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Dissertations / Theses on the topic 'Metabolism in Toxoplasma Gondii'

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Published: 6 September 2023

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Roohi, Aysha. "Toxoplasma gondii infection and the host cell metabolism." Thesis, University of Cambridge, 2013. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.648369.

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Ren, Bingjian. "Physiological importance of phospholipid biogenesis in Toxoplasma gondii." Doctoral thesis, Humboldt-Universität zu Berlin, 2019. http://dx.doi.org/10.18452/20721.

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Toxoplasma gondii ist ein obligater intrazellulärer Parasit, der bei Menschen und Nutztieren Toxoplasmose verursacht. Die Phospholipid-Biosynthese ist entscheidend für das erfolgreiche intrazelluläre Überleben und die Replikation des Parasiten, da sie eine wichtige Rolle bei der Biogenese von Membranorganellen, der Signal- transduktion und anderen zellulären Prozessen spielt. Hier untersuchten wir die physiologische Bedeutung von zwei Synthesewegen, die für PtdEtn und PtdIns verantwortlich sind. Wir zeigen das Vorhandensein einer neuartigen PtdIns-Synthase (PIS) in T. gondii, die als TgPIS bezeichnet wird und ein funktionelles Enzym mit einem katalytisch wichtigen CDP-Alkohol- Phosphotransferase-Motiv codiert, das sich ausschließlich im Golgi-Apparat befindet. Der Parasit importiert Myoinosit aus dem Milieu und verwendet es zusammen mit de novo synthetisiertem CDP- Diacylglycerin, um PtdIns zu produzieren. Eine durch Auxin induzierbare bedingte Unterdrückung von TgPIS schaltet den Lysezyklus des Parasiten aufgrund von Defekten in der Replikation, Motilität und Austritt in Säugetierzellen aus. Das Lipidom-Profiling der PIS-Mutante zeigt eine selektive Reduktion bestimmter PtdIns- und PtdThr-Spezies, wohingegen ausgewählte PtdGro-, PtdSer- und BMP-Spezies erhöht sind, was auf eine enge Interregulation und Homöostase von anionischen Phospholipiden zur Aufrechterhaltung der Membranintegrität hindeutet. Zusätzlich identifizierten wir eine Ethanolamin-Cytidyltransferase (TgECT), das geschwindigkeitsbestimmende Enzym des Kennedy-Signalwegs, das im Cytosol lokalisiert ist. Das Enzym ist eindeutig für den Lysezyklus essentiell, da seine genetische Ablation nicht durchführbar ist und der durch Auxin meditierte bedingte Abbau des Proteins das Parasitenwachstum in Plaqueassays stark beeinträchtigt. Die Lipidomanalyse der Mutante identifizierte eine wichtige Rolle bei der Biogenese ausgewählter Arten von PtdEtn, PtdSer und PtdThr. Darüber hinaus haben wir festgestellt, dass TgECT für die Erzeugung von Ethanolamin-Phosphor-Ceramid (EPC) erforderlich ist, einem seltenen Sphingolipid, das nur eine begrenzte Anzahl von Organismen enthalten. Toxoplasma gondii is an obligate intracellular parasite that causes Toxoplasmosis in human and livestock. Phospholipid biosynthesis is crucial for the successful intracellular survival and replication of the parasites, as the phospholipids have important roles in the biogenesis of membrane organelles, signal transduction and other cellular processes. Here, we dissected the physiological importance of two pathways accounting for the synthesis of PtdEtn and PtdIns. We demonstrated the presence of a novel PtdIns synthase (PIS) in T.gondii termed TgPIS, expressing a functional enzyme with a catalytically vital CDP-alcohol phosphotransferase motif, which resides exclusively in the Golgi body. The parasite imports myo-inositol from milieu, and co-utilizes de novo-synthesized CDP-diacylglycerol to produce PtdIns. An auxin-inducible conditional repression of TgPIS abrogated the lytic cycle of the parasite in mammalian cells due to defects in the replication, motility and egress. Lipidomic profiling of the PIS mutant demonstrated selective reduction of certain PtdIns and PtdThr species, whereas selected PtdGro, PtdSer and BMP species were increased, which suggested a tight inter-regulation and homeostasis of anionic phospholipids to maintain the membrane integrity. In addition, we identified an ethanolamine cytidyltransferase (TgECT), the rate-limiting enzyme of Kennedy pathway, which is localized in the cytosol. The enzyme is clearly essential for the lytic cycle as its genetic ablation was not feasible, and auxin-meditated conditional knockdown severely impaired the parasite growth in plaque assays. Similarly, lipidomic analysis of the mutant identified an important role in the biogenesis of selected species of PtdEtn, PtdSer and PtdThr. Moreover, we discovered that TgECT is required for the generation of ethanolamine-phosphory ceramide (EPC), a rare sphingolipid present only a limited number of organisms.

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Nitzsche, Richard. "Genetic dissection of the central carbon metabolism in the intracellular parasite Toxoplasma gondii." Doctoral thesis, Humboldt-Universität zu Berlin, Lebenswissenschaftliche Fakultät, 2017. http://dx.doi.org/10.18452/17744.

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Toxoplasma gondii ist ein weit verbreiteter einzelliger Parasit, der fast alle warmblütigen Organismen infizieren kann. Asexuelle Fortpflanzung des Parasiten in seiner Wirtszelle wird durch aufeinanderfolgende lytische Zyklen erreicht, was die Bereitstellung einer signifikanten Menge an Energie und Biomasse erforderlich macht. Diese Arbeit zeigt, dass Glukose und Glutamin die beiden wichtigsten physiologischen Nährstoffe für die Synthese von Makromolekülen (ATP, Nukleinsäure, Proteine und Lipide) in T. gondii sind. Die Verfügbarkeit einer der beiden Kohlenstoffquellen reicht aus, um das Überleben des Parasiten sicherzustellen. Der Parasit kann durch Erhöhen des Flusses von Glutamin-abstammendem Kohlenstoff durch den TCA-Zyklus und durch gleichzeitige Aktivierung der Gluconeogenese, eine stetige Biogenese von ATP und Biomasse zur Wirtszellinvasion und Replikation gewährleisten, bzw. der genetischen Deletion des Glukosetransporters entgegenwirken. Der Wachstumsdefekt in der Glykolyse-Mutante wird durch eine kompromittierte Synthese von Lipiden verursacht, die durch Glutamin nicht ausgeglichen werden kann. Die Zugabe von exogenem Acetat kann diesen Wachstumsdefekt allerdings kompensieren. In dieser Arbeit konnten darüber hinaus zwei unterschiedliche Phosphoenolpyruvat-Carboxykinase (PEPCK) Enzyme im Parasiten identifiziert werden, von denen eines im Mitochondrium lokalisiert ist (TgPEPCKmt), während das andere Protein nicht in Tachyzoiten (TgPEPCKnet) exprimiert wird. Parasiten mit intakter Glykolyse können die Deletion von TgPEPCKnet, als auch die genetische Deletion von TgPEPCKmt tolerieren, was ihre Redundanz für das Überleben der Tachyzoiten zeigt. TgPEPCKnet kann auch in der Glykolyse-defizienten Mutante deletiert werden, während TgPEPCKmt für das Überleben des Parasiten in dieser Mutante essentiell ist. Dies zeigte sich durch ein konditionelles Knockdown von TgPEPCKmt, das zu einer Inhibierung des Wachstums des Parasiten führte. Toxoplasma gondii is a widespread protozoan parasite, infecting nearly all warm-blooded organisms. Asexual reproduction of the parasite within its host cells is achieved by consecutive lytic cycles, which necessitates biogenesis of significant energy and biomass. This work shows that glucose and glutamine are the two major physiologically important nutrients used for the synthesis of macromolecules (ATP, nucleic acid, proteins and lipids) in T. gondii, and either of them is sufficient to ensure the parasite survival. The parasite can counteract genetic ablation of its glucose transporter by increasing the flux of glutamine-derived carbon through the TCA cycle and by concurrently activating gluconeogenesis, which guarantee a continued biogenesis of ATP and biomass for host-cell invasion and parasite replication, respectively. Growth defect in the glycolysis-impaired mutant is caused by a compromised synthesis of lipids, which cannot be counterbalanced by glutamine, but can be restored by acetate. Consistently, supplementation of parasite cultures with exogenous acetate can amend the lytic cycle of the glucose transport mutant. Furthermore, this work revealed two discrete phosphoenolpyruvate carboxykinase (PEPCK) enzymes in the parasite, one of which resides in the mitochondrion (TgPEPCKmt), whereas the other protein is not expressed in tachyzoites (TgPEPCKnet). Parasites with an intact glycolysis can tolerate genetic deletions of TgPEPCKmt as well as of TgPEPCKnet, indicating their nonessential roles for the tachyzoite survival. TgPEPCKnet can also be ablated in glycolysis-deficient mutant, whereas TgPEPCKmt is refractory to deletion. In accord, the lytic cycle of a conditional mutant of TgPEPCKmt in the glycolysis-impaired strain was aborted upon induced repression of the mitochondrial isoform, demonstrating its essential role for the glucose-independent survival of tachyzoites.

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Sampels, Vera. "Plasticity of the phosphatidylcholine biogenesis in the obligate intracellular Parasite Toxoplasma gondii." Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2012. http://dx.doi.org/10.18452/16491.

