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Network server performance when communicating with a great number of clients
Mašín, Jan ; Pelka, Tomáš (referee) ; Komosný, Dan (advisor)
The aim is to create a simulation system between two computers of the server-client type to generace the process to server while usany UDP protocol: to set different parameters of the generated process (size of paquet, interval of sending) and evaluace the server-sources consumption (memory), to work out the outcomes in a well-organized manner. There have been tested two servers (each of different output) and operational systéme Windows XP, Windows Server 2003, Linux Mandriva 8.0 and Linux Fedora 7, each of these had been installed on the servers with different outputs. The generated process is carried out with usaje of the paquet generator made in the programming language C++.Two different programmes needed have been created, one of them for klient(paquet generation in OS Windows), its aim is to generate the paquets, the sekond on efor server (functional version working in OS Linux had had to be made) receiving the generated paquets. Evaulation of the source-consumption was carried out by two monitoring programs, one of them suited for OS Windows, the other for OS Linux. Both programs safe the information about procesor-work and operational memory into a text dokument which is very simple and fast way for searching the information. The measured values show charts and graphs that express the efficiency of separate OSs or, in case of need, servers of different outputs are able to handle the generated paquets and how do the alterations of the paquet generator affect the source efficiency( paquet size and paquet-sending interval).
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Localization of nodes in Internet using Vivaldi system with adaptive time step
Mašín, Jan ; Balej, Jiří (referee) ; Komosný, Dan (advisor)
The aim of this thesis was to identification with the principles of logical evaluation of the position of stations on the Internet. Read up on the localization algorithm called Vivaldi with adaptive time step and subsequently to its implementation in the operating system GNU/Linux CentOS distribution. Do one's homework the PlanetLab experimental network (http://www.planet-lab.org/). At selected stations from the network transfer created by the application and verify its function on the real servers located at various places around the globe and assess the accuracy achieved by estimating the distance between stations on the PlanetLab network. In this scope of activity, the application was created to measure the delay prediction using Vivaldi algorithm with adaptive time step which is on principle of operation a client-server where the client performs the steps of the algorithm, Vivaldi and the server only listens, collects the resulting data Vivaldi algorithm and stores them neatly file. Furthermore, the application was developed for direct measurement of the delay, which also functions as a client-server. These applications have been transferred to the selected nodes from the PlanetLab experimental network. Subsequently, these nodes were running, to carry out the necessary measurements. The resulting values were work into tables of using Microsoft Excel. These values were then compared with direct measurements and competitive positioning by the King. Vivaldi localization methods with adaptive time step and the King, were compared based on calculated estimates of both real estate errors and measurement using distribution function of the relative errors of both methods. All this information was evaluated to compare accuracy of both the localization methods and direct measurements.
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Identification of residues of acylated domain of RTX toxins involved in acyltransferase binding
Grobarčíková, Michaela ; Mašín, Jiří (advisor) ; Černý, Ondřej (referee)
Both adenylate cyclase toxin (CyaA) and α-hemolysin (HlyA) are members of Repeats in ToXins (RTX) cytolysins that play key roles in the virulence of Bordetella pertussis and Escherichia coli, respectively. Bacterial RTX toxins represent a growing group of proteins produced by gram- negative bacteria. These pore-forming RTX toxins share several notable common features: (1) they require post-translational activation by attachment of fatty acid chains to two lysine residues; (2) they contain a hydrophobic domain that forms cation-selective pores in target cell membranes; (3) they are secreted by a type I secretion system; (4) after secretion, they become biologically active by binding of Ca2+ to the nonapeptide glycine- and aspartate-rich repeats. CyaA translocates a unique AC enzyme to the cytosol of phagocytes and subverts their bactericidal functions by unregulated conversion of ATP to cAMP. CyaA and HlyA also permeabilize the cell membrane of eukaryotic cells through cation-selective pores. Both toxins preferentially bind to cells expressing β2 integrins but can also interact with a variety of cells that do not express integrins or with naked lipid membranes. Both toxins are activated from protoxin form by post- translational acylation mediated by a specific acyltransferase. CyaA is activated by...
