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Molecular characterization of selected strains of amoebae of the genus Acanthamoeba, the potential human parasites.
ŠTAUBEROVÁ, Kamila
The genus Acanthamoeba belongs to a group of organisms generally called "amoebae". These amoebae can cause fatal disease in humans, being able to enter e.g. through skin in bloodstream and then spreading through the body. In the body they further parasitize in various tissues including the brain there they cause a little known disease called "granulomatous amoebic encephalitis". The disease is most prevalent in people previously ill with, e.g., systemic lupus erythematosus. Granulomatous amoebic encephalitis leads especially to a brain inflammation, which evolves in brain tissue necrosis. Most prominent symptoms are headaches, nausea and vomiting, aphasia and ataxia. This work mentions also some other amoebae as Neagleria fowleri, Balamuthia mandrillaris, Sappinia diploidea a Entamoeba histolytica. Neagleria fowleri is known as a causative agent of an illness called primary amoebic encephalitis. First, I had isolated DNA using Genomic Mini Kit from five amoeba strains. The next step was PCR amplification it is a method depending on alternation of high and lower temperatures in thermocycler. It was followed by electrophoresis, where the presence of bands in agarose gel, which I had prepared myself, showed the presence of absence of amoebic DNA fragments. In case of successful amplification (presence of PCR product) the fragment was excised from gel with scalpel and transferred in a test-tube. The DNA was then cleaned via extraction. The extraction was done with Gel/DNA Fragments Extraction Kit. An intermediate step before sequencing, we tried to clone the product it became obvious that this way is not very effective. The sequencing was provided by the company Seqme. Following work with sequences was conducted in programs BioEdit and alignment in ClustalX. Then, phylogenetic trees were computed in the program PAUP and viewed in TreeView. The results were interpreted in the terms of relationships between the amoebae majority of them belonged to the most common genotype, T4, but their relations within the genotype were not so tight. Concerning the clinical samples available to me (both from human eyes), one of them (O1) belonged to T4 genotype, whereas the other one (O2) belonged to the genotype T3. Another result was evaluation of the effectivity of different primers: specific primers AcaJDP1 and AcaJDP2 were much more efficient than originally used "eukaryotic" primers ERIB1 and ERIB10.
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Diversity, phylogeny and biology of various strains of Cryptosporidium muris
HAVRDOVÁ, Nikola
The morphological, biological, and molecular characteristics of various isolates of Cryptosporidium muris, namely HZ206 and TS03 were described. Oocysts of C. muris TS03 measuring 7.77 ? 0.23 × 5.20 ? 0.08 ?m with a length to width ratio of 1.48 ? 0.02 (n = 100) were morphometrically larger than C. muris HZ206 oocysts measuring 7.62 ? 0.40 × 6.63 ? 0.09 ?m; 1.32 ? 0.02 (n = 100) (p=0.034). Oocysts of both C. muris TS03 and HZ206 isolates obtained from naturally infected Tachyoryctes splendens and Mus musculus musculus, respectively, were infectious for na?ve 8-week-old Mastomys coucha. While the prepatent period of C. muris TS03 was 19-21 days post infection (DPI) and the animals did not lost infection within 120 DPI, animals infected with HZ206 start to shed oocysts 13-15 DPI with patent period 63-112 DPI. The accumulated value of infection intensity (AUC) in TS03 infected animals ranged from 250,000,000 to 800,000,000 oocysts per patent period (OPP) and HZ206 from 11,000,000 to 35,000,000 OPP. Histologically, a massive infection of cryptosporidia was detected in the glandular epithelium of stomach. Histopathological changes had a non-inflammatory character and included distinctive dilatation of infected parts of the glands with atrophy and metaplasia of the glandular epithelial cells. Infection with TS03 influenced the size of the stomach. While physiological weight of stomach of na?ve 12-week old mastomys and those infected with HZ206 were 0.61 ? 0.06 g and 0.70 ? 0.17 g,respectively, abnormal enlargement of stomach was observed in animals infected with TS03 (3.23 ? 0.51 g). There was no effect on feed intake among tested groups. Phylogenetic analyses based on small subunit rRNA, actin, MS1, MS2, MS3 and MS16 gene sequences revealed that C. muris TS03 is genetically distinct from other C. muris isolates. In conclusion, morphological, genetic, and biological data support the establishment of Cryptosporidium muris TS03 as a new species.
