads to a downshift of the energy levels of the W2(hpp)4 molecule. Apart from the properties of the organic materials, other effects such as the impact of different purification systems on the material purity as well as the dependence of material purity on the OLED lifetime has been investigated. No correlations between the purification grade and the amount of impurities were found. OLEDs which contain N,N\'-di(naphthalen-1-yl)-N,N\'-diphenyl-benzidine (alpha-NPD) purified in a vertically interlaced stainless steel sublimation systems shows slightly higher external quantum efficiencies compared to tube-based vacuum sublimation systems. The devices which contain alpha-NPD purified by a sublimation system have an extended lifetime. Finally, the impact of residual gases during device fabrication on OLED lifetime and electrical characteristics was investigated. It was found that water vapor introduces an additional series resistance to the OLED, while the other gases do not influence the electric characteristics. The presence of nitrogen or oxygen impacts the lifetime of the OLEDs by the same amount. Nitrogen is non-reactive, this leads to the conclusion that the influence of nitrogen and oxygen on the OLED lifetime is of non-chemical nature, such as changes in the morphology of the organic layers. Water vapor introduces an additional, even faster degradation process within the first hours of OLED operation. As major sources of device degradation, the dimerization of 4,7-diphenyl-1,10-phenanthroline (BPhen) as well as the complexation reaction of alpha-NPD with a bis(1-phenylisoquinoline)iridium(III) (Ir(piq)2) fragment was identified.

Introduktion Antibiotikaresistens är ett globalt växande problem. Till gruppen β-laktamantibiotika hör piperacillin-tazobaktam och cefotaxim som båda verkar genom att försvaga cellväggen med kovalenta bindningar till peptidoglykanlagret som lyserar cellen. E. coli och K. pneumoniae tillhör gruppen Enterobacteriaceae, som är en del av den humana tarmfloran och ofta förekommande vid urinvägsinfektion och sepsis. Utvidgat Spektrum β-Laktamas (ESBL) är ett enzym som finns hos Enterobacteriaceae och som hydrolyserar β-laktamantibiotika. Matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) är en kvalitativ analysteknik för detektion av kemiska föreningar i avseende på massa och laddning. Kännedom om antibiotikametaboliters molekylvikt vid hydrolys möjliggör detektion. Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS) är en högsensitiv kvantifieringsmetod som separerar molekyler i avseende på polaritet för vidare detektion i avseende på massa och laddning. Syfte Syftet med denna studie var att vidareutveckla en snabb och effektiv metod för att påvisa nedbrytning av piperacillin-tazobaktam och cefotaxim i blodplasma med LC-MS/MS. Material och Metod Tiofaldigt sjunkande koncentrationer av piperacillin-tazobaktam från 2000 till 2 µg/ml, och cefotaxim med koncentrationerna 500 till 0,5 µg/ml analyserades med MALDI-TOF MS, dels intakt men även med bakterierna E. coli och K. pneumoniae med uttryck av olika resistensmekanismer. Vid optimerade koncentrationer spikades plasmaprover med nedbrutet antibiotika som sedan kvantifierades med LC-MS/MS. Resultat Lägsta detektionsgräns med MALDI-TOF MS för intakt och hydrolyserat piperacillin-tazobaktam var 20/2,5 µg/ml. För cefotaxim var lägsta gränsen 5 µg/ml. Med kliniskt relevanta blodkoncentrationer gick hydrolys inte att detektera för. Med tre bakteriekolonier/50 µl kunde dock hydrolys detekteras och kvantifieras med LC-MS/MS. Slutsats Detektion av β-laktamantibiotika är möjligt med både MALDI-TOF MS och LC-MS/MS. För att påvisa hydrolys krävdes större mängder bakterier än förväntat med LC-MS/MS. / Introduction Antibiotic resistance is a global growing problem. Piperacillin-tazobactam and cefotaxime are parts of the group β-lactam antibiotics. The common feature is to inhibit the cell wall synthesis by covalent bindings to the