Thesis (M.S.)--University of Wisconsin--Madison, 1967. / eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references.

Thesis (Ph. D.)--Oregon State University, 2007. / Printout. Includes bibliographical references (leaves 101-109). Also available on the World Wide Web.

Thesis (M.S.)--Brigham Young University. Dept. of Chemical Engineering, 2005. / Includes bibliographical references (p. 101-110).

Thesis (Ph. D.)--University of Canterbury, 2005. / Typescript (photocopy). Includes bibliographical references (leaves 140-159). Also available via the World Wide Web.

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Spores are fundamental to the reproductive success of all land plants. The success of a spore lies in its recalcitrant multi-layered spore wall commonly made of sporopollenin, cellulose, and pectin. However, other polysaccharides may be associated with the intine of spores, and their patterns of deposition vary across taxa. Callose, a plant 1-3-β-glucan polysaccharide, has unsubstantiated accounts of its presence and absence in association with the spore mother cell wall of hornworts for more than a century. To address this conundrum, I used aniline blue, a fluorochrome that has high specificity of binding to beta glucans and when excited with ultraviolet light, it will fluoresce yellow-green. However, due to the limited resolution power of that technique, I also used transmission electron microscopy (TEM) with immunogold labeling to observe the ultrastructural localization of callose using anticallose, a monoclonal antibody. I also bioinformatically probed the genomes and transcriptomes of hornworts to elucidate the callose synthase genes and enzymes which may be responsible for the putative callose deposition. Because of the asynchronous spore development, each sporophyte has a continuum of spores at various developmental stages. Subsequently, the sporophyte of many hornworts makes an ideal system to study all aspects of sporogenesis. For the first time, I provide unequivocal, correlative evidence for callose involvement in spore wall development in hornworts. Here we report on the spatiotemporal deposition of callose in sporogenesis of Phaeoceros carolinianus, and we show that callose is a common wall constituent of the spore intine in three other genera (Anthoceros, Notothylas, and Nothoceros). Callose deposition in hornworts is post-meiotic and begins during early spore wall development after a white lined lemella is formed and during expansion of the exine. As sporopollenin is deposited and the spore wall thickens, callose remains localized in the intine during the remainder of sporogenesis. The occurrence of callose in hornwort spores is a first record of this polysaccharide in the inner spore wall (intine or endospore) of any embryophyte. This suggests that callose may serve the same or similar roles in hornwort intine development and function as pectin-cellulose does in later diverging taxa. Bioinformatic tblastn techniques and molecular high through put Illumina genome sequencing combined with blast techniques for orthologs to callose synthase genes from Arabidopsis thaliana and Physcomitrella patens did not provide any evidence as to if callose synthase genes are present in hornworts This was due to database contamination, sample contamination, and sample quality.

Orientador: Ana Paula Fortuna Perez / Resumo: A família Lygodiaceae sempre teve uma taxonomia contraditória, o número de espécies aceitas no Brasil variando de duas a doze ao longo das décadas. Sendo Lygodiaceae uma família antiga, acredita-se que essas contradições possam ser causadas por híbridos. Os híbridos surgem pelo cruzamento de duas ou mais espécies e, geralmente, são primeiro identificados por suas características morfológicas intermediárias entre as espécies parentais. Normalmente, os híbridos recentes de samambaia são detectados ocorrendo junto com suas espécies parentais. Os esporos de indivíduos híbridos geralmente são mal formados e / ou abortados, portanto, inviáveis, mas os poucos esporos viáveis poderiam iniciar o processo de especiação de uma nova espécie. Considerando a sobreposição de características morfológicas, a hibridação poderia ter ocorrido entre duas espécies de Lygodium no Brasil, L. venustum e L. volubile gerando o hipotético híbrido L. × fayae. Devido à difícil identificação desses táxons, nossos estudos analisaram dois caracteres diferentes (tamanho e morfologia dos esporos e estômatos) por microscopia eletrônica de varredura, estudamos a germinação dos esporos e criamos novas descrições e uma chave de identificação às espécies aceitas para ajudar na delimitação taxonômica dos dois táxons de Lygodium e seu híbrido putativo. De acordo com nossos resultados, as espécies classificadas como L. × fayae na região sudeste do Brasil não são híbridas, mas uma variação morfológica de Lygodium venus... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The family Lygodiaceae has always had a contradictory taxonomy, the number of accepted species in Brazil varying from two to twelve through the decades. Lygodiaceae being an old family, this contradictions are believed to be caused by hybrids. Hybrids arise by the crossing of two or more species, and, usually, they are first identified by its intermediate morphological characteristics among the parental species. Normally, early fern hybrids are detected occurring along with its parental species. The spores of hybrid individuals generally are badly formed and/or aborted, thus, not viable, but the few balanced spores could starts the speciation process of a new species. Considering the overlapping of morphological characteristics, it could have occurred in two species of Lygodium in Brazil, L. venustum and L. volubile and the hypothetical hybrid L. ×fayae. Due to the difficult in identification of these taxa, our studies herein analysed two different characters (spore and stomata size and morphology) through scanning electronic microscopy, we studies the spore germination and new descriptions and an identification key were done to the accepted species to help clearing the taxonomic delimitation of the two taxa of Lygodium and their putative hybrid. According to our results, the species herein classified as L. ×fayae on the Brazilian south-east region is not a hybrid, but a morphological variation of Lygodium venustum. Additional studies on morphometric analysis with the measure... (Complete abstract click electronic access below) / Mestre

