@phdthesis{Koenig2024,
  author    = {K{\"o}nig, Sebastian Thomas},
  title     = {Temperature-driven assembly processes of Orthoptera communities: Lessons on diversity, species traits, feeding interactions, and associated faecal microorganisms from elevational gradients in Southern Germany (Berchtesgaden Alps)},
  doi       = {10.25972/OPUS-35460},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-354608},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2024},
  abstract  = {Chapter I: Introduction Temperature is a major driver of biodiversity and abundance patterns on our planet, which becomes particularly relevant facing the entanglement of an imminent biodiversity and climate crisis. Climate shapes the composition of species assemblages either directly via abiotic filtering mechanisms or indirectly through alterations in biotic interactions. Insects - integral elements of Earth's ecosystems - are affected by climatic variation such as warming, yet responses vary among species. While species' traits, antagonistic biotic interactions, and even species' microbial mutualists may determine temperature-dependent assembly processes, the lion's share of these complex relationships remains poorly understood due to methodological constraints. Mountains, recognized as hotspots of diversity and threatened by rapidly changing climatic conditions, can serve as natural experimental settings to study the response of insect assemblages and their trophic interactions to temperature variation, instrumentalizing the high regional heterogeneity of micro- and macroclimate. With this thesis, we aim to enhance our mechanistic understanding of temperature-driven assembly processes within insect communities, exemplified by Orthoptera, that are significant herbivores in temperate mountain grassland ecosystems. Therefore, we combined field surveys of Orthoptera assemblages on grassland sites with molecular tools for foodweb reconstruction, primarily leveraging the elevational gradients offered by the complex topography within the Berchtesgaden Alpine region (Bavaria, Germany) as surrogate for temperature variation (space-for-time substitution approach). In this framework, we studied the effects of temperature variation on (1) species richness, abundance, community composition, and interspecific as well as intraspecific trait patterns, (2) ecological feeding specialisation, and (3) previously neglected links to microbial associates found in the faeces. Chapter II: Temperature-driven assembly processes Climate varies at multiple scales. Since microclimate is often overlooked, we assessed effects of local temperature deviations on species and trait compositions of insect communities along macroclimatic temperature gradients in Chapter II. Therefore, we employed joint species distribution modelling to explore how traits drive variation in the climatic niches of Orthoptera species at grassland sites characterized by contrasting micro- and macroclimatic conditions. Our findings revealed two key insights: (1) additive effects of micro- and macroclimate on the diversity, but (2) interactive effects on the abundance of several species, resulting in turnover and indicating that species possess narrower climatic niches than their elevational distributions might imply. This chapter suggests positive effects of warming on Orthoptera, but also highlights that the interplay of macro- and microclimate plays a pivotal role in structuring insect communities. Thus, it underscores the importance of considering both elements when predicting the responses of species to climate change. Additionally, this chapter revealed inter- and intraspecific effects of traits on the niches and distribution of species. Chapter III: Dietary specialisation along climatic gradients A crucial trait linked to the position of climatic niches is dietary specialisation. According to the 'altitudinal niche-breadth hypothesis', species of high-elevation habitats should be less specialized compared to their low-elevation counterparts. However, empirical evidence on shifts in specialization is scarce for generalist insect herbivores and existing studies often fail to control for the phylogeny and abundance of interaction partners. In Chapter III, we used a combination of field observations and amplicon sequencing to reconstruct dietary relationships between Orthoptera and plants along an extensive temperature gradient. We did not find close but flexible links between individual grasshopper and plant taxa in space. While interaction network specialisation increased with temperature, the corrected dietary specialisation pattern peaked at intermediate elevations on assemblage level. These nuanced findings demonstrate that (1) resource availability, (2) phylogenetic relationships, and (3) climate can affect empirical foodwebs intra- and interspecifically and, hence, the dietary specialisation of herbivorous insects. In this context, we discuss that the underlying mechanisms involved in shaping the specialisation of herbivore assemblages may switch along temperature clines. Chapter IV: Links between faecal microbe communities, feeding habits, and climate Since gut microbes affect the fitness and digestion of insects, studying their diversity could provide novel insights into specialisation patterns. However, their association with insect hosts that differ in feeding habits and specialisation has never been investigated along elevational climatic gradients. In Chapter IV, we utilized the dietary information gathered in Chapter III to characterize links between insects with distinct feeding behaviour and the microbial communities present in their faeces, using amplicon sequencing. Both, feeding and climate affected the bacterial communities. However, the large overlap of microbes at site level suggests that common bacteria are acquired from the shared feeding environment, such as the plants consumed by the insects. These findings emphasize the influence of a broader environmental context on the composition of insect gut microbial communities. Chapter V: Discussion \& Conclusions Cumulatively, the sections of this dissertation provide support for the hypothesis that climatic conditions play a role in shaping plant-herbivore systems. The detected variation of taxonomic and functional compositions contributes to our understanding of assembly processes and resulting diversity patterns within Orthoptera communities, shedding light on the mechanisms that structure their trophic interactions in diverse climates. The combined results presented suggest that a warmer climate could foster an increase of Orthoptera species richness in Central European semi-natural grasslands, also because the weak links observed between insect herbivores and plants are unlikely to limit decoupled range shifts. However, the restructuring of Orthoptera communities in response to warmer temperatures depends on species' traits such as moisture preferences or phenology. Notably, we were able to demonstrate a crucial role of microclimate for many species, partly unravelling narrower climatic niches than their elevational ranges suggest. We found evidence that not only Orthoptera community composition, specialisation, and traits varied along elevational gradients, but even microbial communities in the faeces of Orthoptera changed, which is a novel finding. This complex restructuring and reassembly of communities, coupled with the nonlinear specialisation of trophic interactions and a high diversity of associated bacteria, emphasize our currently incomplete comprehension of how ecosystems will develop under future climatic conditions, demanding caution in making simplified predictions for biodiversity change under climate warming. Since these predictions may benefit from including biotic interactions and both, micro- and macroclimate based on our findings, conservation authorities and practitioners must not neglect improving microclimatic conditions to ensure local survival of a diverse set of threatened and demanding species. In this context, mountains can play a pivotal role for biodiversity conservation since these offer heterogeneous microclimatic conditions in proximity that can be utilized by species with distinct niches.},
  subject      = {Heuschrecken},
  language  = {en}
}
@phdthesis{Hein2004,
  author    = {Hein, Silke},
  title     = {The survival of grasshoppers and bush crickets in habitats variable in space and time},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-9140},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2004},
  abstract  = {Die zunehmende Nutzung von Landschaften f{\"u}hrt zu einer steigenden Fragmentierung sch{\"u}tzenswerter Fl{\"a}chen. Damit verbunden ist eine Zerschneidung von großen Populationen in Metapopulationen. In solchen F{\"a}llen bestimmt das Gleichgewicht zwischen Aussterben und Besiedlung von Habitaten die regionale {\"U}berlebenswahrscheinlichkeit von Arten. Um diese bestimmen, braucht man ein gutes Verst{\"a}ndnis der Habitatanspr{\"u}che der Arten, sowie Informationen {\"u}ber ihr Ausbreitungsverhalten. Ziel dieser Arbeit war es, geeignete Fl{\"a}chen f{\"u}r Heuschrecken in einer Landschaft identifizieren zu k{\"o}nnen, sowie einen Beitrag zur Quantifizierung der Erreichbarkeit einzelner Fl{\"a}chen durch Individuen zu leisten. Der erste Teil dieser Arbeit besch{\"a}ftigt sich mit der Quantifizierung der Habitateignung von Fl{\"a}chen f{\"u}r Heuschrecken. Dazu habe ich statistische Habitateignungsmodelle mittels logistischer Regression erstellt, evaluiert und validiert. Es zeigte sich, dass die Habitatwahl der Heuschrecken auf einer mittleren r{\"a}umlichen Skalenebene erfolgt. Dies steht mit der beobachteten Ausbreitungsdistanz der Tiere im Einklang. Neben dem nur grob klassifizierten Landschaftsfaktor „Biotoptyp" korrelieren vor allem strukturelle Faktoren sowie abiotische Faktoren mit dem Vorkommen der Heuschreckenarten. Bei der Bestimmung eines gemeinsamen Models f{\"u}r alle drei Heuschreckenarten erwies sich das Model der Art S. lineatus mit den Parametern Biotoptyp und Vegetationsh{\"o}he als am besten geeignet zur Vorhersage der Vorkommen der anderen Heuschreckenarten. Um zu testen, ob auch die Vorkommen von Arten unterschiedlicher Tiergruppen mittels eines gemeinsamen Modells vorhergesagt werden k{\"o}nnen, habe ich sowohl die Heuschreckenmodelle zur Prognose von Faltervorkommen getestet, als auch Modelle f{\"u}r Falter auf Heuschrecken {\"u}bertragen. Dabei erwiesen sich die Heuschreckenmodelle zur Prognose der anderen Arten weniger geeignet als das Modell f{\"u}r das Widderchen Z. carniolica in das der Anteil an geeignetem Habitat sowie die Vorkommen der beiden Saugpflanzen C. jacea und S. columbaria einfließen. Diese Art wird als standorttreu eingestuft und repr{\"a}sentiert damit auch die anderen Arten, die typisch f{\"u}r S{\"a}ume und Halbtrockenrasen sind. Die erh{\"o}hte Mobilit{\"a}t von Z. carniolica im Vergleich zu den Heuschrecken garantiert gleichzeitig auch die Erreichbarkeit aller geeigneten Fl{\"a}chen im Gebiet und damit ein Modell, das nur unwesentlich durch Zufallseffekte bei der Besiedlung beeinflusst wird. Neben der Habitatqualit{\"a}t/-quantit{\"a}t spielt vor allem der Austausch zwischen Fl{\"a}chen eine entscheidende Rolle f{\"u}r das {\"U}berleben der Metapopulation. Im zweiten Teil meiner Arbeit habe ich mich sowohl theoretisch als auch empirisch, mit dem Ausbreitungsverhalten von Heuschrecken besch{\"a}ftigt. In Freilandexperimenten konnte ich zeigen, dass die Annahme eines dichotomen Bewegungsverhaltens f{\"u}r Heuschrecken in einer realen Landschaft nicht zutrifft. Vielmehr wird die Bewegung in einer Fl{\"a}che besser als Kontinuum beschrieben das durch strukturelle Resistenz, Temperatur, Mortalit{\"a}tsrisiko und Ressourcenverf{\"u}gbarkeit bestimmt wird. Die jeweilige Kombination dieser Parameter veranlasst