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Growth and control of Pseudomonas aeruginosa in a multi-species biofilm
Hutlas, Andrej ; Kučerová, Marta (advisor) ; Konečná, Klára (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Cardiff University, School of Pharmacy and Pharmaceutical Sciences Study program: Pharmacy Candidate: Andrej Hutlas Consultants: PharmDr. Marta Kučerová, Ph.D. (Charles); Prof. Jean-Yves Maillard (Cardiff); BSc. Katarzyna Ledwoch Ph.D. (Cardiff) Biofilms are a default mode of growth for most bacteria. Microbes encapsulate themselves within a matrix, composed mainly of extracellular polymeric substances. A biofilm can be composed by multiple species. Matrix environment induces various physiological shifts, such as switch to dormant state or expression of biofilm-specific genes. Mature biofilm's heterogeneous, due to differences in spatial microbe distribution and spatial nutrient utilization across the matrix. Matrix provides many advantages to bacteria, like nutrient capture and transfer or protection against extreme conditions. Washbasin U-bend multi-species biofilms were investigated with a special focus on Pseudomonas aeruginosa. This opportunistic pathogen causes nosocomial infections, mainly in immunocompromised patients, with considerable health and socio-economic impacts. A link between sinks and environmental P. aeruginosa contamination has been established. Environmental decontamination may be an approach to lessen aforementioned...
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Advanced methods of antimicrobial activity screening
Dolinská, Dagmar ; Janďourek, Ondřej (advisor) ; Konečná, Klára (referee)
Charles University, Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Candidate: Dagmar Dolinská Supervisor: PharmDr. Ondřej Janďourek, Ph.D. Work title: Advanced methods for testing antimicrobial activity Goal: The aim of the work in the theoretical part was to summarize methods of antimicrobial screening focusing on antimycotics and in the practical part to evaluate the antifungal activity of 47 tested substances using the microdilution broth method. Methods: The substances were tested using quantitative microdilution broth method on eight strains of yeast and fibrous fungi. These were Candida albicans, Candida krusei, Candida parapsilosis, Candida tropicalis, Aspergillus fumigatus, Aspergillus flavus, Absidia corymbifera and Trichophyton interdigitale. Results: The highest antimycotic activity was demonstrated by compound Ac-0a, where we recorded after 24 hours of incubation activity against all eight strains. However, it was not able to stop the growth of three strains after 48 hours of incubation, the strains concerned were Candida parapsilosis, Aspergillus fumigatus and Aspergillus flavus. Other samples which have shown activity were Cl-2, where activity was observed for Candida krusei, Aspergillus fumigatus and Trichophyton interdigitale strains, Cl-3 for...
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Phylogenetic diversity of culturable prokaryotes from hypersaline environments.
Straková, Dáša ; Jílek, Petr (advisor) ; Konečná, Klára (referee)
Candidate: Dáša Straková Title of diploma thesis: Phylogenetic diversity of culturable prokaryotes from hypersaline environments. Charles University, Faculty of Pharmacy Department of Biological and Medical Sciences University of Sevilla, Faculty of Pharmacy Department of Microbiology and Parasitology Study program: Pharmacy Hypersaline environments are widely distributed habitats characterized by elevated concentrations of salts, mainly represented by aquatic systems and hypersaline soils. Their study has enabled the isolation and characterization of a large number of halophilic microorganisms during the last decades, which are microorganisms that require a high concentration of salts in order to be capable of life and growth. The aim of this work was to continue the initial characterization of halophilic Archaea and Bacteria, previously isolated by Ana Durán-Viseras in 2016 from several water samples from Isla Cristina and Isla Bacuta solar salterns (Huelva, Spain) and from a hypersaline soil located in Odiel Saltmarshes (Huelva, Spain), and to carry out a biodiversity study of these samples. Ana Durán-Viseras started this project and I continued with it during my stay in Sevilla. As a part of the characterization, we sequenced the partial 16S rRNA gene of the isolated strains. For those that...