peptidoglycan layer and thereby causing lysis of the bacterial cell. E. coli and K. pneumoniae are members of the Enterobacteriaceae which is a part of the human normal flora but also are commonly associated with urinary tract infections which sometimes develops into to sepsis. Extended Spectrum β-Lactamases (ESBLs) are enzymes with hydrolytic abilities acting on β-lactam antibiotics, expressed by Enterobacteriaceae. The qualitative, Matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF MS) can be used to detect chemical compounds in the ratio of mass to charge in accordance to their molecular weight. Liquid Chromatography Tandem Mass Spectrometry (LC-MS/MS) is a highly sensitive two-step method of quantification which first separate molecules by their polarity attraction force and then by the ratio of mass to charge. Aim The aim of this study was to develop a fast and efficient method to determine degradation of piperacillin-tazobactam and cefotaxime in blood plasma by LC-MS/MS. Method Tenfold dilution of piperacillin-tazobactam in concentrations of 2000 to 2 µg/ml, and cefotaxime in concentrations of 500 to 0,5 µg/ml where analyzed by MALDI-TOF MS, intact and also with the bacteria E. coli and K. pneumoniae with different expression of antibiotic resistance. Optimized concentrations where fixed in blood plasma and then quantified by LC-MS/MS. Result The detection limit by using MALDI TOF MS of hydrolyzed as well as non-hydrolyzed piperacillin-tazobactam was 20/2,5 µg/ml. The detection limit in cefotaxime was 5 µg/ml. Hydrolysis could not be detected in clinically fixed blood concentrations. Detection and quantification of hydrolysis by LC-MS/MS was possible in a concentration of three bacteria colonies/50 µl. Conclusion It is possible to detect hydrolysis in both MALDI TOF MS and LC-MS/MS. A larger amount of bacteria than expected was needed to demonstrate hydrolysis In LC-MS/MS.

Wie in anderen Zellen sind auch bei Säugerspermien spezifische Lipide und Proteine der Zellmembran aufgrund ihrer heterogenen lateralen Verteilung in speziellen Domänen angereichert, die in unterschiedlichen räumlichen und zeitlichen Dimensionen existieren und der Zelle funktionale Variabilität ermöglichen. Aufgrund der fehlenden aktiven Proteinbiosynthese bietet dies den Spermien eine Möglichkeit, auf unterschiedliche Anforderungen zu reagieren. In der vorliegenden Arbeit wurden daher sogenannte detergenzresistente Membrandomänen (DRMs) aus Eberspermien unterschiedlicher Reifestadien präpariert und untersucht. Dabei stieg bereits in den Dichtegradienten mit zunehmender Reife die Dichte, bei der die opaleszenten Banden auftraten. Eine Analyse dieser mittels 31P-NMR zeigte mit zunehmender Reife eine Anreicherung an Glycerophosphatidylethanolamin und Phosphatidylinositol bei den Glycerophospholipiden, der Gehalt an Sphingomyelin hingegen nahm während der Nebenhodenreifung und auch nach der Ejakulation ab. Diese Veränderungen könnten auf eine Destabilisierung von Membrandomänen hindeuten, um eine Zusammenlagerung zu größeren Domänenclustern zu erleichtern, möglicherweise in Vorbereitung auf Kapazitation und Akrosomenreaktion. Zunächst werden die destabilisierten Membrandomänen jedoch durch die Anlagerung von Seminalplasmaproteinen geschützt, was vermutlich für das verringerte Lipid- zu Proteinverhältnis der DRMs bei Ejakulatspermien sorgt. Aufgrund der generellen Kälteempfindlichkeit von Eberspermien findet ihre Lagerung üblicherweise bei 16°C statt. Dies ist aus mikrobiologischer Sicht nachteilig gegenüber einer kälteren Lagerungstemperatur. Eine Untersuchung der Spermien von 64 Ebern zeigte jedoch bei 10% der Ejakulate eine individuum-spezifische Resistenz gegenüber der Lagerung bei 4°C. Die DRMs der kälteresistenten Spermien hatten einen erhöhten Anteil an langkettigen, mehrfach ungesättigten Fettsäuren, wie 31P-NMR und MALDI-TOF MS Analysen zeigten. / The lateral distribution of lipids and proteins in the plasma