die Tiere dann zu einem entsprechenden Bewegungsmuster, das sich zwischen den beiden Extremen gerichteter und zuf{\"a}lliger Lauf bewegt. In Experimenten zum Grenzverhalten von Heuschrecken best{\"a}tigte sich dieses Ergebnis. F{\"u}r verschiedene Grenzstrukturen konnte ich unterschiedliche {\"U}bertrittswahrscheinlichkeiten nachweisen. Weiterhin konnte ich feststellen, dass Heuschrecken geeignete Habitate aus einer gewissen Entfernung detektieren k{\"o}nnen. Da das Ausbreitungsverhalten von Tieren in theoretischen Modellen eine wichtige Rolle spielt, k{\"o}nnen diese empirischen Daten zur Parametrisierung dieser Modelle verwendet werden. Zus{\"a}tzlich zum Einfluss des Laufmusters der Tiere auf die Erreichbarkeit geeigneter Habitate, zeigte sich in den von mir durchgef{\"u}hrten Simulationsstudien deutlich, dass der landschaftliche Kontext, in dem die Ausbreitung stattfindet, die Erreichbarkeit einzelner Habitate beeinflusst. Dieser Effekt ist zus{\"a}tzlich abh{\"a}ngig von der Mortalit{\"a}tsrate beim Ausbreitungsvorgang. Mit den Ergebnissen aus den Untersuchungen zur Habitateignung lassen sich die f{\"u}r Heuschrecken geeigneten Habitate in einer Landschaft identifizieren. Somit l{\"a}sst sich die potentielle Eignung einer Fl{\"a}che als Habitat, basierend auf Vorhersagen {\"u}ber die {\"A}nderung des Biotoptyps durch ein Managementverfahren, vorhersagen. Diese Information allein reicht aber nicht aus, um die regionale {\"U}berlebenswahrscheinlichkeit einer Art bestimmen zu k{\"o}nnen. Meine Untersuchungen zum Ausbreitungsverhalten zeigen deutlich, dass die Erreichbarkeit geeigneter Fl{\"a}chen von der r{\"a}umlichen Anordnung der Habitate und der Struktur der Fl{\"a}chen, die zwischen Habitaten liegen, abh{\"a}ngt. Zus{\"a}tzlich spielen individuenspezifische Faktoren wie Motivation und physiologische Faktoren eine ausschlaggebende Rolle f{\"u}r die Erreichbarkeit von geeigneten Fl{\"a}chen.},
  subject      = {Naturschutzgebiet Hohe Wann},
  language  = {en}
}
@phdthesis{Boetzl2022,
  author    = {B{\"o}tzl, Fabian Alexander},
  title     = {The influence of crop management and adjacent agri-environmental scheme type on natural pest control in differently structured landscapes},
  doi       = {10.25972/OPUS-24140},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-241400},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2022},
  abstract  = {Summary Chapters I \& II: General Introduction \& General Methods Agriculture is confronted with a rampant loss of biodiversity potentially eroding ecosystem service potentials and adding up to other stressors like climate change or the consequences of land-use change and intensive management. To counter this 'biodiversity crisis', agri-environment schemes (AES) have been introduced as part of ecological intensification efforts. These AES combine special management regimes with the establishment of tailored habitats to create refuges for biodiversity in agricultural landscapes and thus ensure biodiversity mediated ecosystem services such as pest control. However, little is known about how well different AES habitats fulfil this purpose and whether they benefit ecosystem services in adjacent crop fields. Here I investigated how effective different AES habitats are for restoring biodiversity in different agricultural landscapes (Chapter V) and whether they benefit natural pest control in adjacent oilseed rape (Chapter VI) and winter cereal fields (Chapter VII). I recorded biodiversity and pest control potentials using a variety of different methods (Chapters II, V, VI \& VII). Moreover, I validated the methodology I used to assess predator assemblages and predation rates (Chapters III \& IV). Chapter III: How to record ground dwelling predators? Testing methodology is critical as it ensures scientific standards and trustworthy results. Pitfall traps are widely used to record ground dwelling predators, but little is known about how different trap types affect catches. I compared different types of pitfall traps that had been used in previous studies in respect to resulting carabid beetle assemblages. While barrier traps collected more species and deliver more complete species inventories, conventional simple pitfall traps provide reliable results with comparatively little handling effort. Placing several simple pitfall traps in the field can compensate the difference while still saving handling effort.   Chapter IV: How to record predation rates? A plethora of methods has been proposed and used for recording predation rates, but these have rarely been validated before use. I assessed whether a novel approach to record predation, the use of sentinel prey cards with glued on aphids, delivers realistic results. I compared different sampling efforts and showed that obtained predation rates were similar and could be linked to predator (carabid beetle) densities and body-sizes (a proxy often used for food intake rates). Thus, the method delivers reliable and meaningful predation rates. Chapter V: Do AES habitats benefit multi-taxa biodiversity? The main goal of AES is the conservation of biodiversity in agricultural landscapes. I investigated how effectively AES habitats with different temporal continuity fulfil this goal in differently structured landscapes. The different AES habitats investigated had variable effects on local biodiversity. Temporal continuity of AES habitats was the most important predictor with older, more temporally continuous habitats harbouring higher overall biodiversity and different species assemblages in most taxonomic groups than younger AES habitats. Results however varied among taxonomic groups and natural enemies were equally supported by younger habitats. Semi-natural habitats in the surrounding landscape and AES habitat size were of minor importance for local biodiversity and had limited effects. This stresses that newly established AES habitats alone cannot restore farmland biodiversity. Both AES habitats as well as more continuous semi-natural habitats synergistically increase overall biodiversity in agricultural landscapes. Chapter VI: The effects of AES habitats on predators in adjacent oilseed rape fields Apart from biodiversity conservation, ensuring ecosystem service delivery in agricultural landscapes is a crucial goal of AES. I therefore investigated the effects of adjacent AES habitats on ground dwelling predator assemblages in oilseed rape fields. I found clear distance decay effects from the field edges into the field centres on both richness and densities of ground dwelling predators. Direct effects of adjacent AES habitats on assemblages in oilseed rape fields however were limited and only visible in functional traits of carabid beetle assemblages. Adjacent AES habitats doubled the proportion of predatory carabid beetles indicating a beneficial role for pest control. My results show that pest control potentials are largest close to the field edges and beneficial effects are comparably short ranged. Chapter VII: The effects of AES habitats on pest control in adjacent cereal fields Whether distance functions and potential effects of AES habitats are universal across crops is unknown. Therefore, I assessed distance functions of predators, pests, predation rates and yields after crop rotation in winter cereals using the same study design as in the previous year. Resulting distance functions were not uniform and differed from those found in oilseed rape in the previous year, indicating that the interactions between certain adjacent habitats vary with habitat and crop types. Distance functions of cereal-leaf beetles (important cereal pests) and parasitoid wasps were moreover modulated by semi-natural habitat proportion in the surrounding landscapes. Field edges buffered assemblage changes in carabid beetle assemblages over crop rotation confirming their important function as refuges for natural enemies. My results emphasize the beneficial role of field edges for pest control potentials. These findings back the calls for smaller field sizes and more diverse, more heterogeneously structured agricultural landscapes. Chapter VIII: General Discussion Countering biodiversity loss and ensuring ecosystem service provision in agricultural landscapes is intricate and requires strategic planning and restructuring of these landscapes. I showed that agricultural landscapes could benefit maximally from (i) a mixture of AES habitats and semi-natural habitats to support high levels of overall biodiversity and from (ii) smaller continuously managed agricultural areas (i.e. smaller field sizes or the insertion of AES elements within large fields) to maximize natural pest control potentials in crop fields. I propose a mosaic of younger AES habitats and semi-natural habitats to support ecosystem service providers and increase edge density for ecosystem service spillover into adjacent crops. The optimal extent and density of this network as well as the location in which AES and semi-natural habitats interact most beneficially with adjacent crops need further investigation. My results provide a further step towards more sustainable agricultural landscapes that simultaneously allow biodiversity to persist and maintain agricultural production under the framework of ecological intensification.},
  subject      = {{\"O}kologie},
  language  = {en}
}
@phdthesis{Neubert2019,
  author    = {Neubert, Franziska},
  title     = {Markierung postsynaptischer Proteine f{\"u}r die hochaufl{\"o}sende Fluoreszenzmikroskopie},
  doi       = {10.25972/OPUS-19239},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-192394},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2019},
  abstract  = {Das menschliche Gehirn ist ein Organ, das aufgrund seiner Komplexit{\"a}t und zellul{\"a}ren Diversit{\"a}t noch am wenigsten verstanden ist. Eine der Ursachen daf{\"u}r sind zahlreiche Herausforderungen in diversen neurobiologischen Bild-gebungsverfahren. Erst seit der Erfindung der hochaufl{\"o}senden Fluoreszenz-mikroskopie ist es m{\"o}glich, Strukturen unterhalb der Beugungsgrenze zu visua-lisieren und somit eine maximale Aufl{\"o}sung von bis zu 20 nm zu erreichen. Zus{\"a}tzlich h{\"a}ngt die F{\"a}higkeit, biologische Strukturen aufzul{\"o}sen, von der Markierungs-gr{\"o}ße und -dichte ab. Derzeit ist die h{\"a}ufigste Methode zur Proteinf{\"a}rbung die indirekte Antik{\"o}rperf{\"a}rbung, bei der ein Fluorophor-markierter Sekund{\"a}rantik{\"o}rper an einen Epitop-spezifischen Prim{\"a}rantik{\"o}rper bindet. Dabei kann der Abstand von Zielstruktur und Fluorophor bis zu 30 nm betragen, was eine Aufl{\"o}sungs-verminderung zur Folge haben kann. Aufgrund dessen wurden in dieser Arbeit alternative Markierungsmethoden getestet, um postsynaptische Proteine sicht-bar zu machen. Zun{\"a}chst wurde der postsynaptische N-Methyl-D-Aspartat (NMDA)-Rezeptor mit Hilfe konventioneller indirekter Antik{\"o}rperf{\"a}rbung markiert. Hier war die NR1-Untereinheit des NMDA-Rezeptors von besonderem Interesse, da diese in der Autoimmunerkrankung Anti-NMDA-Rezeptor-Enzephalitis invol-viert ist. Patienten dieser seltenen Krankheit bilden Autoantik{\"o}rper gegen die NR1-Untereinheit, wodurch ein schneller reversibler Verlust der NMDA-Rezeptoren auf der Postsynapse induziert wird. Wichtige Informationen k{\"o}nnen nicht mehr ausreichend weitergegeben werden, was psychiatrische und neurologi-sche St{\"o}rungen zur Folge hat. In dieser Arbeit wurden sowohl kommerzielle NR1-Antik{\"o}rper, als auch rekombinante monoklonale NR1-Antik{\"o}rper von Patien-ten mit Anti-NMDA-Rezeptor-Enzephalitis getestet. In konfokalen und in hochaufgel{\"o}sten SIM- (engl. structured illumination microscopy) und dSTORM- (engl. direct stochastic optical reconstruction microscopy) Messun-gen konnten kommerzielle NR1-Antik{\"o}rper keine erfolgreichen F{\"a}rbungen erzielen. Dagegen erwiesen sich