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Comparative study of Trichomonas vaginalis strains with various level of metronidazole resistance
Havelka, Michal ; Tachezy, Jan (advisor) ; Kopáček, Petr (referee)
Trichomonas vaginalis is an anaerobic protozoan that causes the human disease trichomoniasis. The most commonly used drug against trichomoniasis is metronidazole. Metronidazole is highly effective, yet in some cases T. vaginalis is resistant to this drug. In T. vaginalis, we distinguish resistance to metronidazole into two types based on the conditions in which it occurs, aerobic and anaerobic resistance. Both types of resistance are associated with changes in the physiology of the parasite. The aim of this study was to characterize the changes in hydrogenosome and cytosol associated with metronidazole resistance at the proteome level in aerobically resistant clinical isolates, the aerobically resistant line MR3 and the anaerobically resistant line MR100. A further aim of this work was to characterize the function of hybrid- cluster protein and to clarify its role in resistance to metronidazole since its production is more abundant in both aerobically and anaerobically resistant T. vaginalis. This work contributed to identification of several potential proteins that could play a role in the development of metronidazole resistance and established a foothold for further metabolomic and biochemical studies of metronidazole resistant T.vaginalis.
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Protists in oral cavity of humans and animals.
Brixí, Kateřina ; Tachezy, Jan (advisor) ; Rada, Petr (referee)
Trichomonas tenax, Trichomonas gallinae, Trichomonas gypaetinii, Trichomonas stableri, Trichomonas equibuccalis, Trichomonas brixi, Tetratrichomonas canistomae, Tetratrichomonas felistomae and Tetratrichomonas empyemagena are unicellular eukaryotes (protists) that may occur in oral cavity of humans and domestic animals. These protozoans are mostly referred as commensals and relatively low attention is paid to them, since they usually are not primary cause of serious health problems. However, based on current knowledge, it can be concluded that more appropriate term for Entamoeba gingivalis and Trichomonas tenax would be an opportunistic pathogen, while Trichomonas gallinae is a primary pathogen. There are very little studies and findings about the rest of oral protozoans and their pathogenicity, therefore it is not possible to assess their significance for a host. Nevertheless, the oral protozoans should not be neglected, as in some cases they can cause serious health complications in their host.
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FeS cluster assembly in cytosol of Trichomonas vaginalis
Stojanovová, Darja ; Tachezy, Jan (advisor) ; Šuťák, Róbert (referee)
Iron-sulfur (FeS) cluster assembly is extensively studied in model organisms, e.g. Saccharomyces cerevisiae, Homo sapiens, and more recently in Trypanosoma brucei. However, little is known about FeS assembly in divergent anaerobic organisms such as Trichomonas vaginalis, which parasites in the human urogenital tract. This parasitic protist possesses anaerobic form of mitochondria, the hydrogenosome, in which some component of FeS cluster assembly machinery (ISC) has been identified, whereas the cytosolic CIA pathway has not been studied so far. Our work deals mainly with TvIscU, a component of ISC pathway, and T. vaginalis CIA pathway. We suggest that both hydrogenosomal and cytosolic FeS cluster assembly pathways of this parasite differ from typical models. We examined possible ISC-CIA relationship. Next, we found homologues for several key components involved CIA machinery, namely Nbp35, Cfd1, Nar1, Cia1 and Cia2. However, we did not identify any homologous proteins to Tah18, Dre2 and Mms19. We expressed identified proteins with HA-tag and localized them by cell fractionation and immunofluorescence microscopy in T. vaginalis. Finally, we immunoprecipitated two Cfd1 paralogues, TvCfd1A and TvCfd1B to search for their interacting partners. The results suggest that these two paralogues interact with...