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Der obligat intrazelluläre Parasit Toxoplasma gondii ist der Erreger der Toxoplasmose, und dient zugleich als wichtiger Modellorganismus für weitere Human- und Tierpathogene, wie z.B. Plasmodium oder Eimeria. Die Vermehrung von T. gondii erfordert eine effiziente Biosynthese von Phospholipiden für die Herstellung neuer Membranen, was durch die de novo Synthese durch den Parasiten, und/oder den Import von Lipiden aus der umgebenden Wirtszelle gewährleistet werden kann. Während der Parasit zahlreiche Möglichkeiten für Synthese oder Import von PtdEtn und PtdSer verwendet, scheint die Biosynthese des abundantesten Membranlipids PtdCho auschließlich über den CDP-Cholin Weg zu erfolgen. Dieser erstreckt sich in T. gondii über 3 zelluläre Kompartimente, mit einer cytosolischen Cholin-Kinase (TgCK), einer im Zellkern lokalisierenden Cholin-Cytidylyltransferase (TgCCT) und einer Cholin-Phosphotransferase (TgCPT) im ER. Anders als die substrat-spezifische Ethanolamin-Kinase (TgEK), kann TgCK neben Cholin außerdem Ethanolamin phosphorylieren. TgCK zeigt eine geringe Affinität zu Cholin (Km ~0.77 mM), während eine verkürzte TgCK (TgCKS), welcher eine als Signalpeptid vorhergesagte N-terminale Sequenz (20 Aminosäuren) fehlt, eine etwa 3-fach höhere Aktivität aufweist (Km ~0.26 mM). Während jedoch die Wildtyp-TgCK cytosolische Cluster in Toxoplasma bildet, zeigt die verkürzte TgCK eine gleichmäßigere cytosolische Lokalisierung. Wir schlussfolgern daraus, dass der hydrophobe N-Terminus nicht notwendig ist für eine funktionale TgCK, sondern eine strukturelle Funktion bei der Protein-Lokalisierung hat. Eine konitionelle Mutante, in welcher der TgCK Promoter gegen den Tetracyclin-regulierbaren Promoter pTetO7Sag4 ausgetauscht wurde (Deltatgcki), zeigt erstaunlicherweise normales Wachstum und PtdCho Biosynthese. Die TgCK Aktivität und die daraus resultierende PtdCho Synthese sind nur zu ~30% regulierbar. Unsere Ergebnisse deuten auf die Verwendung eines alternativen Startcodons bzw. Promoters hin, welcher zur Expression einer verkürzten (~53-kDa) aber vermutlich aktiven Cholin Kinase führt, wodurch der Verlust der TgCK (~70-kDa) kompensiert wird. Der konditionelle Knockout von TgCCT, dem regulatorischen Enzym des CDP-Cholin Wegs, hatte einen 50%igen Wachstumsdefekt zur Folge. Diese Studie zeigt eine erstaunliche Flexibilität des Parasiten bezüglich seiner Membranzusammensetzung, und bestätigt zugleich die Annahme, dass PtdCho nicht von der Wirtszelle importiert werden kann. Diese Anpassungsfähigkeit stellt einen möglichen Faktor dar, der es T. gondii erlaubt sich in einem breiten Spektrum von Wirten zu vermehren. Toxoplasma gondii is an obligate intracellular apicomplexan parasite that causes life-threatening disease in neonates and in immunocompromised people. Successful replication of Toxoplasma requires substantial membrane biogenesis, which must be satisfied irrespective of the host-cell milieu. Like in other eukaryotes, the two most abundant phospholipids in the T. gondii membrane are phosphatidylcholine (PtdCho) and phosphatidylethanolamine (PtdEtn). Bioinformatics and precursor labeling analyses confirm their synthesis via the CDP-choline and CDP-ethanolamine pathway, respectively. This work shows that the 3-step CDP-choline pathway, involving the activities of TgCK, TgCCT and TgCPT, localizes to the cytosol, nucleus and ER membrane, respectively. The initial reaction is catalyzed by a dual-specificity choline kinase (TgCK, ~70-kDa), capable of phosphorylating choline as well as ethanolamine. The purified full-length TgCK displayed a low affinity for choline (Km ~0.77 mM). TgCK harbors a unique N-terminal hydrophobic peptide that is required for the formation of enzyme oligomers in the parasite cytosol but not for activity. The displacement of the TgCK promoter in a conditional mutant of T. gondii (deltatgcki) attenuated the enzyme expression by ~80%. Unexpectedly, the ?tgcki mutant was not impaired in intracellular growth, and exhibited a normal PtdCho biogenesis. To recompense for the loss of full-length TgCK, the mutant appears to make use of an alternative promoter and/or start codon, resulting in the expression of a shorter but active TgCK isoform identified by the anti-TgCK antiserum, which correlated with its persistent choline kinase activity. Accordingly, the ?tgcki showed an expected incorporation of choline into PtdCho, and susceptibility to dimethylethanolamine (a choline analog). Interestingly, the conditional mutant displayed a regular growth in off state despite a 25% decline in PtdCho content, which suggests a compositional flexibility in T. gondii membranes and insignificant salvage of host-derived PtdCho. The two-step conditional mutagenesis of TgCCT, which caused a reduced growth rate to about 50%, further substantiated this finding. The enzymatic activity of TgCCT and its role in PtdCho synthesis remain to be proven, however. Taken together, the results demonstrate that the CDP-route is likely essential in T. gondii. The competitive inhibition of choline kinase to block the parasite replication appears a potential therapeutic application.The work also reveals a remarkably adaptable membrane biogenesis in T. gondii, which may underly the evolution of Toxoplasma as a promiscuous pathogen.

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Sundaram, Lalitha Sridevi. "Toxoplasma gondii-mediated host cell transcriptional changes lead to metabolic alterations akin to the Warburg effect." Thesis, University of Cambridge, 2017. https://www.repository.cam.ac.uk/handle/1810/267958.

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Toxoplasma gondii is an obligate intracellular parasite, that is able to infect any nucleated cell. An important global pathogen, T. gondii can cycle between primary and secondary hosts, thus enabling widespread penetrance. Within its intracellular niche – a membrane-bound parasitophorous vacuole – T. gondii is nevertheless able to subvert a variety of host cell processes to allow its continued survival and replication. This includes modulation of host signalling processes as well as the scavenging of nutrient macromolecules. In recent years, microRNAs have emerged as important regulators of cellular processes including inflammation, tumorigenesis and metabolism, as well as development. It has become increasingly clear that this species of non-coding RNA is of great importance in ‘fine tuning’ many cellular responses. I hypothesise in this work that host cell miRNAs may be yet another means by which T. gondii manipulates its host upon infection. Using high-throughput-sequencing, I examine host cell transcriptional responses to infection both at the mRNA and microRNA level, using two strains of T. gondii at a variety of Multiplicities of Infection over a time course of 43 hours. Through these transcriptional analyses I identify a number of dysregulated pathways common in tumorigenesis, and contemplate the hypothesis that T. gondii may be behaving as an ‘intracellular tumour’, subverting host cell metabolic processes to mimic a long-known feature of cancer metabolism – that of aerobic glycolysis (the Warburg effect) – in order to satisfy its own energetic and metabolic needs.

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Blume, Martin. "Characterization of the differential significance of sugar Import in the apicomplexan parasites Toxoplasma gondii and Plasmodium." Doctoral thesis, Humboldt-Universität zu Berlin, Mathematisch-Naturwissenschaftliche Fakultät I, 2011. http://dx.doi.org/10.18452/16396.

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Toxoplasma gondii und Plasmodium Spezies sind obligat intrazelluläre Parasiten, die Zucker zur Energiehomöostase als auch für die Synthese lebenswichtiger Makromoleküle verwenden. Die hier vorgestellten Daten zeigen, dass der Glukosetransporter von T. gondii, TgGT1, und die homologen Transporter von P. falciparum und P. berghei, PfHT1 und PbHT1, neben Glukose auch Mannose, Fructose und Galactose transportieren. Toxoplasma Tachyzoiten exprimieren neben TgGT1 noch einen weiteren putative Zuckertransporter (TgST2) an der Parasitenoberfläche. Beide Proteine sind nicht essentiell, wie durch ihre individuelle und gleichzeitige Gendeletion belegt wird. Die Deletion von TgGT1 bewirkt einen geringen Wachstumsdefekt. Die Mutante ?tggt1 zeigt keine Glukoseaufnahmeaktivität und folglich eine verminderte glukoseabhängige Motilität. In ?tggt1 Parasiten wird ein verstärkter Glutaminstoffwechsel nachgewiesen, der ausreichend ist dessen Motilität und Replikationsaktivität zu erhalten. Die ?tggt1 Mutante gewährt Einblick in die Anpassungsfähigkeit von T. gondii an unterschiedliche Wirtszellen. Im Gegensatz zu T. gondii benötigen erythrozytäre Plasmodien Glukose und der Transporter PfHT1 wird derzeit als drug-target eingestuft. Hier wird gezeigt, dass das PfHT1-Homolog, PbHT1, essentiell in Blutstadien des Nagerparasiten Plasmodium berghei ist, jedoch auch während des gesamten Lebenszyklus des Parasiten exprimiert wird. Ein PfHT1- und PbHT1-spezifischer Inhibitor (Compound 3361) kann die Entwicklung von P. berghei Leberstadien und Okineten stark hemmen. Um zukünftig PfHT1-Inhibitoren im Hochdurchsatzverfahren zu identifizieren und testen zu können, wurden auf Saccharomyces cerevisiae und P. berghei basierende Expressionssysteme für PfHT1 entwickelt. Abschließend stellt diese Arbeit die Unterschiedlichkeit des zentralen Kohlenstoffwechsels von Toxoplasma und Plasmodium Parasiten durch bisher unbekannter Aspekte heraus. Toxoplasma gondii and Plasmodium species are obligate intracellular pathogens that utilize host sugars for energy homeostasis and macro molecular synthesis. Here, we report that the T. gondii glucose transporter, TgGT1, and of its homologs of P. falciparum and P. berghei (PfHT1 and PbHT1) transport glucose, mannose, galactose and fructose. Besides TgGT1, Toxoplasma harbours one additional surface localized putative sugar transporter (TgST2). Surprisingly both Proteins are nonessential and only the deletion of TgGT1 inflicts a mild defect in the parasite replication. The ?tggt1 mutant is unable to import glucose and consequently displays an attenuated glucose-dependent motility, which is completely rescued by glutamine. ?tggt1 performs increased glutamine metabolism that is sufficient to sustain motility and replication. The ?tggt1 strain provides a model for further investigating its adaptation to disparate host cells. In contrast to T. gondii, erythrocytic stages of Plasmodium species critically depend on glucose uptake, and the PfHT1 transporter is considered as a drug target against human malaria. Here, we report that PbHT1 (a PfHT1 homolog) is also essential for blood stage development in the rodent malaria parasite P. berghei. PbHT1 is expressed throughout the life cycle. Moreover, a PfHT1- and PbHT1-specific sugar analogue, compound 3361, can inhibit the hepatic development and ookinete formation in P. berghei. These results signify that PbHT1 and exogenous glucose are also required during the ex-erythrocytic stages of P. berghei. To permit a high-throughput screening of selective PfHT1 inhibitors and their subsequent in vivo assessment, we have established a PfHT1-expressing Saccharomyces cerevisiae mutant and generated a PfHT1-dependent ?pbht1 of P. berghei strain. This thesis underscores various previously unknown aspects of sugar metabolism in Toxoplasma and Plasmodium, and unravel their metabolic differences.

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Molan, Aus. "Involvement of Toxoplasma gondii and associated inflammatory markers in the pathogenesis of type 2 diabetes mellitus." Thesis, Edith Cowan University, Research Online, Perth, Western Australia, 2020. https://ro.ecu.edu.au/theses/2338.

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Type 2 diabetes mellitus (T2DM) continues to be a major challenge for public health authorities worldwide due to its growing prevalence rates. This chronic disorder is characterised by persistent hyperglycaemia with disturbances in fat, carbohydrate, and protein metabolism resulting from abnormalities in insulin secretion, action, or both. While potential causes such as obesity, physical inactivity, and dietary patterns have been proposed to explain the growing epidemic, there may also be additional unidentified novel risk factors, such as subclinical inflammation caused by infectious agents, that contribute to this rising prevalence. The intracellular protozoan parasite Toxoplasma gondii has been identified as an understudied pathogen of interest. Infecting approximately one third of the world’s population, T. gondii is considered one of the most successful human parasites. However, while more than 25,000 original research articles, several books, and book chapters have been published on this parasite, there are still many aspects of its biology, natural life cycle, and epidemiology of which we know relatively little about, including the prevalence rate in the general Australian human population. An emerging field of research is beginning to investigate the potential of this pathogen to cause low-grade inflammation that may subsequently increase the risk and development of various metabolic conditions, particularly T2DM. The main objectives of the present study were to: 1) determine seroprevalence of T. gondii infection in a representative Australian human population [Study One] ; 2) determine the seroprevalence of T. gondii in subjects with T2DM compared with control subjects [Study One]; 3) identify additional risk factors for T. gondii infection [Study One]; 4) examine cat ownership as an alternative risk factor for T2DM [Study Two]; 5) determine if T. gondii infection participates in the inflammatory process leading to the development of T2DM [Study Three]; and, 6) determine whether an inflammatory 11-biomarker panel could discriminate between controls, subjects with T. gondii infection, and T2DM patients, independently of other diagnostic indicators [Study Three]. Our group undertook the first set of case-control studies of their kind in Australia by utilising sera samples and survey data collected by the Busselton Population Medical Research Institute during the 2005-2007 Busselton Health Survey. The cohort comprised of 150 participants with T2DM and 150 age- and gender- matched control subjects. Commercial in-vitro diagnostic enzyme-linked immunosorbent assays were used to measure the IgG and IgM seroprevalence rates for study one and high-sensitivity multiplex methods were used to simultaneously measure the 11 biomarkers in study three. From the 300 subjects tested, 64% had T. gondii IgG antibodies detected and 7.7% had IgM antibodies detected. IgG seropositivity increased rapidly in all age groups remaining at over 70% in the 65–74, 75–84, and >84 age groups (p <0.05). No significant difference was observed in the T. gondii IgG seroprevalence rates between the T2DM group (62%) and the healthy control group (66%) (OR: 0.84; p = 0.471). Toxoplasma gondii IgM antibodies were detected in 5% of the T2DM patients and in 10% of the control subjects (OR: 0.51; p = 0.135). There were no significant differences between male and female IgG seroprevalence rates for both the T2DM and the control groups (OR: 0.88 and 0.80; p = 0.723 and p = 0.507, respectively). The IgG seropositivity rate increased significantly in T2DM patients aged 45 to 84 years in comparison to those aged 18 to 44 years (p <0.05). This trend was not observed in the healthy non-T2DM subjects. No risk factors were associated with T. gondii seropositivity in both T2DM patients and healthy controls. IL-β, IL-4, IL-6, and IL-12(p40) were not being produced or were below the levels of detection in the majority of subjects. Serum levels of leptin, IL-4, IL-10 and PAI-1 (total) were positively associated with age in the control T. gondii seronegative group. In contrast, decreasing levels of leptin, IL-10, and PAI-1 (total) was observed in the control seropositive and T2DM seronegative groups. Age was also positively associated with IFN-γ levels in control subjects and negatively associated in the T2DM group. Increased weight, BMI, waist circumference, glucose, insulin, triglycerides, white cell count, leptin, and PAI-1 (total) results were significantly associated with T2DM (p <0.05), while no significant associations with T. gondii infection were identified (p >0.05). Strong positive associations between IL-6 and IL-4; and IL-1RA and IL-β were observed in T. gondii seropositive subjects regardless of which group they belonged to. Likewise, IL-10 showed a strong positive correlation with IL-1RA only in seronegative subjects. High-density lipoprotein displayed significant correlation (p <0.01) with: IL-6 (r = 0.506); IL-4 (r = 0.492); IL-1β (r = 0.456); IL-1RA (r = 0.507); and, IFN-γ (r = 0.456) exclusively in seropositive subjects belonging to the T2DM group. We conclude that T. gondii infection in Western Australia is highly prevalent. Our results also showed that there is no serological evidence of an association between T. gondii infection and T2DM in the studied subjects. Although some biomarkers displayed strong association with T. gondii and/or T2DM, the differences were not significant enough in terms of discriminatory power hence the 11-biomarker panel selected for the present study is not suited for use in clinical practice independently of other diagnostic indicators. In addition, no evidence of an inflammatory association between T. gondii infection and T2DM was found. Further research is warranted to evaluate the potential of additional biomarkers as alternative and/or additional diagnostic targets to provide results with better clinical applicability. It is recommended that Australian health authorities should focus on raising awareness of toxoplasma infection and target T. gondii transmission control.