die rekombinanten monoklonalen NR1-Patientenantik{\"o}rper als sehr spezifisch, sowohl in prim{\"a}ren Neuronen als auch im Hippocampus von murinen Gehirnschnitten und lieferten gute Kolokalisati-onen mit dem postsynaptischen Markerprotein Homer. Um die optische Aufl{\"o}sung zu verbessern, wurde eine neue Markierungs-methode mit sog. „Super-Binde-Peptiden" (SBPs) getestet. SBPs sind modifi-zierte Peptide, die erh{\"o}hte Affinit{\"a}ten und Spezifit{\"a}ten aufweisen und mit ei-ner Gr{\"o}ße von ~ 2,5 nm wesentlich kleiner als Antik{\"o}rper sind. In dieser Arbeit best{\"a}tigte sich ein kleines hochspezifisches SPB, das an den Fluoreszenzfarb-stoff Tetra- methylrhodamin (TMR) gekoppelt ist, als effektiver Marker f{\"u}r das Ankerpro-tein Gephyrin. Gephyrin ist f{\"u}r die Lokalisation und Verankerung einiger post-synaptischer Rezeptoren zust{\"a}ndig, indem es sie mit dem Cytoskelett der Zelle verbindet. SIM-Messungen in prim{\"a}ren Neuronen zeigten eine bessere Clus-terrepr{\"a}sentation bei der F{\"a}rbung von Gephyrin mit SBPs, als mit Antik{\"o}rper-f{\"a}rbung. Zus{\"a}tzlich wurden Kolokalisationsanalysen von Gephyrin zusammen mit dem inhibito-rischen pr{\"a}synaptischen vesikul{\"a}ren GABA-Transporter VGAT durchgef{\"u}hrt. Eine weitere F{\"a}rbemethode stellte die bioorthogonale Click-F{\"a}rbung durch die Erweiterung des eukaryotischen genetischen Codes (engl. genetic code ex-pansion, GCE) dar. Dabei wurde eine unnat{\"u}rliche, nicht-kanonische Amino-s{\"a}ure (engl. non-canonical amino acid, ncAA) ins Zielprotein eingebaut und in Kombination mit der Click-Chemie ortsspezifisch mit organischen Tetrazin-Farbstoff-Konjugaten angef{\"a}rbt. Organische Fluorophore haben den Vorteil, dass sie mit einer Gr{\"o}ße von 0,5 - 2 nm sehr klein sind und damit die nat{\"u}rli-chen Funktionen der Proteine in der Zelle kaum beeinflussen. In dieser Arbeit wurde zum ersten Mal gezeigt, dass der tetramere postsynaptische NMDA-Rezeptor durch die Amber-Supres-sionsmethode bioorthogonal angef{\"a}rbt werden konnte. Aus sieben verschiede-nen Amber-Mutanten der NR1-Untereinheit stellte sich die Y392TAG-NR1-Mutante als diejenige mit der besten Proteinexpression, F{\"a}rbeeffizienz und rezeptorfunktionalit{\"a}t heraus. Dies konnte durch Fluoreszenzmikroskopie- und Whole-Cell Patch-Clamp-Experimenten gezeigt werden. Die bioorthogo-nale Click-F{\"a}rbung durch GCE eignete sich f{\"u}r die F{\"a}rbung des NMDA-Rezeptors in verschiedenen Zelllinien, mit unterschiedlichen Tetrazin-Farbstoff-Konjugaten und f{\"u}r Lebendzellexperimente. In dSTORM-Messungen erwies sich das Tetrazin-Cy5-Farbstoff-Konjugat als ideal aufgrund seiner Gr{\"o}-ße, Photostabilit{\"a}t, Helligkeit und seines geeigneten Blinkverhaltens, sodass eine homogene NMDA-Rezeptorverteilung auf der Zellmembran gezeigt wer-den konnte. NR1-Antik{\"o}rperf{\"a}rbungen wiesen dagegen starke Clusterbildun-gen auf. Die Ergebnisse konnten belegen, dass kleinere Farbstoffe eine deut-lich bessere Zug{\"a}nglichkeit zu ihrem Zielprotein haben und somit besser f{\"u}r die hochaufl{\"o}sende Fluoreszenzmikroskopie geeignet sind.},
  subject      = {hochaufl{\"o}sende Fluoreszenzmikroskopie},
  language  = {de}
}
@phdthesis{Hess2013,
  author    = {Heß, Michael},
  title     = {Vaccinia virus-encoded bacterial beta-glucuronidase as a diagnostic biomarker for oncolytic virotherapy},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-86789},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2013},
  abstract  = {Oncolytic virotherapy represents a promising approach to revolutionize cancer therapy. Several preclinical and clinical trials display the safety of oncolytic viruses as wells as their efficiency against solid tumors. The development of complementary diagnosis and monitoring concepts as well as the optimization of anti-tumor activity are key points of current virotherapy research. Within the framework of this thesis, the diagnostic and therapeutic prospects of beta-glucuronidase expressed by the oncolytic vaccinia virus strain GLV-1h68 were evaluated. In this regard, a beta-glucuronidase-based, therapy-accompanying biomarker test was established which is currently under clinical validation. By using fluorescent substrates, the activity of virally expressed beta-glucuronidase could be detected and quantified. Thereby conclusions about the replication kinetics of oncolytic viruses in animal models and virus-induced cancer cell lysis could be drawn. These findings finally led to the elaboration and establishment of a versatile biomarker assay which allows statements regarding the replication of oncolytic viruses in mice based on serum samples. Besides the analysis of retrospective conditions, this test is able to serve as therapy-accompanying monitoring tool for virotherapy approaches with beta-glucuronidase-expressing viruses. The newly developed assay also served as complement to routinely used plaque assays as well as reference for virally expressed anti-angiogenic antibodies in additional preclinical studies. Further validation of this biomarker test is currently taking place in the context of clinical trials with GL-ONC1 (clinical grade GLV-1h68) and has already shown promising preliminary results. It was furthermore demonstrated that fluorogenic substrates in combination with beta-glucuronidase expressed by oncolytic viruses facilitated the optical detection of solid tumors in preclinical models. In addition to diagnostic purposes, virus-encoded enzymes could also be combined with prodrugs resulting in an improved therapeutic outcome of oncolytic virotherapy. In further studies, the visualization of virus-induced immune reactions as well as the establishment of innovative concepts to improve the therapeutic outcome of oncolytic virotherapy could be accomplished. In conclusion, the results of this thesis provide crucial findings about the influence of virally expressed beta-glucuronidase on various diagnostic concepts in the context of oncolytic virotherapy. In addition, innovative monitoring and therapeutic strategies could be established. Our preclinical findings have important clinical influence, particularly by the development of a therapy-associated biomarker assay which is currently used in different clinical trials.},
  subject      = {Vaccinia-Virus},
  language  = {en}
}
@phdthesis{KuhngebBach2015,
  author    = {Kuhn [geb. Bach], Julia Elisa},
  title     = {Design und Etablierung von Next Generation Sequencing-Methoden zur Diagnostik verschiedener Erbkrankheiten},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-116854},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2015},
  abstract  = {Innerhalb des letzten Jahrzehnts entstanden zahlreiche neue Anreicherungs- und Sequenzier-technologien der zweiten (und dritten) Generation, die in rasantem Tempo weiterentwickelt und schon jetzt in vielen Bereichen als neuer Goldstandard f{\"u}r molekulargenetische For-schung und Diagnostik angesehen werden. Als Hochdurchsatz-Verfahren erm{\"o}glichen diese Next Generation Sequencing-Methoden (NGS) in immer k{\"u}rzerer Zeit die parallele Analyse zahlreicher Proben und immer gr{\"o}ßerer Zielregionen bis hin zum ganzen Genom und f{\"u}hrten in der Humangenetik dadurch zu Forschungsans{\"a}tzen in neuen Dimensionen. In dieser Doktorarbeit, die im molekulargenetischen Diagnostik-Labor der Humangenetik W{\"u}rzburg durchgef{\"u}hrt wurde, wurden in f{\"u}nf Projekten NGS-Ans{\"a}tze unterschiedlicher Stufen bzw. Gr{\"o}ßenordnungen f{\"u}r verschiedene erblich bedingte Erkrankungen konzipiert und etabliert und in Forschungsprojekten sowie der Routinediagnostik eingesetzt. Dabei wurden verschiedene Methoden zur Anreicherung der Zielsequenzen und zur NGS-Sequenzierung erprobt und auf ihre Effizienz beurteilt. Die Ergebnisse des NGS und darauf basierender Nachweis-Experimente wurden in sieben Ver{\"o}ffentlichungen dokumentiert, auf denen diese Dissertation aufbaut. In den drei ersten Projekten wurden das Access Array-System (Fluidigm) zur Anreicherung der Zielsequenzen und der GS Junior (Roche) zur Erzeugung der Sequenzen verwendet. In Projekt 1 wurde COL4A6 als neues Kandidatengen f{\"u}r nicht-syndromale H{\"o}rst{\"o}rungen identifiziert. Um m{\"o}gliche weitere Mutationstr{\"a}ger zu detektieren, wurde erfolgreich ein kleiner NGS-Ansatz f{\"u}r das z{\"u}gige Screening dieses Gens bei knapp 100 weiteren Patienten etabliert. Diese und weitere Ergebnisse best{\"a}tigten die Kausalit{\"a}t der COL4A6-Mutation eines Index-Patienten mit schwerer, X-chromosomal-rezessiver H{\"o}rst{\"o}rung. Ein geeigneter NGS-Ansatz f{\"u}r die Analyse des großen RYR1-Gens wurde in Projekt 2 ge-sucht. Der erste Ansatz mit Access Array-System und GS Junior f{\"u}hrte zwar bei 39 von 87 Patienten mit Maligner Hyperthermie und/oder Central Core Disease zu dem Auffinden einer (potentiell) pathogenen Variante, allerdings mit hohen Ausfallquoten. Mit der zweiten Methode (Anreicherung: SureSelect-System custom design, Agilent; Sequenzierung: HiSeq, Illumina) wurden neben RYR1 noch 63 weitere Gene analysiert, was zu deutlich besseren Ergebnissen und vier Mutationsfunden f{\"u}hrte. Projekt 3 beinhaltete die Etablierung zwei kleiner Panels f{\"u}r Muskelkrankheiten. Ein Panel f{\"u}r drei Gene f{\"u}r Gliederg{\"u}rteldystrophien wurde sogar erfolgreich in die akkreditierte Rou-tinediagnostik {\"u}bernommen. Mit dem zweiten Panel f{\"u}r acht Kandidatengene myofibrill{\"a}rer Myopathien (MFM) wurde u.a. eine neue Mutation im BAG3-Gen identifiziert. Das Exom eines MFM-Patienten wurde in Projekt 4 nach Anreicherung mit dem SureSelect-System (Agilent) auf dem HiSeq (Illumina) sequenziert. Nach Auswertung und Beurteilung der identifizierten Varianten wurde ein neuer Erbgang f{\"u}r Myotilinopathien entdeckt. Verschiedene Nachweisexperimente best{\"a}tigten die Kausalit{\"a}t der Mutation im Myotilin-Gen. In Projekt 5 wurde die komplette genomische Sequenz des F8-Gens nach tiefen intronischen Mutationen bei H{\"a}mophilie-Patienten abgesucht (Anreicherung SureSelect custom design, Agilent; Sequenzierung MiSeq, Illumina). Bei jedem der analysierten Patienten konnte min-destens eine verd{\"a}chtige Variante identifiziert werden, die zu ver{\"a}ndertem Spleißverhalten f{\"u}hren k{\"o}nnte. Drei Mutationen waren schon durch Publikationen bekannt, bei einer weite-ren konnten in vitro-Spleißanalysen die Kausalit{\"a}t best{\"a}tigen. Die Ergebnisse dieser Arbeit zeigen, dass die zur Verf{\"u}gung stehenden Methoden zur An-reicherung von Zielsequenzen aus dem menschlichen Genom und zu deren Sequenzierung je nach Komplexit{\"a}t der Fragestellung, d.h. der Anzahl und Gr{\"o}ße der Gene sowie der Anzahl der zu untersuchenden Proben, sinnvoll und effizient kombiniert werden k{\"o}nnen. Im Verlauf der Arbeit haben sich die NGS-Techniken rasant weiterentwickelt. So sind PCR-basierte Ans{\"a}tze zur Anreicherung der Zielsequenzen f{\"u}r die meisten Anwendungen von hybridisierungs-basierten Methoden verdr{\"a}ngt worden. Von den urspr{\"u}nglich drei konkur-rierenden Verfahren zur Hochdurchsatzsequenzierung hat sich die Methode des „sequen-cing-by-synthesis" (Illumina) weitgehend durchgesetzt. Diese Entwicklung spiegelt sich auch in den w{\"a}hrend dieser Arbeit erhobenen Daten wider.},
  subject      = {Diagnostik},
  language  = {de}
}
@phdthesis{Wegert2010,
  author    = {Wegert, Jenny},
  title     = {WTX-Mutationsscreen und funktionelle Analyse des Retins{\"a}ure-Signalwegs in Wilms Tumoren},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-52822},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2010},
