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Isolierung und Charakterisierung eines phagenähnlichen Bacteriocins und eines virulenten Phagen und deren therapeutische Einsatzmöglichkeiten gegen Yersinia enterocolitica-InfektionenKaspar, Heike Maria 17 December 2004 (has links) (PDF)
Durch die wachsende Anzahl von multiresistenten Bakterien, die auch durch den Mißbrauch von Antibiotika als Masthilfsmittel in der Tierzucht entstanden sind, erlangen alternative Methoden zur Bekämpfung bakterieller Infektionen ihre Bedeutung zurück. Diese Arbeit befaßt sich mit zwei Substanzen, um Infektionen mit Yersinia (Y.) enterocolitica einzudämmen. Es wurde in dieser Arbeit ein Bacteriocin aus Y. enterocolitica isoliert und charakterisiert. Das Reinigungschema folgte den Strategien der Phagenaufreinigung, angeschlossen wurde zur Überprüfung der Reinheit ein Gelfiltrationsschritt. Die Eigenschaften des gereinigten Enterocoliticins wurden in vitro und in vivo getestet. Im Zellkulturversuch zeigte sich das Enterocoliticin in der Abtötung von an eukaryonte Zellen adhärierten Bakterien als sehr wirksam, in eukaryonte Zellen eingewanderte Bakterien wurden hingegen nicht abgetötet. Aufgrund dieser vielversprechenden Ergebnisse wurde der Therapieansatz im Mausmodell angewendet. Das Mausmodell ist für Y. enterocolitica ein bereits erprobtes Modell. Die Tiere wurden oral infiziert, um den natürlichen Infektionsweg nachzustellen, das Enterocoliticin wurde ebenfalls oral verabreicht. Die Infektion wurde durch die Enterocoliticingabe nur unwesentlich beeinflußt, auch gelang der Nachweis des Enterocoliticins weder im Gastrointestinaltrakt noch in den Faeces. Die Therapie der infizierten Mäuse gelang auf diese Weise nicht. Weiterhin wurde ein Yersinia-Phage aus Schweinegülle isoliert, gereinigt und charakterisiert. Es handelt sich um einen T4-ähnlichen, virulenten Phagen mit einer Genomgröße von ca. 50 kbp und einem weiten Wirtsspektrum in Yersinia, das sogar speziesübergreifend ist. Da der Phage bei 37°C die Wirtszelle lysiert und durch seine hohe Wirksamkeit in vitro erschien der Phage von seinen Eigenschaften her zur Phagentherapie als geeignet. Es wurden analog zum Enterocoliticin-Tierexperiment Mäuse mit Y. enterocolitica oral infiziert, diesen Tieren wurde der Phage auf unterschiedlichen Wegen und zu unterschiedlichen Zeitpunkten appliziert. Die Tiere zeigten bei der parenteralen Gabe keinerlei Unverträglichkeitserscheinungen, bei der oralen Gabe wurde der Magensaft zuvor abgepuffert. Der Therapieerfolg im Vergleich zur Kontrollgruppe war wenig vielversprechend, es zeigten sich sehr ähnliche Infektionsverläufe in Kontroll-und Therapiegruppe. Diese beiden untersuchten Alternativwege zur Behandlung von Yersiniosen erwiesen sich bei den angewendeten Methoden bislang als nicht erfolgreich, dennoch muß auf diesem Gebiet weitergeforscht werden, wie erfolgreiche Therapieansätze aus anderen Tiermodellen zeigen. Es wirken nur wenige Phagen im Organismus als Therapeutikum, dennoch müssen diese Untersuchungen unternommen werden, um im Organismus wirksame, antibakterielle Substanzen zu finden und um einen Alternativweg zur Antibiotikumtherapie zu entwickeln. / The increasing number of multi-resistant bacteria, which resulted also from the abuse of antibiotics as mast additives in animal breeding, alternative methods regain importance for the combat at bacterial infections back. In this work two substances were investigated to restrict infections with Yersinia (Y.) enterocolitica. In this study a bacteriocin from Y. enterocolitica, designated enterocoliticin, was isolated and characterized. The purification strategy followed protocols of phage isolation. The purified preparations were examined by final gel filtration step. The properties of the purifed enterocoliticin were tested in vitro and in vivo. In a cell culture assay enterocoliticin was able to kill bacteria adherent to eukaryontic cells very effectively, however, bacteria invaded into eukaryotic cells were not affected. Due to these results enterocoliticin was applied in a mouse-infection-model in a therapeutic attempt. The mouse infection model is a well established system for infections with Y. enterocolitica. The animals were orally infected with Yersinia, and the enterocoliticin was orally applied, too. The infection was only insignificantly influenced by enterocoliticin. In addition of enterocoliticin was not detected succeeded in the gastro-intestinal-tract or in the faeces. The therapy of the infected mice did not succeed in this way. Furthermore, a Yersinia phage from pig manure was isolated and characterized. It is a T4 phage like virulent phage, containing a genom of approx. 