|
|
Study of Sterilization Effects Initiated by Dielectric Barriere Discharge
Slámová, Jitka ; Pekárek, Stanislav (referee) ; doc. RNDr. Karol Hensel, Ph.D., oponent (referee) ; Krčma, František (advisor)
The overall goal of the presented dissertation thesis was to study the sterilization efficiency of dielectric barrier discharge operated at atmospheric pressure. The fungi Aspergillus niger, gram-positive bacteria Bacillus subtilis and in some experiments also gram-negative bacteria Escherichia coli were used as a bio-indicator enabling to evaluate the effect of plasma assisted microbial inactivation. The samples of microorganism were placed on paper Whatman 1 or PET foil and exposed to plasma. The plasma was generated in argon, nitrogen, synthetic dry/humid air with frequency up to 10 kHz and plasma power density in the range of 1,2-2,9 W/cm3 (according to the process gas). The influence of process gas, plasma power density, plasma exposition time, type of microorganism and material of the substrate on the sterilization effect of dielectric barrier discharge was evaluated. Furthermore the contribution of each single mechanism (UV radiation, temperature and reactive species) to the sterilization effect of plasma and influence of gas humidity was evaluated. The DBD was analysed by means of optical emission spectroscopy, thermocouple was used to measure temperature during a sterilization process. In order to verify the mechanical damage of the microbial cell or the substrates during the plasma process the samples were studied by scanning electron microscopy. Generally, on the basis of experimental results, at increasing treatment times, the remaining number of spores (CFU) decreased. Similarly at increasing the plasma power input, the sterilization rate increased. When sterilising the spores of A. niger in plasma using different process gasses, the efficiency of plasma sterilization decreased as follows: argon, humid synthetic air, nitrogen and dry synthetic air. The results observed in argon plasma using different microorganism demonstrated that the sensitivity of vegetative cells resp. spores to DBD decreased as follows: A. niger spores, B. subtilis vegetative cells, E. coli vegetative cells and B. subtilis spores. Simultaneously results observed for sterilization of spores and vegetative cells of B. subtilis and A. niger demonstrated that the spores are generally more resistant to plasma than are the corresponding vegetative cells. Combining the results of contribution of each single mechanism, optical emission spectroscopy and inactivation characteristic it was found out that the reactive species significantly contribute to the plasma sterilization in all process gasses. Furthermore the inactivation process can be partly assisted by UV radiation and also the temperature can contribute in limited extent to inactivation process in some gasses. The contribution of UV radiation to the plasma sterilization decreased as follows: nitrogen, argon, dry syntetic air and humid syntetic air. Moreover it was found out that the contribution of each single mechanism can be species dependent, this is due to the different response of microorganism to the unfavorable external conditions. SEM analysis of the substrates prooved the etching actions of the plasma generated in all process gasses on the surface of the PET foil. The several minute plasma exposition of the PET foil resulted in the occurence of the „hole corrosion“ on the PET surface. Contrary to these there were no visible changes observed in the paper structure.
|
|
|
Nucleic acids as terapeutic agent
Ráčková, Lucie ; Španová, Alena (referee) ; Rittich, Bohuslav (advisor)
With the development of molecular biology development of oncotherapy proceeds. The major progress of modern medicine is gene therapy. In the gene therapy are two categeories, namely, viral vectors and nonviral vectors which are used mainly. Nonviral vectors include plasmids. Plasmid DNA used in medicine must be perfectly purified. Chromatographic methods are mainly used at present. Research and development deals with other methods for example two-phase aqueous systems and magnetic carriers. In experimental part of this thesis, isolation of pUC 19 plasmid DNA from Escherichia coli JM 109 (pUC 19) cell culture was performed via method of alkaline lysis. Quality of isolated plasmid DNA was verified spectrophotometrically and by agarose gel electrophoresis. Isolated plasmid DNA was purified using three methods: RNA in plasmid DNA was precipitated by lithium chloride, RNA was degraded by immobilized RNase A and plasmid DNA was purified using two-phase aqueous system.