  abstract  = {Der Wilms Tumor (WT), auch Nephroblastom genannt, ist einer der h{\"a}ufigsten b{\"o}sartigen Tumoren im Kindesalter. Er entsteht aus embryonalem undifferenziertem Nierengewebe und tritt meist als unilateraler und sporadischer Tumor auf. In 10-15\% der Wilms Tumoren finden sich WT1- und/oder CTNNB1-Mutationen. W{\"a}hrend diese schon l{\"a}nger als genetische Ursachen des Nephroblastoms bekannt sind, wurde erst k{\"u}rzlich WTX als drittes Gen beschrieben, welches eine Rolle in der Tumorentstehung spielt. F{\"u}r einen Großteil der WT ist die genetische Ursache jedoch unklar. Da die bisher publizierten WTX-Mutationsraten auf Untersuchungen kleiner Gruppen basieren und sich stark unterscheiden, sollten in dieser Arbeit WTX-, CTNNB1- und WT1-Mutationen in einem großen WT-Set bestimmt werden. Verluste genetischen Materials in der WTX-Region traten in 17\% der F{\"a}lle auf und waren zwischen den Geschlechtern gleich verteilt. Die Sequenzierung von WT-Proben zeigte, dass nur 2\% von WTX-Punktmutationen betroffen sind. In weiteren 11,5\% der Proben konnte keine WTX-Expression nachgewiesen werden. Die WTX-Ver{\"a}nderungen traten z. T. gemeinsam mit WT1- und/oder CTNNB1-Mutationen auf. Die unvollst{\"a}ndige WTX-Deletion in einigen WT legte die Vermutung nahe, dass innerhalb eines Tumors eine Heterogenit{\"a}t in Bezug auf den WTX-Status m{\"o}glich ist. Dieser Verdacht konnte durch die detaillierte Untersuchung verschiedener Regionen solcher Tumoren erh{\"a}rtet werden: Hierzu wurden histologisch unterschiedliche Bereiche auf den Anteil einer WTX-Mutation bzw. eines WTX-LOH hin untersucht. Obwohl alle Regionen des jeweiligen Tumors einen kompletten LOH auf Chromosom 11 aufwiesen, waren die WTX-Ver{\"a}nderungen unterschiedlich stark ausgepr{\"a}gt. Diese Ergebnisse deuten darauf hin, dass WTX-Ver{\"a}nderungen keine notwendigen und fr{\"u}hen Ereignisse in der Tumorentstehung sind, sondern erst sp{\"a}ter auftreten und nur einen Teil der Tumorzellen betreffen k{\"o}nnen. Die Vermutung, dass WTX-Mutationen keinen direkten Einfluss auf die Tumorentwicklung und prognose haben, wird durch das Fehlen eines signifikanten Zusammenhangs zwischen WTX-Deletion bzw. WTX-Expression und den klinischen Eigenschaften der WT gest{\"u}tzt. Um die Rolle von Genen, die potentiell an der Entstehung und Entwicklung des Nephroblastoms beteiligt sind, zu untersuchen oder m{\"o}gliche neue Therapiestrategien zu {\"u}berpr{\"u}fen, sind in vitro-Modelle n{\"o}tig. Da ein solches f{\"u}r Wilms Tumoren nicht etabliert ist, wurden Prim{\"a}rkulturen aus verschiedenen WT-Proben angelegt. Kulturen aus Tumorgewebe von 12 Patienten mit unterschiedlichen genetischen Ver{\"a}nderungen konnten als echte Tumorzellen validiert werden. Zwei Zelltypen ließen sich morphologisch und immunhistochemisch unterscheiden: Zum einen runde, langsam wachsende Zellen mit Epithelcharakter und zum anderen fibroblasten{\"a}hnliche Zellen, welche weniger differenziert waren und h{\"a}ufig f{\"u}r viele Passagen kultiviert werden konnten. Somit wurde ein Set verschiedener WT-Prim{\"a}rkulturen etabliert, welches nun f{\"u}r in vitro-Experimente zur Untersuchung grundlegender Mechanismen der WT-Entstehung oder zum Test neuer Therapieans{\"a}tze eingesetzt werden kann. Fr{\"u}here Microarray-Analysen deuteten auf eine Deregulation des Retins{\"a}ure (RA)-Signalwegs in fortgeschrittenen Wilms Tumoren hin. Diese Ergebnisse sollten in einem großen unabh{\"a}ngigen Proben-Set mittels Realtime-RT-PCR validiert werden. Eine Deregulation des RA-Signalwegs und die {\"U}berexpression von NMYC wurden f{\"u}r Tumoren der Hochrisikogruppe im Vergleich zu Tumoren mit geringem/mittlerem Risiko nachgewiesen. So stellte sich die Frage, ob Patienten mit fortgeschrittenem WT von einem Retins{\"a}ure-Einsatz in der Therapie profitieren k{\"o}nnten. Um dies zu beantworten, wurde der Effekt von verschiedenen Retinoiden auf WT-Prim{\"a}rkulturen untersucht. Die WT-Zellen wurden mit all-trans RA (ATRA), 9cisRA, dem synthetischen Retinoid Fenretinid (4HPR) und Kombinationen von ATRA bzw. 4HPR und einem HDAC-Inhibitor (SAHA) behandelt. Gene, welche in Hochrisiko-WT differenziell reguliert waren, wurden untersucht und zeigten nach RA-Behandlung eine entgegengesetzte Expression. In sechs der sieben verwendeten Prim{\"a}rkulturen wurde eine RA-vermittelte Proliferationsreduktion nachgewiesen. F{\"u}r die Kombinationen von Retinoiden mit SAHA wurden keine synergistischen Effekte beobachtet. W{\"a}hrend Fenretinid in den meisten Kulturen Apoptose induzierte, verursachten ATRA und 9cisRA morphologische Ver{\"a}nderungen, welche auf Differenzierungsvorg{\"a}nge hindeuteten. Eine Microarray-Analyse ATRA-behandelter WT-Zellen zeigte die differenzielle Regulation vieler Gene, welche eine Rolle in der Bildung der extrazellul{\"a}ren Matrix oder bei Differenzierungsvorg{\"a}ngen von Knochen-, Knorpel-, Nerven- oder Muskelgewebe spielen. Diese Befunde bieten einen weiteren Hinweis darauf, dass Retinoide f{\"u}r den Einsatz in der Therapie des Nephroblastoms geeignet sein k{\"o}nnten.},
  subject      = {Nephroblastom},
  language  = {de}
}
@phdthesis{Vogel2022,
  author    = {Vogel, Cassandra Ezra},
  title     = {The effects of land-use and agroecological practices on biodiversity and ecosystem services in tropical smallholder farms},
  doi       = {10.25972/OPUS-29066},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-290661},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2022},
  abstract  = {Biodiversity is in rapid decline worldwide. These declines are more pronounced in areas that are currently biodiversity rich, but economically poor - essentially describing many tropical regions in the Global South where landscapes are dominated by smallholder agriculture. Agriculture is an important driver of biodiversity decline, through habitat destruction and unsustainable practices. Ironically, agriculture itself is dependent on a range of ecosystem services, such as pollination and pest control, provided by biodiversity. Biodiversity on fields and the delivery of ecosystem services to crops is often closely tied to the composition of the surrounding landscape - complex landscapes with a higher proportion of (semi-)natural habitats tend to support a high abundances and biodiversity of pollinators and natural enemies that are beneficial to crop production. However, past landscape scale studies have focused primarily on industrialized agricultural landscapes in the Global North, and context dependent differences between regions and agricultural systems are understudied. Smallholder agriculture supports 2 billion people worldwide and contributes to over half the world's food supply. Yet smallholders, particularly in sub-Saharan Africa, are underrepresented in research investigating the consequences of landscape change and agricultural practices. Where research in smallholder agriculture is conducted, the focus is often on commodity crops, such as cacao, and less on crops that are directly consumed by smallholder households, though the loss of services to these crops could potentially impact the most vulnerable farmers the hardest. Agroecology - a holistic and nature-based approach to agriculture, provides an alternative to unsustainable input-intensive agriculture. Agroecology has been found to benefit smallholders through improved agronomical and food-security outcomes. Co-benefits of agroecological practices with biodiversity and ecosystem services are assumed, but not often empirically tested. In addition, the local and landscape effects on biodiversity and ecosystem services are more commonly studied in isolation, but their potentially interactive effects are so far little explored. Our study region in northern Malawi exemplifies many challenges experienced by smallholder farmers throughout sub-Saharan Africa and more generally in the Global South. Malawi is located in a global biodiversity hotspot, but biodiversity is threatened by rapid habitat loss and a push for input-intensive agriculture by government and other stakeholders. In contrast, agroecology has been effectively promoted and implemented in the study region. We investigated how land-use differences and the agroecological practices affects biodiversity and ecosystem services of multiple taxa in a maize-bean intercropping system (Chapter 2), and pollination of pumpkin (Chapter 3) and pigeon pea (Chapter 4). Additionally, the effects of local and landscape scale shrub- to farmland habitat conversion was investigated on butterfly communities, as well as the potential for agroecology to mitigate these effects (Chapter 5).},
  language  = {en}
}
@phdthesis{Joschinski2018,
  author    = {Joschinski, Jens},
  title     = {Is the phenology of pea aphids (\(Acyrthosiphon\) \(pisum\)) constrained by diurnal rhythms?},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-148099},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2018},
  abstract  = {The rotation of the earth leads to a cyclic change of night and day. Numerous strategies evolved to cope with diurnal change, as it is generally advantageous to be synchronous to the cyclic change in abiotic conditions. Diurnal rhythms are regulated by the circadian clock, a molecular feedback loop of RNA and protein levels with a period of circa 24 hours. Despite its importance for individuals as well as for species interactions, our knowledge of circadian clocks is mostly confined to few model organisms. While the structuring of activity is generally adaptive, a rigid temporal organization also has its drawbacks. For example, the specialization to a diurnal pattern limits the breadth of the temporal niche. Organisms that are adapted to a diurnal life style are often poor predators or foragers during night time, constraining the time budget to only diurnal parts of the day/night cycle. Climate change causes shifts in phenology (seasonal timing) and northward range expansions, and changes in season or in latitude are associated with novel day length - temperature correlations. Thus, seasonal organisms will have some life history stages exposed to novel day lengths, and I hypothesized that the diurnal niche determines whether the day length changes are beneficial or harmful for the organism. I thus studied the effects of day length on life-history traits in a multi-trophic system consisting of the pea aphid Acyrthosiphon pisum and predatory larvae of Chrysoperla carnea (common green lacewing) and Episyrphus balteatus (marmalade hoverfly). In order to identify the mechanisms for phenological constraints I then focused on diurnal rhythms and the circadian clock of the pea aphid. Aphids reacted to shorter days with a reduced fecundity and shorter reproductive period. Short days did however not impact population growth, because the fitness constraints only became apparent late in the individual's life. In contrast, E. balteatus grew 13\% faster in the shorter day treatment and preyed on significantly more aphids, whereas C. carnea grew 13\% faster under longer days and the elevation of predation rates was marginally significant. These results show that day length affects vital life-history traits, but that the direction and effect size depends on species. I hypothesized that the constraints or fitness benefits are caused by a constricted or expanded time budget, and hence depend on the temporal niche. E. balteatus is indeed night-active and C. carnea appears to be crepuscular, but very little data exists for A. pisum. Hence, I reared the pea aphid on an artificial diet and recorded survival, moulting and honeydew excretion. The activity patterns were clearly rhythmic and molting and honeydew excretion were elevated during day-time. Thus, the diurnal niche could explain the observed, but weak, day length constraints of aphids. The diurnal niche of some organisms is remarkably flexible, and a flexible diurnal niche may explain why the day length constrains were relatively low in A. pisum. I thus studied its circadian clock, the mechanism that regulates diurnal rhythms. First, I improved an artificial diet for A. pisum, and added the food colorant Brilliant Blue FCF. This food colorant stained gut and honeydew in low concentration without causing mortalities, and thus made honeydew excretion visible under dim red light. I then used the blue diet to raise individual aphids in 16:08 LD and constant darkness (DD), and recorded honeydew excretion and molting under red light every three hours. In addition, we used a novel monitoring setup to track locomotor activity continuously in LD and DD. Both the locomotor rhythm and honeydew excretion of A. pisum appeared to be bimodal, peaking in early morning and in the afternoon in LD. Both metabolic and locomotor rhythm persisted also for some time under constant darkness, indicating that the rhythms are driven by a functional circadian clock. However, the metabolic rhythm damped within three to four days, whereas locomotor rhythmicity persisted with a complex distribution of several free-running periods. These results fit to a damped circadian clock that is driven by multiple oscillator populations, a model that has been proposed to link circadian clocks and photoperiodism, but never empirically tested. Overall, my studies integrate constraints in phenological adaptation with a mechanistic explanation. I showed that a shorter day length can constrain some species of a trophic network while being beneficial for others, and linked the differences to the diurnal niche of the species. I further demonstrated that a flexible circadian clock may alleviate the constraints, potentially by increasing the plasticity of the diurnal niche.},