membrane is heterogeneous. Therefore specific lipids and proteins in membranes of mammalian spermatozoa are enriched in special domains of varying size and different time scales enabling the cell’s membrane functional variability. Being transcriptional inactive this is especially relevant for spermatozoa in responding to multiple challenges on their way to fertilization. Therefore so called detergent resistant membrane domains (DRMs) from boar spermatozoa of different developmental stages were investigated. Already in the sucrose density gradients differences were visible, so the opalescent bands of more maturated sperm had a higher density. An analysis of these bands by 31P-NMR showed an enrichment of glycerophosphatidylethanolamine and phosphatidylinositol during maturation and a decrease of sphingomyelin during maturation in the epididymis and even after ejaculation. This suggests destabilization of DRMs and hence of putative membrane domains. This could enable clustering to bigger membrane domain platforms in preparation for capacitation and acrosome reaction. First, however, seminal fluid proteins cover the spermatozoa protecting the membrane with the destabilized membrane domains. This could have led to the detected decrease of the lipid to protein ratio in DRMs of ejaculated sperm. Boar spermatozoa are sensitive to storage at cold temperatures and are therefore usually stored at 16°C, which is especially disadvantageous with regard to growing of bacteria. A screening of sperm from 64 boars showed a ratio of 10% individuals with cold resistant sperm which could be stored at 4°C without quality loss. The DRMs of cold resistant sperm had a higher proportion of longchained, polyunsaturated fatty acids, as shown by analysis with 31P-NMR und MALDI-TOF MS.

Candida albicans is an opportunistic fungal pathogen that may cause mucutaneous infection and/or disseminated candidasis if the host defense system is impaired (such as those in HIV patients). Cell surface of C. albicans is the frontier where initial interplay between host-pathogen takes place and therefore is of great importance in understanding the mechanism of hostpathogen interaction. MALDI-TOF-MS analysis of intact fungal cells yielded mass signatures for rapid species differentiation, strain grouping and yeast morphogenesis monitoring. Cell surface biotinylations at low temperature (4°C), enzymatic digestion of the intact fungal cell surface proteins ("whole cell shaving"), biotin-avidin affinity enrichment of biotinylated peptides, liquid chromatography mass spectrometry (LC-MS) based proteomic approach were employed for unambiguous identification of cell wall/cell wall associated proteins and the exposed peptide segments of these proteins. SILAC (Stable Isotope Labeling by Amino acids in Cell Culture) based CWP quantification analyses were performed to monitor CWP accumulation level change in response to hyphae induction. Information on surface exposed peptide segments and regulation of cell wall/surface protein during morphogenesis provided new candidates to the pool of potential peptide targets for protective vaccine development. A New type of "fluorous" (fluorinated alkane) affinity gained popularity due to its low level nonspecific protein/peptide binding. Fluorous labeling reagents that target primary amine groups in proteins/peptides were synthesized and characterized. The acid labile linker in the labeling reagents allows cleavage of the bulky fluorous tag moiety and the long oligo ethylene glycol (OEG) spacer after fluorous affinity purification. Upon collision induced decomposition, the labeled peptide ion yielded a characteristic fragment that could be retrieved from the residual portion of fluorous affinity tag, and serve as a marker to indicate that the relevant peptide had been successfully labeled. Results showed that both the protein/peptide labeling and affinity enrichment/separation process were highly efficient.

