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Ontogenesis of trematode larval stages of the family Fasciolidae in the intermediate snail hosts.
Pankrác, Jan ; Kašný, Martin (advisor) ; Soldánová, Miroslava (referee)
The family Fasciolidae is an important group of trematodes including serious pathogens of humans and livestock. The life cycle is divided into two phases - sexual reproduction in the definitive host body (large land mammals) and an asexual reproduction in the intermediate host body (aquatic snails of the family Lymnaeidae and Planorbidae). Development within the snail host is characterized by production of large amounts of parasite larvae (sporocyst, rediae, cercariae). Cercariae released from the snail immediately start to transform into metacercariae, the larval stages infectious for definitive host. Snail phase of infection is generally accompanied by number of mostly negative symptoms (massive pathological changes, often followed by reduction of fertility). This summary reveals that current knowledge concerning the ontogenetic development of fasciolids in the intermediate host is uncomplete and unequally investigated. According to this summary is also obvious that some of the published findings are universally valid for all members of the family Fasciolidae and other are characteristic only for particular species.
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Localization and quantification of mRNA coding digestive peptidases of Fascioloides magna
Peterková, Kristýna ; Kašný, Martin (advisor) ; Sojka, Daniel (referee)
Trematode peptidases are important molecules responsible for biocatalysis in many basal biological processes and are crucial in host-parasite interactions. Therefore, these enzymes are intensively studied in order to characterize their biological functions and to use them as potential diagnostic or therapeutic targets. Lately, investigation of transcriptome and secretome revealed, that adult Fascioloides magna (giant liver fluke) expresses and secretes a variety of peptidases. Thus, this thesis focuses on three newly identified enzymes: cathepsin L (FmCL), cathepsin B (FmCB) and cathepsin D (FmCD). In other trematode species, these cathepsins are being linked mainly with the digestion of host blood. We applied quantitative PCR (qPCR) to investigate relative expression levels of the three enzymes among three developmental stages - egg, miracidium and adult. It was revealed that all cathepsins have the highest expression in adult flukes in comparison to eggs and miracidia. We also localized the place of transcription of FmCL, FmCB and FmCD in adult fluke using RNA in situ hybridization. All of the peptidases were detected in gastrodermis, and in addition, they were localized in the reproductive system. The latter surprising finding is suggesting that these enzymes might have multiple functions in adult F....
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Fasciolid flukes: from genes to diagnostic tools
Ježková, Monika ; Leontovyč, Roman (advisor) ; Sojka, Daniel (referee)
Liver flukes of the family Fasciolidae are parasites of mammals including human. Fascioloides magna and Fasciola hepatica are considered as a veterinary and medically important species occurring also in the Czech Republic. Fascioloides magna and F. hepatica infect wide spectrum of wild and domestic ruminants and in case of F. hepatica human can be also infected. Both flukes are responsible for damage of liver tissue and/or bile-ducts of their definitive hosts causing weight lose, anemia, reduced productivity and in specific cases the death of the host. Effective diagnosis plays the key role in control of F. hepatica and F. magna infections. Current diagnostics is predominantly based on serodiagnostic methods using specific antigens e.g. from excretory-secretory products (ESPs). Due to heterogenity of ESPs, such diagnostic markers can lack the specificity and also the reproducibility of the method is poor. Particular proteins of ESPs are often used in diagnostics of fasciolid flukes. Such approach requires biological material and laboratory procedures associated with identification, purification and antigenicity testing of selected proteins. Recent development of parallel sequencing technologies results in huge amount of genomic, transcriptomic and proteomic data, which are publicly available. Such...
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The expression profile of cathepsin L in developmental stages of Fascioloides magna
Šašková, Romana ; Kašný, Martin (advisor) ; Mareš, Michael (referee)
Our experimental organism Fascioloides magna is a digenetic liver fluke from Fasciolidae family which parasitizes in domestic and free-living ruminants of North America and Central Europe including Czech Republic. In Czech Republic this highly pathogenic worm causes a severe liver damage to cervids and bovids and the prevalence locally reaches up to 95%. The biology of F. magna including e.g. the characteristics of host-parasite molecular interaction and the functions of particular molecules produced by the parasite are not fully understood. According to results of our previous research the excretory-secretory products of F. magna adults contain number of molecules which play the crucial role in host tissue invasion, digestion and evasion of the host immune response. One of the most abundant is cysteine peptidase cathepsin L (FmCL). FmCL is supposed to play various key roles in biological processes of all stages during a life cycle and therefore we can suppose its different expression level in particular life stages. In order to define the expression level of FmCL we performed the pilot study with miracidia and adults where the qPCR method was applied. The results of this experiment revealed much higher expression level of FmCL1 in adults than in miracidia. The attempt to in situ localize the mRNA...