50 kbp and posessing a wide host-range in Yersinia. Because of lytic properties of the phage at 37°C and his high effectiveness in vitro the phage appeared to be for phage therapy experiments. Similarly to the enterocoliticin experiment mice were infected with Y. enterocolitica orally, these animals were treated with the phage on different application routes and different time points. The animals did not show any incompatibilities upon parenteral gift. Before oral administration of the phage the gastric juice was buffered. Therapy outcome in comparison to the control group was little promising, it revealed a very similar infection process in control group and therapy group. These two investigated alternative ways for the control of Yersinia infections did not prove successful with the applied methods, however, further research must be carried out as successful therapeutical experiments from other animal models showed. It has to be considered that not all phages are appropriate as therapeutical agent however, more studies must be conducted to find more appropriate substances, which may work as effective antibacterial substances to develop alternative ways to antibiotic therapy.
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Multiple twists in the molecular tales of YopD and LcrH in type III secretion by Yersinia pseudotuberculosis /Edqvist, Petra J., January 2007 (has links)
Diss. (sammanfattning) Umeå : Umeå universitet, 2007. / Härtill 5 uppsatser.
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Microbial biofilm attachment to Caenorhabditis elegansDrace, Kevin. January 2008 (has links) (PDF)
Thesis (Ph. D.)--University of Alabama at Birmingham, 2008. / Title from first page of PDF file (viewed June 6, 2008). Includes bibliographical references.
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Infecção por Yersinia pestis na Bahia: controle efetivo ou silêncio epidemiológico?Saavedra, Ramon da Costa January 2010 (has links)
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Previous issue date: 2010 / Apesar de a peste encontrar-se silente em todo o território brasileiro, seu agente etiológico, a bactéria Yersinia Pestis, permanece firmemente arraigada em seus focos naturais. Desta forma, e tendo em vista a existência de fatores condicionantes, não se pode desconsiderar que a doença, apesar de controlada, continua oferecendo riscos à população. A ocorrência de sorologia positiva para peste em carnívoros domésticos de algumas regiões pestígenas da Bahia nos últimos anos implica na necessidade de uma avaliação mais criteriosa, no intuito de verificar se ainda existe circulação do bacilo pestoso nessas áreas. Analisou-se, neste estudo, a presença de infecção por Y. pestis através do inquérito de soroprevalência em humanos, cães e roedores; e detecção da bactéria em roedores e pool de pulgas. A partir da aplicação de um questionário estruturado avaliou-se a associação existente entre fatores ambientais, sócio-econômicos e biológicos e a soroprevalência da infecção em humanos. Os 630 soros examinados (88 de humanos, 480 de cães, 62 de roedores) apresentaram-se não-reagentes para peste e as análises bacteriológicas realizadas em 14 roedores e 2 pool de pulgas não identificaram a bactéria. No entanto, tais resultados não configuram erradicação da doença no Estado. A natureza cíclica da peste indica que ela pode passar por longos períodos de silêncio e depois ressurgir acometendo um grande número de pessoas. Portanto, a manutenção de uma vigilância ativa e permanente se faz necessária para a detecção precoce da doença e desenvolvimento oportuno das medidas de controle pertinentes. / Salvador
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Specificity of the Yersinia Pestis biotype orientalis in the natural history of plague / Spécificité de Yersina Pestis orientalis biotype dans l'histoire naturelle de pesteAyyadurai, Saravanan 02 July 2010 (has links)
