|
|
Utilization of advanced techniques of electron microscopy in altrastructural analysis of microbial cells and microbial biotechnology products
Havlíčková, Anna ; Sedláček, Petr (referee) ; Krzyžánek,, Vladislav (advisor)
In the theoretical part, this thesis deals with direct and indirect mechanisms of plant growth-promoting rhizobacteria, which is followed by a section on the selected model organism Azotobacter vinelandii. The second part of the theoretical section deals with electron microscopy starting with the possible interactions of the electron beam with the sample, through types of electron microscopes to various methods of preparing biological samples. The experimental part discusses the cultivation of selected bacterial cultures, the gelation of the cell cultures using chosen cross-linking agents (calcium chloride, malic acid, glucono--lactone with calcium carbonate) and, above all, various methods of preparation and analysis of pure and encapsulated cultures by the means of electron microscopy. In particular, cryogenic scanning electron microscopy and scanning transmission electron microscopy imaging were selected. Using both techniques, poly-3-hydroxybutyrate granules, which were confirmed to be elastic even at cryogenic temperatures, were observed inside the bacteria. Furthermore, it was found that cross-linking of extracellular alginate occurs even in samples without the addition of cross-linking agents. However, the hydrogel network present in these samples was significantly thinner than in samples with the addition of all cross-linking agents used. In this work, the density of the hydrogel network cross-linked with glucono--lactone was also compared. When using this agent with the addition of calcium carbonate, a significantly denser hydrogel network was observed in comparison with using the agent alone. Last but not least, when comparing the obtained results with the literature, it was found that the A. vinelandii bacteria form capsules, during the formation of the capsules the shape of the cells changes from an oval to a spherical bacterium and also the size of the given cell is reduced. From the resulting images, it was found that these capsules are made up of two layers, to which literature refers as electron-transparent intine and electron-dense exine.
|
guest ::