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Costa, Beatriz Guerreiro Basilio. "Estudo da modulação do metabolismo lipídico de células dendríticas humanas na infecção por Toxoplasma gondii." Instituto Oswaldo Cruz, 2012. https://www.arca.fiocruz.br/handle/icict/7036.

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Submitted by Alessandra Portugal (alessandradf@ioc.fiocruz.br) on 2013-10-03T11:40:07Z No. of bitstreams: 1 Beatriz Guerreiro Basilio Costa.pdf: 1448715 bytes, checksum: 9fad325ac3484b88b47961c78ef5db34 (MD5) Made available in DSpace on 2013-10-03T11:40:07Z (GMT). No. of bitstreams: 1 Beatriz Guerreiro Basilio Costa.pdf: 1448715 bytes, checksum: 9fad325ac3484b88b47961c78ef5db34 (MD5) Previous issue date: 2012 Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, Brasil A toxoplasmose é uma doença de alta prevalência no mundo, causada pelo Toxoplasma gondii, um parasita intracelular obrigatório. O T. gondii possui mecanismos de modulação do metabolismo do seu hospedeiro que garantem sua sobrevivência e a instalação da infecção crônica. Um dos tipos celulares mais permissivos à infecção e multiplicação do T. gondii são as células dendríticas (DC), paradoxalmente as células apresentadoras de antígeno mais eficazes, com capacidade de deflagrar uma resposta imune protetora eficaz e duradoura. Neste trabalho, estudamos a modulação do metabolismo lipídico de células dendríticas humanas na infecção por T. gondii. Mostramos que o parasita induziu a maturação da célula e um padrão misto de resposta inflamatória, com altas concentrações das citocinas pró-inflamatórias IL-6 e TNF- e da citocina antiinflamatória IL-10. Após 3 horas de infecção, verificamos que T. gondii induziu a expressão gênica de ciclooxigenase-2 (COX-2). De forma importante, observamos por qRT-PCR que a infecção com T. gondii regulou positivamente a expressão do gene do receptor nuclear (RN) regulado por lipídios PPAR, mas não do LXR. Já a expressão do mRNA das moléculas envolvidas no transporte e estoque de lipídios, FABP4 e ADRP, alvos do PPAR, e ABCA1, alvo do LXR, foram aumentadas pela infecção de 3 horas com o parasita. Utilizando duas técnicas distintas (BODIPY e coloração com ósmio) avaliamos a biogênese de corpúsculos lipídicos (CL) após infecção com o T. gondii e constatamos que essas organelas não foram induzidas após 3 horas de infecção. Entretanto, após 24 horas, 90% das DC apresentaram CL e o número de CL por DC foi estatisticamente maior. Observamos a presença de CL em DC não infectadas pelo parasita, mostrando que a indução da biogênese de CL é um fenômeno parácrino, não dependente da infecção celular. Avaliamos também a importância do PPAR na infecção por T. gondii, através do tratamento das DC com seu agonista ou antagonista. Após 3 horas, apenas os genes ADRP, FABP4 e ABCA1, alvos dos receptores nucleares, foram modulados. Por último, investigamos a influência do T. gondii na expressão das moléculas apresentadoras de antígenos lipídicos. Por citometria de fluxo, constatamos que não há alteração na expressão de membrana dessas moléculas. Contudo, por qRT-PCR, observamos que o T. gondii regula negativamente a expressão dos genes cd1d e cd1e. Em conclusão, mostramos que T. gondii foi capaz de regular positivamente o metabolismo lipídico das DC e negativamente as moléculas apresentadoras de lipídios CD1, sem a participação essencial de PPAR nesses processos. Toxoplasmosis is a worldwide high prevalence disease caused by Toxoplasma gondii, an obligate intracellular parasite. T. gondii developed mechanisms for modulating the metabolism of its host, ensuring their survival and the chronic infection. Dendritic cells (DC) are one of the most permissive cell types to T. gondii infection and replication and are, paradoxically, considered the most effective antigen presenting cells, triggering an effective and long-lasting protective immune response. In this work, we studied the modulation of lipid metabolism in human dendritic cells infected with T. gondii. We showed that the parasite induced cell maturation, and a mixed pattern of inflammatory response with high levels of proinflammatory cytokines IL-6 and TNF-, and anti-inflammatory cytokine IL-10. After 3h of infection, we found that T. gondii induced increased gene expression of cyclooxygenase-2 (COX-2). Importantly, we observed by qRT-PCR that T. gondii infection upregulated the nuclear receptor (RN) PPAR gene expression, while the levels of LXR were unchanged. Furthermore, the mRNA expression of molecules involved in the transport and storage of lipids, FABP4 and ADRP, PPARtargets, and ABCA1, LXR target, were also increased at 3h of infection with the parasite. In parallel, using two different techniques, (BODIPY and osmium staining), we evaluated the biogenesis of lipid droplets (LD) after infection with T. gondii, and observed that these organelles were not induced after 3 hours of infection. However, after 24 hours, 90% of DC showed LD and the number of LD per DC was statistically enhanced. We observed the presence of LD in DC not infected with the parasite, showing that induction of biogenesis of LD is a paracrine phenomenon not dependent on cell infection. We also evaluated the role of PPAR in T. gondii infection by treating DC with its agonist or antagonist. After 3 hours of infection only the nuclear receptor target genes ADRP, FABP4 and ABCA1 were modulated. Finally, we investigated the influence of T. gondii in the expression of lipid antigens presenting molecules. By flow cytometry, we observed no variation in expression of these molecules in cell membrane. However, by qRT-PCR, we found that the T. gondii downregulated gene expression of cd1d and cd1e. In conclusion, we showed that T. gondii was able to upregulate the DC lipid metabolism, and downregulate CD1 lipid presentation. Apparently, the RN PPAR has no essential role in this process.

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Silva, Camila Luna da. "Estudo do metabolismo mitocondrial e da resposta anti-apoptótica de células endoteliais humanas durante a evolução da infecção por taquizoítos de Toxoplasma gondii." Universidade do Estado do Rio de Janeiro, 2013. http://www.bdtd.uerj.br/tde_busca/arquivo.php?codArquivo=8172.

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior A toxoplasmose é uma zoonose amplamente distribuída que afeta mais de um terço da população mundial e de grande importância na saúde pública. A maioria das infecções em humanos por Toxoplasma gondii é assintomática. A toxoplasmose é amplamente investigada visto que se apresenta como uma doença grave em pessoas imunodeprimidas (portadores da síndrome da imunodeficiência adquirida (SIDA), não tratados, indivíduos transplantados, paciente em tratamento quimioterápico ou em uso de drogas supressoras e gestantes). A toxoplasmose congênita frequentemente pode levar ao aborto espontâneo ou até mesmo resultar na formação de crianças com algum grau de atraso no desenvolvimento mental e/ou físicos, deste modo, a transmissão congênita pode ser muito mais importante do que se pensava, pois os parasitos encontrados na circulação sanguinea são capazes de infectar as células endoteliais dos vasos e os tecidos circunjacentes, podendo resultar no encistamento do T. gondii. Atualmente a toxoplasmose vem sendo investigada devido a sua associação a inúmeras outras doenças, assim, estudos sobre a evolução da infecção por T. gondii em diferentes tipos de células hospedeiras se fazem necessários para uma abordagem terapêutica adequada. Ao invadir a célula hospedeira o parasito possui a capacidade de recrutar as mitocôndrias promovendo mudanças na organização mitocondrial ao longo da progressão da infecção, garantindo um ambiente favorável a sua multiplicação. Diante disso, investigamos se o parasito possui a capacidade de interferir no metabolismo mitocondrial e na resposta apoptótica da célula endotelial. O presente trabalho teve como objetivo analisar o metabolismo mitocondrial através da respirometria de alta-resolução e da resposta apoptótica através do western blotting das células endoteliais da veia umbilical humana (HUVEC) infectadas por 2, 6 e 20 horas por taquizoítos de T. gondii. A respirometria de alta-resolução revelou que o parasito interfere no metabolismo energético da célula hospedeira. A análise do conteúdo de proteínas da família Bcl-2 por western blotting revelou maior estímulo apoptótico no tempo inicial de infecção, quando comparado aos demais tempos. Os resultados dos conteúdos de caspase 3, proteína efetora da apoptose, não demonstrou diferença nos tempos iniciais de infecção Entretanto, em tempos mais tardios, o conteúdo de caspase 3 mostrou-se significativamente aumentado quando comparado às HUVEC não infectadas. A dinâmica de replicação do parasito foi observada através do monitoramento pelo sistema Time-Lapse Nikon BioStation IMQ em tempo real das células infectadas por T.gondii. Portanto, nossos resultados sugerem que o protozoário ao recrutar as mitocôndrias da célula hospedeira interfere no metabolismo mitocondrial e na modulação da apoptose para garantir um ambiente favorável a sua multiplicação. Toxoplasmosis is a widespread zoonosis that affects more than a third of the world population and of great public health importance. Most human infections with Toxoplasma gondii are asymptomatic. Toxoplasmosis is widely investigated since it presents itself as a serious disease in immunocompromised persons (holders of acquired immunodeficiency syndrome (AIDS), untreated, transplant recipients, patients undergoing chemotherapy or suppressing drugs and pregnant). Congenital toxoplasmosis can often lead to miscarriage or even result in the formation of children with some degree of developmental delay mental and / or physical, thus congenital transmission may be much more important than previously thought, because the parasites found In the bloodstream are able to infect endothelial cells of blood vessels and surrounding tissues, which may result in encystment T. gondii. Currently toxoplasmosis has been investigated because of their association with other diseases, so, studies of the evolution of T.gondii infection in different types of host cells are necessary for an adequate therapeutic approach. To invade the host cell, the parasite has the ability to recruit mitochondria promoting changes in mitochondrial organization along the progression of infection, ensuring a favorable environment for their multiplication. Therefore, we investigated whether the parasite has the ability to interfere with mitochondrial metabolism and apoptotic response of endothelial cells. This study aimed to analyze the mitochondrial metabolism by high-resolution respirometry and apoptotic response by western blotting of endothelial cells of human umbilical vein (HUVEC) infected for 2, 6 and 20 hours per tachyzoites of T. gondii. The high-resolution respirometry revealed that the parasite interferes with the energy metabolism of the host cell. The analysis of the family protein content of Bcl-2 by western blotting revealed higher apoptotic stimulus at the initial time of infection, as compared to other times. The results of the contents of caspase 3 protein effector of apoptosis, showed no difference in the initial days of infection However, in more recent times, the content of caspase 3 was significantly increased when compared to non-infected HUVEC. The dynamics of parasite replication was observed by monitoring the system Time-Lapse Nikon BioStation IMQ in real time from infected cells by T. gondii. Therefore, our findings suggest that mitochondria in recruiting protozoan host cell interfere with mitochondrial metabolism and in the modulation of apoptosis to ensure a favorable environment for multiplication.

