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Molecular characterization of the factors participating in the formation of Giardia intestinalis cyst wall.
Pastyříková, Aneta ; Doležal, Pavel (advisor) ; Hrdý, Ivan (referee)
1 Abstract The surface of Giardia intestinalis cysts is covered by a thick fibrillar wall providing the parasite with the necessary protection against the external environment. The cyst wall consists of proteins and carbohydrates, among which is a unique β-1,3-N-acetylgalactosamine homopolymer (known as Giardan). This carbohydrate is synthesized from glucose by cytosolic enzymes induced during encystation. This thesis focuses on the biosynthesis of Giardan and the functional characterization of UDP-N-acetylglucosamine 4'-epimerase (UA4E), one of the enzymes involved in the synthetic pathway. For these purposes, the corresponding gene was targeted using the CRISPR/Cas9 system. A stable cell line was obtained in which all four copies of the UA4E gene were deleted. The success of gene knock-out was confirmed on the basis of proteomic analysis. Using fluorescence microscopy, we observed that ΔUA4E cells induced to encystation are able to initiate encystation processes and cell differentiation. The presence of ESV vesicles and typical morphological changes corresponding to cysts were observed in the cells. However, these stages had a thin surface layer and were unable to form typical thick wall with fibrillar appearance at their surface leading to the subsequent production of non-viable cysts. Key words Giardia...
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Current treatment options of intestinal and extra-intestinal amebiasis.
Pastyříková, Aneta ; Nohýnková, Eva (advisor) ; Stejskal, František (referee)
Amoebiasis is a parasitic infection caused by the pathogenic intestinal amoeba Entamoeba histolytica, which multiplies as a minute form in the human lumen of the colon. Under not yet defined conditions, the minute form can be transformed into a magna form that attacks the colon wall and is able to spread out of the intestine by blood and infect internal organs, especially the liver. Amoebiasis is primarily treated with chemotherapeutics, which are divided into luminal and tissue depending on the site of action. Luminal drugs target the stages of E.histolytica in the lumen of the intestine and are therefore used to treat asymptomatic and symptomatic forms of intestinal amoebiasis. Tissue drugs, especially the 5-nitroimidazole derivative, metronidazole, which is the drug of choice in the treatment of amoebiasis, penetrate the tissues and are used to treat invasive intestinal and extra-intestinal infections. If chemotherapy is ineffective or there is a risk of rupture of abscess and sepsis, surgical treatment is performed. This thesis summarizes therapeutic approaches to individual clinical forms of amoebiasis, possibilities of chemotherapy at present, but also before the use of 5-nitroimidazole derivates, and alternatives to current drugs. Key words Intestinal amebae of humans, Entamoeba histolytica,...
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