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The detection of factors virulence and resistance genes bacterial pathogens in patients with cystic fibrosis
Jandová, Oldřiška ; Melter, Oto (advisor) ; Nyč, Otakar (referee)
Cystic fibrosis is an autosomal recessive genetic disorder, which is caused by mutation in CFTR gene (Cystic Fibrosis Transmembrane Conductance Regulator). This gene encodes protein with the same name, which is responsible for pathogenesis of CF. Cystic fibrosis is characteristic for frequent infection of respiratory system, which causes, destruction of lung tissue. These infections are characterized by occurrence of typical bacterial pathogenes, for example: S. aureus, P. aeruginosa etc. S. aureus is one of the most typical opportunistic pathogens, which causes serious difficulties in patients with the cystic fibrosis. Strains of S. aureus are characterized by production of multiple virulence factors and resistance to broad spectrum of antibiotics. Besides common mechanisms of resistance there is also possibility of emergence of so called Small Colony Variants in chronically infected patients. These resistant subpopulation is relatively common among S. aureus isolates of patients with CF. The aim of this work was to study isolates from three patients with cystic fibrosis, who are chronically infected by S. aureus. Our goal was to determine changes in the pattern of the antibiotic resistance and occurrence of virulence factors together with description of SCV strains. Powered by TCPDF (www.tcpdf.org)
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Bacterial REP elements: origins, variability and application.
Nunvář, Jaroslav ; Lichá, Irena (advisor) ; Pačes, Jan (referee) ; Melter, Oto (referee)
4 ABSTRACT (English) This thesis is based on three published research papers studying bacterial REP (repetitive extragenic palindrome) elements. REP elements are one of the best-characterized groups of bacterial DNA repeats, distributed mostly in gammaproteobacteria, including enterobacteria. They are present in noncoding parts of host genomes, usually occurring in hundreds of copies. REPs are typically aggregated in higher order repeats. In the Gram-negative model Escherichia coli, interactions of several proteins important for cell's physiology with REPs were described, indicating significant role for these elements for host cells. The first work (Nunvar et al. 2010) presents the discovery of a protein class, related to IS200/IS605 transposases. These proteins, termed RAYTs (REP-associated tyrosine transposases), contain characteristic motifs in their amino acid sequences, which are absent in canonical IS200/IS605 transposases. Another attribute of RAYTs is the arrangement of their encoding genes. These are single copy genes, always flanked at both termini by at least two REPs in inverted orientation. Based on the similarity between the REP-rayt-REP unit and insertion sequences of the IS200/IS605 family, between RAYTs and tyrosine transposases and between REPs and subterminal sequences of the IS200/IS605...
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Introduction of microbiological diagnostic agents of invasive infections - Bartonella henselae and Bartonella quintana
Nováková, Marie ; Melter, Oto (advisor) ; Chrenková, Vanda (referee)
Bartonellae are agents of infections with varied clinical symptomalogy, including serious, such as endocarditis. Direct detection of Bartonella so far in the Czech Republic and therefore does not need to address this issue. Proper diagnosis of these infections is a prerequisite for targeted antibiotic therapy for patients. The genus Bartonella today are more than 20 kinds of bacteria. They are small Gram-negative rod-shaped bacteria that cause the new emerging infections such as cat scratch disease (CSD), showing the typical regional lymphadenitis, as well as bacillary angiomatosis (BA), whose typical symptoms are skin and subcutaneous nodules (nodules). They also cause a disease called trench fever or South American bartonelosis. In the experimental part of the work was performed genotypic and phenotypic methods for the diagnosis of these bacteria, specifically on two types of Bartonella, Bartonella henselae and Bartonella quintana . The results show that by suitable methods to diagnose Bartonella are amplification genus- specific genes ribC and ftsZ, further embodiment of a method pulsed electrophoresis (PFGE) as typing method and the determination of susceptibility to antibiotics, the most appropriate testing showed E-test. This work was carried out optimization of genotypic and phenotypic...
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The introduction of diagnosis of new bacterial pathogens Inquilinus and Pandoraea isolated from patients with cystic fibrosis and determining their sensitivity to antibiotics
Šulcová, Romana ; Melter, Oto (advisor) ; Nyč, Otakar (referee)
Cystic fibrosis is one of the most common autosomal recessive hereditary disease. Nowadays, people, with this disease, because early diagnosis and treatment of other associated symptoms a better prognosis than a few years ago. CF is a disease that is subject to mutation of the CFTR (Cystic Fibrosis Transmembrane Conductance Regulator) on the seventh chromosome. Defect in the CFTR protein leads to failure of resorption and secretion of electrolytes chloride channel and thereby creating viscous mucus in the lungs, which is the main cause of chronic bacterial infections of the lower airways. Among the most common pathogens causing severe respiratory infections in CF patients include: Staphylococcus aureus, Pseudomonas aeruginosa, Burkholderia cepacia complex. In patients with CF is now showing in the airways of new bacterial species whose biological properties are not yet known and their role in the etiopathogenesis of the disease and epidemiology are not exactly known. In these days their detection is used by specific amplification methods, or sequencing of the 16S rRNA gene. Emerging pathogens such as the genus Pandoraea and Inguilinus that belong non-fermenting Gram-negative groups in the rods, and therefore are often exchange for other bacterial species that are phenotypically similar to them. In the...
