|
|
Expression and purification of N-terminal fragment of filamentous hemagglutinin from Bordetella Pertussis in E. Coli
Jurnečka, David ; Stiborová, Marie (advisor) ; Kavan, Daniel (referee)
: Whooping cough is highly contagious disease caused by gram-negative bacteria Bordetella pertussis. During infection the bacteria produces many types of toxins and adhesive molecules including a filamentous hemagglutinin(FHA). FHA is 220 kDa surface-exposed and secreted protein, which plays a key role in host-cell interactions. The project aims at construction of heterologous expression system for production of the N-terminal part of B. pertussis FHA (FHA1-862) in E. coli. The expression vector is composed of system of two independent T7/Lac promoters and enables secretion of FHA1-862 into the culture media. Downstream of the first promoter is fhaB gene encoding FHA1-862 and the letter is followed by fhaC gen encoding the FhaC transport protein, which allows translocation of FHA from periplasmic space to extracellular milieu. FHA1-862 was successfully secreted in E. coli strain BL21 carrying plasmid pMM100 (Laclq) at 30 řC and purified by affinity chromatography on Cellufine resin. These results indicate that FHA1-862 protein can be produced in E. coli, however, the system is inefficient and the yield of the protein is very low. (In Czech)
|
|
|
The effect of sampling point on hygienically significant water indicators in natural bathing establishment
Zelinková, Myra ; Benešová, Libuše (advisor) ; Popovský, Jiří (referee)
In this diploma thesis, I deal with the influence of the sampling site on water quality in natural bathing lakes (Hostivar reservoir, Seberak pond and Vyzlovka pond) in this. I have established that hygienically significant kinds of phytoplankton may differ in terms of water quality especially where there are cyanobacteria constituting water bloom. Surface water bloom may be moved by wind to the lee side of the water body (Microcystis sp.). Aphanizomenon flos-aquae water bloom floating in water column can be affected by wind and by water flow (which may be partly affected by wind as well). With the prevailing fibrous Planktothrix agardhii which does not connstitute water bloom the concentration of chlorophyll-a and cyanobacterial cell abundance in individual sampling sites are similar although the concentration of chlorophyll-a a is about 200 g.l-1 . Microcystis sp. and Scenedesmus sp. survive under eutrophic conditions in competition. From the microbiological perspective, water quality can differ within a single sampling site in places at a distance of less than 100 m. Microbial contamination can be caused by bathers, water birds, farm animals and probably by the removing of microorganisms from sand, mud and sediments on the beach or shore. Rain episodes probably increase the abundance of E.coli...
|
|
|
Extending of shelf life of chilled fish products
PFLUG, Róbert
This diploma thesis was focused on the possibilities of extending the shelf-life of fish products by dipping containing seven commercial additives. ANTIBAK, MIC STAB, Bakont, SEA-F75, Misocarine LR, SAFE A Plus and AMX liquid. The effectiveness of these substances on the extending of shelf-life was evaluated on the basis of tests of TVC (total viable count), level of fat and protein oxidation, determination of nutritional parameters of muscle, and finaly sensory analysis. Experimental species were 2 important commodities for the Czech aquaculture rainbow trout (Oncorhynchus mykiss) and common carp (Cyprinus carpio). The project was divided into 6 sub-stages. During testing was the initial number of aditives (7) limited by stepwise selection for 2 preparations. Namely Bakont and AMX liquid, which were tested further on trout (filet with skin and scales) and carp (filets with skin without scales) chilled bulk and chilled packaged under vakuum. For trout was studied antimicrobial effect of dipping on eviscerated fish with the head with- or without gills. Treated trout fillets in bulk and vakuum-packed showed significantly less abundance of muscle mikroflora. In the case of carp fillets chilled bulk we can not say that the bath had influence on the CMP in meat. However, the combination of dipping and vakuum packaging was singnificantly different between the control and product Bakont. AMX liquid was not applied in this case in sufficient dose or in sufficient time to carp muscle. A positive finding is that the application of the aditives to the product "eviscerated trout with head" it does not matter, whether the gills are left in fish or not. However, in all cases the analysis of the presence of pathogens Escherichia coli, Salmonella spp. or Listeria monocytogenes were negative. From the results of sensory analysis can be concluded, that the substances contained in aditives are not reflected in the sensory properties of tested fish.
