Changing institutions can have a profound and lasting effect on individuals and societies, while at other times are lost to history. Reward systems and punishments shape our decision-making processes, and these experiences are influenced by the institutions, culture, and relationships within a given society at a given time. Additionally, power dynamics are an important factor to consider, as institutions can be used for repression and maintaining power, status, and wealth; however, they also promote freedom and emancipation.
The research designs in subsequent chapters demonstrate empirical approaches to comparative institutional analysis. Each essay identifies changes to unique institutional settings with distinct governance structures or contractual agreements, to provide the conditions for meaningful comparison. The central themes include persistence, path dependence, formal and informal rules, transaction costs, social order and embeddedness, in the broader context of both short-run and long-run time continuum.
Once the institutional settings are established, identification strategies are employed to estimate the effects of variation on decision making and economic outcomes. The essays illustrate the influence of institutional economics on contemporary research designs and demonstrate the merits of applied empirical research. This dissertation has been developed to contribute to a deeper understanding of institutional preconditions and the effects they have on political and economic behavior. By addressing these paradigms, we enhance our understanding of human behavior and the intricate relationship between institutions, decision-making, and outcomes.View less
Increased anthropogenic activities lead to severe global biodiversity loss, requiring effective conservation and management, particularly for terrestrial animals. Long-term wildlife monitoring and genetic research associated are of paramount importance, allowing for characterizing how and why a given ecosystem and wildlife inhabiting it are changing in response to environmental changes and conservation interventions applied. A variety of information thus is needed, such as population dynamics, species distribution, genetic variation, and disease cross-species transmission. However, in-field monitoring and biological sample collection are often hindered due to logistic barriers and ethical issues, especially for rare and threatened species. In my doctoral work, I evaluated the applicability of non-invasive environmental DNA (eDNA) based methods in terrestrial wildlife monitoring in combination with target capture and metabarcoding. Furthermore, I investigated the nucleic acid preservation of minimally-invasive dried blood spots (DBS) samples stored for up to 15 years in sub-standard storage conditions in comparison with blood samples stored in a cold chain. In the meanwhile, I explored the applications of DBS samples in genomic research and viral pathogen surveillance. I revealed sampling from waterholes during the dry season in tropical forests can provide representative environmental samples to apply eDNA-based methods for terrestrial animal monitoring. Target capture prior to sequencing retrieved more genomic information, while metabarcoding tend to be more sensitive to species shedding large amounts of DNA in terms of species detection. Integration of target capture and metabarcoding should be considered to improve the overall performance of eDNA-based methods. In terms of DBS sampling strategy, I provided evidence showing although nucleic acid degradation persisted in comparison to blood samples stored in a cold chain, relatively large DNA fragments (~20,000 bp) were preserved in DBS samples even for over 9 years of storage. DBS samples were suitable for complete mitochondrial genome related research for various species, while its application in viral pathogen research may be limited, particularly RNA viruses. Overall, my work emphasized the value of eDNA-based methods in terrestrial animal surveys and demonstrated the great promise of DBS samples in wildlife genetic or genomic research even for those being stored over a decade.View less
Zona Pellucida Like Domain 1 Protein (ZPLD1) is a main component of the cupula, a gelatinous structure located in the labyrinth organ of the inner ear and involved in vestibular function. The N-glycosylated protein is likely able to organize high-molecular-weight polymers via its zona pellucida (ZP) module, which is common for many extracellular proteins that self-assemble into matrices. In this work, I confirmed that ZPLD1 can form multimers while setting up a cellular model leveraging Mardin-Darby canine kidney (MDCK) cells to study protein polymerization. I identified two motifs within ZPLD1 which regulate protein polymerization and follow previously published conserved regions, identified across ZP proteins. Mutational depletion of either one of these modules led to diminished or abnormal polymer formation outside of the cells, likely due to altered processing at the plasma membrane. Further, intracellular polymer formation was observed. Proteolytic cleavage during secretion, separating the regulatory motif located distinct of the ZP module from the mature monomer, seems to be necessary to enable proper polymerization. This separation requires a transmembrane domain as well as the consensus cleavage site, which were as well located and confirmed using the established model and corresponding ZPLD1 expression variants. I was in addition able to proof that a serine type protease, potentially membrane bound hepsin, executes protein maturation. While the molecular interactions of the identified motifs and the exact processing protease and mechanism remain to be proven, my findings suggest that ZPLD1 is a polymer forming ZP protein, following a well-orchestrated mechanism. This mechanism is likely shared by other homopolymerizing ZP family proteins, includes cleavage of a pro-peptide, conformational changes, and the formation of precursor protein dimers. Finally, the mechanism leads to building up the gelatinous hydrogel of the cupula, thereby being of high significance for proper vestibular function. Ultimately, the variety of pathologies with reported impact of altered ZPLD1 expression indicate potential further functionalities such as cell adhesion, migration or segregation adding significance of this protein as future research target.View less
Energy and charge transfer processes in aqueous solution play important roles in biochemical processes, where the mechanisms of such phenomena are affected, and even driven, by aqueous-phase reactant-water interactions. Such processes and interactions have been probed using the Liquid-Jet (LJ)-based Photoelectron Spectroscopy (PES) technique. Energy-tunable, ionizing-radiation sources and the non-resonant, resonant, steady-state, and ultrafast-time-resolved PES techniques have been applied to probe the surface or bulk and valence or core-level electronic, and by extension molecular, structure of several archetypal, highly-soluble, aqueous-phase molecular systems: triiodide, pyrimidine, pyridazine, and pyrazine. The chemically-important, lowest vertical ionization energies and characteristic, atom- and isomer-specific core-level (I 3d/4d, C and N 1s) spectra of the four studied systems are presented here. Ultrafast time-resolution studies of the excited-state dynamics of aqueous pyrazine solutions are also presented.
With the help of electronic structure theory and spectral simulations, the experimental triiodide X-ray PES results suggest a near-linear geometric structure and increased asymmetry of the anion in aqueous solution, compared to in ethanol and methanol. This asymmetry was found to be greater at the surface-vacuum interface than in the aqueous solution bulk. Additionally, non-resonant and C and N pre-K-edge resonant X-ray PES experiments were performed on the aqueous diazine molecules, revealing the vertical ionization potentials, bonding character and atomic parentage, intra- and intermolecular charge re-arrangement, and degree of localization of the valence band Molecular Orbitals (MOs).
Having measured the ground-state ionization energetics, UV-pump (267 nm), EUV-probe (32.1 nm) femtosecond time-resolved LJ-PES measurements were performed on aqueous pyrazine solutions under Pump-Induced-Space-Charge (PISC) minimized conditions. Global fit analyses of the LJ-PES data revealed an initial 40 ± 20 fs internal conversion (IC) timescale, with subsequent population transfer to lower-lying states occurring in 35 ± 10 ps and 120 ± 30 ps. Slower relaxation behaviors were found in aqueous pyrazine solution in comparison to previously reported gas-phase studies. Associated solvent-induced relaxation mechanisms have been proposed. Furthermore, it is suggested that the relaxation dynamics of aqueous pyrazine are altered when similar experiments are performed under PISC conditions.