|
|
|
Antimicrobial activity of selected natural extracts
Šandová, Lucie ; Veselá, Mária (referee) ; Zemanová, Jana (advisor)
The aim of the bachelor thesis was to determine antimicrobial activity of selected natural extracts, specifically three model extracts made from purple coneflower (Echinacea purpurea) and four commercial extracts – true lavender (Lavandula angustifolia), lemon balm (Melissa oficinalis), echinacea herbal drops (Echinacea purpurea) and echinacea extract (Echinacea purpurea) with ginger, against selected microorganisms. Theoretical part is focused on description of bioactive substances that are found in plants. It also describes extraction methods used for obtaining extracts from plant materials, methods for determining antimicrobial activity and then the use of natural extracts in food and cosmetic industry. In experimental part, antimicrobial effect of the extracts was tested against gram-negative bacteria Escherichia coli, gram-positive bacteria Bacillus cereus and yeast Candida glabrata. For determination of the antimicrobial activity, two methods (hole method and disc diffusion method) were used and compared. The highest antimicrobial activity against selected microorganisms was showed by commercial extract of lemon balm, which contains lot of geraniol and citroneloll. Significant antimicrobial activity had also been shown by commercial extract of true lavender. On the contrary, extracts from Echinacea had not shown any antimicrobial activity against selected microorganisms.
|
|
|
Use of genotoxicity tests to characterization of some natural substances and particles
Tilšarová, Kamila ; Bokrová, Jitka (referee) ; Márová, Ivana (advisor)
In production of drugs, cosmetics, in food industry and other biotechnologies, testing of genotoxicity (that are changes in genetic information which are not inhereted) is one of the common steps in the process before distribution to the market. This bachelor´s thesis is focused on the characterization of extracts from chosen natural substances with respect to the content of polyfenols, flavonoides and antioxidant activity. These extracts were encapsulated into liposomes and chitosan particles. The aim was testing possible genotoxicity of whole extracts and particles on possible genotoxicity on the basis of SOS Chromotest performed on bacteria E. coli. This genotoxic assay did not show genotoxic effect of neither pure extracts nor tested liposomes.
|
|
|
Effects of ultrasound on biological materials
Peprníčková, Anna ; PhD, Vladan Bernard, (referee) ; Vlk,, Daniel (advisor)
In this bachelor thesis I deal with effects of ultrasound on biological materials. In theoretical part is write up a literature search about effects of ultrasound on bimacromolecules namely deoxyribonucleic acid (DNA), proteins (BSA) and bacterial cells. Effects of ultrasound was investigated practically on genetically modified bacterias of Escherichia coli. These bacterias were sounded by ultrasound by various frequences for the same time and then their viability was measured on luminometer. The results for ever frequency were compared.
|
|
|
Influence of antibacterial nanomaterials on the development of bacterial resistance
Bábíková, Daniela ; Lucie,, Bozděchová (referee) ; Šmerková, Kristýna (advisor)
Bacterial resistance to antibiotics is a long-discussed problem. The use of antibiotics in primary agricultural production, especially in pig breeding. High levels of zinc subsequently seeps into the soil in the form of secondary agricultural products. Doses of effect the intestinal bacteria of pigs, but also the microorganisms in the environment which can subsequently lead to the development of resistance to metals also antibiotics. This thesis deals with the effect of ZnO and newly used nanoparticles (ZnO NPs) on bacteria S. aureus, methicilin-rezistant S. aureus and E. coli. Acquired data point to the effect of ZnO on minimal inhibition concentration of antibiotics and the development of resistance in E. coli. Based on biochemical tests, the change of metabolic pathways reacting with selected substrates was confirmed. Repeated subcultur of bacteria with tested ZnO and ZnO NPs has on the susceptibility of bacteria to antibiotics and changes their biochemical pathways. Studies dealing with the issue of long-term effect on medicinal substances are essential for the development of modern antibacterial components.