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HIRAMOTO, ROBERTO M. "Efeitos da radiacao ionizante sobre a estrutura, metabolismo e infecciosidade de um protozoario patogenico, Toxoplasma gondii (Nicolle and Manceaux, 1908)." reponame:Repositório Institucional do IPEN, 1998. http://repositorio.ipen.br:8080/xmlui/handle/123456789/10660.

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Made available in DSpace on 2014-10-09T12:42:49Z (GMT). No. of bitstreams: 0 Made available in DSpace on 2014-10-09T14:08:23Z (GMT). No. of bitstreams: 1 06121.pdf: 5715540 bytes, checksum: ca3626a80eb62b032ade08a2bfe344c4 (MD5) Dissertacao (Mestrado) IPEN/D Instituto de Pesquisas Energeticas e Nucleares - IPEN/CNEN-SP

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Nitzsche, Richard [Verfasser], Nishith [Gutachter] Gupta, Kai [Gutachter] Matuschewski, and Maik [Gutachter] Lehmann. "Genetic dissection of the central carbon metabolism in the intracellular parasite Toxoplasma gondii / Richard Nitzsche ; Gutachter: Nishith Gupta, Kai Matuschewski, Maik Lehmann." Berlin : Lebenswissenschaftliche Fakultät, 2017. http://d-nb.info/1130217469/34.

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Mason, Sam. "Toxoplasma gondii in sheep." Thesis, University of Leeds, 2010. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.556024.

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Toxoplasma gondii infects sheep horizontally (from cat faeces) or vertically (transplacentally). Vertically infected lambs sometimes die. Here, transmission and performance impacts were considered in one Charollais flock and one Swaledale flock. B1-PCR was performed on umbilical cord, heart and brain. MAT was performed on blood and pleural effusion. IgG-ELISA was performed on colostrum. B1-PCR was more sensitive than four other methods, producing a band in 50% of replicates when each replicate contained 0.02 parasite genome copies. 16/243 (6.6%) viable Charollais, 30/263 (11.4%) viable Swaledale, 3/54 non-viable Charollais and 0116 non-viable Swaledale were PCR-positive, showing no difference between flocks. At age four months 64/524 (12.2%) Charollais and 10/329 (3.0%) Swaledale were seropositive, showing relatively high seroprevalence in Charollais. 5/44 non-viable Charollais and 1114 non-viable Swaledale were seropositive. Colostrum ELISA was 75% sensitive and 100% specific relative to serum MAT. 15/408 (3.7%) Charollais and 31139 (2.2%) Swaledale were colostrum ELISA-positive, showing no difference between flocks. PCR positivity was not associated with seropositivity. PCR positivity was randomly dispersed between litters. In Charollais seropositivity was overdispersed between litters, seroprevalence was higher than PCR prevalence, young ewes' lambs were frequently PCR-positive and large litters frequently contained seropositive lambs. Those results might have been due to vertical transmission. In Swaledale, PCR positivity was not associated with ewe age and seropositivity was rare. Those results suggested little transmission. Lamb seroconversion, and colostrum ELISA positivity, were not associated with ewe age. Overall, it is suggested that ewes ingested oocysts but vertical transmission was sometimes interupted by lambing, especially in Swaledale. In eight cases clinical toxoplasmosis was suspected. No evidence was found suggesting subclinical effects of T. gondii leading to reduced lamb survival. Charollais born PCR-positive were relatively light at age two months but that association was not found in Swaledale. Serology did not confirm any stunting effect.

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Paugam, André. "Protéasome de "Toxoplasma gondii"." Paris 5, 2002. http://www.theses.fr/2002PA05P611.

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Le protéasome est un complexe protéique cellulaire dont le rôle essentiel est la protéolyse des protéines anormales. Notre objectif était de caractériser le protéasome du toxoplasme (tachyzoi͏̈tes de la souche RH). Par immunoblot, nous avons montré que des extraits cellulaires de toxoplasmes étaient marqués par l'anticorps anti-protéasome MCP231. Par immunofluorescence et microscopie confocale, nous avons observé une localisation essentiellement cytoplasmique du protéasome de T. Gondii, résultat confirmé par la microscopie électronique. L'étude des activités protéolytiques du protéasome a montré que l'activité chymotrypsine-similaire du toxoplasme est proche de celle des cellules de mammifères, alors que l'activité trypsine-similaire est 20 fois plus faible. A l'aide d'un modèle d'infection de fibroblastes murins nous avons étudié l'effet du traitement des toxoplasmes par la gliotoxine, inhibiteur naturel du toxoplasme. L'invasion é été appréciée, 2h après l'infection, par le pourcentage de cellules infectées détectées par cytofluorimétrie de flux (utilisation d'une souche de toxoplasme RH mutée pour exprimer la protéine fluorescente GFP). La multiplication intracellulaire a été quantifiée par le pourcentage d'incorporation d'uracile tritié de cultures de 24h. Bien que la gliotoxine ne modifie pas la pénétration dans les cellules hôtes, elle diminue fortement la multiplication intracellulaire des parasites (CI50 de 0,5m[mu grec]M). L'effet inhibiteur de la gliotoxine sur l'activité chymotrypsine-similaire est 5 fois plus faible pour les toxoplasmes que pour les cellules HeLa. Pour caractériser la structure du protéasome du toxoplasme, nous avons étudié ses sous-unités par électrophorèse bi-dimensionnelle. La comparaison des profils des sous-unités colorés au nitrate d'argent du protéasome de toxoplasme avec le protéasome humain (placenta) a montré des profils différents. L'immunomarquage (anticorps antiprotéasome) montre qu'au moins une des différences protéiques observées concerne bien le protéasome. La caractérisation de cette sous-unité par spectrométrie de masse est en cours d'étude. La caractérisation du protéasome de toxoplasme et sa comparaison avec celui de la cellule hôte, en mettant en évidence des différences significatives, pourrait ouvrir la voie à de nouvelles perspectives thérapeutiques.

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Xia, Dong. "Proteomics of Toxoplasma gondii." Thesis, University of Liverpool, 2009. http://livrepository.liverpool.ac.uk/1276/.

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The Apicomplexan parasite Toxoplasma gondii is an obligate intracellular parasite. Infection by T .gondii causes the disease toxoplasmosis, which is one of the most prevalent parasitic diseases of animals and humans. It has been 100 years since the first discovery of the parasite in 1908; research on T. gondii has been carried out in many scientific disciplines consistently expanding the understanding of this parasite. In the last ten years, the developments of EST, microarray, genome sequencing and continuing efforts towards genome annotation has centralized the focus of T. gondii research on the understanding of gene expression and gene functions on the genome scale. Equipped with the technical advances in mass spectrometry and bioinformatics, proteomics has become established as an integral component in the post-genomics era by providing first-hand data on the functional products of gene expression. In this study, three complementary proteomic strategies, 1-DE, 2-DE and MudPIT, have been used to characterise the proteome of T. gondii tachyzoites. Protein identifications have been acquired for more than two thousand (2252) unique release 4 genes, representing almost one third (29%) of the predicted proteome of all life cycle stages. Functional predictions for each protein were carried out, which provided valuable insights into the composition of the expressed proteome and their potential biological roles. The T. gondii proteomic data has been integrated into the publically accessible ToxoDB, where 2477 intron-spanning peptides provided supporting evidence for correct splice site annotation of the release 4 genome annotation. The incompleteness of the release 4 genome annotation has been highlighted using peptide evidence, confirming 421 splice sites that are only predicted by alternative gene models. Analysis has also been carried out on the proteomic data in the light of other genome wide expression data. The comparison of the proteome and transcriptome of Toxoplasma and other Apicomplexa parasites has revealed important discrepancies between protein and mRNA expression where interesting candidates have been highlighted for further investigation. A preliminary DIGE study has been developed to characterize protein expression changes in T. gondii grown in the presence or absence of glucose. In conclusion, this study has demonstrated the importance of proteomic applications in understanding gene expression profiles and regulation in T. gondii and highlighted the importance and potential of proteogenomic approaches in genome annotation process. The importance of temporal and quantitative proteomics as well as the future of systems biology has been discussed.

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Roch-Deries, Florance Candolfi Ermanno. "Choriorétinite à toxoplasma gondii." [S.l] : [s.n.], 2003. http://www.scd.uhp-nancy.fr/docnum/SCDMED_T_2003_ROCH_DERIES_FLORENCE.pdf.

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com, Nevi Parameswaran@gmail, and Nivethitha (Nevi) Parameswaran. "Toxoplasma gondii in Australian Marsupials." Murdoch University, 2008. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20100203.145857.

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Diagnostic tools were developed and utilised to detect Toxoplasma gondii infection in a range of Australian marsupial species and identify epidemiological trends. An ELISA was developed to detect anti-T. gondii IgG in macropod marsupials. When compared with the commercially available MAT (modified agglutination test), the ELISA was in high agreement and yielded a ê coefficient of 0.96. Of 18 western grey kangaroos (Macropus fuliginosus) tested for the presence of T. gondii DNA by PCR, the 9 ELISA positive kangaroos tested PCR positive and the 9 ELISA negative kangaroos tested PCR negative indicating that the ELISA protocol was both highly specific and sensitive and correlated 100% with the more labour intensive PCR assay. A T. gondii seroprevalence study was undertaken on free ranging Australian marsupials. There was a T. gondii seroprevalence of 15.5% (95%CI: 10.7-20.3) in western grey kangaroos located in the Perth metropolitan area. The T. gondii seroprevalence in male western grey kangaroos was significantly less than their female counterparts (p=0.038), which may be related to behavioural differences causing differences in exposure to oocysts or recrudescence of T. gondii infection in pregnant females. Marsupial populations located in islands free from felids had a low overall T. gondii seroprevalence. A case control study determined that marsupials located in areas where felids may roam are 14.20 (95%CI: 1.94-103.66) times more likely to be T. gondii seropositive than marsupials located on felid-free islands. PCR, immunohistochemistry and serological techniques were used to detect T. gondii infection in marsupial dams and their offspring. T. gondii DNA was detected in the pouch young of chronically infected western grey kangaroos and a woylie (Bettongia penicillata). T. gondii DNA was also identified in the mammary gland of the woylie dam suggesting that infection of the woylie pouch young was from suckling milk from the mammary gland. Results of the study demonstrate that vertical transmission of T. gondii occurs in Australian marsupials and may be of importance in the maintenance of T. gondii infection in Australian marsupial populations. Animal tissue and meat from Australia, predominately from Australian marsupials, were screened for T. gondii DNA using PCR primers for the multi-copy, T. gondii specific B1 gene. Sequencing of the B1 gene revealed atypical genotypes in 7 out of 13 samples from Australia. These 7 isolates contained single nucleotide polymorphisms (SNPs) in the B1 gene that could not be matched with known sequences from strains I, II, III and X. Six unique genotypes were identified out of the 7 atypical isolates; two out of the 7 isolates had the same unique sequence at the B1 gene whereas the other 5 isolates each had different combinations of SNPs at the B1 gene. A majority of T. gondii isolates sampled from native Australian marsupials were of an atypical genotype. The discovery of atypical strains of T. gondii in Australia leads to further questions regarding the origin and transmission of these atypical strains. Additional studies linking atypical strains with their clinical manifestation are also warranted.