In general, this thesis highlights the sensitivity of LJ-PES to solution-phase electronic structure, molecular geometries, and dynamic photophysicochemical processes. The experimental results provide the necessary information to enhance the implemented experimental techniques, model general polyatomic molecular behaviors in aqueous solutions, and better understand the photochemical processes that underly numerous real-world applications.View less
The present thesis was designed to assess the question whether lead-free hunting rifle bullets represent an adequate surrogate for conventional lead-based bullets in terms of animal welfare. The study was initiated as a result of the stakeholder conference on lead intoxications in white-tailed sea eagles organised by the IZW in 2005. Fragments of conventional lead core hunting bullets had been recognised as a major source of lead poisoning in white-tailed sea eagles and many other wildlife species. Reservations concerning the alleged reduced killing efficacy of lead-free bullets as an alternative to lead bullets were widely raised by hunters and the ammunition industry. For this study different approaches were used in order to assess the wounding capacity – the killing efficacy – and to describe fragmentation patterns of lead-free deforming and lead-free partially fragmenting bullets compared to lead-based bullets: 297 wild ungulates shot during regular hunting events in Germany were radiographed between 2006 and 2009 and the number of fragments as well as the maximum distance of fragments to the wound tract were evaluated. Fragmentation patterns significantly differed between lead-free and lead-based bullets. The use of lead-based bullets was associated with both a significantly higher number of fragments and a significantly larger maximum distance of fragments in relation to the wound tract. We showed that the use of lead core bullets results in a broad contamination of the carcass and the offal from shot ungulates and therefore should be avoided with regard to biological conservation, consumer protection and the prevention of food waste. The amount of tissue destruction in bodies of shot ungulates displays the wounding capacity associated with a bullet under field conditions. Comparable locations of the wound tract within the body should result in comparable wound dimensions. To answer the question whether wounds caused by lead-free bullets are less severe, thereby potentially causing superfluous pain to the animal by delayed entry of death, we conducted CT scans and post-mortem macroscopic examinations of 34 shot wild ungulates. The comparison of wound channel diameters along the bullet path and the determination of the maximum cross-sectional area of the wound tract did not reveal significant differences between lead-free and lead-based bullets. Additionally, we assessed the morphology of the wounds. All animals showed severe injuries to vital organs, which were likely to have caused rapid death due to exsanguination. No hints were found that lead-free bullets would produce less severe or smaller wounds. We conclude that lead-free rifle bullets perform as good as conventional lead core bullets under normal German hunting conditions. Wound ballistic simulation using glycerine soap as a tissue surrogate is widely used in ballistic research. The wounding potential of a given bullet corresponds to the cavity volume in the soap block. In order to compare lead-free and lead-based hunting rifle bullets under standardised conditions we analysed the cavities associated with three lead core and eight lead-free bullets. We did not find a superiority of lead core bullets in terms of cavity diameters in ballistic soap. The shape of the cavity, though, was significantly influenced by the bullet type. The maximum cavity diameter was positioned within the front part of the block for all bullets but situated significantly more distant to the target surface (“deeper”) for lead-free deforming bullets than lead core bullets or lead-free partially fragmenting bullets. So, the first bullet type might be more suitable for hunting larger wildlife species. Bullet-tissue-interaction varied substantially within bullet types, especially within the lead-free partially fragmenting bullets. Our results clearly showed that the actual bullet construction exerts a stronger influence on the wounding potential than the presence or absence of a lead core. This suggests that a standardised regime for official testing of all hunting bullet constructions prior to their licencing as official hunting bullets is desirable. Taking into consideration the results of all three parts of this thesis, lead-free hunting rifle bullets have to be regarded as an adequate surrogate for conventional lead-based bullets in terms of animal welfare. As the toxicity of lead for both humans and wildlife is well known and safety concerns have been shown to be unfounded, the phasing-out of conventional lead core bullets for hunting purposes should be strongly encouraged.View less
Diese Dissertation untersucht auf theoretischer, methodologischer und methodischer Ebene Möglichkeiten und Grenzen der kritischen Reflexion von Zukünften aus Perspektive der Kritischen Zukunftsforschung. Unter kritischer Zukunftsforschung werden jene Ansätze verstanden, die sich in einem deliberativen und ggf. transformativen Impuls mit den Konstruktionsbedingungen und -kontexten gegenwärtiger Zukünfte auseinandersetzen. Die Arbeit baut auf dem sozialen Konstruktionismus auf und vertieft die dort angelegte Betonung von Bedeutungen in der Auseinandersetzung mit sprachphilosophischen Positionen, insbesondere mit der Semiotik Ecos sowie poststrukturalistischem Denken. Kritische Reflexion wird als Störung von Denkrahmen begriffen, d.h. von naturalisierten Bedeutungen, über die Zukünfte durch spezifische Wirklichkeitskonstruktionen präfiguriert werden und die in der Rezeption direkt anschlussfähig sind. Diese Störung ermöglicht es, Denkrahmen zu denaturalisieren und Kontingenz wieder in Bedeutungsprozesse einzuführen. Dies erlaubt die Deliberation und ggf. Transformation von Bedeutungen. Auf methodologischer Ebene fokussiert sich diese Arbeit auf semiotische Zugänge zu kritischer Reflexion, d.h. auf solche, die, in Abgrenzung zu diskursorientierten Ansätzen, die Denkrahmen innerhalb von Zukunftsbildern aus der Perspektive des Bedeutungssystems untersuchen. Dazu wird auf Entautomatisierungsstrategien für die kritische Reflexion im Sinne der Störung eingegangen, die Rolle des hermeneutischen Verstehens für die Konzeption des Reflexionsprozesses erläutert und die Nähe zu Spiel und Spekulation diskutiert. In den veröffentlichten Artikeln werden zudem konkrete methodische Ansätze exploriert.View less