|
|
|
Potenciální antimikrobiální účinky vybraných přírodních látek
MAŠKOVÁ, Hana
This study was focused on potential antiviral and antibacterial effects of viniferin and some derivatives of adamantane. The most suitable concentration was determined using flow cytometer. The antiviral effect on tick-borne encephalitis virus infection in glioblastoma cells was determined. Virus concentration was determined using plaque assay. The antibacterial effect was detected by the incubation of bacterial strains with suitable concentration of selected natural substances. The rate of bacterial growth was determined using absorbance measurement at 600 nm and plating on agar plates.
|
|
|
Analýza kvality pitné vody v jižních Čechách
VRÁNEK, Ivo
The goal of diploma thesis was a comparison of potable water indicators in representative areas of the south Bohemia. Monitored period had been from January 2012 till December 2014. Monitored parameters were microbiological indicators (Escherichia coli, coliform microbes, number of colonies at 22°C, number of colonies at 36°C, organoleptic indicators (color, turbidity, smell, taste). Physical and chemical indicators (ammonium ions, nitrates, nitrites, chemical consumption of oxygen, chlorides, conductivity, manganese, pH, sulphates, calcium and magnesium, iron). From the total complex of 1165 data, there were 23 values over the limit (1,97%) of evaluation of all monitored quality indicators (microbiological, organoleptic, physical and chemical). In evaluation of microbiological indicators, there were 6 values (1,97%) over the limit, from total 304 data. In organoleptic indicators, there was 1 value (0,44%) over the limit, from total 227 data. In evaluation of physical and chemical indicators, there were 16 values (2,52%) over the limit, from total 634 data. In total, there are 23 values which don´t meet determined limits from The Regulation No. 252/2004 and EU directives. There are these values of limits - number of colonies at 22°C, number of colonies at 36°C, turbidity, manganese, calcium and iron.
|
|
|
Electrophotocatalytic wather disinfection
Štefancová, Eva ; Drbohlavová, Jana (referee) ; Veselá, Mária (advisor)
This work focuses on the photoelectrocatalytic water disinfection by applying electrical voltage onto an electrode system. Working electrode consist of electrical conductor covered by thin film TiO2, because of its very important photoactivity. Disinfection effect was monitored on Gram-negative bacteria E. coli under different conditions. Inactivation of bacterial cells in contamined aqueous solution, first experiment was observed only with presence of oxygen, later reactor was irradiated with UV-A compact blacklight with intensity of 4 mW• m-2 or electrical voltage of 1V was applied to electrode. By adding both effects at the same time I was verifying photoelectrocatalysic water disinfection on chosen microorganism.
|
|
|
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.
|
|
|
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.
|
|
|
Study of elementary inactivation processes acting during sterilization of procaryotic systems in dielectric barrier discharge
Bittnerová, Zuzana ; Sťahel, Pavel (referee) ; Kozáková, Zdenka (advisor)
Presented master’s thesis is focused on the study of the elementary inactivation processes acting during sterilization of procaryotic systems in dielectric barrier discharge (DBD). Sterilization is an important biomedical and food-industry application and plasma sterilization is one of the methods, which are suitable for sterilization of heat and chemical sensitive materials. Biologically contaminated samples were treated in dielectric barrier discharge operated at atmospheric pressure. The discharge was generated in argon and in nitrogen. The plasma power density was 2725,93 mW cm–3 in argon and 2325,93 mW cm–3 in nitrogen. Gram positive bacteria Bacillus subtilis and gram negative bacteria Escherichia coli were used as a bioindicator. Bacteria were spread onto the surface of Whatman No.1 filtration paper. The influence of UV radiation, reactive species, heat and plasma discharge where the synergistic function of all of the agents was studied. Effects of UV radiation and temperature were studied separately. In order to separate the effect of UV radiation generated by DBD the quartz glass transmitting UV radiation was employed. During the plasma exposition selected samples were covered with the quartz window while other samples were directly exposed to the plasma. Covered samples were exposed to UV radiation and temperature (which cannot be eliminated), samples without quartz window were directly plasma exposed (treated). Results show that for covered samples the lower inactivation was reached than by the samples directly exposed to plasma. When studying the effect of temperature, the temperature between the DBD electrodes was measured by means of a thermocouple. Afterwards the samples were placed in an oven and exposed to the same temperature as was measured between the electrodes. By comparing the results of heat treated samples and plasma treated samples it can be assumed that the influence of the temperature during the sterilization process in DBD is very low. The discharge parameters were studied by means of the Optical Emission Spectroscopy. Plasma treated samples were assessed employing Scanning Electron Microscopy (SEM). Damage of Bacillus subtilis cell wall due to the effect of plasma was observed while no effect of plasma on the structure of filtration paper was detected.
|
guest ::
