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Web Application for Primer Selection for 16S rRNA Amplicon Sequencing
Jurča, Jan ; Martínek, Tomáš (referee) ; Smatana, Stanislav (advisor)
Main goal of this thesis was an implementation of web application, that can evalutate and recommend primer pairs for 16S rRNA amplicon sequencing, according to users specific needs and by this simplify the procces of primer pair selection. Primer pair evaluation is based on database of primer pairs, which contains data about senzitivity and specifity of each primer pair. Part of the work was also the performance optimization of the primer pair analysis algorithm, that computes senzitivity and specifity data. This optimization helped in an integration of algorithm into the application, which means that users can submit their own primer pair sequences, run analysis and get informations about position of amplified region, sensitivity and specifity.
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Variability in selected haematological traits related to gastrointestinal microbiota in parrots (Psittaciformes)
Dlugošová, Sylvie ; Vinkler, Michal (advisor) ; Volf, Jiří (referee)
Thousands of parrots all over the world suffer from illnesses and medical complications that can result from interactions between their immune system and bacteria in their digestive tract. The aim of this master's thesis is to understand the link between symptoms of these medical issues, the composition of blood and gastrointestinal microbiota in parrots. Using the hematological methods, 198 blood samples representing 53 parrot species were analyzed. The composition of microbiome was defined by combination of a molecular approach using bacterial 16S rRNA gene sequencing in 132 fecal samples, 12 intestine samples, 228 cloacal swabs and 236 beak swabs representing in total 61 parrot species and a diagnostic approach by psittacine fecal Gram's stain method. Significant association of hematological parameters with individual, environmental and clinical factors was observed, as well as its considerable interspecific variability. Absolute heterophile and lymphocyte counts have been shown more useful for infectious and autoimmune disease monitoring than H/L ratio. Relative numbers of basophiles were the best indicator for behavioral disorders. In relation to hematological parameters, the effect of the bacterial family Flavobacteriaceae, as part of the oral microbiota, and the bacteria Escherichia or...
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Taxonomic revision of the genera Anisus in the Czech Republic (Mollusca: Planorbidae)
Zavoral, Tomáš ; Juřičková, Lucie (advisor) ; Beran, Luboš (referee)
The aim of this work is to critically review the anatomical and morphological characters being currently used in the determination of Central European species of the genus Anisus and to confront them with molecular characters. For the molecular analysis mitochondrial genes for 16S rRNA and cytochrome c oxidase - subunit I (COI) were used. DNA analysis showed that known species occuring in the Czech Republic form well distinguishable genetic lines. Subsequent revisions of the anatomical characters of these lines have proven that these characters are due to their variability not suitable for determination, especially for the differentiating of the species A. spirorbis and A. leucostoma. The conchological characters have proven more suitable, especially the ratio of the size of the last and penultimate whorl. With the help of this character, we can safely determine a population within which there are transitional forms in other morphological and anatomical characters.
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Karyotype differentiation of Euscorpius scorpions (Scorpiones: Euscorpiidae) in Europe
Novotný, Tomáš ; Šťáhlavský, František (advisor) ; Ráb, Petr (referee)
The aim of presented work is to provide characteristics of the karyotypes of scorpions of the genus Euscorpius. Genus Euscorpius is a typical representative of scorpions in Europe. Its occurrence is wide throughout Europe. Until now, 18 species of this genus have been described. In this work six species were karyologically analyzed and one species was shown to possess only basic diploid number of chromosomes: E. carpathicus - 2n=90, E. concinnus - 2n=88, E. hadzii - 2n=68, E. sicanus - 2n=66, E. tergestinus - 2n=60, E. naupliensis - 2n=60, E. italicus - 2n=36. Description of the karyotypes revealed that all species studied exhibit achiasmatic meiosis; no presence of sex chromosomes was detected. The basic hypothesis of phylogenetic relationships and karyotype evolution of the genus Euscorpius was outlined. High interspecies variability in chromosome total count was found and by analysis of the 16S rRNA gene the taxonomic status of the species was confirmed. Hence, it seems that cytogenetic methods can contribute to the understanding of species diversity and differentiation of possible cryptic species within the genus Euscorpius.
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Influence of commercial probiotic preparations on human intestinal microbiome composition
Balatka, Štěpán ; Hodek, Petr (advisor) ; Osičková, Adriana (referee)
The intestinal microflora is an extensive ecosystem of microorganisms that consists of symbiotic and pathogenic species. The microflora is responsible for many important functions in the human body. An unquestionable function is that it affects the health state of the host. The higher the biodiversity, the greater the benefit for the host. However, it is necessary to point out that this should not include a high diversity of pathogenic bacterial species. There are many "beneficial" species, especially from the Bifidobacterium and Lactobacillus families. In recent decades, the popularity of supplementing these "beneficial" species with various supplementary diets (e.g. probiotics) has been growing a lot. The presented diploma thesis deals with pilot studies of liquid commercial probiotic preparations from American companies Ascended Health (not available on the Czech market) and their effects on the human microbiome. The study involved 9 volunteers who provided 70 fecal samples before, during, and after use of the studied products. Two methods were used to determine the biodiversity of intestinal bacterial species. Both are based on the identification by bacterial DNA that encodes gene 16S rRNA. The first method uses PCR-DGGE technique and then identification by Sanger sequencing. The second method...