  subject      = {Tagesrhythmus},
  language  = {en}
}
@phdthesis{Krueger2014,
  author    = {Kr{\"u}ger, Alice},
  title     = {Die Entwicklung regenerativer Implantatmatrices auf der Basis von Kollagen Typ I zur Anwendung bei degenerativen Bandscheibenerkrankungen},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-106504},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2014},
  abstract  = {Degenerative Bandscheibenerkrankungen wie Protrusionen oder vorgefallenes Nukle-usgewebe f{\"u}hren h{\"a}ufig zu chronischen Schmerzen und schr{\"a}nken die Bewegungsmo-bilit{\"a}t sehr ein. Operative Behandlungsm{\"o}glichkeiten wie die Nukleotomie oder die Fusion von Wirbelk{\"o}rpern stellen traumatische Eingriffe in das komplexe System der Wirbels{\"a}ule dar. Biologische Verfahren, durch die eine Regeneration des gesch{\"a}digten Gewebes erzielt werden kann, sind klinisch bisher nicht etabliert. Ziel dieser Arbeit ist die Entwicklung, Herstellung und Testung regenerativer azellul{\"a}-rer Implantatmatrices auf der Basis von Kollagen Typ I, die den degenerierten Nukleus pulposus ersetzen sollen. Insbesondere eine H{\"o}henminderung der Bandscheibe kann zu Anschlussdegenerationen benachbarter Segmente f{\"u}hren. Dies soll durch die Implan-tatmatrix ausgeglichen werden. Nach der Konstruktion und dem Bau eines Reaktors aus dem Hochleistungskunststoff Polytetrafluorethylen (PTFE), der allen Anforderungen eines CE-Konformit{\"a}tsbewertungsverfahrens entspricht, wird eine hoch verdichtete Kollagen Typ I Matrix mit einer St{\"a}rke von 1 mm hergestellt. Diese kann {\"u}ber den Pro-zess der Lyophilisation auf 0,6 mm weiter reduziert werden. Es gelingt, die Matrix in einer Edelstahlh{\"u}lse zu platzieren, {\"u}ber die mit Hilfe eines passgenauen F{\"u}hrungssta-bes die endoskopische Implantation in die Nukleuskavit{\"a}t erfolgen soll. Im Rahmen der Interkorporellen Fusionstage des Diakonie Klinikums Stuttgart wird das operative Handling an einem humanem Pr{\"a}parat simuliert. Die Implantation erfolgt offen {\"u}ber einen transforaminalen Zugang in zwei nukleotomierte Segmente der lumbalen Wir-bels{\"a}ule. Die anwesenden Wirbels{\"a}ulenchirurgen beurteilen die M{\"o}glichkeit der endo-skopischen Applikation als positiv und machbar. Durch den Zusatz des Polysaccharids Hyalurons{\"a}ure gelingt es, die Quelleigenschaften der hoch verdichteten Matrix zu steigern, so dass diese wie natives Nukleusgewebe in der Lage ist, Fl{\"u}ssigkeit in Ruhe wieder aufzunehmen. Das Quellpotential und die da-mit einhergehende Volumenzunahme nach Kompression sind f{\"u}r ein Nukleusersatzma-terial essentiell. Die hier verwendete Hyalurons{\"a}ure geht jedoch im offenen System der in vitro Inkubation innerhalb von 11 Tagen verloren. Dennoch zeigen sich weitere Vorteile gegen{\"u}ber der Matrix ohne Hyalurons{\"a}ure-Zusatz innerhalb der Testungen heraus. Diese sind neben dem erh{\"o}hten Quellpotential z. B. eine gesteigerte Rate der Zellproliferation der verwendeten bovinen und humanen Bandscheibenzellen (bBSZ und hBSZ) sowie humanen mesenchymalen Stammzellen (hMSC), die {\"u}ber die Be-stimmung der Zellzahl und Viabilit{\"a}t ermittelt wird. Zudem zeigt sich eine gesteigerte mechanische Stabilit{\"a}t, die {\"u}ber die Spannungs-Kompressions-Messungen evaluiert wird. {\"U}ber Lebend-/ Totf{\"a}rbungen und Zytotoxizit{\"a}tstests an Monolayerkulturen kann zudem nachgewiesen werden, dass die notwendige Endsterilisation durch γ-Bestrahlung zu keinen zytotoxischen Ver{\"a}nderungen der Matrix f{\"u}hrt. Da die verdich-tete Implantatmatrix azellul{\"a}r als Medizinprodukt der Klasse III eingesetzt werden soll, wird als erg{\"a}nzende Matrix zur F{\"u}llung kleinster Hohlr{\"a}ume die zun{\"a}chst fl{\"u}ssige ChondroFillerliquid Matrix (ein Knorpelersatzmaterial der Firma Amedrix GmbH, Esslin-gen) durch den Zusatz von Hyalurons{\"a}ure modifiziert und in der Zellkultur getestet. Da es sich hierbei um ein Zweikammerspritzensystem handelt, ist die Verwendung von Additiva wie z. B. Stammzellen technisch m{\"o}glich. Die Ermittlung der maximalen Inku-bationszeit von Zellen in verschieden konzentrierten hyperosmotischen Neutralisations-l{\"o}sungen ergibt eine Dauer von 5 min, bis irreversible Zellsch{\"a}den auftreten. In Migra-tionsversuchen kann gezeigt werden, dass die ChondroFillerliquid Matrix als Konektiv zwischen nativem Nukleusgewebe und verdichteter Implantatmatrix fungiert. Des Wei-teren synthetisieren bBSZ, hBSZ und hMSC sulfatierte Glykosaminoglykane und behal-ten dabei ihr charakteristisches Genexpressionsprofil. Die chondrogene Differenzie-rung durch die Verwendung eines chondrogenen Differenzierungsmediums gelingt bei den hMSC bereits nach einer Kultivierungsdauer von 14 d. Die Zellverteilung in den Implantatmatrices und deren Morphologie entspricht dem nativen Nukleusgewebe. Die biomechanische Testung an einem international anerkannten Modellsystem f{\"u}r humane Wirbels{\"a}ulen - der Kalbswirbels{\"a}ule - ergibt, dass die Nukleotomie zu einer Erh{\"o}hung des Range of Motion (RoM) in alle Richtungen nach Flexion/Extension, Seit-neigung rechts/links und axiale Rotation rechts/links sowie zu einer H{\"o}henreduktion des Segments im Vergleich zum Intaktzustand f{\"u}hrt. Nach der Implantation der ver-dichteten Implantatmatrix wird der RoM deutlich reduziert. Das Segment weist dadurch eine hohe Steifigkeit {\"a}hnlich dem Intaktzustand auf. Die H{\"o}henreduktion kann durch die Implantation beinahe vollst{\"a}ndig wieder ausgeglichen werden. Im Rahmen der zyklischen Dauerbelastungen treten jedoch Implantatextrusionen auf. Zudem nimmt die Steifigkeit deutlich ab, der RoM hingegen wieder zu. Da das bovine Modell jedoch nicht der in vivo Situation entspricht und beispielsweise eine zunehmende In-tegration des Implantats durch Einwachsen nicht erm{\"o}glicht, ist die hohe Extrusionsra-te als nicht realistisch zu werten. Klinische Studien am Tier und Mensch m{\"u}ssen zeigen, inwieweit derartige Extrusionen ohne die Verwendung eines Anulusverschlußsystems auftreten. Im Rahmen der vorliegenden Arbeit ist es gelungen, einen geeigneten Reaktor zu ent-wickeln und mit diesem eine biokompatible, stabile und quellf{\"a}hige Matrix herzustel-len, die den H{\"o}henverlust nach einer Nukleotomie auszugleichen vermag. Die modifi-zierte ChondroFillerliquid Matrix stellt eine ideale Erg{\"a}nzung dar, da {\"u}ber diese Zellen oder andere Additiva verabreicht werden k{\"o}nnen und deren konektive Wirkung die Zellbesiedlung der azellul{\"a}ren Matrix beg{\"u}nstigt.},
  subject      = {Bandscheibenkrankheit},
  language  = {de}
}
@phdthesis{Reimer2017,
  author    = {Reimer, Anastasija},
  title     = {Search for novel antimicrobials against \(Neisseria\) \(gonorrhoeae\) and \(Chlamydia\) \(trachomatis\)},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-143168},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2017},
  abstract  = {The obligate human pathogen Neisseria gonorrhoeae is responsible for the widespread sexually transmitted disease gonorrhoea, which in rare cases also leads to the development of disseminated gonococcal infection (DGI). DGI is mediated by PorBIA-expressing bacteria that invade host cells under low phosphate condition by interaction with the scavenger receptor-1 (SREC-I) expressed on the surface of endothelial cells. The interaction of PorBIA and SREC-I was analysed using different in vitro approaches, including surface plasmon resonance experiments that revealed a direct phosphate-independent high affinity interaction of SREC-I to PorBIA. However, the same binding affinity was also found for the other allele PorBIB, which indicates unspecific binding and suggests that the applied methods were unsuitable for this interaction analysis. Since N. gonorrhoeae was recently classified as a "super-bug" due to a rising number of antibiotic-resistant strains, this study aimed to discover inhibitors against the PorBIA-mediated invasion of N. gonorrhoeae. Additionally, inhibitors were searched against the human pathogen Chlamydia trachomatis, which causes sexually transmitted infections as well as infections of the upper inner eyelid. 68 compounds, including plant-derived small molecules, extracts or pure compounds of marine sponges or sponge-associated bacteria and pipecolic acid derivatives, were screened using an automated microscopy based approach. No active substances against N. gonorrhoeae could be identified, while seven highly antichlamydial compounds were detected. The pipecolic acid derivatives were synthesized as potential inhibitors of the virulence-associated "macrophage infectivity potentiator" (MIP), which exhibits a peptidyl prolyl cis-trans isomerase (PPIase) enzyme activity. This study investigated the role of C. trachomatis and N. gonorrhoeae MIP during infection. The two inhibitors PipN3 and PipN4 decreased the PPIase activity of recombinant chlamydial and neisserial MIP in a dose-dependent manner. Both compounds affected the chlamydial growth and development in epithelial cells. Furthermore, this work demonstrated the contribution of MIP to a prolonged survival of N. gonorrhoeae in the presence of neutrophils, which was significantly reduced in the presence of PipN3 and PipN4. SF2446A2 was one of the compounds that had a severe effect on the growth and development of C. trachomatis. The analysis of the mode of action of SF2446A2 revealed an inhibitory effect of the compound on the mitochondrial respiration and mitochondrial ATP production of the host cell. However, the chlamydial development was independent of proper functional mitochondria, which excluded the connection of the antichlamydial properties of SF2446A2 with its inhibition of the respiratory chain. Only the depletion of cellular ATP by blocking glycolysis and mitochondrial respiratory chain inhibited the chlamydial growth. A direct effect of SF2446A2 on C. trachomatis was assumed, since the growth of the bacteria N. gonorrhoeae and Staphylococcus aureus was also affected by the compound. In summary, this study identified the severe antichlamydial activity of plant-derived naphthoquinones and the compounds derived from marine sponges or sponge-associated bacteria SF2446A2, ageloline A and gelliusterol E. Furthermore, the work points out the importance of the MIP proteins during infection and presents pipecolic acid derivatives as novel antimicrobials against N. gonorrhoeae and C. trachomatis.},