The human body can be targeted by many man-made and natural-occurring chemicals, potentially leading to adverse effects. This includes the endocrine system, essentially consisting of the EATS modalities (estrogen, androgen, thyroid, and steroidogenesis). Substances that interfere with the endocrine systems are called endocrine active substances and can finally be assessed as endocrine disruptors. Endocrine disruptors cannot be registered in the EU, unless exposure to the environment is negligible. The thyroid hormone system (THS) is important in regulating various physiological processes, including metabolism, energy regulation, and organ function. The THS controls gene expression and pathways that impact metabolism, thermoregulation, cardiovascular function, and neural connectivity. Perturbations in the THS can lead to increased or decreased thyroid hormone (TH) signalling as well as thyroid-related diseases including cancer. The developing foetus is dependent on maternal TH supplementation, and very susceptible towards alterations in TH supply. Resulting developmental changes are also called developmental neurotoxicity. Testing for THS parameters is routinely done in animal experiments. The EU wants to accelerate the switch from animal to non-animal methods (NAMs). Currently, there is no OECD validated NAM for the THS available. Newly developed NAMs should be standardized and reproducible. They should adhere to the Guidance Document on Good In Vitro Method Practices (GIVIMP) that provides guidance on critical aspects for the development of reliable and reproducible in vitro methods. The THS is a complex signalling pathway that includes production, secretion, transport, uptake, signalling and metabolization of TH as well as a negative feedback loop to control TH release. The iodothyronine deiodinases (DIO) are one important key event (KE) in the THS and is linked to adverse neurodevelopmental outcomes in mammals. DIOs regulate TH signalling by metabolizing TH to more or less active TH metabolites. Historically, testing for DIO1 inhibition was done in animals. A novel in vitro method using mouse liver enzyme as a source for DIO1 and the Sandell-Kolthoff (SK) reaction to quantify released iodide from a substrate was published in 2014. This method is part of a validation study led by the European Union Reference Laboratory for Alternatives to Animal Testing (EURL ECVAM) with the goal to develop NAMs that cover different endpoints in the THS. The validation consists of two parts where the reproducibility of the method is demonstrated (part 1) and the method is checked for its relevance and reliability (part 2). In this work, the DIO1-SK assay was optimized and standardized. Mouse liver microsomes were substituted by human liver microsomes to adhere to the human situation. Variability of generated results was reduced by implementing measures to control batch-specific microsome activity, inherent activity of assay constituents in the SK reaction and optimal enzymatic iodide release during microsome incubation. Linear reaction kinetics in the SK reaction was ensured by optimizing incubation time and variable SK reagent concentrations. The reproducibility of the DIO1-SK assay was demonstrated by testing of six described DIO1 inhibitors in at least five independent assay runs (part 1). Guidance for establishing the method at other laboratories was provided; an initial Standard Operation Procedure (SOP) as well as acceptance criteria to assess the validity of future runs were defined. Using the derived SOP, the DIO1-SK assay was tested for its predictivity using a described class of DIO inhibitors, gold-containing substances. As such, organic, inorganic gold substances as well as gold nanoparticles were tested. A strategic approach using the mentioned gold substances as well as substances lacking the gold moiety was chosen to verify that the observed DIO1-inhibition was mediated by the gold. All tested organic and inorganic gold substances produced DIO1 inhibition at concentrations comparable to the literature. Structural analogues lacking the gold moiety did not produce DIO1 inhibition. Interestingly, small gold nanoparticles (5 nm) caused DIO1 inhibition, while bigger nanoparticles (>30 nm) did not. It was verified that the observed inhibition was not caused by dissolved gold in the incubation media. This study verified the potential of the DIO1-SK assay to predict DIO1 inhibition and has shown the applicability of the method towards nanoparticles. For the part 2 of the validation study of EURL ECVAM, the predictivity of the assay was assessed. A blinded set of substances, provided by the EU, was tested in the method. This included thorough solubility testing as well as two newly introduced methods to assess the specificity of observed DIO1 inhibition. Upon experimental completion, the data was deblinded and compared to available literature data. A data interpretation procedure (DIP) using the variability of the assay was used to categorize the results. Multiple substances produced high inhibition. The introduced specificity methods identified two substances as method-interfering leading to exclusion for further analysis. Inhibition data was consistent with available in vitro and in silico literature data showing the methods relevance. A DIP was derived using three categories classifying substance into full, partial or no inhibitor based on efficacy. Additionally, full inhibitors were subcategorized into potent and weak full inhibitors using the substances IC50 (“potency”). An updated SOP was derived and published to allow other laboratories to use the method. In this thesis, an in vitro method was developed that predicts the KE of DIO1 inhibition optimized, standardized, and reproducibly. It includes methods to exclude undissolved substance, observed effects that are not specific to inhibition of DIO1 and a method-specific DIP that classifies the generated data based on assay-inherent variability. The work presented is the basis for the acceptance of the method at OECD level and is a forerunner in the validation study coordinated by EURL ECVAM. Once methods covering other KEs in the THS are validated, they must be combined with the DIO1-SK assay to form an in vitro test battery. This includes computational methods that can predict different KEs and modelling that extrapolates enzymatic data to an organismal level. The current acceptance process for methods on an OECD level takes multiple years. Acceptance processes for NAMs at the OECD must be accelerated to make validated methods available more quickly and further drive the switch from animal to non-animal testing.View less
Weltabgewandter Sprachspieler oder »radikaler Realist«? Fremd- und Selbstzuschreibung gehen in Bezug auf den Autor Ror Wolf weit auseinander. Dies gründet auf einer nur scheinbaren Paradoxie: In Wolfs vielgestaltigem Werk vollzieht sich der Zugriff auf Wirklichkeit gerade im Modus der Sabotage, Unterbrechung, Irritation oder Verzerrung – kurz: im Modus der Störung. Ausgehend von der langen Prosa fragt Barbara Bausch nach möglichen Formen literarischer Referenzialität. Dabei konturiert sie Wolfs experimentelle und zugleich engagierte Poetik des ästhetisch produktiven Störens als Kreuzungspunkt verschiedenster Suchbewegungen des Prosaschreibens in den 1950er bis 1980er Jahren.View less
This thesis explores the spectroscopic properties and chemistry of diatomic molecules, which hold significant promise for applications in areas like quantum information and ultracold chemistry.
Firstly, the Diatomic Molecular Spectroscopy Database, accessible through a dynamic website, has been implemented. This database predominantly consolidates spectroscopic information while enabling the computation and visualization of Franck-Condon factors, and is adaptable for user contributions. Based on this database, machine learning models have been built to effectively reveal relationships among spectroscopic constants, with input features based on constituent atoms' group and period. Similarly, a comprehensive dataset of contemporary experimental electric dipole moments has been created. Utilizing this dataset, it has been shown that a machine learning model can accurately predict dipole moments using spectroscopic constants.
The availability of precise spectroscopic data allows for a rigorous assessment of advanced quantum chemistry methods. Specifically, we investigated the accuracy of coupled-cluster with single, double, and perturbative triple excitations [CCSD(T)] in predicting electric dipole moments when combined with different basis sets. Additionally, the hyperfine constants for the a$^3\Pi$ state of aluminum monofluoride (AlF) have been computed and compared to experimental values. Our study underscores the significance of a thorough evaluation encompassing both experimental and theoretical methodologies.