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Application of human monocytic cell line THP-1 for study of pathogenesis in whooping cough agent Bordetella pertussis
Čurnová, Ivana ; Petráčková, Denisa (advisor) ; Mašín, Jiří (referee)
Bordetella pertussis is strictly human pathogen that causes severe infection of the respiratory tract known as whooping cough, which is currently on the rise. B. pertussis was considered as an extracellular pathogen for a very long time. Recently it was shown the ability of B. pertussis to survive inside early endosomes of macrophages. This ability is studied in the human monocytic cell line THP-1 and also in primary macrophages from human donors. This diploma thesis is focused on THP-1 infectious model and mainly for the early phase of infection. A previously performed transcriptomic study showed significantly affected genes of B. pertussis during intracellular survival in THP-1 macrophages. In this study, we selected genes that are in some way related to intracellular survival inside human macrophages or have significantly effect for intracellular survival. The effect of the mutation in these genes was tested both on the level of cytotoxicity to THP-1 cells and the related number of surviving bacteria inside the macrophages. The deletion strain in two genes for cysteine dioxygenase (BP2871 and BP3011) and the mutant strain allocated in the Bvg+ phase were less cytotoxic than the control strain. Monitoring the effect of opsonization to intracellular survival have not such clear results. The effect...
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Relationship between the structure of synthetic organic molecules, their behavior in the phospholipid membrane and their effect on bacteria
Brzobohatá, Hana ; Fišer, Radovan (advisor) ; Mašín, Jiří (referee)
Lipophoshopnoxins (LPPOs) are organic compounds with antimicrobial activity, which take place on the baterial membrane by forming pores. LPPOs are promising bactericidal molecules with low MIC (minimal inhibition concentration) and high HC50 (hemolytic concentration) values. Structurally, LPPOs belong to the so-called synthetic antimicrobial peptidomimetics, which do not contain any peptidic bonds, but imitate the structure and function of membrane active oligopeptides (e.g. by copying the distribution of the surface charge of the molecule). The aim of this study was to describe the antimicrobial properties of the molecules DR 7072, DR 34P1, DR 362 and DR 236, which represent LEGO-LPPOs as a new generation of this antimicrobials. The selected molecules differ in the length of linker module, in the structure of hydrophobic module or both. All tested molecules have comparable antimicrobial activity with MIC between 2 and 16 μg/ml and have the ability to permeabilize bacterial membrane of both Gram negatives (G- ; Pseudomonas aeruginosa CCM 3955) and Gram positives (G+ ; Staphylococcus aureus CCM 4223). LEGO-LPPO have different mechanism of action on S. aureus and P. aeruginosa. The molecules show different mechanism of action against the mentioned bacteria. The bactericidal effect on G+ is faster and...
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Mode of action of the 4th generation of antibacterial compounds lipophosphonoxins
Helusová, Michaela ; Mikušová, Gabriela (advisor) ; Mašín, Jiří (referee)
Lipophosphonoxins are small synthetic antimicrobials targeting the cytoplasmic membrane of bacteria. This thesis focuses on comparison of three lipophosphonoxins which differ in the number of carbons in its modules and in antimicrobial and hemolytic activity. The most promising candidate compound is lipophosphonoxin 7072 showing good antimicrobial activity as well as low hemolytic activity. Other two tested lipophosphonoxins are 7070 displaying high hemolytic activity and weakly antibacterial lipophosphonoxin 7107. The pore-forming activity of lipophosphonoxins is investigated using model membranes as well as living bacteria Staphylococcus aureus and Escherichia coli. The results show that small difference in structure can fundamentally affect the activity of these molecules. Lipophosphonoxins 7072 a 7070 display equal antibacterial activity against tested bacteria by forming pores in the bacterial membrane. Bacteria rapidly die of loss of membrane potential caused by lipophosphonoxins. The high hemolytic activity of the compound 7070 is probably related with its preference for uncharged membranes. The weak antimicrobial activity of 7107 is caused by its capability to form only small pores and its incapability to overcome and disrupt the outer membrane. Key words: antimicrobial agents,...
