Diss. (sammanfattning) Umeå : Univ., 2005. / Härtill 4 uppsatser.

Thesis supervisor: Bradley D. Jones. Includes bibliographic references (p. 136-151).

Francisella tularensis, a Gram-negative coccobacillus, is an extremely virulent intracellular pathogen. Infection of humans with this pathogen results in tularemia, a life-threatening disease. An approximately 35 kb region found in the F. tularensis genome exhibits many features of a pathogenicity island. This region has a lower G+C content than the average G+C content of the F. tularensis genome, and is surrounded by transposable elements. Results from both a previous study and our present study demonstrated that at least four genes located on the Francisella pathogenicity island (FPI) are required for virulence. This represents the first description of a pathogenicity island in F. tularensis. The FPI-encoded proteins, however, have no significant similarities to any known bacterial proteins. Therefore, we believe that the FPI genes may encode a cluster of novel virulence factors, although the mechanism and their characteristics remain to be determined.

Tularemia is a one of the most complex zoonotic diseases. Francisella tularensis McCoy and Chapin, the causative agent of tularemia is considered endemic in Texas, but outbreaks are rare and there are few human cases each year. Tularemia is listed as a Category A biological weapon and air samples are taken daily in select major metropolitan areas, including Houston, to monitor for its presence. I determined the potential risk for tularemia introduction and spread in southeast Texas through field surveillance for the pathogen and its major arthropod vector in the region, Amblyomma americanum (L.); completion of a habitat capability map for A. americanum, based on landscape analysis of the study area; and potential movement and long-term establishment of tularemia through development of a spatially explicit, agent-based, simulation model. Field and laboratory investigations resulted in the identification of two samples positive for F. tularensis. A feral cat tested positive for Type B tularemia using a new aptamer-based assay, and one sample returned positive in Amblyomma maculatum by polymerase chain reaction. This work sheds light on a complex host-pathogen-vector interaction in the rural to urban interface and establishes a framework for future tularemia field work and pathogen modeling in the rural to urban interface.

Tularemia is a one of the most complex zoonotic diseases. Francisella tularensis McCoy and Chapin, the causative agent of tularemia is considered endemic in Texas, but outbreaks are rare and there are few human cases each year. Tularemia is listed as a Category A biological weapon and air samples are taken daily in select major metropolitan areas, including Houston, to monitor for its presence. I determined the potential risk for tularemia introduction and spread in southeast Texas through field surveillance for the pathogen and its major arthropod vector in the region, Amblyomma americanum (L.); completion of a habitat capability map for A. americanum, based on landscape analysis of the study area; and potential movement and long-term establishment of tularemia through development of a spatially explicit, agent-based, simulation model. Field and laboratory investigations resulted in the identification of two samples positive for F. tularensis. A feral cat tested positive for Type B tularemia using a new aptamer-based assay, and one sample returned positive in Amblyomma maculatum by polymerase chain reaction. This work sheds light on a complex host-pathogen-vector interaction in the rural to urban interface and establishes a framework for future tularemia field work and pathogen modeling in the rural to urban interface.

Thesis (Ph.D.)--University of Missouri-Columbia, 2007. / The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Title from title screen of research.pdf file (viewed on March 23, 2009) Vita. Includes bibliographical references.

Thesis (Ph. D.)--Virginia Commonwealth University, 2009. / Prepared for: Dept. of Microbiology & Immunology. Title from resource description page. Includes bibliographical references. Unavailable until August 5, 2014.

Th.--Méd.--Nancy 1, 1984. N°: 210.

Master of Science / Food Science Institute / Randall K. Phebus / Francisella tularensis is a Gram-negative bacterium that can cause tularemia in humans. The disease can be acquired through several routes, one of which is the ingestion of contaminated food and water. The pathogen has the potential to be used as a biological weapon, and its intentional introduction in the food supply is a possible way to cause public harm. The purpose of this study was to evaluate the growth and/or survival of F. tularensis in food matrices under various storage conditions. Cystine Heart Agar enriched with 2% hemoglobin and supplemented with antibiotics was used to aid the enumeration of the target organism by suppressing the levels of indigenous microflora in foods. During the first portion of the study, the growth/survival of highly virulent F. tularensis subsp. tularensis SCHU S4 strain was investigated in prepared bagged iceberg lettuce stored at two temperatures. F. tularensis counts were significantly different (p≤0.05) among temperature levels for days 1-3, but not day 4. After the first 24 h of incubation, the mean estimates of F. tularensis counts were 1.00 log cfu g⁻¹ lower in lettuce stored at 23±1°C compared to lettuce stored at 6±1°C. Lower recovery rates at higher temperatures are likely due to the more rapid proliferation of naturally present bacteria which can inhibit the growth of F. tularensis. After 48 and 72 h, differences in F. tularensis counts between temperatures were 0.55 log cfu g⁻¹ and 0.3 log cfu g⁻¹, respectively. For the second portion of the experiment, the ability of F. tularensis to grow or survive was evaluated in nine food matrices with variable compositional content (red delicious apples, green bell pepper, shredded iceberg lettuce, strawberries, whole liquid eggs, boneless ham steak, beef hot dogs, 80-20 ground beef, and 2% UHT milk) at 4, 21, and 37°C. F. tularensis grew well in pasteurized whole liquid eggs stored at 21 and 37°C. With the exception of ham, liquid whole eggs, and bell pepper, the pathogen was detected infrequently throughout the duration of the study. Very low recovery rates were obtained for shredded lettuce, hot dogs, and ground beef.