Yesinia pestis est l'agent de la peste, maladie infectieuse spontanément mortelle, et une bactérie classée parmi les agents de bioterrorisme de groupe A [http://www.bt.cd.gov/agent/plague]. Les cas sporadiques ont été rapportés dans plusieurs pays d'Asie, d'Afrique, et d'Amérique et la peste reste endémique en Afrique (République Démocratique du Congo; Madagascar) qui déclare le plus grand nombre de cas annuels. La majorité de cas de peste chez les humains et les animaux sauvages se manifeste dans les régions délimitées géographiquement et appelées communément les foyers de la peste. Les mécanismes de la résistance de la peste dans le sol des foyers reste de nos jours un sujet de recherche alors que la peste est maintenant considérée comme une maladie re-émergente. Au cours de notre travail, nous avons développé un outil pour l'identification de Y. pestis par spectrométrie de masse MALDI-TOF MS. Cette méthode s'est avérée très simple et efficace pour l'identification au niveau des espèces, et constitue une méthode de première ligne d'identification. Nous avons ensuite montré que Y. pestis survivait et maintenait sa virulence pendant au moins neuf mois dans le sol stérilisé par la vapeur et humidifié, dépourvu d'éléments nutritifs ajoutés et d'invertébrés du sol. Afin de contribuer à l'étude de l'épidémiologie de la peste, nous avons démontré que seul le biovar Oriantalis est transmis dans un modèle animal par les poux d'homme (Pediculus humanus), les biovars Antiqua et Medievalis de Y. pestis n'étant pas transmissibles par les poux de corps. Le mécanisme impliqué dans la transmission de la peste par les poux de corps reste inconnu, ce qui voudrait dire que le mécanisme de l'adaptation de Y. pestis Orientalis à des nouveaux vecteurs qui sont corrélés aux circonstances de l'épidémie mortelle provoquée par la peste bubonique, reste aussi inconnu. Au cours d'un dernier travail, nous avons étudié des nouveaux composés pour la prophylaxie de la peste. Notamment, nous avons évalué le potentiel du lovastatine dans la prévention de la mortalité pendant la peste. Il a été démontré sur un modèle d'expérimentation avec les souris que la lovastatine réduisait considérablement le taux de mortalité associée à la peste. Toutes les données que nous avons rapportées dans ce rapport de thèse sont destinées à mieux comprendre le cycle épidémiologique de la peste. / Yersinia pestis is the agent of deadly plague and a bacterium listed in the group A of potential bioterrorism agents [http://www.bt.cdc.gov/agent/plague/]. Sporadic cases are reported in several countries in Asia, Africa and America. Majority of human plague cases and enzootic animals occur in the geographical areas of so-called plague foci. The mechanisms sustaining geographical foci of plague remain poorly understood and plague been classified as a currently re-emerging disease. As first step, we established new front line tool for Y. pestis identification by using MALDI-TOF MS. This method was demonstrated to be simple and effective for Y. pestis identification at species level. Second step, we demonstrated that Y. pestis survived fully virulent for at least 9 months in a steam sterilized, humidified soil devoid of any nutritional supplements or any soil invertebrates. In third step we successfully demonstrated that the human louse (Pediculus humanus) as vector of plague and the body lice transmission of plague was restricted to Orientalis biovar; Antiqua and Medievalis biovars of Y. pestis were not able to transmit by body lice. This result shows that a un- explained mechanism is involved in the body lice transmission of plague and Y. pestis Orientalis adaptation to newly described vectors which effectively correlates the mass death caused by bubonic plague in Black Death individuals. Finally we conclude our study by exploring new compounds for the plague prophylaxis. The potential role of lovastatin in the prevention of mortality during plague was assessed. Lovastatin could significantly reduce the mortality associated with plague in an experimental mouse model. All These data herein we reported in our study may help to better understanding the epidemiology of plague.