AlF and calcium monofluoride (CaF), among other metal monofluorides, have emerged as highly promising options for experiments involving laser cooling and trapping of cold molecules. We have compared the efficiency of different fluorine-donor molecules producing AlF and CaF through metal atom ablation in a buffer gas cell. Additionally, we present an efficient machine learning method for fitting the potential energy surface of AlF-AlF system, trained on relevant configurations from molecular dynamics simulations at the CCSD(T) level.View less
Hydrogels are widely used as biomaterials for cell culture and tissue engineering. They consist of natural biopolymers (e.g., collagen, elastin, hyaluronic acid) or synthetic polymers (e.g., polyethylene glycol, polylactic acid, polyacrylamide). The mechanical properties of hydrogels, such as stiffness, toughness and stress relaxation, depend primarily on the molecular building blocks and the spatial organization of polymeric network chains and cross-links (network topology). Star-shaped, 4-arm polyethylene glycol (sPEG) is a widely used hydrogel building block. sPEG displays a very low polydispersity and carries exactly one terminal cross-linking site per arm. As a result, almost ideal covalently cross-linked networks have been synthesized from sPEG building blocks. In this thesis, my goal was to replace sPEG with a purely protein-based, 4-arm polymer building block, based on the tetrameric fluorescent protein DsRed and random coil polypeptides (RCPP). This strategy allowed for the recombinant production of fusion proteins consisting of DsRed, RCPP and a covalent or dynamic cross-linking unit thus providing purely protein-based hydrogels. More importantly, it gives access to sequence-controlled RCPPs with an exactly determined number of monomer units per chain (i.e., monodispersity). Employing a modular gene design, the length of the RCPPs can be easily varied and also polymer lengths beyond commercially available sPEGs can easily be obtained. In a first series of experiments, three different variants of DsRed were tested for their suitability to serve as the tetrameric core of the fusion proteins. Key criteria where their expression level in an E. coli expression system as well as their stability in a hydrogel network. The different variants were engineered so that they carry exactly one Cys residue per monomeric unit. The Cys residues were used to cross-link the DsRed proteins with maleimide-terminated sPEG to obtain a hydrogel network. After cross-linking, DsRed becomes the mechanically weakest link in the network and is expected to show force-induced dissociation when the hydrogel experiences strain. To probe possible pulling geometry effects, Cys was introduced either at the C-terminus or at an internal, surface-exposed position. Hydrogels were formed for all tested DsRed variants whereas no cross-linking took place for a Cys-free mutant. A simple stress-strain experiment was performed where the fluorescence signal of DsRed was monitored in parallel. It has been proposed that a shift from red to green fluorescence may occur upon tetramer dissociation; however, no such change was observed over a large range of applied strains. These experiments suggest that DsRed possesses a high stability and is well suited as a tetrameric core. Continuing with one DsRed variant, I genetically fused elastin-like polypeptides (ELPs) of controlled length to the C-terminus of the fluorescent protein. The fusion protein was further extended with either SpyCatcher or SpyTag to test the possibility of using this protein ligation system as a permanent covalent cross-link. I systematically varied the length of the ELP chain and tested the influence of this parameter on the resulting hydrogel properties, in combination with the total protein concentration. Rheology experiments show purely elastic behavior with a stiffness of around 10,000 Pa for the best hydrogel obtained (ELP length of 132 nm and protein concentration of 2 mM). This is a clear improvement over other protein-based hydrogels, which often show a stiffness less than 1000 Pa. The linear viscoelastic region scaled with ELP length, which is a clear indication that the hydrogels are indeed cross-linked via the terminal SpyCatcher/SpyTag fusion partners. Overall, these results show that purely protein-based hydrogels can be synthesized with a high level of molecular control, similar to what has already been obtained with sPEGs in other experiments. For most cell culture applications, dynamically cross-linked hydrogels are of advantage. It has been shown that cells respond to the viscoelastic properties of the hydrogel and the presence of dynamic bonds further allows cell migration and proliferation. In the next step, I thus replaced the SpyCatcher/SpyTag system with reversible coiled coil (CC) cross-links. To prevent hydrogel formation during recombinant protein expression, a heterodimeric CC (AB) was used. In contrast to the SpyCatcher/SpyTag system, expression and purification of these fusions proteins was more challenging. The DsRed-ELP-A fusion protein showed a low expression yield. DsRed-ELP-B had a strong tendency to aggregate, which most likely originated from the tendency of the B sequence to form homodimers. Preliminary experiments, where the B peptide was modified to reduce homodimer stability, did not show a significant improvement. Further optimization of the sequences is thus necessary before a sufficient amount of protein can be obtained for hydrogel synthesis. The above results have shown that ELPs are excellent RCPPs for hydrogel synthesis; however, ELPs also undergo a phase transition in response to salt concentration and temperature. As a possible replacement for ELPs, I tested another protein-based repeat sequence, consisting of serine, alanine and proline. Even though this PAS sequence has already been used as a replacement for PEG in protein therapeutics, its application as a hydrogel building block is novel. Initial expression and purification tests with the fusion proteins DsRed-PAS-A and DsRed-PAS-B also show a low yield, even lower than for the corresponding ELP fusion proteins. Overall, in this thesis I have tested several protein-based modules that can be used as tetrameric cores, random coil polymers and cross-linking modules for the synthesis of protein-based hydrogels. The results obtained from the above work show the potential of protein-based hydrogels but also highlight that it can be a challenge to obtain sufficient concentrations and overall quantities of the required fusions proteins. Despite the clear need for optimization of sequences and procedures, the building blocks introduced and tested will aid the future design of novel molecularly controlled hydrogels. Protein-based hydrogels are powerful candidates to serve as extracellular matrix mimicking, mechano-responsive networks with tunable mechanical properties.View less
In einer durch Diversität und Technisierung gekennzeichneten Welt bleiben Wertegefüge und Normvorstellungen in Bezug auf das Älterwerden nicht unberührt. Die Herausforderungen, die der demografische Wandel an die Gesellschaft stellt, sind immens und lassen sich nicht durch zu kurz greifende Maßnahmen adressieren. Die sich verändernden Rahmenbedingungen in Bezug auf das Älterwerden haben tiefgreifende Folgen für eine alternde Gesellschaft. Aus diesem Grund müssen im Rahmen der Zukunftsforschung neue Visionen oder Zukunftsbilder bzw. Szenarien entworfen werden, um sich einer wünschenswerten Zukunft des Älterwerdens anzunähern und so die diesbezügliche politische, wissenschaftliche und gesellschaftliche Diskussion zu unter-stützen. In Bezug auf die notwendige normativ-evaluative, aber auch deskriptiv-evaluative Bewertung dieser Zukunftsbilder gibt es zum gegenwärtigen Zeitpunkt jedoch im Rahmen der Zukunftsforschung keine konsistente theoretische Bewertungsgrundlage, um die unterschiedlichen Zukunftsbilder bzw. Szenarien zu vergleichen. Da Szenarien elaboriertere „Was wäre, wenn“-Sätze sind und diese in der theoretischen Philosophie als kontrafaktische Konditionale diskutiert werden, können Szenarien bzw. Zukunftsbilder auf die gleiche Weise analysiert werden, wie einzelne Philosophen es im Zusammenhang mit kontrafaktischen Konditionalen vorgeschlagen haben. Daraus folgt, dass auch