Introduction : L'hybridation in situ en fluorescence (FISH) est souvent utilisée pour l'étude des populations microbiennes hétérogènes. Toutefois, cette méthode n'est pas adaptée pour détecter des bactéries sous forme de spores, vue leur grande résistance aux traitements de perméabilisation conventionnels. Cependant, les bactéries formant des spores ont un rôle écologique, économique et médical important. Le but de cette étude est de développer des protocoles de perméabilisation rapides afin de détecter des spores de Bacillus par FISH. Méthodes : Un protocole pour les spores de B. megaterium a été adapté et optimisé pour les modèles choisis (B. cereus, B. atrophaeus et B. megaterium). Des sondes universelles et spécifiques ont été utilisées lors de l'hybridation. L'effet du traitement de perméabilisation a été évalué à Laide de la microscopie électronique à transmission et à balayage. Par la suite, les protocoles ont été adaptés pour permettre l'entrée de grosses molécules (comme la streptavidine) afin de permettre l'utilisation de méthode d'amplification de signal. Résultats : Avec les protocoles développés, les spores de Bacillus ont été détectées avec des sondes par FISH. La microscopie électronique à balayage a permis d'observer les différences de surface entre les spores traitées et non traitées. Des spores de Bacillus ont été détectées avec les protocoles adaptés pour la streptavidine. Conclusion : Des protocoles efficaces ont été développés pour détecter rapidement des spores de Bacillus par FISH. En utilisant cette technique, il est possible de détecter des spores de Bacillus en moins d'une heure.

Many endospore-forming bacteria cause diseases such as anthrax and food poisoning. Spores however also contribute to various agricultural and industrial processes. Spores possess extreme resistance properties, notably to chemical, et and dry heat, desiccation, and UV damage. For pathogenic spore formers, this poses an issue as spores are resistant to most decontamination methods currently in use. This work focuses on characterizing proteins thought to contribute to spore stability and efficient spore germination. Understanding how spores can remain stable for long periods of dormancy and against various insults and rapidly initiate germination could allow for the development of techniques that induce germination early and rapidly, promoting inexpensive decontamination. Physiological studies found that a family of spore-associated lipoproteins is needed for efficient spore germination and influences membrane fluidity in dormant spores. All the members of the lipoprotein family serve the same function, as each can fulfill the role of another. In vivo cross-linking was used to characterize protein-protein interactions found on the inner spore membrane. Glutaraldehyde crosslinking revealed that the four lipoproteins appear to interact. Bacterial two-hybrid analysis on individual protein domains further suggests the lipoproteins seem to interact through their predicted ring-building motif within their otherwise uncharacterized domains. Additionally, the absence of the spore lytic enzyme SleB seems to alter the crosslinking pattern of the lipoproteins, suggesting either it's interacting or helping facilitate lipoprotein interactions. Fluorescence microscopy reveals an unequal spatial distribution of the lipoproteins on the spore membrane, which seems to be supported by preliminary super-resolution microscopy studies. Further work aiming to characterize the entire inner spore membrane interactome is currently being conducted. The presented research used many methods and built many collaborations with the goal of providing insight to spore dormancy and efficient spore germination with an additional goal of understanding inner spore membrane protein behavior and how it leads to the highly resistant properties native to bacterial endospores. / Doctor of Philosophy / Certain species of Gram-positive bacteria can form a dormant cell called a bacterial endospore. Endospores, or spores, are highly resistant to insults such as noxious chemicals, wet and dry heat, and UV irradiation. These resistance properties make spores immune to standard sanitation methods, and result from various anatomical structures innate to the spore. In medicine, this poses a problem as spore forming bacteria can be causative agents of diseases such as food poisoning, anthrax, infant botulism, hospital-acquired diarrhea, and others. In agriculture and other industries, spore forming bacteria can be used as insecticides or fungicides, and have a long shelf life, making them ideal for long term storage. Research in the following document aims to answer questions relating to how spores transition from dormant spore to a typical cell. Understanding of these processes can inform novel decontamination techniques, better more stable spore-based products, and subversion of disease, depending on which process/ structure in the spore is altered.

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