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Galleria mellonella in vivo model for the study of antimicrobial drug vs. pathogen interaction
Polová, Dominika ; Konečná, Klára (advisor) ; Kudláčková, Zděnka (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Study program: Healthcare Bioanalytics Author: Bc. Dominika Polová Supervisor: RNDr. Klára Konečná, Ph.D. Title of thesis: Galleria mellonella in vivo model for the study of antimicrobial drug vs. pathogen interaction Background: This study focuses on establishment of the in vivo invertebrate mo- del Galleria mellonella primarily for the study of in vivo efficacy and toxicity of newly synthetized compounds with proved antiinfective potential. Major part of the study is devoted to optimization of conditions for laboratory insect rearing, determination of lethal infectious doses after infection of Galleria mellonella larvae by methicillin re- sistant Staphylococcus aureus, implementation of methodical approach for assessing the relative acute toxicity of candidate antiinfective compounds, and establishment of laboratory manuals for routine histopathological studies. Methods: In the study Galleria mellonella larvae were reared in stable laboratory con- ditions (37 ◦ C) and in the presence of rich nutrition. Sowing method was used in or- der to obtain infectious LD50. In the preliminary study we focused on acute toxicity of tested compounds and on methodology optimization leading to histopathological...
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In vitro screening of novel potentially active antimycotic compounds II
Comová, Kateřina ; Janďourek, Ondřej (advisor) ; Konečná, Klára (referee)
Charles University Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Study program: Specialist in Laboratory Methods Candidate: Bc. Kateřina Comová Supervisor: PharmDr. Ondřej Janďourek, Ph.D. Title of diploma thesis: In vitro screening of novel potentially active antimycotic compounds II Key words: yeasts, fungi, antimycotics, resistance, microdilution broth metod, minimal inhibition concentration. The main focus of this thesis is on antifungal resistance and the mechanism of action of selected antifungal agents. The theoretical part of the thesis gives a general overview of the characteristics of microscopic fungi and the diseases they cause. Attention is also paid to antifungal agents that are commonly used in the treatment of fungal infections, as well as those that are currently at various stages of clinical trials. This ties in with the mechanism of action of these compounds, which is also described here. The theoretical part also summarizes the development and principles of antifungal resistance. The theoretical part concludes with the description of the methods used in the determination of in vitro susceptibility of fungal agents to antifungals. The practical part of the thesis describes the procedure of the quantitative broth microdilution method, which...
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Atopic dermatitis ańd progress in therapy
Krymová, Lucie ; Rathouská, Jana (advisor) ; Konečná, Klára (referee)
Atopic dermatitis is a multifactorial chronic inflammatory disease which is one of the most common skin diseases. The prevalence is relatively high and is still growing. In former times, atopic dermatitis was considered primarily a childhood disease but today, the prevalence is increasing in adult patients too. Atopic dermatitis is a chronic incurable disease with a significant negative impact on patients' lives. The negative impact on the quality of life often leads to a lack of adherence of these patients to the standard treatment procedure, which implies the need to combine different therapies in the field of complementary and alternative medicine. While there is still not enough data available to conclude that adjuvant therapies are effective in the conventional sense, there are already promising results that suggest that these therapeutic strategies might be an alternative to treating this disease. Although mild and moderate forms of disease can be well controlled using external treatments, such as emollients, topical corticosteroids, and local calcineurin inhibitors, the treatment of severe atopic dermatitis is still a major challenge. Systemic treatment with corticosteroids is a relatively effective treatment in many cases, but cannot be used for a long time due to numerous side effects. And...
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Overview of methodological approaches leading to qualitative anad quantitative microbial biofilm evaluation
Holmanová, Pavlína ; Konečná, Klára (advisor) ; Vejsová, Marcela (referee)
Charles Univerzity Faculty of Pharmacy in Hradec Králové Study program: Medical Laboratory Technician Author: Pavlína Holmanová Supervisor: RNDr. Klára Konečná, Ph.D. Title of bachelor thesis: Overview of methodological approaches leading to qualitative and quantitative microbial biofilm evaluation Background: The aim of this work was to compose an overview of methodical approaches leading to qualitative and quantitative evaluation of microbial biofilms. To optimize the methods leading to preparation of mounts with biofilm-forming microorganisms evaluated by epifluorescence or confocal laser scanning microscopy. Methods: Optimization of microbial biofilm staining procedures for subsequent evaluation by epifluorescence microscopy. Formation of microbial biofilms - cultivation of microorganisms in various cultivation media in a microtiter plate, preparation of solutions with fluorescent dyes. Observation of coloured, dispersed biofilms by fluorescence microscope. Results: Using fluorescence microscopy, optimal target concentrations of fluorescent dyes in various combinations were verified. The combination of Syto 9 (SYTO™ 9 Green Fluorescent Nucleic Acid Stain) and propidium iodide was confirmed to be effective for the determination of viability of biofilm-forming microbial cells. The combination of...
