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Cryptic diversity of free-living trichomonads and their phylogenetic position within Parabasalia
Céza, Vít ; Čepička, Ivan (advisor) ; Hampl, Vladimír (referee)
Trichomonads (Parabasalia) are anaerobic microeukaryotes classified in the supergroup Excavata. Inclusion of parabasalids within Excavata is exclusively based on the molecular- phylogenetic evidence. Over 400 species of parabasalids have been described so far, and the vast majority of them are endobiotic. In contrast, only few species of free-living parabasalids forming four independent lineages have been described (Pseudotrichomonas keilini, Ditrichomonas honigbergii, Monotrichomonas carabina, Honigbergiella sp., Tetratrichomonas undula, and Lacusteria cypriaca). Lacusteria cypriaca is a new species and genus described in our recent paper. In this paper we published the first two sequences of SSU rDNA from Pseudotrichomonas keilini as well. All of these lineages are likely secondarily free-living, and they developed from endobiotic ancestors. In addition to the already published Lacusteria cypriaca and Pseudotrichomonas keilini strains, we have recently obtained seven another isolates of free-living trichomonads (LAGOS2D, E2NT, CK, LAGOS2M, GR8, GOU23 LIVADIAN, and VAV1A1); from all of these isolates we sequenced SSU rDNA and performed phylogenetic analyses. These isolates split into four independent evolutionary lineages, which indicate that free-living parabasalids are more diversed and...
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Evolution of Retortamonadida (Eukaryota: Excavata: Fornicata)
Smejkalová, Pavla ; Kostka, Martin (referee) ; Čepička, Ivan (advisor)
Retortamonads (Retortamonadida; genera Chilomastix and Retortamonas) are a small group of protists comprising intestinal commensals of both vertebrates and invertebrates and one free-living species of the genus Chilomastix. Molecular phylogenetic studies showed that retortamonads are closely related to diplomonads, Carpediemonas, Dysnectes, Hicanonectes, Kipferlia and two undescribed lineages of free-living Carpediemonas-like organisms, together forming the monophyletic excavate group Fornicata. For a long time Retortamonadida have been assumed to be a monophyletic group. However, first molecular phylogenetic study including sequence data from both Retortamonas and Chilomastix suggested that Retortamonadida are paraphyletic and that diplomonads branch within Retortamonadida. Retortamonads still remain poorly studied protist group and their phylogeny is unclear. We sequenced and analysed SSU rDNA of ten Retortamonas and four Chilomastix SSU rDNA sequences. In addition, we sequenced SSU rDNA of an undescribed enteromonad lineage. The phylogenetic tree of Fornicata was largely unresolved and the phylogenetic position of the genus Chilomastix remained unclear. On the other hand, the genus Retortamonas and diplomonads formed a robust clade. Retortamonas sequences split into three host-specific lineages. The...
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Evolutionary strategy in biomineralization of mollusc shells: Micromechanical properties of shells and their relationship with the environment
Hrabánková, Iva ; Frýda, Jiří (advisor) ; Marika , Marika (referee)
This thesis deals with the micromechanical properties (microhardness and elastic modulus) of selected molluscan shell microstructures and their relationship to crystallographic textures and environment. Results of the study suggest that extremely high hydrostatic pressure has a relatively weak influence on the mechanical properties of calcitic as well as aragonitic shell layers. No statistically significant difference was found in microhardness and elastic modulus of nacre between bivalvian species living in marine and freshwater environments. On the other hand, the study reveals a statistically significant positive correlation between the micromechanical properties and thickness of aragonitic platelets forming the nacre. Unexpected result is a negative correlation of the degree of ordering of aragonitic platelets with both elastic modulus as well as microhardness. Thus the degree of ordering explains about 80% of the variability of these micromechanical properties. Statistically significant but weaker correlation was also found between micromechanical properties and textural type of bivalvian nacre. The nacre with uniaxial texture is harder and more elastic than the nacre with quasimonocrystallic texture. The data suggests that higher hardness and elasticity corresponds to a primitive state of nacre and that during the subsequent evolution values of these micromechanical properties were decreasing. This trend contradicts with generally accepted hypothesis of increassing predation pressure in the marine ecosystem during the Paleozoic and Mesozoic.
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Evolution and genomics of symbionts in Hippoboscidae
ŠOCHOVÁ, Eva
Obligately blood-sucking parasites harbour symbiotic bacteria providing them B-vitamins and cofactors missing from their blood diet. Within Hippoboscoidea group (parasites of birds and mammals), tsetse flies as medically important vectors have been studied extensively while bat flies and louse flies tend to be neglected. This thesis is composed of two complementary manuscripts focused on phylogeny and origin of bacterial symbionts in Hippoboscidae family (manuscript 1) and their genome evolution (manuscript 2). First, phylogenetic approach was employed to determine lineages of obligate and facultative symbionts present in this group. Second, genomic and phylogenomic analyses were carried out to better understand evolution of obligate endosymbionts from the Arsenophonus genus in this group. Results of the two studies indicate that relationships between Hippoboscoidea and their symbionts are extremely dynamic with frequent replacements of obligate symbionts. This hypothesis is supported by both phylogenetic and genomic evidence, in particular, Arsenophonus endosymbionts of Hippoboscidae represent several distinct lineages (of likely different ages) with noticeable differences in genome features and metabolic capabilities. The data presented in this thesis thus greatly extend our knowledge about evolution and genomics of symbiotic bacteria in Hippoboscidae and bloodsucking hosts in general.
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Detection of Correlated Mutations
Ižák, Tomáš ; Bendl, Jaroslav (referee) ; Martínek, Tomáš (advisor)
Tato práce zkoumá existující možnosti a metody detekce korelovaných mutací v proteinech. Práce začíná teoretickým úvodem do zkoumané problematiky. Využití informací o korelovaných mutacích je především při predikci terciální struktury proteinu či hledání oblastí s významnou funkcí. Dále následuje přehled v současnosti používaných metod detekce a jejich výhody a nevýhody. V této práci jsou zkoumány zejména metody založené na statistice (například Pearsonově korelačním koeficientu nebo Pearsonově chi^2 testu), informační teorii (Mutual information - MI) a pravděpodobnosti (ELSC nebo Spidermonkey). Dále jsou popsány nejdůležitější nástroje s informací o tom, které metody používají a jakým způsobem. Také je diskutována možnost návrhu optimálního algoritmu. Jako optimální z hlediska úspěšnosti detekce je doporučeno využít více zmíněných metod. Také je doporučeno při detekci využít fyzikálně-chemických vlastností aminokyselin. V praktické části byla vyvinuta metoda využívající fyzikálně-chemických vlastností aminokyselin a fylogenetických stromů. Výsledky detekce byly porovnány s nástroji CAPS, CRASP a CMAT.
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Phylogeny of amoebae of family Flabellulidae
LISNEROVÁ, Martina
This thesis deals with one of little groups of amoeboid protozoa family Flabellulidae of Amoebozoa group (Lobosa, Tubulinea). This group has only one cell with one or many nuclei. Body is not covered by shell or scales, but only with glycocalyx. They move by using lobopodias, the movement being based on actin-myosin complex. Family Flabellulidae includes currently the genera Flabellula and Paraflabellula, historically genus Flamella. We lack sufficient information about phylogeny of family Flabellulidae. New sequences of actin genes, SSU rRNA and COX1 from members of family Flabellulidae were obtained in this study and the relationships between family Flabellulidae and other hierarchical groups are also discussed.
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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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