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Kremer, Katrin. "Vesicular trafficking in Toxoplasma gondii." Thesis, University of Glasgow, 2013. http://theses.gla.ac.uk/4753/.

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Toxoplasma gondii is an obligate intracellular protozoan parasite with a worldwide prevalence. Together with the causative agent of malaria (Plasmodium falciparum) and other medically important pathogenic parasites it belongs to the phylum of the Apicomplexa. Besides identifiable eukaryotic organelles, apicomplexan parasites differ from other eukaryotic cells by an extra set of specialised secretory organelles (micronemes, rhoptries and dense granules), that are sequentially secreted during invasion of the host cell. Upon host cell contact the apically located micronemes are the first organelles to be released and contain crucial virulence factors that are secreted. In order to systematically analyse vesicular traffic with a special focus on the secretory pathway of rhoptry and microneme proteins the ddFKBP system was used to perform a systematic analysis of Rab proteins in Toxoplasma gondii. Rab proteins are small GTP- binding proteins that are involved in targeting and fusion of vesicles from a donor to an acceptor membrane. Whereas higher eukaryotes like human cells encode more than 60 different Rab proteins apicomplexan parasites possess only a reduced core set of Rab proteins. Performing co-localisation studies with generated parasite lines expressing ddFKBPmyc-tagged versions of Rab1A, 1B, 2, 4, 5A, 5C, 7, 18 and Rab5B-ddFKBPHA revealed, that all these Rabs localise to the early secretory pathway (Rab1B, 2 and 18), the Golgi (Rab4), or the late secretory pathway (Rab5A, Rab5B, Rab5C and Rab7). No exact localisation could be defined for Rab1A. Rab5A and Rab5C, normally involved in endocytic uptake, were identified as important regulators of traffic to micronemes and rhoptries in Toxoplasma gondii, using an overexpression screen of Rabs and the analysis of trans-dominant mutants of promising candidates. Intriguingly, some microneme proteins could be found to traffic independently on functional Rab5A and Rab5C, indicating the existence of independent transport routes to micronemes, which again indicates that apicomplexans have remodelled Rab5-mediated vesicular traffic into a secretory system that is essential for host cell invasion. By using two-colour super-resolution stimulated emission depletion (STED) microscopy, distinct localisations of independent microneme proteins could be verified. This demonstrated that micronemal organelles are organised in distinct subsets or subcompartments. Given these results, it can be assumed that apicomplexan parasites modify classic regulators of the endocytic system to carry out essential parasite-specific roles in the biogenesis of their unique secretory organelles.

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Bhalla, Mayank. "Molecular studies on Toxoplasma gondii." Thesis, University of Reading, 2000. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.339977.

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Parameswaran, Nevi. "Toxoplasma gondii in Australian marsupials /." Murdoch University Digital Theses Program, 2008. http://wwwlib.murdoch.edu.au/adt/browse/view/adt-MU20100203.145857.

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Parameswaran, Nivethitha (Nevi). "Toxoplasma gondii in Australian Marsupials." Thesis, Parameswaran, Nivethitha (Nevi) (2008) Toxoplasma gondii in Australian Marsupials. PhD thesis, Murdoch University, 2008. https://researchrepository.murdoch.edu.au/id/eprint/1680/.

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Parameswaran, Nivethitha (Nevi). "Toxoplasma gondii in Australian Marsupials." Parameswaran, Nivethitha (Nevi) (2008) Toxoplasma gondii in Australian Marsupials. PhD thesis, Murdoch University, 2008. http://researchrepository.murdoch.edu.au/1680/.

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Chaichan, Patcharee. "Epidemiology of Toxoplasma gondii in Thailand." Thesis, Limoges, 2017. http://www.theses.fr/2017LIMO0014/document.

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Toxoplasma gondii est un parasite intracellulaire obligatoire. L'infection par T. gondii est largement répandue dans le monde entier. Néanmoins, elle est peu étudiée dans les pays d'Asie du Sud-est dont la Thaïlande.Nous avons réalisé 3 travaux sur le terrain en Thaïlande pour essayer de comprendre la circulation de ce parasite à travers une étude de séroprévalence chez des poulets en zone rurale et des essais d’isolement de souches chez les animaux en vue d’un génotypage. Lors des deux premières missions de terrain dans deux villages de la province de Kanchanaburi, nous avons cherché à déterminer la séroprévalence de l’infection chez des poulets (Gallus domesticus) en utilisant 2 tests sérologiques, Modified-Agglutination Test (MAT et immunofluorescence indirecte (IFAT) puis à isoler des souches de T.gondii à partir des animaux séropositifs. Lors de la troisième mission réalisée dans 3 autres provinces thaïlandaises(Nakhonratchasima, Lopburi et Saraburi), nous avons essayé d’isoler directement le parasite à partir de carcasses de poulets vendues sur les marchés ou d’autres animaux trouvés morts.La séroprévalence globale pour les 2 premières misions sur 600 poulets du Kanchanaburi était de 17,7% (IC 95% :14,6-20,7) et 33,0% (IC 95% : 29,2-36,8), par MAT et IFAT respectivement. Le calcul du coefficient κ montre une absence de concordance entre les deux tests.Au total, 162 essais d'isolement ont été effectués par inoculation à des souris, mais aucune souche viable de T. gondii n'a été isolée pendant ces 3 travaux sur le terrain. Cependant, nous avons détecté la présence d’ADN toxoplasmique en qPCR ciblant le gène 529 bp dans 13 culots de digestion d’organes de poulets, pigeon, caille et dans des cerveaux ou coeurs de souris inoculés par 16 autres poulets. Les Ct observés en qPCR étaient ≥33 indiquant une faible quantité d’ADN parasitaire dans nos échantillons qui n’a pas permis une caractérisation génétique par marqueurs microsatellites.Ce travail a démontré l'importance et les difficultés du travail de terrain pour l'étude de séroprévalence ainsi que l'étude d'isolement. L'isolement des souches de T. gondii a demandé un travail d'échantillonnage intensif, complexe dans l’environnement tropical et humide de la Thaïlande. Les différents paramètres ayant pu avoir un impact négatif sur nos résultats sont discutés. Ils expliquent l’absence d’isolement de souches chez des animaux séropositifs Toxoplasma gondii is an obligate intracellular parasite. Toxoplasma gondii infection is widespread throughout the world. Nevertheless, it is poorly studied in Southeast Asian countries including Thailand. We carried out 3 field works in Thailand to try to understand the circulation of T. gondii through a seroprevalence study in chickens in rural areas and strain isolation attempts in animals. During the two first field works, performed in Kanchanaburi province, we determined the seroprevalence in chickens (Gallus domesticus) using 2 serological tests, a Modified-Agglutination-Test (MAT) and an immunofluorescence assay (IFAT) and subsequently tried to isolate the strains of T. gondii from seropositive animals. During the third field work carried out in 3 other Thai provinces (Nakhonratchasima,Lopburi and Saraburi), we attempted to isolate strains directly from chicken carcasses sold in different markets or other dead animals.The overall seroprevalence for 600 chickens sampled over the two field works in Kanchanaburi was 17.7% (95%CI: 14.6% -20.7) and 33.0% (95% CI: 29.2-36.8), by MAT, and IFAT, respectively. The κ coefficient indicated an absence of concordance between the 2 serological tests.A total of 162 isolation attempts were performed by mouse bioassays, but no viable strain of T. gondii was isolated during these 3 field works. However, a qPCR targeting 529 bp T. gondii gene was positive for 13 digestion pellets of organs of chickens, pigeon, quail and in brains or hearts of mice inoculated with 16 other chickens. These qPCR were weaklly positive (Ct ≥33) indicating a low amount of parasite DNA in our samples that did not allow genotyping T. gondii with microsatellite markers.This work demonstrated the importance and difficulties of field work for the seroprevalence study as well as strain isolation. The isolation of T. gondii strains required intensive and complex sampling in the tropical and humid environment of Thailand. The diverse factors that could have a negative impact on our results are discussed. They might explain the absence of strain isolation from seropositive animals

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Woods, Stuart. "Immunological control of toxoplasma gondii infection." Thesis, University of Strathclyde, 2012. http://oleg.lib.strath.ac.uk:80/R/?func=dbin-jump-full&object_id=19276.

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Prevalent worldwide, the protozoan parasite, Toxoplasma gondii, is an important cause of spontaneous abortion, ocular disease, mental retardation and encephalitis. Currently there are no human vaccines available. The first major aim of this study was to test potential HLA restricted peptide vaccines, previously shown to be protective in HLA-transgenic mice, against oocyst infection. The ability of entrapment within non-ionic surfactant vesicles to improve the efficacy of the HLA-B*0702 restricted vaccine was also studied. In parallel we tested the novel T. gondii ΔRPS13 live-attenuated vaccine against oocyst challenge. As determined by survival, only ΔRPS13 provided a measure of protection against oocyst challenge. We also demonstrated that the live vaccine induced a greater CD8+ T cell effector response than the adjuvanted peptide vaccine. Successful vaccination is in large part dependent on inducing an appropriate response in the primary host cell populations that consequently influences the development of adaptive immunity. Parasite induced macrophage arginase-1 expression, for example, has been shown to be influential during T. gondii infection. Arginase-1 expression is negatively regulated by Map Kinase Phosphatase-2 (MKP-2), the second major aim of the project was to study the effect of MKP-2 deficiency on T. gondii infection. MKP-2-/- mice were found to be more susceptible to infection with increased parasite growth and increased mortality compared with wild type mice. Increased susceptibility was associated with reduced serum nitrite levels and enhanced tissue arginase-1 expression although the Th1 response was unaltered. In vivo inhibition of iNOS and arginase-1 revealed that while NO production is of paramount importance in controlling parasite growth arginase-1 could also limit parasite growth independently. In vitro studies utilising macrophages confirmed a role for arginase-1 in parasite control. Results highlight a complex interaction between iNOS and arginase-1 and T. gondii in L-arginine metabolism but indicate that manipulation of early infection events influence disease outcome.

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Cirelli, Kimberly M. "Rodent inflammasome activation by Toxoplasma gondii." Thesis, Massachusetts Institute of Technology, 2016. http://hdl.handle.net/1721.1/105635.

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Thesis: Ph. D., Massachusetts Institute of Technology, Department of Biology, 2016. Cataloged from PDF version of thesis. Includes bibliographical references. Toxoplasma gondii is an obligate intracellular pathogen capable of chronically infecting nearly all warm-blooded animals, including humans. The chronic stage is characterized by the presence of semi-dormant cysts in brain and muscle tissues. These cysts are crucial in the success of Toxoplasma as they are orally infectious and allow for the transmission of the parasite between hosts. As the host immune response drives cyst formation, the establishment of this chronic infection relies on the parasite's ability to find a balance between activation of a host immune response and evasion of parasiticidal mechanisms. This balance is achieved through the modulation of host cell processes by parasite proteins secreted from specialized secretory organelles known as rhoptries and dense granules. Here, we report that Toxoplasma activates the inflammasomes in mice and rats. The inflammasomes are a set of cytoplasmic pattern recognition receptors (PRRs). Activation of the inflammasomes results in caspase-1 activation and the cleavage and release of the pro-inflammatory cytokines, Interleukin (IL)-1[beta] and IL-18. IL-1p is an important mediator of local inflammation and neutrophil recruitment. IL- 18 induces Interferon (IFN)-[gamma], which is a critical cytokine in the control of Toxoplasma. A form of cell death, termed pyroptosis, can accompany inflammasome activation. The NLRP3 inflammasome is activated in mouse macrophages, leading to the secretion of IL-1[beta] in vitro. The NLRP1 and NLRP3 inflammasomes play a major role in mouse survival and control of parasite replication in vivo. The NLRPI inflammasome is activated in infected macrophages from rats that are able to completely clear infection. Toxoplasma infection leads to the secretion of active IL-I[beta] and IL-18. Activation of the NLRP1 inflammasome leads to pyroptosis, a programmed form of cell death. Pyroptosis prevents parasite replication within the host cell and likely promotes clearance by nearby immune cells. Using a chemical mutagenesis screen, we identified three Toxoplasma dense granule proteins (GRAs), GRA18, GRA27 and GRA28, essential for NLRP1 inflammasome activation and pyroptosis in rat macrophages. Our work has identified Toxoplasma gondii as a novel activator of the rodent inflammasomes and demonstrated host cell death as a mechanism to control parasite replication. We have also identified three novel parasite proteins required for this activation, providing insight into interactions between parasite and host, which may aid in the treatment of human infection. by Kimberly M. Cirelli. Ph. D.