|
|
|
Využití různých typů povrchově modifikovaných magnetických nanočástic pro izolaci a purifikaci bakteriální DNA/RNA
Bednářová, Eliška
In the submitted thesis "The use of different types of surface modified magnetic nanoparticles for isolation and purification of bacterial DNA/RNA", magnetic nanoparticles modified with tetraethoxysilane (TEOS) and (3-aminopropyl)triethoxysilane (APTES) were used. The aim was to optimize the nucleic acid isolation protocol using magnetic particles from both gram negative and gram positive bacteria in order to achieve the highest possible RNA yield. In the theoretical part of the thesis, the structure and composition of bacterial cells were summarized, especially the composition of the cell wall, which allows the bacteria to be divided into gram positive and gram negative. Furthermore, the different options for bacterial detection used in clinical practice were presented. Nucleic acid isolation is often an essential step in the detection of bacteria using molecular methods. A method based on magnetic particles is one of them, and the principle was explained here. The composition and functionalisation of the particles used for nucleic acid isolation have been described. In the practical part of the thesis, magnetic particles were characterized by several methods to specify their properties. Thus, the most suitable particles for nucleic acid isolations was selected. Next, the particles were used to optimize the procedure for nucleic acid isolation from cell lysate. The lysis buffer with 30% Triton X-100 detergent was determined to be the most effective and similar results were obtained with the 30% EcoSurf detergent. Subsequently, a method for efficient bacterial cell lysis was optimalized. Several methods were used and the best results were achieved by mechanical homogenization with 0.1 mm zirconia beads in combination with lysis buffer with 1% EcoSurf detergent. RNA yields from isolations by magnetic particles were compared with those from column isolations using a commercial isolation kit. Approximately 350 ng/µl of RNA yield was achieved for S. aureus using the column method, while 70 ng/µl less was achieved using magnetic particles. RNA yields were significantly higher than those using custom buffers, therefore, further optimization of the isolation protocol and composition of custom buffers in nucleic acid isolation would be advisable. Magnetic particles with both TEOS and APTES modifications have great potential for use in the isolation of nucleic acids from bacteria.
|
|
|
Escherichia coli jako alimentární patogen a příčina kažení potravin
Smejkalová, Martina
The main aim of the bachelor’s thesis is to characterize the bacterium Escherichia coli. The thesis focuses on the general characteristics of this bacterium and also the whole genus Escherichia and also the bacterial cell. The bacterium Escherichia coli is described in terms of morphology, biochemical properties, antigenic properties and epidemiology. However, the main focus was its pathogenic properties, which are also related to prevention and treatment. Pathogenic Escherichia coli is divided into extraintestinal and intestinal. In thesis, the different representatives are characterized as well as the diseases they cause. This is related to the topic of alimentary diseases, which is also explained in the thesis. The last issue that is discussed in the thesis is food spoilage and the characteristics of meat products, which are related to the practical part of the thesis. The practical part focuses on the analysis of Escherichia coli, as well as coliforms and CPM in raw meat and sausage samples. In the raw meat samples, the prevalence of all groups of microorganisms was the highest, whereas in the sausages the prevalence was very low and Escherichia coli and coliforms were not present at all. Raw meat is very susceptible to contamination and proliferation of micro-organisms, whereas sausages have been properly cooked, which means that this result was expected.
|
|
|
Hodnocení mikrobiologických rizik jedlého hmyzu
Hedvičáková, Adéla
Eating edible insects is an activity that has recently become increasingly important. In order to use edible insects safely, it is necessary to carry out, among other things, an assessment of the risks associated with consumption. This work focused on the microbiological risk assessment of edible insects. The theoretical part of the thesis provides information on the nutritional properties of edible insects and the risks associated with insect consumption. It also mentions legislation related to the inclusion of insects in food. The thesis describes the impact of insects and their consumption on the environment and presents factors that need to be further addressed in the future. The thesis provides a list of important pathogenic microorganisms. In the practical part it is possible to find a statistical evaluation of the measured values of the pathogens present in the insects and in the feed. Total counts of microorganisms, Escherichia coli, Enterobacteriaceae, moulds, Staphylococcus aureus and lactic acid bacteria were determined. The results were compared with those of studies on similar topics.
|
|
|
Vliv kovů v potravinářství na bakteriální virulenci
Pavezka, Pavel
The bachelor's thesis deals with the issue of gene transfer of resistance of β-lactam antibiotics by means of horizontal gene transfer, specifically transformation. It deals with the transfer of resistance genes through the food chain, when Escherichia coli can subsequently cause foodborne illness. It also points to the current risks of resistance to β-lactam antibiotics and an alternative method of replacing antibiotics with an iron complex together with an antibiotic. The experimental part is demonstrating the antibacterial effect of the Fe16+AMP complex on the ampicillin-resistant bacterial strain E. coli CCM 4225 and the sensitive strain E. coli CCM 3953. The minimum inhibitory concentration was experimentally determined for these samples. Morphological changes upon exposure of the Fe16+AMP complex to E. coli CCM 3953 were also observed using a cryo-SEM scanning electron microscope. The development of resistance to the model bacterium E. coli TOP10 by transformation was also investigated and verified by polymerase chain reaction PCR and gel electrophoresis. Based on the results, it was proven that the Fe16+AMP complex has an inhibitory effect on the bacterium E. coli CCM 3953 and does not have a significant effect on the resistance spreading via transformation mechanism.
|
guest ::