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Isolierung und Charakterisierung eines phagenähnlichen Bacteriocins und eines virulenten Phagen und deren therapeutische Einsatzmöglichkeiten gegen Yersinia enterocolitica-InfektionenKaspar, Heike Maria 10 November 2003 (has links)
Durch die wachsende Anzahl von multiresistenten Bakterien, die auch durch den Mißbrauch von Antibiotika als Masthilfsmittel in der Tierzucht entstanden sind, erlangen alternative Methoden zur Bekämpfung bakterieller Infektionen ihre Bedeutung zurück. Diese Arbeit befaßt sich mit zwei Substanzen, um Infektionen mit Yersinia (Y.) enterocolitica einzudämmen. Es wurde in dieser Arbeit ein Bacteriocin aus Y. enterocolitica isoliert und charakterisiert. Das Reinigungschema folgte den Strategien der Phagenaufreinigung, angeschlossen wurde zur Überprüfung der Reinheit ein Gelfiltrationsschritt. Die Eigenschaften des gereinigten Enterocoliticins wurden in vitro und in vivo getestet. Im Zellkulturversuch zeigte sich das Enterocoliticin in der Abtötung von an eukaryonte Zellen adhärierten Bakterien als sehr wirksam, in eukaryonte Zellen eingewanderte Bakterien wurden hingegen nicht abgetötet. Aufgrund dieser vielversprechenden Ergebnisse wurde der Therapieansatz im Mausmodell angewendet. Das Mausmodell ist für Y. enterocolitica ein bereits erprobtes Modell. Die Tiere wurden oral infiziert, um den natürlichen Infektionsweg nachzustellen, das Enterocoliticin wurde ebenfalls oral verabreicht. Die Infektion wurde durch die Enterocoliticingabe nur unwesentlich beeinflußt, auch gelang der Nachweis des Enterocoliticins weder im Gastrointestinaltrakt noch in den Faeces. Die Therapie der infizierten Mäuse gelang auf diese Weise nicht. Weiterhin wurde ein Yersinia-Phage aus Schweinegülle isoliert, gereinigt und charakterisiert. Es handelt sich um einen T4-ähnlichen, virulenten Phagen mit einer Genomgröße von ca. 50 kbp und einem weiten Wirtsspektrum in Yersinia, das sogar speziesübergreifend ist. Da der Phage bei 37°C die Wirtszelle lysiert und durch seine hohe Wirksamkeit in vitro erschien der Phage von seinen Eigenschaften her zur Phagentherapie als geeignet. Es wurden analog zum Enterocoliticin-Tierexperiment Mäuse mit Y. enterocolitica oral infiziert, diesen Tieren wurde der Phage auf unterschiedlichen Wegen und zu unterschiedlichen Zeitpunkten appliziert. Die Tiere zeigten bei der parenteralen Gabe keinerlei Unverträglichkeitserscheinungen, bei der oralen Gabe wurde der Magensaft zuvor abgepuffert. Der Therapieerfolg im Vergleich zur Kontrollgruppe war wenig vielversprechend, es zeigten sich sehr ähnliche Infektionsverläufe in Kontroll-und Therapiegruppe. Diese beiden untersuchten Alternativwege zur Behandlung von Yersiniosen erwiesen sich bei den angewendeten Methoden bislang als nicht erfolgreich, dennoch muß auf diesem Gebiet weitergeforscht werden, wie erfolgreiche Therapieansätze aus anderen Tiermodellen zeigen. Es wirken nur wenige Phagen im Organismus als Therapeutikum, dennoch müssen diese Untersuchungen unternommen werden, um im Organismus wirksame, antibakterielle Substanzen zu finden und um einen Alternativweg zur Antibiotikumtherapie zu entwickeln. / The increasing number of multi-resistant bacteria, which resulted also from the abuse of antibiotics as mast additives in animal breeding, alternative methods regain importance for the combat at bacterial infections back. In this work two substances were investigated to restrict infections with Yersinia (Y.) enterocolitica. In this study a bacteriocin from Y. enterocolitica, designated enterocoliticin, was isolated and characterized. The purification strategy followed protocols of phage isolation. The purified preparations were examined by final gel filtration step. The properties of the purifed enterocoliticin were tested in vitro and in vivo. In a cell culture assay enterocoliticin was able to kill bacteria adherent to eukaryontic cells very effectively, however, bacteria invaded into eukaryotic cells were not affected. Due to these results enterocoliticin was applied in a mouse-infection-model in a therapeutic attempt. The mouse infection model is a well established system for infections with Y. enterocolitica. The animals were orally infected with Yersinia, and the enterocoliticin was orally applied, too. The infection was only insignificantly influenced by enterocoliticin. In addition of enterocoliticin was not detected succeeded in the gastro-intestinal-tract or in the faeces. The therapy of the infected mice did not succeed in this way. Furthermore, a Yersinia phage from pig manure was isolated and characterized. It is a T4 phage like virulent phage, containing a genom of approx. 