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Ravindran, Sandeep. "Effector protein secretion by toxoplasma gondii /." May be available electronically:, 2009. http://proquest.umi.com/login?COPT=REJTPTU1MTUmSU5UPTAmVkVSPTI=&clientId=12498.

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Brennan, Anthea. "Toxoplasma gondii infection in Australian felines." Thesis, The University of Sydney, 2015. http://hdl.handle.net/2123/15060.

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Toxoplasma gondii is a significant human and animal parasite with worldwide occurrence. The population structure and prevalence in definitive hosts and intermediate hosts has been well studied in a number of regions including North and South America and Europe. While Toxoplasma gondii has been previously reported in a number of mammalian species in Australia, there is a surprising lack of information regarding the population structure of T.gondii in Australia and current rates of exposure in domestic companion animal hosts. Members of the felidae family are the only known definitive host of T. gondii in which sexual reproduction of the parasite occurs.Sexual reproduction results in shedding of environmentally resillient oocysts in faeces. All strains of T. gondii can be traced back to a feline host. As such, an understanding of T. gondii infection among feline hosts is an important first step to understanding general prevalence and population structure of the parasite in a particular area. The seroprevalence of T. gondii in cats worldwide is estimated to be 30-40%. In addition to being a definitive host of T. gondii, domestic cats are susceptible to clinical disease due to T.gondii. Although uncommon, feline toxoplasmosis occurs in immunosuppressed cats, congenitally infected kittens and occasionally in otherwise clinically healthy individuals. Little is known about whether parasite genotype is associated with severity of disease in naturally occurring toxoplasmosis of domestic cats. The first aim of this research was to expand knowledge of feline T. gondii infections in Australia by determining the seroprevalence of T. gondii in owned Australian cats and identifying riskfactors for infection. The second aim was to determine whether the genotype of T. gondii is a significant determinant of whether cats develop clinical toxoplasmosis. To estimate seroprevalence of T. gondii in Australian owned cats, Toxoplasma specific IgG ELISAs were performed on sera from 425 owned domestic Australian cats. A multivariate 12 analysis of the results from a questionnaire given to the owners was used to evaluate lifestyle factors which could contribute to increased likelihood of infection. Of the 425 cats tested in this study 38% (n=162) were seropositive. The prevalence in different geographic regions ranged from 16-71%. Cats fed raw beef or raw kangaroo in their diet or cats that hunted rodents were significantly more likely to be seropositive To identify the genotypes associated with latent and active infections in Australian cats, tissue samples were collected from cats undergoing routine post mortem examination at the University Veterinary Teaching Hospital (n=28). Serology to detect T. gondii specific IgG was performed after collection of heart blood from cats of unknown T. gondii serostatus. Cases were chosen based on the likelihood of a cat being exposed to the parasite. Results of a PCR targeting the B1 gene to detect T. gondii DNA were positive in tissue samples from 11 of 17 (65%) seropositive cats tested including four with clinical toxoplasmosis and seven with latent infections, as determined by serology, histologic findings and immunohistochemistry. Three of the four cats with clinical toxoplasmosis were immunosuppressed. T. gondii type II (ToxoDB genotype #3) was determined in four cats with clinical toxoplasmosis and three cats with latent toxoplasmosis using PCR-RFLP at 12 loci (SAG1, 5’SAG2 and 3’SAG2, altSAG2, SAG3, BTUB, GRA6, c22-8, c29-2, L358, PK1 and Apico) and direct sequencing of the multicopy B1 gene. Novel T. gondii B1 gene polymorphisms were detected in two strains (at nucleotide posititions 233, 366 and 595) and a B1 gene polymorphism unique to Australia was identified in another (guanine/adenine at nucleotide position 378). One cat was coinfected with two or more type-II like strains at 3’SAG2. The results of this study suggest that the infecting T. gondii genotype is not a determinant of clinical disease in cats naturally infected with T. gondii and type II strains are the most prevalent in Australia. 13 The ToxoDB#3 genotype has been previously identified in Australian hosts. Given the high prevalence in the present study, it appears that this genotype may be endemic to Australian hosts and this discovery warrants further investigation of a larger sample set using high resolution techniques such as those presented in this thesis to confirm this. Naturally infected cats with active and latent infection were both infected with the same strain, indicating the infecting strain is not the major determinant of disease progression and that other host factors may be of greater importance. Throughout this program of research, epidemiological aspects of feline T. gondii infections in Australia were elucidated further. Key findings include a seroprevalence of 38% among owned Australian cats, the identification of a cat which was infected with more than one strain of T. gondii, the discovery that infections with the same ToxoDB#3 genotype can cause both clinical and latent infection in cats and the confirmation of diversity at the B1 locus in felines, which has been previously found in Australian wildlife species. This is the first survey of feline infections in owned Australian cats to evaluate lifestyle factors which contribute to the likelihood of exposure and it is also the first study to genetically characterise infecting strains in cats with clinical toxoplasmosis. Further studies are needed to fully understand the population structure and also the prevalence of T. gondii in Australia as current data is lacking in other domestic and wildlife species.

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Burrells, Alison Clair. "Toxoplasma gondii in animal and human hosts." Thesis, University of Edinburgh, 2014. http://hdl.handle.net/1842/9628.

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The protozoan parasite Toxoplasma gondii (T. gondii) is an important zoonotic pathogen, which has the ability to infect all warm blooded mammals including humans, with approximately one third of the human population predicted to be infected. Transmission of the parasite to the foetus during pregnancy can result in miscarriage, however, a child infected during pregnancy may go on to develop clinical symptoms such as retinochoroiditis (ocular toxoplasmosis), hydrocephalus or learning difficulties in later life. Post-natally acquired infection in humans is generally asymptomatic, however, individuals who are immunocompromised may develop ocular toxoplasmosis or toxoplasmic encephalitis. T. gondii type II is reported to be the predominant genotype in Europe and the United States, but currently very little information exists about the prevalence and genotypes present within Great Britain. Consumption of T. gondii tissue cysts from raw or undercooked meat is a main source of infection for humans, with infected pork being considered a high risk. Currently the “gold standard” for assessing the viability of infective T. gondii tissue cysts is by an in vivo mouse bioassay. However, more recent ethical requirements to reduce, refine or replace experimental animals raises the question as to whether molecular technologies could be incorporated into these studies to reduce mouse numbers. The main aims of this PhD were to: (i) determine the prevalence and genotypes of T. gondii within different wildlife populations and humans in Great Britain; (ii) determine whether vaccination of pigs with a live attenuated strain of T. gondii would reduce the load of viable T. gondii tissue cysts within this species; (iii) study the viability and dissemination of tissue cysts from oocyst and bradyzoite infected pigs and (iv) to compare mouse bioassay with molecular detection of T. gondii DNA from experimentally infected pigs. The main findings of this work show that the prevalence of T. gondii within carnivorous wildlife varied from 6.0% to 44.4% depending on the host species with type II being the predominant lineage identified, however, type III and two alleles for type I were also present. In humans, serological detection of the parasite from a group of Scottish blood donors from Glasgow and Dundee (n=1403) was determined at 13.0%, molecular detection of T. gondii in human brains (n=151) from the Sudden Death Brain Bank show a prevalence of 17.9%. A correlation between increasing age and an increase in the detection of parasite was identified from both study groups. T. gondii strain genotyping using DNA extracted from human brains identified alleles for type I and III, however, no direct link between cause of death and detection of parasite DNA could be made. Live vaccination and subsequent oocyst challenge of pigs showed a significant reduction in the establishment of viable T. gondii tissue cysts. Mouse bioassay clearly demonstrates this result, where 100% of mice that were inoculated with homogenised tissues from vaccinated/challenged pigs survived, compared to the survival of only 51% of mice, which received homogenised tissues from non-vaccinated/oocyst challenged animals. In addition, porcine tissues from pigs challenged with either oocysts or bradyzoites did not show a significant difference in mouse survival following bioassay of these tissues. Challenge with either stage of the parasite (oocysts or bradyzoites) showed a preference to form tissue cysts in brains and highly vascular muscles (tongue, diaphragm, heart or masseter) of pigs. The findings, comparing mouse bioassay with molecular detection of parasite DNA from homogenised porcine tissue (prior to inoculation into mice), showed similar levels of detection. However, mouse bioassay was more sensitive and also provides evidence of parasite viability. In conclusion, this research not only provides current figures for prevalence and genotypes of T. gondii in both wildlife and humans in Great Britain, it also successfully answers the question as to whether live vaccination of pigs with the S48 strain can reduce the tissue cyst burden. These promising results show the potential of a vaccine against T. gondii in producing safer pork for human consumption. Although the mouse bioassay still remains the most sensitive method for the detection and viability assessment of tissue cysts, further research should be carried out in this area, perhaps incorporating a technique such as magnetic capture qPCR, to enable an effective in vitro technique to be developed.

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Riahi, Dehkordi Hamayoun. "Hammondia hammondi : études comparatives avec Toxoplasma gondii." Limoges, 1997. http://www.theses.fr/1997LIMO105E.

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Liu, Elizabeth. "The Autophagy Pathway and Toxoplasma gondii Infection." Case Western Reserve University School of Graduate Studies / OhioLINK, 2015. http://rave.ohiolink.edu/etdc/view?acc_num=case1428103561.

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Bertin, Brigitte. "Etude immunochimique des antigènes de Toxoplasma gondii." Bordeaux 2, 1985. http://www.theses.fr/1985BOR22001.

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Hartmann, Jan. "Golgi and centrosome cycles in Toxoplasma gondii." [S.l. : s.n.], 2005.

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Bertin, Brigitte. "Etude immunochimique des antigènes de Toxoplasma gondii." Grenoble : ANRT, 1985. http://catalogue.bnf.fr/ark:/12148/cb37594507t.

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Jesus, Rogerio Fernando de. "Infeccção natural por Toxoplasma gondii em quirópteros." Universidade Federal da Bahia, 2015. http://repositorio.ufba.br/ri/handle/ri/20375.