  subject      = {Neisseria gonorrhoeae},
  language  = {en}
}
@phdthesis{Busch2013,
  author    = {Busch, Rhoda},
  title     = {Redundancy and indispensability of NFATc1-isoforms in the adaptive and innate immune system},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-91096},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2013},
  abstract  = {Peritonitis is a common disease in man, frequently caused by fungi, such as Candida albicans; however, in seldom cases opportunistic infections with Saccharomyces cerevisiae are described. Resident peritoneal macrophages (prMΦ) are the major group of phagocytic cells in the peritoneum. They express a broad range of surface pattern recognition receptors (PRR) to recognize invaders. Yeast infections are primarily detected by the Dectin-1 receptor, which triggers activation of NFAT and NF-κB pathways. The transcription of the Nfatc1 gene is directed by the two alternative promoters, inducible P1 and relatively constitutive P2 promoter. While the role of P1-directed NFATc1α-isoforms to promote survival and proliferation of activated lymphocytes is well-established, the relevance of constitutively generated NFATc1β-isoforms, mainly expressed in resting lymphocytes, myeloid and non-lymphoid cells, remains unclear. Moreover, former work at our department indicated different roles for NFATc1α- and NFATc1β-proteins in lymphocytes. Our data revealed the functional role of NFATc1 in peritoneal resident macrophages. We demonstrated that the expression of NFATc1β is required for a proper immune response of prMΦ during fungal infection-induced acute peritonitis. We identified Ccl2, a major chemokine produced in response to fungal infections by prMΦ, as a novel NFATc1 target gene which is cooperatively regulated through the NFAT- and canonical NF-κB pathways. Consequently, we showed that NFATc1β deficiency in prMΦ results in a decreased infiltration of inflammatory monocytes, leading to a delayed clearance of peritoneal fungal infection. We could further show that the expression of NFATc1β-isoforms is irrelevant for homeostasis of myeloid and adaptive immune system cells and that NFATc1α- (but not β-) isoforms are required for a normal development of peritoneal B1a cells. In contrast to the situation in myeloid cells, NFATc1β deficiency is compensated by increased expression of NFATc1α-isoforms in lymphoid cells. As a consequence, NFATc1ß is dispensable for activation of the adaptive immune system. Taken together our results illustrate the redundancy and indispensability of NFATc1-isoforms in the adaptive and innate immune system, indicating a complex regulatory system for Nfatc1 gene expression in different compartments of the immune system and likely beyond that.},
  subject      = {Immunsystem},
  language  = {en}
}
@phdthesis{Pfeuffer2000,
  author    = {Pfeuffer, Thilo},
  title     = {Interaktion des Proteins ActA von Listeria monocytogenes mit dem Wirtszellprotein LaXp180},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-1859},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2000},
  abstract  = {Listeria monocytogenes, ein fakultativ intrazellul{\"a}rer Krankheitserreger, besitzt die F{\"a}higkeit, Wirtszellen zu penetrieren, sich in ihnen zu vermehren, sich intrazellul{\"a}r zu bewegen und auch benachbarte Zellen direkt zu infizieren. Die intrazellul{\"a}re Fortbewegung erfolgt durch Polymerisation von zellul{\"a}rem Aktin, wodurch charakteristische Aktinschweife an einem Pol der Bakterien entstehen. Der einzige bakterielle Faktor, der f{\"u}r die Aktinpolymerisation notwendig ist, ist das Oberfl{\"a}chenprotein ActA. ActA allein ist aber nicht in der Lage, Aktin zu polymerisieren, sondern kann dies nur in Assoziation mit Proteinen der Wirtszelle. Die einzigen bisher bekannten Wirtszellproteine, die direkt mit ActA interagieren, sind das Phosphoprotein VASP und der Arp2/3-Komplex. VASP bindet an den zentralen prolinreichen Bereich von ActA und beschleunigt durch die Rekrutierung von Profilin den Prozeß der Aktinpolymerisation. Der Arp2/3-Komplex interagiert mit dem N-terminalen Bereich von ActA und initiiert die eigentliche Aktin-Polymerisation. Um weitere eukaryotische, mit ActA interagierende Proteine (AIPs) zu isolieren, wurde {\"u}ber einen "Yeast Two-Hybrid"-Test mit ActA als K{\"o}der eine embryonale Maus-cDNA-Genbank getestet. Dabei wurden drei verschiedene AIPs identifiziert, von denen eines identisch mit dem humanen Protein LaXp180 (auch "CC1" genannt) ist. LaXp180 ist ein 180 kDa Protein mit {\"u}ber 50 theoretischen Phosphorylierungsstellen in der N-terminalen H{\"a}lfte, w{\"a}hrend die C-terminale H{\"a}lfte "coiled-coil"-Strukturen ausbilden kann. Dar{\"u}berhinaus enth{\"a}lt LaXp180 eine Kern-Lokalisations-Sequenz und ein Leucin-Zipper-Motiv. Die Bindung von LaXp180 an ActA wurde in vitro unter Verwendung von rekombinantem His6-Tag-LaXp180 und rekombinantem ActA best{\"a}tigt, da rekombinantes ActA nur an einer Ni-Agarose-S{\"a}ule gebunden wurde, wenn diese vorher mit His6-Tag-LaXp180 beladen war. {\"U}ber RT-PCR konnte zum ersten Mal die Expression LaXp180-spezifischer mRNA in verschiedenen S{\"a}ugerzellen nachgewiesen und mit einem polyklonalen anti-LaXp180-Serum durch Immunopr{\"a}zipitation erstmals ein 194 kDa großes Protein in S{\"a}ugerzellextrakten detektiert werden. Die intrazellul{\"a}re Lokalisation von LaXp180 wurde {\"u}ber Immunfluoreszenzmikroskopie untersucht. Immunfluoreszenzf{\"a}rbungen von Fibroblasten mit dem anti-LaXp180-Serum zeigten eine starke F{\"a}rbung der Zellkerne und definierter Bereiche direkt neben den Kernen, w{\"a}hrend das restliche Zytoplasma schwach gef{\"a}rbt war. {\"U}ber Immunfluoreszenzmikroskopie mit dem anti-LaXp180-Serum an mit L. monocytogenes infizierten Zellen konnte gezeigt werden, daß LaXp180 mit der Oberfl{\"a}che vieler, aber nicht aller intrazellul{\"a}rer, ActA-exprimierender Listerien kolokalisiert. Dagegen wurde nie eine Kolokalisation mit intrazellul{\"a}ren, aber ActA-defizienten Mutanten beobachtet. Dar{\"u}berhinaus ist LaXp180 asymmetrisch auf der Bakterienoberfl{\"a}che verteilt und schließt sich gegenseitig mit der F-Aktin-Polymerisation aus. LaXp180 ist ein putativer Bindungspartner von Stathmin, einem 19 kDa Phosphoprotein, das die Mikrotubuli-Dynamik reguliert. {\"U}ber Immunfluoreszenz konnte gezeigt werden, daß auch Stathmin mit intrazellul{\"a}ren, ActA-exprimierenden L. monocytogenes kolokalisiert.},
  subject      = {Listeria monocytogenes},
  language  = {de}
}
@phdthesis{Duque2022,
  author    = {Duque, Laura Maria Ribeiro},
  title     = {Effects of ozone on plants and plant-insect interactions},
  doi       = {10.25972/OPUS-27798},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-277983},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2022},
  abstract  = {Anthropogenic activities are causing air pollution. Amongst air pollutants, tropospheric ozone is a major threat to human health and ecosystem functioning. In this dissertation, I present three studies that aimed at increasing our knowledge on how plant exposure to ozone affects its reproduction and its interactions with insect herbivores and pollinators. For this purpose, a new fumigation system was built and placed in a greenhouse. The annual plant Sinapis arvensis (wild mustard) was used as the model plant. Plants were exposed to either 0 ppb (control) or 120 ppb of ozone, for variable amounts of time and at different points of their life cycle. After fumigation, plants were exposed to herbivores or pollinators in the greenhouse, or to both groups of insects in the field. My research shows that ozone affected reproductive performance differently, depending on the timing of exposure: plants exposed at earlier ages had their reproductive fitness increased, while plants exposed later in their life cycle showed a tendency for reduced reproductive fitness. Plant phenology was a key factor influencing reproductive fitness: ozone accelerated flowering and increased the number of flowers produced by plants exposed at early ages, while plants exposed to ozone at later ages tended to have fewer flowers. On the other hand, the ozone-mediated changes in plant-insect interactions had little impact on plant reproductive success. The strongest effect of ozone on plant-pollinator interactions was the change in the number of flower visits received per plant, which was strongly linked to the number of open flowers. This means that, as a rule, exposure of plants to ozone early in the life cycle resulted in a higher number of pollinator visits, while exposure later in the life cycle resulted in fewer flower visits by potential pollinators. An exception was observed: the higher number of visits performed by large syrphid flies to young ozone-exposed plants than to the respective control plants went beyond the increase in the number of open flowers in those plants. Also, honeybees spent more time per flower in plants exposed to ozone than on control plants, while other pollinators spent similar amounts of time in control and ozone-exposed plants. This guild-dependent preference for ozone-exposed plants may be due to species-specific preferences related to changes in the quality and quantity of floral rewards. In the field, ozone-exposed plants showed only a tendency for increased colonization by sucking herbivores and slightly more damage by chewing herbivores than control plants. On the other hand, in the greenhouse experiment, Pieris brassicae butterflies preferred control plants over ozone-exposed plants as oviposition sites. Eggs laid on ozone-exposed plants took longer to hatch, but the chances of survival were higher. Caterpillars performed better in control plants than in ozone-exposed plants, particularly when the temperature was high. Most of the described effects were dependent on the duration and timing of the ozone exposure and the observed temperature, with the strongest effects being observed for longer exposures and higher temperatures. Furthermore, the timing of exposure altered the direction of the effects. The expected climate change provides ideal conditions for further increases in tropospheric ozone concentrations, therefore for stronger effects on plants and plant-insect interactions. Acceleration of flowering caused by plant exposure to ozone may put plant-pollinator interactions at risk by promoting desynchronization between plant and pollinator activities. Reduced performance of caterpillars feeding on ozone-exposed plants may weaken herbivore populations. On the other hand, the increased plant reproduction that results from exposing young plants to ozone may be a source of good news in the field of horticulture, when similar results would be achieved in high-value crops. However, plant response to ozone is highly species-specific. In fact, Sinapis arvensis is considered a weed and the advantage conferred by ozone exposure may increase its competitiveness, with negative consequences for crops or plant communities in general. Overall, plant exposure to ozone might constitute a threat for the balance of natural and agro-ecosystems.},
  subject      = {Plant},
  language  = {en}
}
@phdthesis{Hartmann2024,
  author    = {Hartmann, Oliver},
  title     = {Development of somatic modified mouse models of Non-Small cell lung cancer},
  doi       = {10.25972/OPUS-36340},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-363401},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2024},