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The intracellular symbionts of blood-feeding Diptera group Hippobosccoidea
ŠOCHOVÁ, Eva
Blood-sucking parasites from the Hippoboscoidea group (Insecta: Diptera) harbor symbiotic bacteria providing them with B-vitamins and cofactors missing from their blood diet. Although including medically important vectors such as tsetse flies, evolutionary history of symbiosis in the whole group is poorly understood. This thesis is focused on analysis of symbionts in the Hippoboscidae family (parasites of birds and mammals). Phylogenetic approach was employed to determine lineages of obligatory and facultative symbionts present in this group. Results of the study indicate that relationships between Hippoboscoidea and their symbionts are extremely dynamic with frequent replacements of obligate symbionts. To which extent is this dynamic evolutionary history of symbiotic systems in Hippoboscoidea caused by features of their parasitic lifestyle (e.g. host switches from mammals to birds) will be an interesting question for further studies.
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Phylogenetic relationship of living Equidae
PERNEROVÁ, Ludmila
This thesis deals with a phylogeny of living Equidae based on an available evidence (morphology, behaviour, chromosomes, mitochondrial and nuclear genes) and using maximum parsiomony and Bayesian approaches; and ILD, PBS analyses which are able to recognise incongruences between specific character partitions. My results are relatively very robust and rational in comparison with published studies. My thesis consists of a review (literature, GenBank) and obtaining of many sequences by myself.
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Species diversity and speciation mechanisms in Crenicichla (Neotropical cichlids)
PIÁLEK, Lubomír
This thesis contributes to the knowledge of the species diversity of the Crenicichla lacustris sp. group in the La Plata River basin with description of three new species. Speciation mechanisms within two different species flocks from the middle Paraná/Iguazu and Uruguay Rivers were studied with a phylogenomic approach applying a novel genotyping method based on a Double-Digest Restriction site Adjacent DNA (ddRAD) sequencing. Our results support a repeated origin of morphological species being evolved several times sympatrically and independently in different drainages. A considerable role of hybridization/introgression as an evolutionary force was also proposed. The thesis further uncovers biogeographic aspects of the southern part of Brazilian shield and adjacent coastal rivers.
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Charakteristics of scales of neotropical cichlids (Tribus Heroini) - phylogenetic interpretation
STOPOVÁ, Barbora
Work has the aim to describe charakteristics of scales selected species fish from the Central America and evaluace relationship with view on the group and compare scalimetric evolutional tree with molecular. Relationship between Heroini and Cichlasomatini is really close and Heroini constitute the second largest tribe of neotropical cichlids. These fish are very varied. Heroini is considered as monofyletic group. Development of the tribe in two branches. Theoretic part of this work is focused on phylogenetic knowledge from the other authors and group description. In the part with own issues there are characteristics of scales, microphotographs.
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Molecular phylogeny and genome evolution of insect symbiotic bacteria
NOVÁKOVÁ, Eva
Since the introduction of advanced molecular methods the research on insect bacterial symbioses underwent a major focus shift towards large scale phylogenetics and comparative genomics. These new fields provided answers to several fundamental questions of symbiont evolution, functional capabilities of the host-associated bacteria, and the role of symbionts in the host?s biology. However, the vast diversity and complexity of symbiotic relationships still leaves gaps in our understanding to a rich mosaic of various symbiont types, effects and transitions from facultative association to obligate mutualism. The presented study focuses on distribution, diversity, phylogenetic patterns, evolutionary transitions and genome evolution of two less known but ecologically diverse bacterial genera, Arsenophonus and Sodalis. The thesis also takes advantage of the knowledge on a well established symbiotic model between aphids and Buchnera and reveals several evolutionary patterns in the host and symbiont.
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The evolution of the social parasitism of Phengaris (Maculinea)
PECH, Pavel
The presented thesis deals with the evolution of the social parasitism of Phengaris (Maculinea) butterflies. It consists of three related problems: I) The phylogeny of Glaucopsyche section (Lepidoptera: Lycaenidae: Polyommatinae) {--} the social parasitism occured only once in the Glaucopsyche section. II) The conditions in which the social parasitism of Phengaris evolved {--} the process was probably facilitated by unique species composition of the myrmecofauna of the biotopes inhabited by the ancestor of Phengaris. III) The host specificity of Phengaris {--} today{\crq}s view on the host specificity of Phengaris is poorly supported.
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