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Circulating tumor DNA in advanced colorectal carcinoma patients
Kopalová, Dominika ; Benešová, Lucie (advisor) ; Saláková, Martina (referee)
Colorectal carcinoma (CRC) is worldwide known as one of the most often tumor diseases. Approximately 50 % of patients are diagnosed with advanced stage of CRC and moreover even after removing all of the tumor bearings, there is high risk of its recurrence. As most of others solid tumors the CRC also releases fragments of DNA also known as the circulating tumor DNA (ctDNA). The ctDNA analysis is a new tool for monitoring continuous tumor burden and for observing treatment response. Due to its minimal invasivity and high specificity is suitable for instance for long period postoperative follow-up of patients with CRC. Within the long period follow-up process an evaluation radicality of operation is conducted as well as an early detection of new tumor foci. This thesis focus on an origin and releasing mechanism of ctDNA into bloodstream, its features and clinical utility options for patients with an advanced colorectal carcinoma. Key words: circulating tumor DNA, ctDNA, advanced colorectal carcinoma, mCRC, marker
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Bats - viral Pandora's box
Mejsnar, Martin ; Saláková, Martina (advisor) ; Kuthan, Martin (referee)
Bats, as a very old order of mammals, are significant reservoirs of viral pathogens and in many cases are also scientifically proven carriers of the viruses. These viruses are becoming more and more known, mainly due to the growing human population and thus the frequent contact of humans and bats. In this work I will follow up only some zoonoses that the Chiroptera family carries and I selected them according to their scientific knowledge and, of course, the impact on human society. However, this area is still full of information unclear or completely unknown, so further research is desirable. The basic methods of virus identification that are used to investigate new viruses are discussed in the second part. Key words: bats, virus, zoonosis, reservoir, epidemics
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Highly multiplexed virus detection in research of multifactorial diseases
Kunteová, Kateřina ; Cinek, Ondřej (advisor) ; Saláková, Martina (referee)
Next generation sequencing, which allows concurrent parallel sequencing of many samples and makes it possible to distinguish the infection from multiple viral types in the sample, is well suited as a detection format for such assays described below. The aim of the thesis was to develop a method that could detect all known types of human adenoviruses, human enteroviruses, and bacteriophages selected for their presence in the intestine. Using the next- generation sequencing. The first step was to design primers capable of detecting all known types of viruses, covering the area that is capable of distinguishing these viruses. This method was tested on a set of 47 human adenovirus samples and 30 human enterovirus samples of known serotype. Samples with two serotypes in different proportions were also created. After amplification of the target genome, the samples were purified and sequenced on MiSeq, Illumina. The method was further used in the typing of adenoviruses, enteroviruses and bacteriophages in pre-diabetic cohorts of DIPP, MIDIA, and a cohort of diabetics from African and Asian countries. The tested sample was RNA / DNA isolated from the stool specimen. We have demonstrated that the method is capable to detect all tested virus types, including infections with two different types, even if the...
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Diagnostics of Tick-borne Encephalitis: Present State and Future
Hustedová, Anna ; Španielová, Hana (advisor) ; Saláková, Martina (referee)
Tick-borne encephalitis is one of the most severe tick-borne neuroinfections in Europe and Asia. This thesis is focused on its source, Tick-borne encephalitis virus (TBEV). Diagnostics of TBEV is usually based on detection of specific anti-TBEV antibodies in patient's serum by using enzyme-linked immunosorbent assay. However antibodies that develop during the infection often cross-react with other flavivirus specific antibodies. As an antigen for ELISA tests the formalin inactivated virus is usually used, its production is restricted to laboratories with biosafety level 3. This thesis is seeking for possibilities of production of specific antigen for serological diagnostic of TBEV using heterologous expression systems. As the new antigens, parts of glycoprotein E and subviral particles were used. Production of antigen in the formo of subviral particles seems to be a good alternative to inactivated virus.
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The role of posttranslational modifications of minor proteins and acetylation of microtubules in mouse polyomavirus infection
Mariničová, Zuzana ; Horníková, Lenka (advisor) ; Saláková, Martina (referee)
Mouse polyomavirus (MPyV) capsid is composed of the main capsid protein VP1 and minor capsid proteins VP2 and VP3. Minor proteins are not essential capsid assembly, but they are key for efficient viral infection. The first part of this thesis studies the modifications of VP2 and VP3, the deamidation of Asn at 253 of VP2 (137 of VP3) and N-terminal acetylation of Ala of VP3, which could be the cause of double bands for VP2 and VP3 on SDS-PAGE. Mutated genomes of MPyV N253D (Asn to Asp) and N253E (Asn to Glu) simulating deamidation and A117V (Ala to Val) with reduced acetylation were prepared previously. We prepared three isolations of the mutant viruses and we confirmed that the deamidation is the cause of the double bands. Mutant viruses were compared to the wild type in terms of efficiency of infection, but the role of deamidation could not be proven. Virus A117V is noninfectious either due to lowered acetylation or the substitution of amino acid at this position. This thesis also studies the role of -tubulin acetylation in the infection of MPyV. The role of -tubulin acetylation in viral infection is being investigated to find new antiviral strategies. Acetylation rises after MPyV infection, but this is not due to a change in mRNA expression of tubulin acetylating (TAT1) or deacetylating enzyme...
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Minor Structural Proteins of Polyomaviruses: Attributes and Interactions with Cellular Structures
Vinšová, Barbora ; Horníková, Lenka (advisor) ; Saláková, Martina (referee)
Even though polyomaviruses have been intensively studied for more than 60 years, the role of minor structural proteins VP2 and VP3 in some important steps of viral life cycle has still not been fully elucidated, explicitly their role in viral genome delivery to the cell nucleus and their involvement in late phases of viral life cycle. This diploma thesis focuses on the study of minor proteins of Mouse polyomavirus (MPyV) and Human polyomavirus BK (BKV). Four rabbit polyclonal antibodies against minor proteins of polyomaviruses MPyV or BKV have been prepared within this diploma thesis. Two of these prepared antibodies target minor proteins of MPyV (α-MPyV VP2/3) or BKV virus (α-BKV VP2/3), other two prepared antibodies recognize C-terminal sequence common to minor proteins VP2 and VP3 of MPyV (α-MPyV C-termVP2/3) or BKV virus (α-BKV C-termVP2/3). In the second part of this diploma thesis we aimed to study toxicity of BKV virus minor proteins during individual production in mammalian cells. Obtained results suggest that minor proteins of BKV virus might not exhibit as high levels of cytotoxicity as minor proteins of MPyV virus. Third part of this diploma thesis is devoted to investigation of interactions of BKV and MPyV minor proteins with cellular proteins and within one another respectively....
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