  abstract  = {In 2020, cancer was the leading cause of death worldwide, accounting for nearly 10 million deaths. Lung cancer was the most common cancer, with 2.21 million cases per year in both sexes. This non-homogeneous disease is further subdivided into small cell lung cancer (SCLC, 15\%) and non-small cell lung cancer (NSCLC, 85\%). By 2023, the American Cancer Society estimates that NSCLC will account for 13\% of all new cancer cases and 21\% of all estimated cancer deaths. In recent years, the treatment of patients with NSCLC has improved with the development of new therapeutic interventions and the advent of targeted and personalised therapies. However, these advances have only marginally improved the five-year survival rate, which remains alarmingly low for patients with NSCLC. This observation highlights the importance of having more appropriate experimental and preclinical models to recapitulate, identify and test novel susceptibilities in NSCLC. In recent years, the Trp53fl/fl KRaslsl-G12D/wt mouse model developed by Tuveson, Jacks and Berns has been the main in vivo model used to study NSCLC. This model mimics ADC and SCC to a certain extent. However, it is limited in its ability to reflect the genetic complexity of NSCLC. In this work, we use CRISPR/Cas9 genome editing with targeted mutagenesis and gene deletions to recapitulate the conditional model. By comparing the Trp53fl/fl KRaslsl- G12D/wt with the CRISPR-mediated Trp53mut KRasG12D, we demonstrated that both showed no differences in histopathological features, morphology, and marker expression. Furthermore, next-generation sequencing revealed a very high similarity in their transcriptional profile. Adeno-associated virus-mediated tumour induction and the modular design of the viral vector allow us to introduce additional mutations in a timely manner. CRISPR-mediated mutation of commonly mutated tumour suppressors in NSCLC reliably recapitulated the phenotypes described in patients in the animal model. Lastly, the dual viral approach could induce the formation of lung tumours not only in constitutive Cas9 expressing animals, but also in wildtype animals. Thus, the implementation of CRISPR genome editing can rapidly advance the repertoire of in vivo models for NSCLC research. Furthermore, it can reduce the necessity of extensive breeding.},
  subject      = {CRISPR/Cas-Methode},
  language  = {en}
}
@phdthesis{Schwarz2023,
  author    = {Schwarz, Jessica Denise},
  title     = {Genome-wide reporter screens identify transcriptional regulators of ribosome biogenesis},
  doi       = {10.25972/OPUS-27901},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-279010},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2023},
  abstract  = {Cellular growth and proliferation are among the most important processes for cells and organisms. One of the major determinants of these processes is the amount of proteins and consequently also the amount of ribosomes. Their synthesis involves several hundred proteins and four different ribosomal RNA species, is highly coordinated and very energy-demanding. However, the molecular mechanims of transcriptional regulation of the protein-coding genes involved, is only poorly understood in mammals. In this thesis, unbiased genome-wide knockout reporter screens were performed, aiming to identify previously unknown transcriptional regulators of ribosome biogenesis factors (RiBis), which are important for the assembly and maturation of ribosomes, and ribosomal proteins (RPs), which are ribosomal components themself. With that approach and follow-up (validation) experiments, ALDOA and RBM8A among others, could be identified as regulators of ribosome biogenesis. Depletion of the glycolytic enzyme ALDOA led to a downregulation of RiBi- and RPpromoter driven reporters on protein and transcript level, as well as to a downregulation of ribosome biogenesis gene transcripts and of mRNAs of other genes important for proliferation. Reducing the amount of the exon junction complex protein RBM8A, led to a more prominent downregulation of one of the fluorescent reporters, but this regulation was independent of the promoter driving the expression of the reporter. However, acute protein depletion experiments in combination with nascent RNA sequencing (4sU-Seq) revealed, that mainly cytosolic ribosomal proteins (CRPs) were downregulated upon acute RBM8A withdrawal. ChIP experiments showed RBM8A binding to promoters of RP genes, but also to other chromatin regions. Total POL II or elongating and initiating POL II levels were not altered upon acute RBM8A depletion. These data provide a starting point for further research on the mechanisms of transcriptional regulation of RP and RiBi genes in mammals.},
  subject      = {Ribosom},
  language  = {en}
}
@phdthesis{Streinzer2013,
  author    = {Streinzer, Martin},
  title     = {Sexual dimorphism of the sensory systems in bees (Hymenoptera, Apoidea) and the evolution of sex-specific adaptations in the context of mating behavior},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-78689},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2013},
  abstract  = {Bees have had an intimate relationship with humans for millennia, as pollinators of fruit, vegetable and other crops and suppliers of honey, wax and other products. This relationship has led to an extensive understanding of their ecology and behavior. One of the most comprehensively understood species is the Western honeybee, Apis mellifera. Our understanding of sex-specific investment in other bees, however, has remained superficial. Signals and cues employed in bee foraging and mating behavior are reasonably well understood in only a handful of species and functional adaptations are described in some species. I explored the variety of sensory adaptations in three model systems within the bees. Females share a similar ecology and similar functional morphologies are to be expected. Males, engage mainly in mating behavior. A variety of male mating strategies has been described which differ in their spatiotemporal features and in the signals and cues involved, and thus selection pressures. As a consequence, males' sensory systems are more diverse than those of females. In the first part I studied adaptations of the visual system in honeybees. I compared sex and caste-specific eye morphology among 5 species (Apis andreniformis, A. cerana, A. dorsata, A. florea, A. mellifera). I found a strong correlation between body size and eye size in both female castes. Queens have a relatively reduced visual system which is in line with the reduced role of visual perception in their life history. Workers differed in eye size and functional morphology, which corresponds to known foraging differences among species. In males, the eyes are conspicuously enlarged in all species, but a disproportionate enlargement was found in two species (A. dorsata, A. florea). I further demonstrate a correlation between male visual parameters and mating flight time, and propose that light intensities play an important role in the species-specific timing of mating flights. In the second study I investigated eye morphology differences among two phenotypes of drones in the Western honeybee. Besides normal-sized drones, smaller drones are reared in the colony, and suffer from reduced reproductive success. My results suggest that the smaller phenotype does not differ in spatial resolution of its visual system, but suffers from reduced light and contrast sensitivity which may exacerbate the reduction in reproductive success caused by other factors. In the third study I investigated the morphology of the visual system in bumblebees. I explored the association between male eye size and mating behavior and investigated the diversity of compound eye morphology among workers, queens and males in 11 species. I identified adaptations of workers that correlate with distinct foraging differences among species. Bumblebee queens must, in contrast to honeybees, fulfill similar tasks as workers in the first part of their life, and correspondingly visual parameters are similar among both female castes. Enlarged male eyes are found in several subgenera and have evolved several times independently within the genus, which I demonstrate using phylogenetic informed statistics. Males of these species engage in visually guided mating behavior. I find similarities in the functional eye morphology among large-eyed males in four subgenera, suggesting convergent evolution as adaptation to similar visual tasks. In the remaining species, males do not differ significantly from workers in their eye morphology. In the fourth study I investigated the sexual dimorphism of the visual system in a solitary bee species. Males of Eucera berlandi patrol nesting sites and compete for first access to virgin females. Males have enlarged eyes and better spatial resolution in their frontal eye region. In a behavioral study, I tested the effect of target size and speed on male mate catching success. 3-D reconstructions of the chasing flights revealed that angular target size is an important parameter in male chasing behavior. I discuss similarities to other insects that face similar problems in visual target detection. In the fifth study I examined the olfactory system of E. berlandi. Males have extremely long antennae. To investigate the anatomical grounds of this elongation I studied antennal morphology in detail in the periphery and follow the sexual dimorphism into the brain. Functional adaptations were found in males (e.g. longer antennae, a multiplication of olfactory sensilla and receptor neurons, hypertrophied macroglomeruli, a numerical reduction of glomeruli in males and sexually dimorphic investment in higher order processing regions in the brain), which were similar to those observed in honeybee drones. The similarities and differences are discussed in the context of solitary vs. eusocial lifestyle and the corresponding consequences for selection acting on males.},
  subject      = {Biene},
  language  = {en}
}
@phdthesis{Schilcher2023,
  author    = {Schilcher, Felix},
  title     = {Regulation of the nurse-forager transition in honeybees (\(Apis\) \(mellifera\))},
  doi       = {10.25972/OPUS-28935},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-289352},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2023},
  abstract  = {Honeybees are among the few animals that rely on eusociality to survive. While the task of queen and drones is only reproduction, all other tasks are accomplished by sterile female worker bees. Different tasks are mostly divided by worker bees of different ages (temporal polyethism). Young honeybees perform tasks inside the hive like cleaning and nursing. Older honeybees work at the periphery of the nest and fulfill tasks like guarding the hive entrance. The oldest honeybees eventually leave the hive to forage for resources until they die. However, uncontrollable circumstances might force the colony to adapt or perish. For example, the introduced Varroa destructor mite or the deformed wing virus might erase a lot of in-hive bees. On the other hand, environmental events might kill a lot of foragers, leaving the colony with no new food intake. Therefore, adaptability of task allocation must be a priority for a honeybee colony. In my dissertation, I employed a wide range of behavioral, molecular biological and analytical techniques to unravel the underlying molecular and physiological mechanisms of the honeybee division of labor, especially in conjunction with honeybee malnourishment. The genes AmOARα1, AmTAR1, Amfor and vitellogenin have long been implied to be important for the transition from in-hive tasks to foraging. I have studied in detail expression of all of these genes during the transition from nursing to foraging to understand how their expression patterns change during this important phase of life. My focus lay on gene expression in the honeybee brain and fat body. I found an increase in the AmOARα1 and the Amforα mRNA expression with the transition from in-hive tasks to foraging and a decrease in expression of the other genes in both tissues. Interestingly, I found the