verschiedene Szenarien bzw. Zukunftsbilder dahingehend auf ihre Glaubwürdigkeit untersucht werden können, indem geprüft wird, ob es in der universalen Menge möglicher Welten einige gibt, die der kleinsten Sphäre möglicher Welten angehören und somit der eigentlichen Welt ähnlich sind und in denen das Antezedens des Szenarios das Konsequens des Szenarios immer nach sich zieht. Auf dem Fundament der Verantwortungsethik kann daran anschließend eine normative Bewertung durchgeführt werden, um die Wünschbarkeit von verschiedenen Szenarien bzw. Zukunftsbildern aus der Perspektive von gesellschaftlichen Entscheidungstragenden zu bewerten. Aus der Sicht der Zukunftsforschung und der Zukunftsethik ist dabei eine wünschenswerte Handlung von Entscheidungstragenden dann gegeben, wenn vorsorgend Verantwortung übernommen wird. Verantwortungsvolles und vorsorgendes Handeln muss von Entscheidungstragenden also forciert werden, damit sich die Situation der (Mitglieder der) Gesellschaft verbessert oder zumindest nicht verschlechtert. Um diese Ansätze für Kriterien einer deskriptiv- und normativ-evaluativen Bewertung von Zukunftsbildern zu prüfen, wird ein expertengestützter Szenario-Prozess durchgeführt und Zukunftsbilder einer alternden Gesellschaft konstruiert. Auf Grundlage der Bewertung von Expertinnen und Experten wird eine Cross-Impact Analyse durchgeführt und die Ergebnisse einer Einflussfak-torenanalyse beschrieben. Als Ergebnis dieser Analysen werden Storyboards der Zukunftsbilder erstellt, um auf dieser Basis die Narration der Szenarien zu erarbeiten. In der zusammenfassenden Bewertung der Zukunftsbilder bzw. Szenarien einer alternden Gesellschaft, kann sowohl die Ähnlichkeit als auch die Wünschbarkeit der einzelnen Narrative gemeinsam betrachtet werden. Die Kombination der beiden Kriteriensätze ergibt dabei für jedes Zukunftsbild einen Schnittpunkt von Ähnlichkeit und Wünschbarkeit und ermöglicht so eine Priorisierung verschiedener Zukunftsbilder.View less
In this thesis, two main application fields for off-axis reflection zone plates (RZPs) in X-ray optics were analyzed: as element for the monochromatization of soft and hard X-ray beams and as spectroscopy element for X-ray spectrometers with synchrotron radiation, free electron lasers, as well as laboratory sources such as laser plasma sources and high harmonic generators. As a big step forward in the design process, a new calculation software was developed, using a modern mathematical description of an ellipse-plane intersection. This new approach reduces the data volume and calculation time to a minimum, while ensuring a high accuracy of the structures and opens many possibilities for the design of multi-channel parallel spectrometers for monochromatic and polychromatic radiation. The spectroscopic properties of off-axis RZPs were systematically investigated in the hard X-ray range. The spatial resolution was found to be better than 2 micrometers at 8.3 keV with a diffraction efficiency of 17.5 %. The experimentally tested spectroscopic arrangement showed a resolving power up to 400 in an energy range of 1000 eV around the central energy. This experiment indicates a possibility of absorption spectroscopy on transition metals in the energy range below 10 keV. Simultaneously, a time elongation in the optical element was estimated as 0.35 fs, which would allow ultra-fast time resolved spectroscopy methods. For the first time in the soft x-ray range, within the framework of this thesis, multi-channel RZPs for an ultra-sensitive parallel X-ray spectrometer were realized. Through this development, the spectrometer could be applied at the free electron laser source LCLS in Stanford (outside of this dissertation). The spectroscopy of highly diluted materials, metallo-enzyme complexes that contain manganese oxides, was performed. The new spectrometer has one order of magnitude better acceptance than conventional grating spectrometers. The results of these measurements are not mentioned in this thesis. The developed design model was applied for the realization of two types of new laboratory source spectrometers based on RZPs: a parallel spectrometer for a scanning electron microscope and a single-channel spectrometer for a laser-plasma X-ray source.View less
Brain function relies on synaptic vesicle (SV) exocytosis and endocytosis. These cell physiological processes are crucial for neurotransmission. Ca2+ triggers the exocytic fusion of synaptic vesicles, which leads to the release of neurotransmitter. To sustain neurotransmission, SV exocytosis must be spatiotemporally coupled to a qualitatively and quantitatively corresponding retrieval of membrane and SV proteins via compensatory endocytosis. Yet, it is unknown how neurons balance SV exo- and endocytosis to maintain presynaptic membrane homeostasis und thereby sustain brain function. This study identifies Synaptotagmin 1 (Syt1), the main Ca2+ sensor and key factor for synchronous neurotransmission, as a homeostatic, post-fusion trigger for compensatory endocytosis. Severe conserved defects in neurotransmission are caused by genetic loss, mutation, or acute inactivation of Syt1. As an SV protein, Syt1 interacts via its two C2 domains with proteins of both, the exo- and endocytic machinery, and furthermore mediates neurotransmission upon binding to charged phospholipids of the membrane. We demonstrate that Syt1 couples SV exocytosis and compensatory endocytosis by triggering the local, activity-dependent synthesis of phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] at presynaptic sites. Exocytosed Syt1 on the neuronal plasma membrane recruits phosphatidylinositol 4-phosphate [PI(4)P] 5-kinase Iγ (PIPKIγ), the main PI(4,5)P2 synthesizing enzyme at the synapse. The Syt1-dependent recruitment of PIPKIγ results in elevated levels of presynaptic PI(4,5)P2. As an important signaling lipid and driving force for SV endocytosis, we detect Syt1-dependent facilitation of SV endocytosis. Genetic interference with Syt1/ PIPKIγ complex formation selectively impairs PI(4,5)P2-triggered SV endocytosis but not exocytic SV fusion. We show the Syt1-triggered synthesis of signaling lipids to couple SV exo- and endocytosis across a wide range of physiological stimulation paradigms. Considering Syt1 and PI(4,5)P2 being associated with various physiological as well as pathophysiological processes, we predict similar mechanisms to couple fusion and retrieval in other cell types undergoing regulated secretion.View less
Ausgangspunkt meiner Arbeit ist die Frage, auf welche Weise Wir-Gefühle in den liberal- demokratischen Ländern wie der BRD und ROK mobilisiert und modelliert werden können. Die vorliegende Arbeit setzt an einem ganz spezifischen Punkt an, dort, wo erstens, nach Rancière, die Frage der Gemeinschaft, die neben Freiheit und Individuum als ein wichtiger Wert der Demokratie ist, bislang zu wenig diskutiert wurde, und zweitens, nach Kappelhoff, das Kino der paradigmatische Ort ist, wo die Frage der Gemeinschaft und Wir-Gefühle unendlich wiederholt in relationalen Verhältnissen zwischen der Welt, audiovisuellen Bilder und Zuschauer(-körper) entwickelt und modelliert werden kann. Die westdeutschen und südkoreanischen Liebesfilme aus den Jahren 1950–1961 boten sich dabei als äußerst pas- sender Forschungsgegenstand an, da an ihnen der intensive Subjektivierungsprozess des Wir beobachtet wurde, der die eigene Nationalidee und -identität in der Ausrichtung nach dem liberal-demokratisch orientierten Wir-Bild unendlich wiederholt aneignet, imaginiert und modelliert.
Vor diesem Hintergrund widmete sich die vorliegende Arbeit anhand der Liebesfilme aus den Jahren 1950–1961 in der BRD und ROK exemplarisch zu untersuchen, in welcher kul- tur- und geschichtsspezifischen Affektpoetiken das eigene Wir-Bild und Wir-Gefühle von sich und für sich modelliert werden konnten. So wird in der vorliegenden Arbeit zunächst ein theoretischer Ansatz entwickelt, der greifbar macht, auf welche Weise Wir-Gestaltung und Wir-Gefühle im Kino modelliert werden können. Und die Verhältnisse der liberal-demokratischen Wir-Gestaltung und Wir-Gefühle werden insbesondere durch die ausführliche Filmanalyse der sechs ausgewählten Liebesfilme festgestellt und angesichts dieser Analysenergebnisse wird die Kultur- und Geschichtsspezifik der Affektpoetik klar herausgestellt.