Francisella tularensis, agent étiologique de la tularémie, classée parmi les bactéries pathogènes les plus infectieuses pour l’homme, possède une palette d’enzymes lui permettant de faire face au stress oxydatif, notamment à l’intérieur des macrophages, sa principale cible cellulaire. Nous nous sommes consacrés à l’étude d’un nouveau locus potentiellement impliqué dans la réponse au stress oxydatif chez F. tularensis LVS : le locus FTL_0200-FTL_0209. L’analyse in silico de ce locus révèle une homologie de la partie proximale avec le locus batL impliqué dans l’aérotolérance chez Bacteroides fragilis. D'une part, le premier gène du locus, FTL_0200, détermine une protéine chaperon putative ATPase AAA+ de la famille MoxR. D'autre part, les gènes FTL_0201 à FTL_0205 présentent des motifs protéiques de type facteur von Willebrand de type A (VWA) et tétratricopeptide (TPR), potentiellement impliqués dans la liaison d’ions métalliques et les interactions protéineprotéine. Or, les gènes déterminant des ATPases AAA+ de type MoxR sont très souvent retrouvés dans des loci codant pour des prot ines à domaines VWA et TPR. Enfin, les données expérimentales disponibles sur ce type de loci suggèrent que les protéines chaperons MoxR s’associeraient avec des protéines à motifs VWA et TPR pour former des complexes impliqués dans l’insertion de cofacteurs métalliques sur leurs cibles dédiées (enzymes ou autres). Ces observations nous ont conduit à étudier le rôle du gène FTL_0200 dans la virulence de F. tularensis, et sa participation à la réponse au stress. L'étude du mutant de délétion LVS!FTL_0200 démontre que le gène FTL_0200 joue un rôle majeur dans la survie intramacrophagique, ainsi que dans la virulence de F. tularensis dans le modèle murin; elle confirme également sa participation à la résistance au stress. Une analyse transcriptionnelle du locus nous a permis d’identifier le site d’initiation de sa transcription ; les régions -10 et -35 correspondent à un site de fixation pour le facteur alternatif "32, intervenant en conditions de stress. Des expériences d’étiquetage moléculaire (TAPtag) sur les protéines FTL_0200, FTL_0201 et FTL_0205 codées par le locus, ont été réalisées afin d’identifier leurs partenaires d’interaction, et démontrer l’association des protéines produites par le locus en un complexe multimérique. L’analyse des résultats obtenus par spectrométrie de masse montre une interaction de FTL_0200 avec FTL_0201, une interaction de FTL_0201 avec FTL_0205 et une interaction de FTL_0205 avec FTL_0207, suggérant l’association de ces protéines en un complexe multimérique. De plus, les trois protéines, plus particulièrement FTL_0205, semblent interagir avec des enzymes proches ou internes au cycle de Krebs : l’-cétoglutarate déshydrogénase (KGDH), la pyruvate déshydrogénase (PDH) et l’acétyl-CoA carboxylase (ACC). Une expérience de double hybride bactérien confirme partiellement ces résultats. La protéine FTL_0205, présentant beaucoup plus d’interactions que les deux autres protéines, le gène correspondant a été muté. Il présente une sensibilité très accrue au stress oxydatif ainsi qu’un défaut de multiplication dans la lignée de macrophages murins primaires issus de moelle osseuse, et une atténuation de virulence dans les souris. L’activité des trois enzymes métaboliques découvertes dans les expériences de TAP, a été mesurée chez les mutants FTL_0200 et FTL_0205, et comparée à celle de la souche sauvage. La KGDH et la PDH présentent une activité significativement diminuée chez les deux mutants, tandis que l’activité de l’ACC reste inchangée. Les enzymes du cycle de Krebs participent à la synthèse de certains acides aminés. Une expérience d’infection cellulaire, avec ajout de casaminoacides, chez le mutant FTL_0205 montre une restauration de sa virulence au niveau de celle de la souche sauvage. / Pas de résumé en anglais