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Rotavirus as an agent of diarrheal diseases
Hampl, Tomáš ; Vejsová, Marcela (advisor) ; Konečná, Klára (referee)
Author: Tomáš Hampl Title: Rotavirus as an agent of diarrheal diseases Bachelor thesis Charles University in Prague, Faculty of Pharmacy, Hradec Králové Field of study: Medical Laboratory Background: This bachelor thesis aims at dealing with rotavirus issues, taxonomically classifying them, describing their structure, properties, origin and course of the diseases they cause. In the experimental part the thesis will deal with the epidemiology of rotavirus infections at the Faculty Hospital in Hradec Králové in the period between 2010-2017 and their evaluation. Main findings: Rotaviruses are very often the cause of serious and dangerous diarrhea in children up to 5 years of age, including infants and toddlers. According to the WHO, more than half a million children worldwide die each year, where children will get ill around 150 million. Therefore, early diagnosis and appropriate therapy are very important. An adult can also become infected, the course of the disease is much milder than children. Conclusions: Important prevention of the origin and spread of rotavirus infections requires careful personal hygiene, especially hand washing. Because infections are the most common in children's groups, disinfection of the environment, toys, furniture and other objects is necessary in case of spread of...
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Optimization of the method for sensitivity evaluation of biofilm-forming staphylococci against candidate antimicrobial compounds
Diepoltová, Adéla ; Konečná, Klára (advisor) ; Jílek, Petr (referee)
Charles University Faculty of Pharmacy in Hradec Králové Study program: Healthcare Bioanalytics Author: Bc. Adéla Diepoltová Supervisor: RNDr. Klára Konečná, Ph.D. Title of thesis: Optimization of the method for sensitivity evaluation of biofilm- forming staphylococci against candidate antimicrobial compounds Background: The aim of this thesis was to optimize approach for in vitro formation of staphylococcal biofilms on the pegs and on the wells of the 96-well panel as an analogous approach to commercially available Calgary Biofilm Device system. The aim of the Experiment 1 was to evaluate incubation conditions (such as impact of a growth medium, incubation mode, optical density of the starting bacterial inoculum and type of surface) leading to maximal biofilm formation of two biofilm producer strains with unknown biofilm phenotype and one staphylococcal strain known as strong biofilm producer. The most advisable conditions were used in incubation process of Experiment 2. This work should propose the approach leading to in vitro formation of the most voluminous staphylococcal biofilms exploitable for candidate drug antimicrobial activity testing. Methods: Spectrophotometric measurement of the crystal violet colour extracted from wells with fixed and stained Staphylococci to evaluate the ability to...
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Vaginal microbiota and its effect in pregnancy
Mašátová, Eva ; Vejsová, Marcela (advisor) ; Konečná, Klára (referee)
Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biological and Medical Sciences Author: Eva Mašátová Trainer: Mgr. Marcela Vejsová, Ph.D. Title: Vaginal microbiota and its effect in pregnancy This work was created to summarize the knowledge of the vaginal microbiota in general and also during the pregnancy. The work deals with the description of individual microorganisms naturally occurring in the vaginal environment and their occurrence in the population. Diagnostic methods and techniques used in gynecology for examination of the vaginal environment are also described. The last part describes the antimicrobial treatment and its changes during the pregnancy. The vaginal microbiota is a diverse ecosystem of every woman that changes during her lifetime with a variety of influences. During the pregnancy, the risk of serious complications, such as premature birth or infections, is significantly increased when the patient is colonized. If the patient is colonized, it is not possible to definitely determine whether it is a pathological condition or not. Representatives of the genus Streptococcus, Staphylococcus, Gardnerella, Candida, Ureaplasma, Mycoplasma and others are naturally present in the vaginal environment. The pathogen is securely proven mainly by the density of...
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