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Submitted by Emanoel Martins Filho (emanoelfilho@ufba.br) on 2016-09-13T00:53:42Z No. of bitstreams: 1 Rogerio_Fernando.pdf: 1011706 bytes, checksum: ec29e66873bd428e74520f5f059408bb (MD5) Approved for entry into archive by Patricia Barroso (pbarroso@ufba.br) on 2016-09-13T20:36:53Z (GMT) No. of bitstreams: 1 Rogerio_Fernando.pdf: 1011706 bytes, checksum: ec29e66873bd428e74520f5f059408bb (MD5) Made available in DSpace on 2016-09-13T20:36:53Z (GMT). No. of bitstreams: 1 Rogerio_Fernando.pdf: 1011706 bytes, checksum: ec29e66873bd428e74520f5f059408bb (MD5) CONS NAC DE DESENVOLVIMENTO CIENTIFICO E TECNOLOGICO - CAPES Toxoplasma gondii é um protozoário coccídeo formador de cistos teciduais, que tem como hospedeiros definitivos os felídeos, e como hospedeiros intermediários mamíferos e aves. É um parasito disseminado em todos os continentes, que infecta aproximadamente um terço da população humana e pode causar encefalite fatal em pacientes imunodeficientes. Nos animais, tem relevância principalmente em pequenos ruminantes, por causar abortos e outras alterações reprodutivas. Quirópteros podem se infectar com T. gondii e atuarem como fonte de infecção para animais silvestres, domésticos e o homem. No Brasil, ocorre uma alta variabilidade genética do parasito, que pode ser explicada pela grande variedade de hospedeiros no ambiente silvestre. Objetivou-se com este estudo determinar a frequência de infecção em quirópteros de vida livre no estado da Bahia por T. gondii e realizar o isolamento in vivo do protozoário a partir desses animais. Foram utilizadas 124 amostras, provenientes de 97 indivíduos de sete espécies de morcegos, capturados entre os anos de 2008 e 2015, sendo encontrados dois indivíduos positivos por meio da PCR de tecidos, correspondendo a 2,06% de positividade. Nenhum isolamento foi realizado uma vez que os tecidos disponíveis para bioensaio apresentaram-se negativos na PCR Toxoplasma gondii is a cyst-forming protozoan coccidia, which has felids as definitive hosts, and mammals and birds as intermediate hosts. It’s distributed in all continents and infects about a third of the human population. T. gondii can cause fatal encephalitis in immunodeficient patients. In animals, it’s relevant mainly in small ruminants, for causing abortions and other reproductive abnormalities. Bats can become infected with T. gondii and act as a source of infection for wild and domestic animals and man. In Brazil, there is a high genetic variability of the parasite, which can be explained by the great variety of hosts in the wild environment. The objective of this study was to determine the frequency of free-living bats infection in Bahia by T. gondii and perform in vivo isolation of the parasite from these animals. A total of 124 samples were used from 97 individuals of seven species of bats, caught between the years 2008 and 2015. Two animals were positive by tissue PCR, corresponding to 2.06% of positivity. No isolation was achieved once the tissue available for bioassay showed to be negative by PCR.

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Carey, Robert Francis IV. "Toxoplasma gondii and behavioral modification in hosts." Thesis, Boston University, 2013. https://hdl.handle.net/2144/12065.

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Thesis (M.A.)--Boston University Toxoplasma gondii is a heteroxenous protozoan parasite that is found in nearly every species of mammal and billions of latently infected humans worldwide. The symptoms and morbidities associated with acute, congenital, and AIDS-associated toxoplasmosis are familiar to many, while those associated with latent toxoplasmosis are not nearly as well known. Behavioral manipulation is a common strategy of parasite and parasitoid species, and recent research into T. gondii has revealed that T. gondii infection alters the way rodents respond to the odor of the urine of its feline predators, which are also the definitive hosts of T. gondii. Humans have been found to be potentially affected by T. gondii as well: associations have been identified between latent T. gondii infection and psychiatric diseases (including schizophrenia), personality changes, and traffic accidents. This review investigates the state of current scientific knowledge related to Toxoplasma gondii, analyzes recent developments, and examines the implications on public health. We also provide critical analysis of the published literature and make suggestions for future research.

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Parigi, Maria <1984&gt. "Toxoplasma gondii in animals and the environment." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2014. http://amsdottorato.unibo.it/6423/1/Parigi_Maria_tesi.pdf.

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Toxoplasma gondii is an obligate intracellular parasite capable of infecting virtually all warm-blooded species, including humans, but cats are the only definitive hosts. Humans or animals acquire T. gondii infection by ingesting food or water contaminated with sporulated oocysts or by ingesting tissue cysts containing bradyzoites. Toxoplasmosis has the highest human incidence among zoonotic parasitic diseases, but it is still considered an underreported zoonosis. The importance of T. gondii primary infection in livestock is related to the ability of the parasite to produce tissue cysts in infected animals, which may represent important sources of infection for humans. Consumption of undercooked mutton and pork are considered important sources of human Toxoplasma gondii. The first aim of this thesis was to develop a rapid and sensitive in- house indirect ELISA for the detection of antibodies against T. gondii in sheep sera. ROC-curve analysis showed high discriminatory power (AUC=0.999) and high sensitivity (99.4%) and specificity (99.8%) of the method. The ELISA was used to test a batch of sheep sera (375) collected in the Forli-Cesena district. The overall prevalence was estimated at 41.9% demonstrating that T. gondii infection is widely distributed in sheep reared in Forli-Cesena district. Since the epidemiological impact of waterborne transmission route of T.gondii to humans is now thought to be more significant than previously believed, the second aim of the thesis was to evaluate PCR based methods for detecting T. gondii DNA in raw and finished drinking water samples collected in Scotland. Samples were tested using a quantitative PCR on 529 bp repetitive elements. Only one raw water sample (0.3%), out of the 358 examined, tested T. gondii positive demonstrating that there is no evidence that tap water is a source of Toxoplasma infection in Scotland.

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Parigi, Maria <1984&gt. "Toxoplasma gondii in animals and the environment." Doctoral thesis, Alma Mater Studiorum - Università di Bologna, 2014. http://amsdottorato.unibo.it/6423/.

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Nolan, Kay. "The transmission dynamics of Toxoplasma gondii in sheep." Thesis, University of Warwick, 2006. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.479313.

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Mohammed, Saleem. "Molecular studies on the protozoan parasite Toxoplasma gondii." Thesis, University of Reading, 1994. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.262211.

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Wheeler, Ruth Belinda. "Studies of the molecular biology of Toxoplasma gondii." Thesis, University of Reading, 1993. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.356985.

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Nickdel, Mohammad Barat. "Role of Th2 cytokines in Toxoplasma gondii infection." Thesis, University of Strathclyde, 2002. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.248258.

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Henriquez, Fiona Luisa. "Recharacterization of Toxoplasma gondii dense granule protein GRA3." Thesis, University of Strathclyde, 2003. http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.273437.

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Wansadhipathi-Kannangra, Nilu Kumari. "Sphingolipid synthases of Toxoplasma gondii and other organisms." Thesis, Durham University, 2011. http://etheses.dur.ac.uk/3204/.

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THE SPHINGOLIPID SYNTHASES OF TOXOPLASMA GONDII AND OTHER ORGANISMS Nilu Kumari Wansadhipathi-Kannangara Toxoplasma gondii is an obligate intracellular protozoan parasite of the phylum Apicomplexa, and toxoplasmosis is an important disease in immuno-compromised individuals - especially AIDS patients, organ transplant recipients and patients receiving anti-cancer chemotherapy. Toxoplasmosis also has economic importance as it results in spontaneous abortion in economically important animals. Due to emerging drug resistance and the possible side effects of the existing therapeutics there is a necessity to explore new drug targets. Sphingolipids are essential components of eukaryotic cell membranes and signal transduction pathways. Mammals produce sphingomyelin (SM) via a SM synthase, whereas yeast, plants and some protozoa utilise an inositol phosphorylceramide (IPC) synthase to produce IPC. IPC synthases have no mammalian equivalent and have been proposed as targets for anti-fungals and anti-protozoals. In this study sphingolipid scavenging from the host was shown to be non-essential for Toxoplasma proliferation indicating de novo synthesis is key. To investigate this pathway in the parasite an orthologue of the SM synthase from the related apicomplexan Plasmodium falciparum was identified in the genome database as being encoded by a single copy gene. Subsequently it was characterized as a functional orthologue of the yeast AUR1p (IPC synthase), with mass spectrometry identifying this lipid species in parasite extracts for the first time. Like AUR1p and the human SM synthase, the Toxoplasma sphingolipid synthase (Tg SLS) is Golgi localized. However, unlike AUR1p Tg SLS is resistant to the inhibitor aureobasidin A and also facilitates production of a minor complex sphingolipid suggested to be SM. Further study characterized the sphingolipid synthases from the Trypanosoma species (protozoan Kinetoplastida) and the plant Arabidopsis. Bioinformatic and phylogenetic analyses of these and previously characterized SM and IPC synthases, indicate them to be a family of evolutionarily related, but functionally diverse, enzymes. Many of these may prove to be targets for pharmaceutical or herbicidal intervention.

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Ghérardi, Arnaud. "Toxoplasma gondii : étude de la purine nucléoside phosphorylase." Lyon 1, 2001. http://www.theses.fr/2001LYO1T208.

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La 4e de couverture indique : "Toxoplasma gondii est un protozoaire Apicomplexa responsable de toxoplasmose congénitale et cérabrale. Cette dernière est une importante cause de morbidité et de mortalité chez l'immunodéprimé. Comme la majorité des protozoaires, T. Gondii est incapable de synthétiser ses propres purines et dépend donc entièrement de celles de son hôte. Il dispose des enzymes pour interconvertir celles-ci de façon à former les bases puriques qui intégreront divers métabolimes et constitueront les acides nucléiques. Le dosage de six enzymes de la voie des purines a montré que la purine nucléoside phosphorylase (PNP Ec 2. 4. 2. 1) présentait l'activité enzymatique la plus élevée dans le cerveau murin et dans la forme tachyzoi͏̈te de la souche ME49 du parasite isolé de culture cellulaire. L'enzyme de cerveaux de souris saines et de souris parasitées par la souche avirulente DUR de T. Gondii a été purifiée d'un facteur 40. La PNP de tachyzoi͏̈tes de la souche virulente RH et avirulente ME49 isolés de culture cellulaire a été purifiée d'un facteur 11 et la détermination des paramètres cinétiques montre une différence entre l'enzyme du parasite et celle de l'hôte. Des composés chimiques inhibiteurs de la PNP de protozoaires et des composés de formules originales, synthétisés par le laboratoire de Chimie Organique, ont été testés, et l'activité enzymatique de l'enzyme a été dosée par chromatographie liquide haute performance afin de déterminer les IC50 et les Ki des inhibiteurs. La révélation histoenzymatique par précipitation du plomb a été réalisée. L'activité PNP sur des coupes de kyste cérébral de souris et sur des tachyzoi͏̈tes de la souche ME49 de T. Gondii isolés de culture cellulaire a été mise en évidence par microscopie électronique. La localisation de la PNP est cytoplasmique et est particulièrement importante dans les tachyzoi͏̈tes par rapport à leur cellules hôtes. Une intense activité intracytoplasmique péri-membranaire a été observée dans ces formes. "

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Liew, Lloyd. "Structural studies on microneme proteins from Toxoplasma gondii." Thesis, Imperial College London, 2013. http://hdl.handle.net/10044/1/11654.

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Toxoplasma gondii is a polyxenous parasite of the phylum Apicomplexa and the etiological agent of the coccidian disease Toxoplasmosis. Toxoplasma gondii is characterized by its unique gliding motility as well as its ability to infect virtually all nucleated cells. The micronemes are a group of secretory organelles that contribute to the ability of the parasite to recognize the surface of the host-cell prior to parasite invasion via the secretion of adhesive protein complexes. Previous attempts at obtaining structural data for several microneme proteins from Toxoplasma gondii such as microneme protein TgMIC 2, 3 and 4 have so far been unsuccessful due to the inability to produce soluble protein samples using the prokaryotic expression system, Escherichia coli. This work is focussed upon the adaptation of a eukaryotic expression system, Pichia pastoris, to producing Apicomplexan microneme proteins which can cause problems in prokaryotic expression systems due to its eukaryotic origin as well as its high cysteine content. Modifications to the fermenter medium and standard screening protocols allowed for the expression of microneme protein domain constructs in Pichia pastoris with yields ranging from 0.1 to 1.2 mg/L in fermenter culture. Soluble protein samples were produced for several constructs expressed in Pichia pastoris including full length samples of TgMIC3 and TgMIC4 as well as the TSR56 pair in TgMIC2. In addition, isotopically enriched soluble and folded protein samples were successfully produced with a yield of 0.02 to 0.12 mg/L and were successfully subjected to analysis via 2D NMR. Full length TgMIC4 was also successfully crystallized and resulted in crystals which diffracted to 8.0Å. Pichia pastoris has been successful in producing soluble microneme protein samples and has the potential to alleviate the long-standing bottleneck in our efforts to understand the mechanisms involved in Toxoplasma gondii host-cell invasion.