50 kbp and posessing a wide host-range in Yersinia. Because of lytic properties of the phage at 37°C and his high effectiveness in vitro the phage appeared to be for phage therapy experiments. Similarly to the enterocoliticin experiment mice were infected with Y. enterocolitica orally, these animals were treated with the phage on different application routes and different time points. The animals did not show any incompatibilities upon parenteral gift. Before oral administration of the phage the gastric juice was buffered. Therapy outcome in comparison to the control group was little promising, it revealed a very similar infection process in control group and therapy group. These two investigated alternative ways for the control of Yersinia infections did not prove successful with the applied methods, however, further research must be carried out as successful therapeutical experiments from other animal models showed. It has to be considered that not all phages are appropriate as therapeutical agent however, more studies must be conducted to find more appropriate substances, which may work as effective antibacterial substances to develop alternative ways to antibiotic therapy.
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Investigation of T3SS regulation inYersinia pseudotuerculosis using a secreted chimeric YopE1-86-Bla reporter systemMohammad, Alif January 2021 (has links)
No description available.
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Avaliação dos locos CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) em estudos epidemiológicos de cepas de Yersinia pestis / Evaluation of CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) loci in epidmiologic study of strains of Yersinia pestisLins, Rosanny Holanda Freitas Benevides January 2011 (has links)
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Previous issue date: 2011 / Fundação Oswaldo Cruz. Centro de Pesquisas Aggeu Magalhães. Recife, PE, Brasil / Yersinia pestis é o agente causador da peste, doença primária de roedores, transmitida por pulgas infectadas, podendo infectar o homem e outros mamíferos. A maioria dos estudos de genotipagem realizada em cepas brasileiras de Y. pestis demonstrou baixo poder discriminatório, revelando padrões genotípicos semelhantes para cepas com diferentes características epidemiológicas. A análise do clustered regularly interspaced short palindromic repeats (CRISPR) vem sendo utilizada para genotipagem e estudos filogenéticos em diferentes gêneros bacterianos, inclusive de Y. pestis. Neste estudo foi realizada a genotipagem de cepas de Y. pestis da coleção de cultura do Serviço Nacional de Referência em Peste do Centro de Pesquisas Aggeu Magalhães - CPqAM/FIOCRUZ, isoladas nos três focos de peste do Estado de Pernambuco, pela análise dos locos CRISPR. Para as amplificações das 51 amostras, foram utilizados primers dirigidos às sequências CRISPR descritas na literatura (YPa, YPb e YPc). Os amplicons obtidos, após PCR, foram sequenciados, a fim de analisar a estrutura de cada loco CRISPR. Dois locos se apresentaram polimórficos: YPa, com alelos de 150pb a 570pb e YPb, com alelos de 330pb a 331pb. O loco YPc se mostrou monomórfico com alelos de 208pb. Os padrões CRISPR obtidos para cepas de Y. pestis de focos de outros países foram os mesmos observados nas cepas brasileiras. Diferentes arranjos dos espaçadores (YPa, YPb e YPc) foram observados e possibilitou que as cepas fossem agrupadas em 12 perfis genotípicos (PG1-PG12). Dois perfis