Auf diesem Weg war das Gemeinsame die heuristische Denkfigur, die uns die relationalen Verhältnisse, die unendlich zu verändernde Kultur und Geschichte und die gleichberechtigt von Einzelnen aufzuteilende Ganzheit zusammen als ein konstruktiven Prozess aufgreifen lässt. So wurde das Konzept der Ausdrucksbewegung als die adäquate Analysenmethode angewandt, die das Zusammenwirken des Gemeinsamen – der kinematografischen Kommunikation, der kultur- und geschichtsspezifischen Ausdrucksformen und der (Moral- /Wert-/Sinn-/Erfahrungs-)Horizont – konkret als spezifische – qualitativ konkretisiert zu er- fahrende -sinnliche Einheit erfassen lässt.
Schließlich dienen die filmanalytisch geführten Patterns der affektpoetischen Ausdrucksfor- men der Wir-Gestaltung und Wir-Gefühle als Leitfaden der Reflexion darüber, inwieweit die allgemeine liberal-demokratische Idee des Zusammenlebens angeeignet und entwickelt werden konnte. Und die komparatistisch herauskristallisierte jeweilige Gestaltungsformen der Wir-Gestaltung und Wir-Gefühle erweisen sich als kultur- und geschichtsspezifische Ex- pressivität, die das bestimmte Eigenbild als Wir imaginiert und es gemeinsam als „attraktiv“ erfährt und weiterhin so selbst aneignet und denkt.View less
Hearing and touch are two of our senses that rely on mechanotransduction to convert mechanical stimulus into electrochemical activity. Several studies have demonstrated that touch and hearing share common genetic determinants; mutations in genes causing deafness are also associated with reduced tactile sensitivity. Furthermore, transcriptome analysis from several studies has revealed that the same genes are expressed in both systems. However, the role of these genes has yet to be determined. In this work, I demonstrated that the calcium and integrin binding protein 2 (CIB2), a key component of mechanotransduction in the auditory system, is also implicated in somatosensory mechanosensation. CIB2 is an auxiliary subunit of the mechanotransduction channel in the auditory system that modulates channel function and determines correct channel positioning. Mutations in Cib2 cause hearing loss in humans and mice, as well as the cessation of mechanotransduction in the auditory system. By applying immunohistochemistry, I demonstrated the expression of CIB2 in a subset of DRG neurons, mainly nociceptors and, to a lesser extent, mechanoreceptors. CIB2 was also present in the terminal endings of sensory neurons that innervate skin end organs. In particular, CIB2 was found in terminal afferents innervating Meissner’s corpuscles and hair follicles, both associated with rapidly adapting mechanoreceptors. The impact of CIB2 on the physiological properties of different types of mechanoreceptors and nociceptors was investigated using an ex-vivo skin nerve preparation from the saphenous and tibial nerve. I showed that genetic ablation of Cib2 had a profound impact on rapidly adapting mechanoreceptors (RAMs). In a velocity and vibration experiment, RAMs lacking Cib2 exhibited dramatically higher frequency sensitivity. Thus, CIB2 influences RAM adaption rate and functions as a brake on their velocity sensitivity. CIB2 exhibits structural similarities with KChIP proteins, which are like CIB2, calcium sensors. Additionally, KChIP proteins have been shown to modulate Kv4 channel surface expression, assembly, and kinetics, leading us to the hypothesis that CIB2 influences RAM adaptation by interacting with and regulating a potassium channel. KCNQ4 inhibition or genetic deletion in RAMs had previously been shown to enhance velocity and vibration responses, similarly to observations in Cib2 knockout mice. I found that inhibiting KCNQ4 did not affect RAM responses in Wnt1-Cre; Cib2fl/ fl mice, showing that in mice lacking Cib2, KCNQ4 does not function effectively in RAMs. We were able to study the influence of CIB2 on KCNQ4 kinetics by employing a whole-cell patch clamp and transiently overexpressing CIB2 and KCNQ4 in the CHO cell line. We discovered that the deactivation time for KCNQ4 was slower after CIB2 overexpression. Furthermore, KCNQ4 mRNA was found in 60% of Cib2-positive neurons. Thus, we conclude that CIB2 interacts with and modulates KCNQ4. The absence of CIB2 in C-fibers, clearly altered their physiological properties. C-fires lacking Cib2 showed signs of sensitization as they displayed a prominent increase in after-charge responses. However, genetic deletion of Cib2 did not affect another type of nociceptor, the Aδ nociceptors. Finally, we assessed the behavioral impact of peripheral Cib2 ablation on light touch and mechanical pain. By using multiple behavioral assays, I found that mice lacking Cib2 exhibit a profound loss of touch sensation. Therefore, I performed an electrical search experiment using an ex-vivo skin nerve preparation to search for mechanically insensitive fibers and found that half of the Aβ-fibers were mechanically silent. These findings suggest that CIB2 is interacting with a mechanosensitive channel, the nature of which is yet to be investigated. This study allowed us to conclude that CIB2 interacts with and modulates ion channels (potassium and mechanosensitive channels) in the somatosensory system and therefore regulates sensory mechanotransduction. We hypothesize that CIB2, as in the auditory system, is an auxiliary subunit of channels in the somatosensory system.View less
The utilization of bumblebees in the laboratory setting is increasing primarily for the purpose of behavioral analysis and neural recordings. Their remarkable physiological resilience and their willingness to adapt to diverse environmental conditions render them exceptionally suitable for a wide range of experimental investigations. Moreover, they represent the vast group of wild bees, which are also struggling with the current problems arising from agricultural mismanagement. To find a suitable setup to perform behavioral tests accompanied by electrophysiological recordings for bumblebees, a novel experimental apparatus was designed and constructed. The bumblebee could autonomously navigate between the colony and the test/feeding arena. Initially, the focus was set on the normal behavior and development of the colonies within this controlled artificial environment. Through a series of iterative refinements, a training protocol was established. The bumblebees had to learn a specific feeding place marked by a local cue and matching panorama. Notably, extinction tests revealed an enhanced behavior towards the learned goals. This change in behavior is proving, that the artificial arena with the connection to the colony and animals foraging on their own motivation adequately fulfills the requisite criteria for conducting such experiments. Furthermore, tests were performed treating animals with Calypso®, an insecticide formulation with neonicotinoid Thiacloprid. The extinction tests resulted in a shift in behavioral patterns towards the border area compared to the controls, displaying a change in exploratory behavior. These findings serve as evidence, that the experimental procedure is suitable for laboratory based navigational experiments researching sub-lethal effects of treatments. Electrophysiological recordings were performed on trained animals using an established setup, yielding stable and sortable data. However, no discernible pattern indicative of memory retrieval was observed. Instead, subtle indications of involvement in exploratory behavior or exploratory learning were detected. The limitations of comparing simultaneously recorded neurons prevented the identification of a network pattern for learned object identification. Nevertheless, the study emphasizes the suitability of the setup for conducting electrophysiological recordings in trained bumblebees. The recommendation to upgrade the outdated recording devices is made to enable long-lasting multi-unit recordings and facilitate further intriguing analyses.View less
To target resources and policies where they are most needed, it is essential that policy-makers are provided with reliable socio-demographic indicators on sub-groups. These sub-groups can be defined by regional divisions or by demographic characteristics and are referred to as areas or domains. Information on these domains is usually obtained through surveys, often planned at a higher level, such as the national level. As sample sizes at disaggregated levels may become small or unavailable, estimates based on survey data alone may no longer be considered reliable or may not be available. Increasing the sample size is time consuming and costly. Small area estimation (SAE) methods aim to solve this problem and achieve higher precision. SAE methods enrich information from survey data with data from additional sources and "borrow" strength from other domains . This is done by modeling and linking the survey data with administrative or register data and by using area-specific structures. Auxiliary data are traditionally population data available at the micro or aggregate level that can be used to estimate unit-level models or area-level models.