O diagnóstico rápido e eficiente da mastite subclínica é importante para reduzir a persistência da doença e os prejuízos decorrentes. O objetivo do estudo foi avaliar a técnica de espectrometria de massas (MS) por ionização e dessorção a laser assistida por matriz - tempo-de-vôo (MALDI-TOF) para identificação de bactérias causadoras de mastite subclínica bovina por dois métodos: 1) a partir de bactérias isoladas por cultivo icrobiológico; 2) a partir da recuperação de bactérias diretamente do leite, visando eliminar completamente a necessidade de cultivo microbiológico para identificação dos patógenos. O estudo foi composto por dois experimentos (1 e 2), no experimento 1 foram utilizadas 33 amostras de leite provenientes de animais das raças Gir e Holandesa de quatro fazendas leiteiras para a identificação icrobiológica e MALDI-TOF MS. As amostras com resultados conflitantes foram confirmadas por sequenciamento do gene 16S rRNA. Os resultados de cultura microbiológica foram Staphylococcus aureus (n = 13), Streptococcus agalactiae (n = 10) e Estafilococos coagulase negativo (ECN) (n = 10). Para todas as amostras de Streptococcus agalactiae, resultados similares foram observados para a identificação microbiológica e por MALDI-TOF MS. De 13 isolados de Staphylococcus aureus, 11 foram igualmente identificados por MALDITOF, 1 isolado foi identificado como Staphylococcus haemolyticus por sequenciamento do gene 16S rRNA, e o outro isolado isolado com resultado conflitante foi caracterizado como cultura mista de Staphylococcus aureus e Enterococcus faecalis. Em relação às amostras de ECN, todas as amostras do grupo foram identificadas por MALDI-TOF em nível de gênero (S. simulans, S. epidermidis, S. Haemolyticus, S. Chromogens e S. aureus coagulase negativa). No experimento 2 foi avaliado o método de recuperação de bactérias presentes em leite e sua identificação por MALDI-TOF MS utilizando o método de contaminação experimental de leite com Escherichia coli, Enterococcus faecalis e Staphylococcus aureus. A identificação de patógenos recuperados diretamente do leite foi possível quando a concentração de E. faecalis e S. aureus foi de 106 ufc/mL, e de 107 ufc/mL para E.coli. Concluímos que o uso de MALDI-TOF MS pode acelerar a identificação de patógenos causadores de mastite subclínica bovina podendo contribuir para a adoção de medidas de controle e tratamento mais adequado. / Rapid and efficient diagnosis of subclinical mastitis is important to reduce the persistence of the disease and its losses. This study aimed to evaluate the technique of mass spectrometry (MS) and desorption ionization by matrix assisted laser time-offlight (MALDI-TOF) for identification of bacteria causing bovine subclinical mastitis by two methods: 1) from bacteria isolated by microbiological culture, 2) from bacteria recovered directly from milk, to eliminate completely the need for microbiological culture for identification of pathogens. The study consisted of two experiments (1 and 2).In experiment 1, 33 milk samples from Gir and Holstein animals collected in four dairy farms were used for microbiological identification and MALDI-TOF MS. The samples with different results were confirmed by sequencing the 16S rRNA. These samples were identified by microbiological culture as Staphylococcus aureus (n = 13), Streptococcus agalactiae (n = 10) and Staphylococcus coagulase negative (SCN) (n = 10). For all strains of Streptococcus agalactiae, similar results were observed for microbiological identification and MALDI-TOF MS. From 13 isolates of Staphylococcus aureus, 11 were also identified by MALDI-TOF, one isolate was identified as Staphylococcus haemolyticus by 16S rRNA sequencing , and the other discrepant sample was charactezided a mixed culture of Staphylococcus aureus and Enterococcus faecalis. Regarding the SCN samples, all samples of this group were identified by MALDI-TOF at gender level (S. simulans, S. epidermidis, S. haemolyticus, S. Chromogens and S. aureus coagulase negative). In experiment 2, we evaluated the method of recovery of bacteria present in milk and their identification by MALDI-TOF MS using experimental method of contamination of milk with Escherichia coli, Enterococcus faecalis and Staphylococcus aureus. The identification of pathogens recovered directly from milk was possible when the concentration of E. faecalis and S. aureus was 106 ufc/mL and 107 ufc/mL for E.coli. We conclude that the use of MALDI-TOF MS can accelerate the identification of pathogens causing bovine subclinical mastitis may contribute to the adoption of control measures and appropriate treatment.