foram distribuídos nos três focos estudados: PG1 perfil mais frequente (38 cepas) e PG3 (seis cepas). Os demais perfis foram específicos de uma determinada região de isolamento: PG2 para o foco de Triunfo e PG4-PG7 para o foco de Araripe. Os perfis PG8 a PG12 representaram o restante das cepas de focos de outros países. As mesmas cepas foram analisadas, anteriormente, através de onze locos VNTR (MLVA), e foram agrupadas em 35 perfis genotípicos / A menor diversidade observada no CRISPR ocorre, provavelmente, devido à região estar envolvida na codificação gênica e com maior pressão seletiva, portanto, com menor risco de sofrer mutação decorrente de estocagem. A análise dos locos CRISPR, complementar ao uso do MLVA, será útil na identificação e rastreamento de novas cepas, contribuindo para o estabelecimento de medidas de controle adequadas nas áreas de foco para prevenção da peste e na investigação de novos casos que possam surgir no Brasil
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MOLECULAR AND GENETIC CHARACTERIZATION OF THE VIRULENCE PLASMID OF YERSINIA ENTEROCOLITICA.DUBEL, JACQUELINE ROBERTA. January 1983 (has links)
Purified DNA from a nalidixic acid resistant derivative of a virulent serotype 0:3 clinical isolate of Yersinia enterocolitica was subjected to transpositional mutagenesis in an effort to construct avirulent mutants. The resulting transpositional mutants, as well as the wild-type virulent strain and its isogenic derivative that had been cured of the virulence plasmid, were analyzed for plasmid DNA content. The plasmid DNA content of each strain was further characterized by restriction endonuclease digestion and the transposon insertion sites for the mutants were located. All of the strains were then tested for pathogenicity by the following assays: calcium dependence, colonization of the mouse gastrointestinal tract, HEp-2 cell adherence and invasion, HEp-2 cell monolayer detachment, autoagglutination, serum resistance, outer membrane protein production and production of V antigen. In addition, the hydrophobic properties of each strain were examined by a rapid polystyrene plate method and hydrophobic interaction chromatography. The results of the tests were compared to plasmid DNA analyses for each strain in an attempt to identify any plasmid-associated genes that are related to virulence. The wild-type strain was virulent, or positive, by all of the assays employed for evaluation of pathogenicity. In contrast, its isogenic derivative that had been cured of the virulence plasmid was negative, or avirulent, for the same assays with one exception. The avirulent plasmidless strain still retained the ability to adhere to and invade HEp-2 cells, supporting the belief that these properties are probably encoded by the bacterial chromosome. In addition, three transpositional mutants were constructed that were no longer calcium dependent, capable of detaching HEp-2 cell monolayers or able to produce three unique outer membrane proteins. Restriction endonuclease analysis confirmed the presence of the transposon on the Hind III "A" fragment of the virulence plasmid and located the region responsible for the lost virulence properties. The gene or set of genes identified were designated cal and the respective calcium independent mutants Cal⁻. Furthermore, the assays for hydrophobicity indicated that the virulence plasmid, specifically the cal gene(s), codes for hydrophobic properties on the surface of the bacterium. The study demonstrated that a virulence-associated region, cal, is located on the virulence plasmid of Y. entrocolitica and responsible for calcium dependence, HEp-2 cell monolayer detachment, the production of the three plasmid-specified outer membrane proteins and cell-surface hydrophobicity.
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The spatial population dynamics of house mice (Mus musculus domesticus) with reference to the potential transmission of zoonosesPocock, Michael James Orlando January 2001 (has links)
No description available.
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