Due to strict privacy regulations, it is often difficult to obtain these data at the micro level. Therefore, models based on aggregated auxiliary information, such as the Fay-Herriot model and its extensions, are of great interest for obtaining SAE estimators. Despite the problem of small sample sizes at the disaggregated level, surveys often suffer from high non-response. One possible solution to item non-response is multiple imputation (MI), which replaces missing values with multiple plausible values. The missing values and their replacement introduce additional uncertainty into the estimate. Part I focuses on the Fay-Herriot model, where the resulting estimator is a combination of a design-unbiased estimator based only on the survey data (hereafter called the direct estimator) and a synthetic regression component. Solutions are presented to account for the uncertainty introduced by missing values in the SAE estimator using Rubin's rules. Since financial assets and wealth are sensitive topics, surveys on this type of data suffer particularly from item non-response. Chapter 1 focuses on estimating private wealth at the regionally disaggregated level in Germany. Data from the 2010 Household Finance and Consumption Survey (HFCS) are used for this application. In addition to the non-response problem, income and wealth data are often right-skewed, requiring a transformation to fully satisfy the normality assumptions of the model. Therefore, Chapter 1 presents a modified Fay-Herriot approach that incorporates the uncertainty of missing values into the log-transformed direct estimator of a mean. Chapter 2 complements Chapter 1 by presenting a framework that extends the general class of transformed Fay-Herriot models to account for the additional uncertainty due to MI by including it in the direct component and simultaneously in the regression component of the Fay-Herriot estimator. In addition, the uncertainty due to missing values is also included in the mean squared error estimator, which serves as the uncertainty measure. The estimation of a mean, the use of the log transformation for skewed data, and the arcsine transformation for proportions as target indicators are considered. The proposed framework is evaluated for the three cases in a model-based simulation study. To illustrate the methodology, 2017 data from the HFCS for European Union countries are used to estimate the average value of bonds at the national level. The approaches presented in Chapters 1 and 2 contribute to the literature by providing solutions for estimating SAE models in the presence of multiply imputed survey data. In particular, Chapter 2 presents a general approach that can be extended to other indicators.
To obtain the best possible SAE estimator in terms of accuracy and precision, it is important to find the optimal model for the relationship between the target variable and the auxiliary data. The notion of "optimal" can be multifaceted. One way to look at optimality is to find the best transformation of the target variable to fully satisfy model assumptions or to account for nonlinearity. Another perspective is to identify the most important covariates and their relationship to each other and to the target variable. Part II of this dissertation therefore brings together research on optimal transformations and model selection in the context of SAE. Chapter 3 considers both problems simultaneously for linear mixed models (LMM) and proposes a model selection approach for LMM with data-driven transformations. In particular, the conditional Akaike information criterion is adapted by introducing the Jacobian into the criterion to allow comparison of models at different scales. The methodology is evaluated in a simulation experiment comparing different transformations with different underlying true models. Since SAE models are LMMs, this methodology is applied to the unit-level small-area method, the empirical best predictor (EBP), in an application with Mexican survey and census data (ENIGH - National Survey of Household Income and Expenditure) and shows improvements in efficiency when the optimal (linear mixed) model and the transformation parameters are found simultaneously. Chapter 3 bridges the gap between model selection and optimal transformations to satisfy normality assumptions in unit-level SAE models in particular and LMMs in general. Chapter 4 explores the problem of model selection from a different perspective and for area-level data. To model interactions between auxiliary variables and nonlinear relationships between them and the dependent variable, machine learning methods can be a versatile tool. For unit-level SAE models, mixed-effects random forests (MERFs) provide a flexible solution to account for interactions and nonlinear relationships, ensure robustness to outliers, and perform implicit model selection. In Chapter 4, the idea of MERFs is transferred to area-level models and the linear regression synthetic part of the Fay-Herriot model is replaced by a random forest to benefit from the above properties and to provide an alternative modeling approach. Chapter 4 therefore contributes to the literature by proposing a first way to combine area-level SAE models with random forests for mean estimation to allow for interactions, nonlinear relationships, and implicit variable selection. Another advantage of random forest is its non-extrapolation property, i.e. the range of predictions is limited by the lowest and highest observed values. This could help to avoid transformations at the area-level when estimating indicators defined in a fixed range. The standard Fay-Herriot model was originally developed to estimate a mean, and transformations are required when the indicator of interest is, for example, a share or a Gini coefficient. This usually requires the development of appropriate back-transformations and MSE estimators. 5 presents a Fay-Herriot model for estimating logit-transformed Gini coefficients with a bias-corrected back-transformation and a bootstrap MSE estimator. A model-based simulation is performed to show the validity of the methodology, and regionally disaggregated data from Germany are used to illustrate the proposed approach. 5 contributes to the existing literature by providing, from a frequentist perspective, an alternative to the Bayesian area-level model for estimating Gini coefficients using a logit transformation.View less
In 1978, Edward Said redefined Orientalism as a Western interpretation of the Middle East best characterized by an inherent cultural hostility. It’s essence, he declared, was the invariable distinction between Western superiority and Eastern inferiority. At the beginning of the twenty-first century, however, cultural critics Darrell Y. Hamamoto (2000), Vijay Prashad (2000), Jane Chi Hyun Park (2010), Jane Naomi Iwamura (2011), and David Weir (2011) have shed light on America’s long fascination with the Far East and more affirmative forms of Orientalism. Building on their work, I map developments of Far Orientalism on American screens at the turn of the century from an evolutionary perspective. In three case studies, I read audiovisual texts in their sociohistorical and media ecological contexts to trace representational shifts from the 1980s to the 2010s. Since the intersection of race and sex is significant for any discussion of Orientalism, I am mostly concerned with narratives featuring interracial romance. The first case study focuses on Michael Cimino’s crime thriller film Year of the Dragon (1985). My analysis is embedded in an examination of contemporary films related to the Vietnam War as well as the emergence of both the Model Minority myth and the redemption narrative. The next chapter is concerned largely with contextualizing Edward Zwick’s epic historical drama film The Last Samurai (2003) within the genre histories of both the American Western and the Japanese Eastern. These efforts culminate in investigations of the ways of how the film relates to the popularization of Buddhism and reworks the White Savior trope. The final case study offers an analysis of Ronald D. Moore’s science fiction television series Battlestar Galactica (2004-09) on the background of the rise of neoliberal economic policy and transhumanist philosophy as well as in relation to assimilation narratives and the genre history of cyberpunk. My research results demonstrate a trend from classic Orientalism to Techno-Orientalism, Spirito-Occidentalism, and outright Occidentalism. In the American imagination, I argue, East Asia has come to represent both the worst expression of modernity and the solution to its dehumanizing side. Neither have stereotypes been shattered nor has the geographical dualism been shed. Older fantasies merely have been complemented by more recent variations. I consider this study to be an extension of Said’s studies of Orientalism as well as a contribution to the fields of American cultural history, Asian American studies and, to a lesser extent, postcolonial studies, gender studies, and media studies.View less