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The introduction of diagnosis of new bacterial pathogens Ralstonia a Achromobacter isolated from patients with cystic fibrosis and determining their sensitivity to antibiotics
Michálková, Alice ; Melter, Oto (advisor) ; Bébrová, Eliška (referee)
Cystic fibrosis (mucoviscidosis) is an incurable genetic disease caused by mutations in the CFTR (Cystic Fibrosis Transmembrane Conductance Regulator Gene). The most common causes of increased mortality and morbidity of patients include bacterial respiratory infections which may occur even due to less frequent pathogens. Bacterial species of the Ralstonia and Achromobacter genera are not considered pathogenic for healthy people, but they have been established as pathogens in the sputum of patients with CF. However, due to their phenotypic similarity to other bacterial pathogens encountered in patients with CF, microbiologists often do not pay attention to them. The aim of this thesis was to propose some methods of identification of the genus Ralstonia, to determine both quantitative and qualitative susceptibility towards antibiotics, and to make a bibliographical search focused on the issue of the Achromobacter genus. Strains of Ralstonia spp. were identified using phenotypic and genotypic methods and were tested for the susceptibility towards antibiotics. The thesis also proposes a new method of genotypic indentification of R. respiraculi.
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Study of biofilm Pseudomonas aeruginosa and introduction of new method for antibiotics susceptibility
Mošničková, Markéta ; Melter, Oto (advisor) ; Chrenková, Vanda (referee)
Biofilm is a complex structure produced by bacterial cells which contain highly hydratated filamentous exopolysacharides. Biofilm exists everywhere in our surroundings or nature. It creates for instance a sticky viscous structure covering the surface of stones or other solid objects underwater. The technology of sewage cleaning remediation is essentially dependent on the presence and function of biofilm producing bacteria. Adhered biofilm in medicine has usually negative consequences for patient health, hence, reduction of its role is a goal of researches in the field. Here biofilm first of all adheres and colonizes catheters or artificial joints or other implants and is a source of a chronic infection which limits the healing process or normal function of the implants. In cytic fibrosis patient biofilm production is also responsible for the fact that acute infection caused by P. aeruginosa could change within a few months in chronic infection when mucoid strain could be cultivated from the patient. In my bachelor thesis I have analyzed clinical material and bacterial isolate of P. aeruginosa of a cystic fibrosis patient which dissociated into mucoid and smooth phenotype using light-, electron- and immunofluorescence microscopy. I have applied an original method using resuspended bacterial culture in solid...
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Mechanisms of MLSB resistance in Staphylococcus aureus in patients with cystic fibrosis
Bukáčková, Eva ; Melter, Oto (advisor) ; Matyska Lišková, Petra (referee)
The aim of this thesis is to summarize the mechanisms of resistance of Staphylococcus aureus to MLSB antibiotics (macrolides, lincosamides and streptogramins B type) which are used to treat respiratory infections in cystic fibrosis patients. This pathogen evolved during time many various strategies of resistance to these proteosynthesis inhibitors. The most common mechanisms are target site modification, modification of the antibiotic itself or antibiotic eflux out of the bacterial cell. Apart from these mechanisms based on acquisition of genes, a mutation of specific genes can also result in resistance of the strain. In the lungs of CF patients, long-term antibiotic treatment together with immune system defects result in development of a unique niche. It is colonized (besides other bacteria) by S. aureus, which is well adapted to this environment and also uses different mechanisms of resistance as hypermutation or switching to dwarf phenotype (small colony variants) enabling intracellular persistence. MLSB antibiotics as well as beta-lactams are being applied as the treatment of choice for respiratory infections in CF patients. Studying the mechanisms of MLSB resistance is therefore of extraordinary importance.
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Study of persistence phenomenon at Staphylococcus aureus
Kubištová, Lucie ; Lichá, Irena (advisor) ; Melter, Oto (referee)
Staphylococcus aureus is human pathogen and is causative factor of many diseases with different character, from light infections of the skin to life-threatening sepsis. S. aureus often colonizes cystic fibrosis patient's lungs and causes long-standing pneumonia, which can cause death in these patients. Infection caused by S. aureus are, as most of the infections, treated with antibiotics. Failure in treatment is caused because of presence of resistant strains. It was shown, that treatment failure can be caused by different type of the antibiotic tolerance - by the persistence. Persisters are phenotypic variants of isogenic bacterial population, that is unaffected by antibiotic treatment even though they don't have genetics determinants of the resistance. The phenomenon of the persistence in bacteria is still relatively poorly understood. The aim of my thesis was to contribute to the characterization of the persistence in S. aureus. We have optimized a method for rapid identification of persisters' amount in the bacterial population after treatment with antibiotics using the method of measuring the killing curves. We have found out, that exposure to osmotic stress causes increase in number of persistence in the bacterial population by 1 - 2,5 order. Using quantitative PCR method we have analyzed the...
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