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The Effect Of Quality Trimming On Joining Paired-End Reads In Microbiome Data Analysis
Heřmánková, Kristýna
The main goal of microbiome research is to determine the microbial composition of atarget sample. In successfully performed research, a cause of a disease can be found or pathogensin an environmental sample can be revealed. The correct evaluation of microorganisms present inthe analysed sample is therefore required, but unfortunately not that often reached. The accuracy ofthe resulting composition can be affected already in pre-processing steps of the analysis. The frequentissue that can negatively affect the reliability of research is data loss. This loss is most commonin the case of the paired-end reading method. The occurrence of data loss can be caused by theneed of joining this pair of sequences into one continuous sequence. The need of joining reads withlow-quality ends together can lead to counts reduction of sequences in an available dataset, andtherefore unrealistic results can be obtained. This paper shows, how quality trimming can affect thenumber of lost sequences during the reads joining step.
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Tool for Visualization of Microbiome Data
Mišáková, Silvia ; Martínek, Tomáš (referee) ; Smatana, Stanislav (advisor)
Táto práca sa zameriava na vytvorenie nového nástroja pre vizualizáciu mikrobiomových dát. Vytvorený nástroj používa pre redukciu dimenzií analýzu hlavných komponent (PCA) a analýzu hlavných súradníc (PCoA). V prípade výpočtu dištančnej matice sú použité metriky Bray-Curtis odlišnosť a UniFrac. Spracované dáta sú následne ofarbené na základe užívateľom zvolených metadát. Výsledky sú prezentované pomocou dvoch typov grafov. Prvý z nich je stĺpcový a zobrazuje podiel každej hlavnej zložky. Druhý, bodový graf, vizualizuje konečný výsledok požadovanej analýzy. V rámci práce bola pridaná možnosť stiahnuť si vypočítanú maticu a taktiež tabuľku prvých N hlavných zložiek vypočítaných danou analýzou.
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Biofilm formation in probiotic cultures and its application in pharmacy
Ryšávka, Petr ; Obruča, Stanislav (referee) ; Vorlová, Lenka (referee) ; Márová, Ivana (advisor)
The work was comprehensively focused on the development of adhesive forms of probiotics in the form of a biofilm on combined carriers with a prebiotic component. The second part dealed with the influence of food on the multiplication and survival of selected types of probiotic bacteria. Subsequently, the effect of individualized probiotic supplements on changes in the human intestinal microbiome was monitored. Suitable adherent probiotic strains for biofilm formation were selected and tested. Methods have been introduced and different variants of carriers for culturing and binding bacteria have been tested. In vitro experiments verified the stability of biofilm stucture and its resistance to low pH, bile and antibiotics in comparison with the planktonic cell form. The antimicrobial effect of probiotic strains in the form of a biofilm was studied. The cultivation of the multispecies biofilm on the combined carrier was optimized and the stability of the biofilm and the final viability of probiotic bacteria were confirmed. Furthermore, the influence of various foods and beverages on the viability of probiotic bacteria was evaluated with emphasis on the simulation of passage through the gastrointestinal tract. Both models, solutions with standardised concentrations of alcohol, sugar, salts, proteins or different pH and different types of real foods and beverages were tested. The effect of food and beverages was tested on monocultures of Lactobacillus acidophilus, Bifidobacterium breve and on probiotic capsules containing a mixed culture of probiotic microorganisms. The survival of probiotics in various food matrices in the simulated gastrointestinal tract was quantitatively different. We managed to define foods suitable for supporting the multiplication of probiotic bacteria. A separate part of the work was focused on the targeted modulation of the intestinal microbiome by individualized probiotics that were prepared on the basis of molecular biological analyzes of the intestinal microbiome aimed at detecting the percentage of lactobacilli, bifidobacteria and phylum Firmicutes and Bacteroidetes. Personalized probiotic supplementation confirmed the positive effect of this approach on microbiome changes, especially on the increase of the content of lactobacilli, bifidobacteria and the overall diversity of the microbiome.
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Bacteria Classification Based on Marker Genes
Pelantová, Lucie ; Hon, Jiří (referee) ; Smatana, Stanislav (advisor)
The aim of this work is proposal of new method for bacteria classification based on sequences of marker genes. For this purpose was chosen 10 marker genes. Resulting MultiGene classifier processes data set by dividing it in several groups and choosing gene for each group which can distinguish this group with best results. This work describes implementation of MultiGene classifier and its results in comparison with other bacteria classifiers and with classification based entirely on gene 16S rRNA.
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Bacteria Classification into Taxonomic Categories Based on Properties of 16s rRNA
Grešová, Katarína ; Hon, Jiří (referee) ; Smatana, Stanislav (advisor)
The main goal of this thesis was to design and implement a tool that would be able to classify the sequences of the 16S rRNA gene into taxonomic categories using the properties of the 16S rRNA gene. The created tool analyzes all input sequences simultaneously, which differs from common classification approaches, which classify input sequences individually. This tool relies on the fact that bacteria contain several copies of the 16S rRNA gene, which may differ in sequence. The main contribution of this work is design, implementation and evaluation of the capabilities of this tool. Experiments have shown that the proposed tool is able to identify the corresponding bacteria for smaller datasets and determine the correct ratios of their abundances. However, with larger datasets, the state space becomes very large and fragmented, which requires further improvements in order for it to search the state space in an efficient way.
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