Livestock-associated methicillin-resistant Staphylococcus aureus (LA-MRSA) are opportunistic zoonotic pathogens that have been globally recognized as a health threat in human and veterinary medicine. They are known for their common multiresistance to antimicrobial agents and may cause severe diseases although normally lacking major virulence factors. Colonized pigs and their related surroundings represent a considerable LA-MRSA reservoir. Due to their low host specificity, LA-MRSA isolates can easily cross species barriers. Therefore, persons with frequent (occupational) exposure to pigs have an increased risk of LA-MRSA colonization and/or infection and they may further transmit these isolates to contact persons and animals. The LA-MRSA prevalence differs geographically: Clonal Complex (CC) 398 dominates among pigs in Europe, Australia and the Americas, whereas CC9 is the most common lineage among pigs in Asia. This thesis revealed that differences in metabolic properties did most likely not play a key role in the development of the aforementioned two different epidemic porcine LA-MRSA clones in Germany and China, respectively. However, it was hypothesized that LA-MRSA CC398 might gradually replace the currently dominating CC9 lineage in China in the future because of many beneficial features that may enable preferential survival of the CC398 lineage under hostile environmental conditions or in the host. Possibly unfavorable metabolic variations and a lower growth rate of the Chinese LA-MRSA CC9 clone may be disadvantageous in niche adaptation and, thus, may support the replacement process. Consequently, the public health risk originating from porcine LA-MRSA might rise because of the increased pathogenicity of the CC398 lineage. Moreover, this thesis provided insight into population dynamics and molecular diversity of the epidemic porcine LA-MRSA CC398 lineage in Germany over 13 years in comparison with the porcine LA-MRSA CC9 lineage dominating in China. In general, both clones exhibited typical individual characteristics, but also some novel features. During comparison, similarities as well as distinct differences were recognized between the two lineages. The German CC398 isolates were widespread within Germany and largely reflected the pig density of the different regions. They displayed a wide molecular diversity and rather close phylogenetic relationships. An ongoing diversification was recognized over time. The CC398 and CC9 lineage both displayed a clear virulence potential, although genes coding for the major virulence factors Panton-Valentine leucocidin and toxic shock syndrome toxin-1 were not detected. Moreover, both lineages exhibited a great variety of AMR properties that mirrored the respective sales figures of veterinary antimicrobial agents in the two countries. The AMR profiles of the Chinese isolates were more extensive. In both lineages, AMR was largely associated with mobile genetic elements (MGEs), which can be transferred easily across strain, species or even genus boundaries via horizontal gene transfer (HGT). However, the German CC398 isolates displayed a greater variety of different MGEs, which points towards the easier integration of foreign genetic material. In addition, clonal and geographical correlations of molecular characteristics, virulence and resistance properties were more frequently observed among the LA-MRSA CC398 from Germany than temporal correlations. Furthermore, multiple novel MGEs were characterized in this thesis. A novel type V variant of the methicillin-resistance mediating SCCmec cassette was identified in a porcine LA-MRSA CC398 isolate from China. Homologies with other type V and XII elements indicate an ongoing evolution of SCCmec elements in LA-MRSA CC398 from pigs. Tn553 and Tn560 represent two new members of the Tn554 transposon family detected in Chinese LA-MRSA CC9 and CC398 isolates, respectively. Tn553 harbored a complete beta-lactam resistance mediating blaZ-blaR1-blaI beta-lactamase operon and preferred a different integration site than former Tn554-related transposons. Tn560 carried a multiresistance gene cluster comprising the spectinomycin resistance gene spcV, a novel spc variant that is most likely the product of a recombination between the related spc and spw genes, the pleuromutilin-lincosamide-streptogramin A resistance gene lsa(E) and the lincosamide resistance gene lnu(B). Plasmid pHKS3860 from a German porcine LA-MRSA CC398 isolate harbored the novel macrolide-lincosamide-streptogramin B resistance gene erm(54), cadmium, mercury and copper resistance genes, as well as an ica gene cluster for biofilm formation. As a consequence of such co-localizations on the same MGE, multiple resistance and/or virulence properties can be transferred in a single event. Even in the absence of a direct selection pressure, the risk for co-selection processes is increased. Finally, this thesis expanded the knowledge on the epidemic porcine LA-MRSA lineages in Germany and China. They represent a considerable AMR repository adding to the global health threat posed by AMR with HGT as major driver of rapid dissemination. Moreover, the understanding of genetic rearrangements in LA-MRSA and how these bacteria exchange genetic material has been improved. The diversity of LA-MRSA further increases and new emerging and possibly more virulent and/or resistant variants should be closely monitored. Due to the zoonotic potential and the risk of further spread among pig husbandries and into the human community, a full-scale LA-MRSA surveillance is vitally important.View less
A large diversity of viruses is transmitted by hematophagous mosquitoes. Among them are several infectious agents of major concern for human health, like dengue virus, chikungunya virus, Zika virus, yellow fever virus, Japanese encephalitis virus, and West Nile virus. Apart from arboviruses that infect vertebrate hosts, mosquitoes also harbor a range of insect-specific viruses that can have an influence on the transmission of other mosquito viruses. In this work, a baseline study was conducted on the mosquitoes of the Namibian Zambezi Region and their accompanying viruses. Mosquito sampling was performed in National Parks and Conservancies across four different land-use types, including undisturbed habitats and areas used for agriculture or pasture. The mosquito community composition was investigated, and infection rates with arboviruses and insect-specific viruses were determined. Most sampling locations showed a high diversity of different mosquito species, which supported an equally high genetic diversity of RNA viruses from all the relevant families of mosquito viruses. Established arboviruses, like West Nile virus, were identified among the virus detections. Further, we could detect a high number of putatively novel viruses, most of which were mosquito-specific viruses from the viral families Flaviviridae and Phenuiviridae. No conclusive correlation could be derived between mosquito host biodiversity and RNA virus infection prevalence. Instead, mosquito community composition was found to have a significant impact on the infection prevalence of the viruses studied.View less