opposite pattern of the AmOARα1 and AmTAR1 mRNA expression in the honeybee fat body during orientation flights. Furthermore, I closely observed juvenile hormone titers and triglyceride levels during this crucial time. Juvenile hormone titers increased with the transition from in-hive tasks to foraging and triglyceride levels decreased. Furthermore, in-hive bees and foragers also differ on a behavioral and physiological level. For example, foragers are more responsive towards light and sucrose. I proposed that modulation via biogenic amines, especially via octopamine and tyramine, can increase or decrease the responsiveness of honeybees. For that purpose, in-hive bees and foragers were injected with both biogenic amines and the receptor response was quantified 1 using electroretinography. In addition, I studied the behavioral response of the bees to light using a phototaxis assay. Injecting octopamine increased the receptor response and tyramine decreased it. Also, both groups of honeybees showed an increased phototactic response when injected with octopamine and a decreased response when injected with tyramine, independent of locomotion. Additionally, nutrition has long been implied to be a driver for division of labor. Undernourished honeybees are known to speed up their transition to foragers, possibly to cope with the missing resources. Furthermore, larval undernourishment has also been implied to speed up the transition from in-hive bees to foragers, due to increasing levels of juvenile hormone titers in adult honeybees after larval starvation. Therefore, I reared honeybees in-vitro to compare the hatched adult bees of starved and overfed larvae to bees reared under the standard in-vitro rearing diet. However, first I had to investigate whether the in-vitro rearing method affects adult honeybees. I showed effects of in-vitro rearing on behavior, with in-vitro reared honeybees foraging earlier and for a shorter time than hive reared honeybees. Yet, nursing behavior was unaffected. Afterwards, I investigated the effects of different larval diets on adult honeybee workers. I found no effects of malnourishment on behavioral or physiological factors besides a difference in weight. Honeybee weight increased with increasing amounts of larval food, but the effect seemed to vanish after a week. These results show the complexity and adaptability of the honeybee division of labor. They show the importance of the biogenic amines octopamine and tyramine and of the corresponding receptors AmOARα1 and AmTAR1 in modulating the transition from inhive bees to foragers. Furthermore, they show that in-vitro rearing has no effects on nursing behavior, but that it speeds up the transition from nursing to foraging, showing strong similarities to effects of larval pollen undernourishment. However, larval malnourishment showed almost no effects on honeybee task allocation or physiology. It seems that larval malnourishment can be easily compensated during the early lifetime of adult honeybees.},
  subject      = {Biene},
  language  = {en}
}
@phdthesis{Hafen2015,
  author    = {Hafen, Bettina},
  title     = {Physical contact between mesenchymal stem cells and endothelial precursors induces distinct signatures with relevance to tissue regeneration and engineering},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-119417},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2015},
  abstract  = {The goal of the project VascuBone is to develop a tool box for bone regeneration, which on one hand fulfills basic requirements (e.g. biocompatibility, properties of the surface, strength of the biomaterials) and on the other hand is freely combinable with what is needed in the respective patient's situation. The tool box will include a variation of biocompatible biomaterials and cell types, FDA-approved growth factors, material modification technologies, simulation and analytical tools like molecular imaging-based in vivo diagnostics, which can be combined for the specific medical need. This tool box will be used to develop translational approaches for regenerative therapies of different types of bone defects. This project receives funding from the European Union's Seventh Framework Program (VascuBone 2010). The present study is embedded into this EU project. The intention of this study is to assess the changes of the global gene expression patterns of endothelial progenitor cells (EPCs) and mesenchymal stem cells (MSCs) after direct cell-cell contact as well as the influence of conditioned medium gained from MSCs on EPCs and vice versa. EPCs play an important role in postnatal vasculogenesis. An intact blood vessel system is crucial for all tissues, including bone. Latest findings in the field of bone fracture healing and repair by the use of tissue engineering constructs seeded with MSCs raised the idea of combining MSCs and EPCs to enhance vascularization and therefore support survival of the newly built bone tissue. RNA samples from both experimental set ups were hybridized on Affymetrix GeneChips® HG-U133 Plus 2.0 and analyzed by microarray technology. Bioinformatic analysis was applied to the microarray data and verified by RT-PCR. This study gives detailed information on how EPCs and MSCs communicate with each other and therefore gives insights into the signaling pathways of the musculoskeletal system. These insights will be the base for further functional studies on protein level for the purpose of tissue regeneration. A better understanding of the cell communication of MSCs and EPCs and subsequently the targeting of relevant factors opens a variety of new opportunities, especially in the field of tissue engineering. The second part of the present work was to develop an ELISA (enzyme-linked immunosorbent assay) for a target protein from the lists of differentially expressed genes revealed by the microarray analysis. This project was in cooperation with Immundiagnostik AG, Bensheim, Germany. The development of the ELISA aimed to have an in vitro diagnostic tool to monitor e.g. the quality of cell seeded tissue engineering constructs. The target protein chosen from the lists was klotho. Klotho seemed to be a very promising candidate since it is described in the literature as anti-aging protein. Furthermore, studies with klotho knock-out mice showed that these animals suffered from several age-related diseases e.g. osteoporosis and atherosclerosis. As a co-receptor for FGF23, klotho plays an important role in bone metabolism. The present study will be the first one to show that klotho is up-regulated in EPCs after direct cell-cell contact with MSCs. The development of an assay with a high sensitivity on one hand and the capacity to differentiate between secreted and shedded klotho on the other hand will allow further functional studies of this protein and offers a new opportunity in medical diagnostics especially in the field of metabolic bone disease.},
  subject      = {Vorl{\"a}uferzelle},
  language  = {en}
}
@phdthesis{Ullrich2014,
  author    = {Ullrich, Melanie},
  title     = {Identification of SPRED2 as a Novel Regulator of Hypothalamic-Pituitary-Adrenal Axis Activity and of Body Homeostasis},
  url       = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-107355},
  school      = {Universit{\"a}t W{\"u}rzburg},
  year      = {2014},
  abstract  = {SPRED proteins are inhibitors of the Ras/ERK/MAPK signaling pathway, an evolutionary highly conserved and very widespread signaling cascade regulating cell proliferation, differentiation, and growth. To elucidate physiological consequences of SPRED2 deficiency, SPRED2 KO mice were generated by a gene trap approach. An initial phenotypical characterization of KO mice aged up to five months identified SPRED2 as a regulator of chondrocyte differentiation and bone growth. Here, the loss of SPRED2 leads to an augmented FGFR-dependent ERK activity, which in turn causes hypochondroplasia-like dwarfism. However, long term observations of older KO mice revealed a generally bad state of health and manifold further symptoms, including excessive grooming associated with severe self-inflicted wounds, an abnormally high water uptake, clear morphological signs of kidney deterioration, and a reduced survival due to sudden death. Based on these observations, the aim of this study was to discover an elicitor of this complex and versatile phenotype. The observed kidney degeneration in our SPRED2 KO mice was ascribed to hydronephrosis characterized by severe kidney atrophy and apoptosis of renal tubular cells. Kidney damage prompted us to analyze drinking behavior and routine serum parameters. Despite polydipsia, which was characterized by a nearly doubled daily water uptake, the significantly elevated Na+ and Cl- levels and the resulting serum hyperosmolality could not be compensated in SPRED2 KOs. Since salt and water balance is primarily under hormonal control of aldosterone and AVP, we analyzed both hormone levels. While serum AVP was similar in WTs and KOs, even after experimental water deprivation and an extreme loss of body fluid, serum aldosterone was doubled in SPRED2 KO mice. Systematic investigation of contributing upstream hormone axes demonstrated that hyperaldosteronism developed independently of an overactivated Renin-Angiotensin system as indicated by halved serum Ang II levels in KO mice. However, aldosterone synthase expression in the adrenal gland was substantially augmented. Serum corticosterone, which is like aldosterone released from the adrenal cortex, was more than doubled in SPRED2 KOs, too. Similar to corticosterone, the production of aldosterone is at least in part under control of pituitary ACTH, which is further regulated by upstream hypothalamic CRH release. In fact, stress hormone secretion from this complete hypothalamic-pituitary-adrenal axis was upregulated because serum ACTH, the mid acting pituitary hormone, and hypothalamic CRH, the upstream hormonal inductor of HPA axis activity, were also elevated by 30\% in SPRED2 KO mice. This was accompanied by an upregulated ERK activity in paraventricular nucleus-containing hypothalamic brain regions and by augmented hypothalamic CRH mRNA levels in our SPRED2 KO mice. In vitro studies using the hypothalamic cell line mHypoE-44 further demonstrated that both SPRED1 and SPRED2 were able to downregulate CRH promoter activity, CRH secretion, and Ets factor-dependent CRH transcription. This was in line with the presence of various Ets factor binding sites in the CRH promoter region, especially for Ets1. Thus, this study shows for the first time that SPRED2-dependent inhibition of Ras/ERK/MAPK signaling by suppression of ERK activity leads to a downregulation of Ets1 factor-dependent transcription, which further results in inhibition of CRH promoter activity, CRH transcription, and CRH release from the hypothalamus. The consecutive hyperactivity of the complete HPA axis in our SPRED2 KO mice reflects an elevated endogenous stress response becoming manifest by excessive grooming behavior and self-inflicted skin lesions on the one hand; on the other hand, in combination with elevated aldosterone synthase expression, this upregulated HPA hormone release explains hyperaldosteronism and the associated salt and water imbalances. Both hyperaldosteronism and polydipsia very likely contribute further to the observed kidney damage. Taken together, this study initially demonstrates that SPRED2 is essential for the appropriate regulation of HPA axis activity and of body homeostasis. To further enlighten and compare consequences of SPRED2 deficiency in mice and particularly in humans, two follow-up studies investigating SPRED2 function especially in heart and brain, and a genetic screen to identify human SPRED2 loss-of-function mutations are already in progress.},
  subject      = {Renin-Angiotensin-System},
  language  = {en}
}