<p>Eosinophil granulocytes are tissue dwelling leukocytes that are implicated in host defence, particularly against helminthic parasites; they also participate in most inflammatory disorders. Although eosinophils have important roles in host defence mechanisms, their actions can also be harmful to the host as in the allergic inflammation where lung epithelium is destructed due to the release of toxic granule proteins. </p><p>The focus of the present thesis work has been to characterize the molecular and functional heterogeneity of the granule protein Eosinophil Cationic Protein (ECP). We investigated a coding ECP gene polymorphism (arg97thr) in an African population endemically exposed to the <i>Schistosoma mansoni</i> parasite and found a correlation between ECP genotype and disease manifestations; ECP^97arg was more effective in terms of host defence against the parasite, but was also correlated to development of liver fibrosis in infected subjects. </p><p>We purified ECP^97arg and ECP^97thr from healthy blood donors and showed that they differ in their cytotoxic activities; ECP^97arg was cytotoxic whereas ECP^97thr was non-cytotoxic. They did not differ in terms of RNase activity or in their ability to stimulate fibroblast-mediated collagen gel contraction. </p><p>We developed a new SELDI-TOF MS assay to enable the study of the structure of ECP in more detail and showed that ECP is produced in several glycosylated forms, and that the degree of glycosylation determines the cytotoxic activity. Enzymatic deglycosylation significantly enhanced the cytotoxic activity of highly glycosylated ECP-variants.</p><p>To summarize, in this thesis we demonstrated that the cytotoxic activity of ECP is dependent on both a gene polymorphism and post-translational modifications, and that the cytotoxic activity is distinct from other functions of ECP. We speculate that ECP is synthesised in heavily glycosylated variants as a means to protect the host from its harmful effects and that ECP is activated by deglycosylation when required at the site of inflammation.</p>

Eosinophil granulocytes are tissue dwelling leukocytes that are implicated in host defence, particularly against helminthic parasites; they also participate in most inflammatory disorders. Although eosinophils have important roles in host defence mechanisms, their actions can also be harmful to the host as in the allergic inflammation where lung epithelium is destructed due to the release of toxic granule proteins. The focus of the present thesis work has been to characterize the molecular and functional heterogeneity of the granule protein Eosinophil Cationic Protein (ECP). We investigated a coding ECP gene polymorphism (arg97thr) in an African population endemically exposed to the Schistosoma mansoni parasite and found a correlation between ECP genotype and disease manifestations; ECP97arg was more effective in terms of host defence against the parasite, but was also correlated to development of liver fibrosis in infected subjects. We purified ECP97arg and ECP97thr from healthy blood donors and showed that they differ in their cytotoxic activities; ECP97arg was cytotoxic whereas ECP97thr was non-cytotoxic. They did not differ in terms of RNase activity or in their ability to stimulate fibroblast-mediated collagen gel contraction. We developed a new SELDI-TOF MS assay to enable the study of the structure of ECP in more detail and showed that ECP is produced in several glycosylated forms, and that the degree of glycosylation determines the cytotoxic activity. Enzymatic deglycosylation significantly enhanced the cytotoxic activity of highly glycosylated ECP-variants. To summarize, in this thesis we demonstrated that the cytotoxic activity of ECP is dependent on both a gene polymorphism and post-translational modifications, and that the cytotoxic activity is distinct from other functions of ECP. We speculate that ECP is synthesised in heavily glycosylated variants as a means to protect the host from its harmful effects and that ECP is activated by deglycosylation when required at the site of inflammation.

Wavelet methods possess versatile properties for statistical applications. We would like to explore the advantages of using wavelets in the analyses in two different research areas. First of all, we develop an integrated tool for online detection of network anomalies. We consider statistical change point detection algorithms, for both local changes in the variance and for jumps detection, and propose modified versions of these algorithms based on moving window techniques. We investigate performances on simulated data and on network traffic data with several superimposed attacks. All detection methods are based on wavelet packets transformations. We also propose a Bayesian model for the analysis of high-throughput data where the outcome of interest has a natural ordering. The method provides a unified approach for identifying relevant markers and predicting class memberships. This is accomplished by building a stochastic search variable selection method into an ordinal model. We apply the methodology to the analysis of proteomic studies in prostate cancer. We explore wavelet-based techniques to remove noise from the protein mass spectra. The goal is to identify protein markers associated with prostate-specific antigen (PSA) level, an ordinal diagnostic measure currently used to stratify patients into different risk groups.