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The role of a conserved tyrosine residue of acylated domain in membrane insertion and penetration of RTX toxins
Lepesheva, Anna ; Mašín, Jiří (advisor) ; Petráčková, Denisa (referee)
Pore-forming RTX toxins are key virulence factors of many Gram-negative pathogens. These proteins share several common structural and functional features: (i) the presence of repetitive sequences rich in glycine and aspartate, which are important for calcium ion binding, (ii) transport from the bacterial cytosol through the type I secretion system (T1SS), (iii) modification by a fatty acid at specific lysines in the acylated domain by a specific acyltransferase, and (iv) the presence of an amphipathic region responsible for the formation of cation-selective pores in the target membrane. The aromatic side chain of the conserved tyrosine residue 940 in the acylated segment of the RTX adenylate cyclase toxin (CyaA, ACT or AC-Hly) of Bordetella pertussis plays a key role in the interaction of the toxin with the target cell membrane. The aim of this study was to determine whether the corresponding conserved residues Y940, Y642, Y643 and Y639 secreted by the homologous RTX toxin CyaA from Bordetella bronchiseptica, HlyA from Escherichia coli, ApxIA from Actinobacillus pleuropneumoniae and RtxA from Kingella kingae play the same critical role in membrane insertion and pore formation. The hemolytic and cytotoxic activities of these toxins were completely impaired only after replacement of the conserved...
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Quality control management of extracorporeal circulation with Data management system
Fichtl, Jaromír ; Rulíšek, Jan (advisor) ; Mašín, Jaroslav (referee)
During extracorporeal circulation are typical fluctuation of parameters of basic life functions.These variations of values can have a direct impact on postoperative recovery. May even lead to temporary or permanent damage to organ function. Mean arterial pressure is one of the basic unit of tissue perfusion. If there is a drop in mean arterial pressure below 50 mmHg, there may be organs hypoperfusion associated with desaturation of venous blood. In our study focused on gathering data in the management of cardiopulmonary bypass during cardiac procedures by using the data collection would Stocker De. We focus on course of the values of mean arterial pressure and actual venous blood oxygen saturation. The group included 45 patients meet the criteria: valve surgery, surgery is an operating room No.3, surgery in normothermia. In 22 patients were recorded during cardiopulmonary bypass hypotension for more than four minutes ( Group A). In the remaining 23 patients the episode of hypotension has not been reported (Group B). Both groups were compared. We compared the average time of hospitalization, the average time of intubation and the incidence of neurological complications. The average time of hospitalization was in Group A - 180 hours, Group B - 210 hours. The average time of intubation was in Group A - 114...
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Adenylate cyclase toxin of bacteria Bordetella pertussis: mechanism of potassium efflux from macrophages
Pospíšilová, Eva ; Mašín, Jiří (advisor) ; Konopásek, Ivo (referee)
The adenylate cyclase toxin-hemolysin (ACT or CyaA) is a key virulence factor of Bordetella pertussis, the agent of the human respiratory disease whooping cough. CyaA penetrates phagocytes expressing the CD11b/CD18 integrin and exhibits two different activities. One toxin conformer oligomerizes in cell membrane and permeabilizes it by forming small cation-selective pores. Another toxin conformer appears to act as monomer. It forms a calcium influx path in the membrane, concomitantly with translocating into cells the adenylate cyclase enzyme domain that binds calmodulin and catalyzes unregulated conversion of cytosolic ATP into cAMP. We show here that CyaA causes efflux of K+ from CD11b+ cells by a mechanism that requires binding of CyaA to integrin CD11b/CD18 and permeabilization of the cellular membrane by pore-forming conformer of CyaA. Intact CyaA and the enzymatically inactive CyaA-AC- toxoid unable to generate cAMP produced the same kinetics of K+ efflux showing that elevation or signaling of cAMP had no role in this activity. The truncated CyaA variant (CyaA-AC) devoid of its invasive AC domain failed to promote K+ efflux despite a normal pore forming activity on erythrocyte and artificial membranes. However, binding of the MAb 3D1, which recognizes the distal segment of the AC domain...
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