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Characterization and function of Factor C from the tick \kur{Ixodes ricinus}
HARTMANN, David
Factor C is a multi-domain serine protease which recognizes Gram-negative bacteria via binding to lipopolysaccharides and triggers hemolymph clotting cascade in the horseshoe crab. A closely related molecule was also found to be present in the genome of the tick Ixodes scapularis. In this work, the full sequence of Factor C ortholog from Ixodes ricinus (IrFC) was determined. IrFC is mainly expressed in tick hemocytes and the heavy chain of the activated molecules is present in tick hemolymph as confirmed by Western blotting with antibodies raised against recombinant fragments of IrFC. The function of the IrFC in tick innate immunity was assessed using its silencing by RNA interference.
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Production and functional characterization of tick salivary protease inhibitors
KOTÁL, Jan
Two cysteine and two serine protease inhibitors from a tick Ixodes ricinus saliva were overexpressed using a prokaryotic overexpression system and refolded to their native state. Both cysteine protease inhibitors were tested as potential antigens for an anti-tick vaccine showing no effect on tick feeding or reproduction. Various immunological methods were employed to test the potential immunomodulatory function of these proteins without success.
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Molecular characterization of NADPH oxidase in the gut of the tick \kur{Ixodes ricinus}
KUČERA, Matěj
This thesis focuses on characterization of one member of NADPH oxidases - dual oxidase (DUOX) which has been described as the main factor of epithelial immunity in the gut of model organism Drosophila melanogaster. We have identified an orthologous gene coding for DUOX in the tick I. ricinus and described its tissue expression profile. The DUOX is mainly expressed in the gut of unfed ticks and seems to be downregulated upon artificial microbial infection. A fragment of tick DUOX was prepared as recombinant protein and used for preparation of specific antibodies to be used for further characterization of the enzyme. Our main aim is to highlight the importance of tick DUOX producing the reactive oxygen species and their role in the defence against pathogenic organisms within the tick gut.
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Presence of tick-borne encephalitis in Pribram district with its level of awareness and general attitude to vaccination against it in selected populations
KUCHAŘOVÁ, Eliška
Central European tick-borne encephalitis is a disease belonging to the group of zoonosis, which is transmitted by the tick Ixodes ricinus. It is an acute neuroinfection caused by the RNA virus that can seriously endanger people?s lives. Tick-borne encephalitis is one of the diseases typical for its natural pocket. The most common way of transmission onto a human being is by the bite of an infected tick. However, there have been cases where the contagion took place by consuming unpasteurized milk or dairy products. Although it is a disease which can be easily prevented, vaccination is still very low in our country. If we do not press people to improve this situation, we can expect frequency of tick-borne encephalities will be increased.
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Preparation of recombinant inhibitor of serine proteases from the tick \kur{Ixodes ricinus}
VLNOVÁ, Ivana
Tick serine protease inhibitors could be important anti-tick vaccines targets because of their properties and functions. The aim of this work was to prepare recombinant inhibitor of serine proteases from the tick Ixodes ricinus in baculovirus expression system. Two tick saliva proteins of the serpine superfamily were selected for this purpose and transformed into plasmids. One recombinant protein was expressed in baculovirus expression system, purified and its biochemical analyses were done.
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Vaccine potentiality of cystatin from tick \kur{Ixodes ricinus}
HARCUBOVÁ, Adéla
Ticks belong to the ectoparasites which are dangerous for the human beings because of the transmission of bacterial, viral, and protozoan pathogens. The development of a vaccine against tick is very important. Cystatins play important role in tick digestion and they have an immunomodulation effects. For this reason cystatins are possible candidates for this vaccines. This thesis focuses on experimental mice vaccination with recombinant cystatin from Ixodes ricinus.
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The effect of tick´s serpin IRS-2 on dendritic cells activated by TLR4 ligand
POSPÍŠILOVÁ, Šárka
IRS-2 is the inhibitor of serine proteases from the Ixodes ricinus tick. My task in this thesis was to find out the effect of the IRS-2 on dendritic cells activated by TLR4 ligand or by Borrelia afzelii. This effect was studied on several levels. I focused on the cytokine production, the expression of costimulatory molecules and cell signaling pathways. The results show that the IRS-2 may inhibit the expression of costimulatory molecules CD-80 a CD-86 on the cell surface, but this finding needs to be confirmed again. The production of cytokines was not affected by the IRS-2. The effect of the IRS-2 on the activity of p38, Erk1/2 nor NF-?B in LPS stimulated cells vas not observed. The fosforylation of STAT 3 in cells activated by the B. afzelii was lowered by the IRS-2.
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Identification and characterization of newly found antimicrobial peptide (IRAMP) from hard tick \kur{Ixodes ricinus}
OUŘEDNÍKOVÁ, Lucie
Antimicrobial proteins (AMPs) are effector molecules and an important part of the innate immune system. AMPs have a broad antimicrobial spectrum and lyse microbial cells by interaction with biomembranes. Besides direct impact in host defence, AMPs are mediators of inflammation with impact on epithelial and inflammatory cells influencing diverse processes such as cytokine release, cell proliferation, angiogenesis, wound healing, chemotaxis, immune induction, and protease-antiprotease balance. AMPs could replace antibiotics which efficiency has decreased due to extensive clinical use. Therefore knowledge of mechanism of action of antimicrobial peptides, their properties and possible usage is essential for their further use as therapeutics. Ticks are blood-feeding ectoparisites that serve as extremely effective vectors of pathogens. Analysis of the ticks molecules that are involved in immune response to the pathogens invasion represent one of the strategies in searching for new compounds that might be used in future as theurapeutic agents. This study represents analysis of newly identified antimicrobial peptide form in the hard tick Ixodes ricinus (IRAMP). IRAMP revealed the high similarity to the recently described antimicrobial peptide isolated from hard tick Ixodes scapularis (protein ISAMP). Analysis and characterization of novel AMP, testing its antimicrobial potential and expression pattern are the main objectives of this study
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