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Bouleau, Anne Pascaline. "Identification et caractérisation de métalloprotéases de Toxoplasma gondii." Thesis, Reims, 2014. http://www.theses.fr/2014REIMM202/document.

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Toxoplasma gondii est un parasite protozoaire intracellulaire obligatoire appartenant à la famille des Apicomplexa. Chez les protozoaires, les protéases possèdent des rôles clés au niveau du cycle parasitaire, et sont ainsi considérées comme des facteurs de virulence. Chez T. gondii, les métallopeptidases pourraient être impliquées dans la traversée des différentes barrières biologiques. A l'heure actuelle, seules cinq métallopeptidases de T. gondii ont été décrites : une aminopeptidase N, deux toxolysines, une leucine aminopeptidase et une FtsH1 peptidase. Lors de l'étude de l'influence de l'invasion de monocytes humains par T. gondii sur le profil d'expression des métalloprotéases matricielles monocytaires, nous avons mis en évidence une protéase parasitaire présentant à la fois des propriétés gélatino- et élastinolytique.Le but de ce travail est de caractériser, purifier et identifier cette gélatinase de T. gondii.Dans un premier temps, nous avons caractérisé la gélatinase d'environ 100 kDa sécrétée par T. gondii comme étant une MMP-9 like (métallo-endopeptidase à zinc) mais ne possédant pas le même processus d'activation que les MMPs humaines.Dans un deuxième temps, après purification partielle par une série de chromatographie chélatrice de zinc, nous avons identifié par spectrométrie de masse la gélatinase comme étant la TGME49_227948 annotée dans ToxoDB. Cette protéase présente les domaines protéiques d'une métalloprotéase à zinc de la sous-famille M16C, et est proche au niveau de sa structure 3D de la chaine A de la 2FGE présente chez Arabidopsis Thaliana. Afin de confirmer l'annotation de ToxoDB, nous avons séquencé l'extrémité 5' de l'ARNm de cette protéase. Cependant, nos résultats expérimentaux ne sont pas en concordance avec l'annotation prédite dans la base de données ToxoDB.La séquence en acides aminés de cette protéase, nous a permis de synthétiser deux anticorps polyclonaux spécifiques afin de mettre en évidence deux formes de 140 kDa et 100 kDa et donc d'émettre l'hypothèse que cette protéase pourrait être clivée afin d'être activée. De plus, cette métalloprotéase a été détectée par western blot dans le cytosol des parasites mais elle est aussi secrétée dans le milieu conditionné. Par immunolocalisation, la protéase est présente au sein du parasite, au niveau du cytosol sans localisation préférentielle dans un organite particulier.Dans ce travail, nous avons montré que la gélatinase parasitaire sécrétée par T. gondii pourrait dégrader des composés de la matrice extracellulaire, d'où son rôle potentiel dans le mécanisme de traversée des barrières biologiques Toxoplasma gondii is an intracellular protozoan parasite which belongs to the Apicomplexa phylum. In protozoans, proteases have key roles in the parasitic cycle. They thereby are considered as virulence factors. In T. gondii, metallopeptidases may be involved in the crossing of biological barriers. Currently, only five metallopeptidases from T. gondii are described: an aminopeptidase N, two toxolysins, one leucine aminopeptidase and one FtsH1 peptidase. During the study of the influence of human monocytes invasion by T. gondii on the monocytic matrix metalloproteases expression profile, we brought out a parasitic protease showing gelatino- and elastinolytic properties.The aim of this study is to characterize, purify and identify this gelatinase from T. gondii.First we characterized the 100 kDa-gelatinase secreted by T. gondii as an MMP-9-like (zinc metalloendopeptidase) but its activation process is different from human MMPs.Then, after the partial purification by a series of zinc-chelate chromatographies, we identified by mass spectrometry the gelatinase as the TGME49_227948 annotated in ToxoDB. This protease has M16C subfamily zinc-metalloprotease protein domains and is close to the 3D structure of the A chain of the 2FGE in Arabidopsis thaliana. In order to confirm the ToxoDB annotation, we sequenced the 5' end of the mRNA of this protease. Nevertheless our experimental results are not in the line with the predicted annotation in ToxoDB database.The amino acids sequence of this protease allowed us to synthesize two specific polyclonal antibodies in order to highlight two forms, 140 kDa and 100 kDa, and to emit the hypothesis that this protease could be cleaved to be activated. Moreover this metalloprotease was detected by Western Blot in the cytosol of the parasites but it can also be secreted in the conditioned medium. By immulocalization, the protease is present in the cytosol of the parasite without any preferential localization in a particular organelle.In this study, we showed that the parasitic gelatinase secreted by T. gondii could degrade extracellular matrix compounds, which could explain its potential role in the mechanism involved in the crossing of biological barriers

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Alruhaili, M. H. B. "Genetic diversity of African isolates of Toxoplasma gondii." Thesis, University of Salford, 2016. http://usir.salford.ac.uk/37753/.

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Toxoplasma gondii is an intracellular protozoan parasite and has the ability to infect all warm-blooded animals, including humans. While the three clonal lineages of T. gondii (I, II and III) predominate in North America and Europe, strains from other regions in the world appear to have more diverse genotypes. The aim of the current research is to analyse the level of genetic variation among local African T. gondii isolates in relation to their phenotype (genotype phenotype relationships). In this study, multi-locus nested PCR sequence analysis of seven Ugandan T. gondii isolates was applied using nine different genetic markers distributed across seven chromosomes and the apicoplast genome of T. gondii, which improved the discrimination power to detect variation among the local Ugandan strains. Although these markers were sufficient to separate global variation between T.gondii strains, they were not adequate to totally resolve within closely related local isolates. To understand the impact of local variation on strain diversity, whole genome sequence was generated for two Ugandan strains type II using Illumina MiSeq paired-end sequencing, revealing variations between these strains and the type II reference strain of T. gondii (TgME49). In this study, we have perhaps the first example of the deeper sequencing of isolates from the same geographical region at the same time point, which showed that they are non-identical. Novel polymorphisms were identified in a virulence associated gene in both Ugandan strains resulting in modification of the protein structure of this gene which could be associated with phenotype variation in the in vitro growth rate of these strains. Comparing the in vitro growth rates of the sympatric Ugandan strains, a cluster of 3 strains had higher growth. These were genotypically identical by using PCR sequencing technique, while the non-identical sympatric strains had lower growth rates, providing evidence that genotype may influence phenotype. An important finding was evidence of recombination between type II and III within three Ugandan strains, revealed through multi-locus PCR sequencing, and in an additional Ugandan strain through deeper whole genome sequencing. Six polymorphic markers were identified via analysis of three biologically relevant genes families (SRS, ROPs and GRA), enhancing the resolution power to identify variations among local type II strains of T. gondii. It is recommended that further study of these polymorphic markers is carried out and that they are added into the MLST analysis of T. gondii, especially between closely related local isolates.

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Frohnecke, Nora. "Funktionelle Charakterisierung des Ferredoxin Redoxsystems von Toxoplasma gondii." Doctoral thesis, Humboldt-Universität zu Berlin, 2018. http://dx.doi.org/10.18452/19075.

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Toxoplasmose ist weltweit eine der am häufigsten auftretenden parasitären Zoonosen mit einer geschätzten Infektionsrate von über 30%. Toxoplasma gondii (Phylum: Apicomplexa) besitzt ein Plastid ähnliches Organell, den Apicoplasten. In diesem befindet sich das einzig bekannte Redoxsystem, welches aus der Ferredoxin-NADP+-Reduktase und Ferredoxin (Fd) besteht. Fd als Elektonendonator liefert Elektronen an verschiedene essentielle Stoffwechselwege, wie der Isoprenoidvorstufen- und Liponsäuresynthese. Um die bei einem Elektronentransfer benötigte direkte Protein-Protein-Interaktion eingehend zu analysieren, wurde ein bakterielles Reverse Two Hybrid System verwendet, womit die Interaktion von TgFd und TgLipA gezeigt werden konnte. Da angenommen wird, dass Fd eine zentrale Rolle in verschiedenen Stoffwechselwegen übernimmt, ist für einen Fd Knockout ein komplexer biochemischer Phänotyp zu erwarten, der möglicherweise zum Absterben der Parasiten führt. Zur Untersuchung dessen wurden zwei komplementäre Wege verfolgt. Eine der Strategien basierte auf dem grundsätzlichen Nachweis, dass Fd unerlässlich für das Überleben von T. gondii ist. Mit Hilfe des DiCre Systems sollte ein definierter genetischer Fd Knockout hergestellt werden, welcher jedoch nicht zweifelsfrei generiert werden konnte. Bei der zweiten Strategie kam ein konditionales Knockdown System zur Anwendung, bei welchem die Expression des Fd Gens nach Induktion herabreguliert wird. Mit Hilfe dessen konnten weitreichende Auswirkungen der Fd Defizienz auf T. gondii gezeigt werden: die Fettsäuresynthese der im Apicoplasten synthetisierten Fettsäuren ist reduziert sowie die Motilität durch eine beeinträchtigte Isoprenoidsynthese verringert, wodurch insgesamt drastische Auswirkungen auf das Parasitenwachstum gezeigt werden konnten. Beide Stoffwechsel sind vom Elektronendonator Fd abhängig und durch die Fd Herabregulation betroffen. Die Ergebnisse unterstreichen die essentielle Rolle des Fd-Redoxsystems von T. gondii. Toxoplasmosis is one of the most common parasitic zoonoses world-wide, around 30% of human beings are infected. Toxoplasma gondii (phylum: Apicomplexa) contains a unique intracellular organelle derived from plastids, called apicoplast. The only known redox system in the apicoplast consists of the ferredoxin NADP+-reductase and its redox partner, ferredoxin (Fd). The latter donates electrons to different essential metabolic pathways in the apicoplast like the last two enzymes of the isoprenoid precursor biosynthesis and the lipoic acid synthesis. To dissect protein protein interactions for an electron transfer a bacterial reverse two hybrid system was used. The physical interaction of both proteins TgFd and TgLipA could be shown. Fd is supposed to play an important role in diverse metabolic pathways, hence a knock-out of the Fd gene is expected to generate a complex biochemical phenotype and be lethal to the parasite. Therefore two complementary approaches were used to analyze the role of TgFd in this context. The first strategy shall verify the essentiality of TgFd for the survival of T. gondii. It is based on the DiCre system whereby a defined genetic knock out of TgFd is produced. Respectives parasites have been generated, but at the end no genetic Fd knock out could be produced. In the second approach a conditional knock-down was generated, where the expression of the TgFd gene is repressed after induction. The Fd deficiency has wide ranging effects on T. gondii: The fatty acid synthesis of the apicoplast-synthesized fatty acids is reduced as well as the motility is decreased due to an affected isoprenoid synthesis. In total this leads to a dramatic inhibition of parasite growth. Both metabolic pathways depend upon the electron carrier Fd and thus are affected by Fd deficiency. The results underline the essential role of the ferredoxin redoxsystem of T. gondii.

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Dardé, Marie-Laure. "Contribution à la caractérisation de Toxoplasma Gondii : étude isoenzymatique." Limoges, 1990. http://www.theses.fr/1990LIMO101A.

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Gastens, Martin. "Virulenz-assoziierte Proteine von Toxoplasma gondii Identifikation und funktionelle Charakterisierung /." [S.l. : s.n.], 2003. http://deposit.ddb.de/cgi-bin/dokserv?idn=968522270.

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Adjogblé, Koku Zikpi. "Biochemische und funktionelle Charakterisierung eines neuen "Dense-granules"- Proteins von Toxoplasma gondii: GRA9." [S.l. : s.n.], 2004. http://deposit.ddb.de/cgi-bin/dokserv?idn=971753792.

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