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Pathogenic trichomonads in cats and dogs
Vobořilová, Pavlína ; Tachezy, Jan (advisor) ; Hampl, Vladimír (referee)
Trichomonads are anaerobic flagellated protists that are either parasites or commensals. They frequently inhabit digestive, respiratory, and urogenital tracts of vertebrates, including domestic cats and dogs. In these hosts, four trichomonad species has been described: Tetratrichomonas canistomae and Tetratrichomonas felistomae that are commensals of the host oral cavity; Pentatrichomonas hominis, a commensal of intestinal tract that could be found in dogs and cats but also in other mammals including humans; and pathogenic Tritrichomonas foetus that causes, in addition to cattle infection, feline intestinal trichomonosis. Although, trichomonads in dogs and cats are probably of cosmopolitan distribution we have no information about their presence in Czech Republic. The first aim of this study was to distinguish types of trichomonads present in the oral cavity of dogs and cats and to get preliminary epidemiological data. The second aim was to demonstrate the presence of Tritrichomonas foetus in cats and dogs in the Czech Republic and to identify potential risk factors for the disease. Cultivation and nested PCR were used to determine the presence of trichomonads in dogs and cats. Sequencing and phylogenetic analysis based on ITS1-5.8rRNA-ITS2 gene sequence was used to identify species of isolated...
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Prokaryotic proteins of antioxidant defense in Trichomonas vaginalis hydrogenosomes
Smutná, Tamara
Parasitic protists with modified mitochondria represent important and exciting group of organisms, not only from the view of eukaryotic cell evolution but also because these parasites are causative agents of serious and widespread diseases. The study and understanding of their biology is thus necessary for the development of new antiparasitic drugs. These organisms reside in host body cavities with low concentrations of oxygen and while they lack typical mitochondria, they possess mitochondrion-related organelles which still integrate many physiologically important processes. Trichomonas vaginalis is an anaerobic flagellate inhabiting mucosal surface of vagina. Instead of canonical mitochondria, T. vaginalis possesses organelles termed hydrogenosomes. These organelles harbor pathways of ATP-generating metabolism via substrate-level phosphorylation, dependent on enzymes prone to oxidative damage, such as pyruvate:ferredoxin oxidoreductase and Fe-Fe hydrogenase. Because the environment of trichomonads is not fully anaerobic, the parasite had to develop complex strategies to cope with both oxygen and reactive oxygen species (ROS) generated by host immune system cells. Recent data from T. vaginalis proteomic and genomic analyses revealed the presence of bacterial-type proteins potentially participating...
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Resistance of Trichomonas vaginalis and other anaerobic pathogens to metronidazole
Havelka, Michal ; Tachezy, Jan (advisor) ; Rada, Petr (referee)
The aim of this bachelor thesis is to desbribe mechanisms of resistance to metronidazole in three anaerobic eukaryotic pathogens (T. vaginalis, G. intestinalis, E. histolytica). Diseases caused by these pathogens belong to the list of important but currently neglected diseases. Metronidazole acts only on microbes with an anaerobic metabolism. The drug enters the cell by passive diffusion and needs to be activated by reduction of the nitro group. Mechanisms of activation are different for every pathogen. Enzymes bound with energetic metabolism in hydrogenosomes, cytosolic thioredoxin reductase and nitroreductases play a major role in the activation. The drug damages cells in three ways - DNA damage, formation of covalent bonds with proteins, and covalent bonds with thiols. Pathogens have specific mechanisms to defend themselves against the drug. They can either down-regulate enzymes that activate metronidazole, reduce it to non-reactive aminoimidazole or they can increase the intracellular concentration of oxygen which leads to deactivation of the drug by futile cycling. These mechanisms are bound with physiological changes and subsequently with lowered viability of these pathogens. Furazolidone and benzimidazole derivatives are the best candidates to become an alternative to metronidazole for the...
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Interaction between hydrogenosomes and endoplasmic reticulum in Trichomonas vaginalis
Kučerová, Jitka ; Tachezy, Jan (advisor) ; Černý, Jan (referee)
Endoplasmic reticulum-mitochondria encounter structure (ERMES) is a protein complex tethering ER and mitochondria. ERMES consists of four core subunits - Mmm1, Mmm2 (Mdm34), Mdm10 and Mdm12. It was first discovered in Saccharomyces cerevisiae and most functional information is based on studies of this organism. ERMES affects mitochondrial distribution and morphology, participates in lipid trafficking and is important for homeostasis of the cell. In Trichomonas vaginalis, the human urogenital parasite, three genes for putative, highly divergent components of ERMES complex were predicted. However, the cell localization of these proteins and their function is unknown. This thesis is focused on investigation of ERMES components in T. vaginalis, their cellular localization, interactions between components and identification of their possible interacting partners.
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Prokaryotic proteins of antioxidant defense in Trichomonas vaginalis hydrogenosomes
Smutná, Tamara
Parasitic protists with modified mitochondria represent important and exciting group of organisms, not only from the view of eukaryotic cell evolution but also because these parasites are causative agents of serious and widespread diseases. The study and understanding of their biology is thus necessary for the development of new antiparasitic drugs. These organisms reside in host body cavities with low concentrations of oxygen and while they lack typical mitochondria, they possess mitochondrion-related organelles which still integrate many physiologically important processes. Trichomonas vaginalis is an anaerobic flagellate inhabiting mucosal surface of vagina. Instead of canonical mitochondria, T. vaginalis possesses organelles termed hydrogenosomes. These organelles harbor pathways of ATP-generating metabolism via substrate-level phosphorylation, dependent on enzymes prone to oxidative damage, such as pyruvate:ferredoxin oxidoreductase and Fe-Fe hydrogenase. Because the environment of trichomonads is not fully anaerobic, the parasite had to develop complex strategies to cope with both oxygen and reactive oxygen species (ROS) generated by host immune system cells. Recent data from T. vaginalis proteomic and genomic analyses revealed the presence of bacterial-type proteins potentially participating...
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Immune system, nitric oxide and parasites
Oreský, Matej ; Hrdý, Ivan (advisor) ; Macháček, Tomáš (referee)
Nitric oxide belongs to smallest known molecules. It has been known since the mid- 17th century, but only in the second half of the 20th century its significance in medicine began to be discovered. Already in the 19th century, nitroglycerin was used to treat heart disease, and the connection with nitric oxide was later found. Nitric oxide works, for example, as a signaling molecule in biosystems, acts as a vasodilator in endothelial tissue but also has a number of pathophysiological effects. Because of its radical nature, it plays an important role in the immune system in the non-specific response. By reaction with certain radicals in the body, strong reactive compounds such as peroxynitrite can form. However, some intracellular parasites have been found to be able to resist the effects of the immune system associated with the NO molecule. They could, besides other things, serve proteins from the family of flavodiiron proteins to this purpose. At the end this review focuses on the group of flavodiiron proteins and the interaction of three parasites with the immune system of the host: Entamoeba, Giardia and Trichomonas. Key words: Nitric oxide, NO synthase, macrophage, Trichomonas vaginalis, Giardia intestinalis, Entamoeba histolytica, flavodiiron protein
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Prokaryotic proteins of antioxidant defense in Trichomonas vaginalis hydrogenosomes
Smutná, Tamara ; Hrdý, Ivan (advisor) ; Horváth, Anton (referee) ; Kopáček, Petr (referee)
Parasitic protists with modified mitochondria represent important and exciting group of organisms, not only from the view of eukaryotic cell evolution but also because these parasites are causative agents of serious and widespread diseases. The study and understanding of their biology is thus necessary for the development of new antiparasitic drugs. These organisms reside in host body cavities with low concentrations of oxygen and while they lack typical mitochondria, they possess mitochondrion-related organelles which still integrate many physiologically important processes. Trichomonas vaginalis is an anaerobic flagellate inhabiting mucosal surface of vagina. Instead of canonical mitochondria, T. vaginalis possesses organelles termed hydrogenosomes. These organelles harbor pathways of ATP-generating metabolism via substrate-level phosphorylation, dependent on enzymes prone to oxidative damage, such as pyruvate:ferredoxin oxidoreductase and Fe-Fe hydrogenase. Because the environment of trichomonads is not fully anaerobic, the parasite had to develop complex strategies to cope with both oxygen and reactive oxygen species (ROS) generated by host immune system cells. Recent data from T. vaginalis proteomic and genomic analyses revealed the presence of bacterial-type proteins potentially participating...
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