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Experimental infection of Oryctolagus cuniculus with fluke Fascioloides magna
Melounová, Klára ; Kašný, Martin (advisor) ; Novobilský, Adam (referee)
Fasioloides magna is a trematode parasitizing in the liver parenchyma of ruminants. Its life cycle is associated with the humid environment and includes intermediate freshwater snail hosts from family Lymnaeidae. According to the ability of host to form a certain type of a pseudocyst during fascioloidosis, they can be,divided in three groups, specific definitive hosts (red deers, fallow deers, roe deers), nonspecific definitive hosts (cattle, wild boars and elks) and atypical hosts (sheeps and goats). Beside the natural infections also the experimental infections of other potential host species has been realized (chamois, llama and bighorn sheep and traditional laboratory animals such as mice, guinea pigs, rats and rabbits). In the context of different diseases, many changes in infected organism can occur. These can be qualitatively and quantitatively evaluated. Similarly, during fascioloidosis the changes associated with the presence of the parasite in the host's body is possible to monitor, e.g. antibody production, increase in the number of eosinophils, release of eggs in faeces, internal bleeding, or the level liver damage. The liver damage is corresponding primarily to biochemical parameters of blood, not only the liver enzymes, but also other blood components, like blood proteins, lipids,...
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Early embryonal development and morphogenesis of selected organ systems of the rediae and cercariae of Fascioloides magna.
Pankrác, Jan ; Kašný, Martin (advisor) ; Macůrková, Marie (referee)
Fascioloides magna (giant liver fluke) is a digenetic trematode with two-host life cycle and high veterinary importancy. Typical definitive host is a deer (Cervidae), but many other species from different families can be accidentally infected, for example sheep, goat or cattle. Very important role in the life cycle of F. magna has the first host - fresh water snail of the family Lymnaeidae. Three different life stages of F. magna, two of them with ability of reproduction - sporocysts and rediae develop in the body of snail. The third stage - cercaria is produced by rediae. Cercariae are able to escape from the snail, encyst and become infective for the definitive host. Since the second half of the 19th century many researchers studied the development of particular stages in the first intermediate host, but many characteristics of this process are still not fully understood. This thesis should reveal some of unanswered questions concerning to the reproduction and ontogenetic development of trematodes, which is presented on the examples of three organ systems - muscles, nerves and excretory system of rediae and cercariae of F. magna.
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Morphology of Fascioloides magna life stages and histopathological changes in selected species of definitive hosts
Košťáková, Monika ; Kašný, Martin (advisor) ; Hodová, Iveta (referee)
Fascioloides magna is a trematode originating in North America. It is a significant pathogen, especially in cervids, but it can also infect ruminants kept for agricultural purposes.Adult trematodes are found in the liver tissue of their definitive hosts, where they can survive for a long period of time and produce large quantity of eggs. Those are passed through bile ducts and intestine with feces into the external environment. This thesis is focused on the morphology of individual developmental stages of F. magna. To this date only very limited number of literary resources concerning the topic had been published. Recorded results are compared with literary resources on Fasciola hepatica, a closely related trematode. The morphological characteristics of the individual developmental stages were studied by using histological, electron-miscroscopical and other methods (fluorescent labelling). The thesis also describes pathological changes of the definitive hosts' liver tissue. By means of the stated methods, the thesis expands the existing knowledge on the morphology of F. magna especially of surface structures and the distribution of the senzoric organs, some of which have been described for the very first time. Powered by TCPDF (www.tcpdf.org)
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Ontogenesis of trematode larval stages of the family Fasciolidae in the intermediate snail hosts.
Pankrác, Jan ; Kašný, Martin (advisor) ; Soldánová, Miroslava (referee)
The family Fasciolidae is an important group of trematodes including serious pathogens of humans and livestock. The life cycle is divided into two phases - sexual reproduction in the definitive host body (large land mammals) and an asexual reproduction in the intermediate host body (aquatic snails of the family Lymnaeidae and Planorbidae). Development within the snail host is characterized by production of large amounts of parasite larvae (sporocyst, rediae, cercariae). Cercariae released from the snail immediately start to transform into metacercariae, the larval stages infectious for definitive host. Snail phase of infection is generally accompanied by number of mostly negative symptoms (massive pathological changes, often followed by reduction of fertility). This summary reveals that current knowledge concerning the ontogenetic development of fasciolids in the intermediate host is uncomplete and unequally investigated. According to this summary is also obvious that some of the published findings are universally valid for all members of the family Fasciolidae and other are characteristic only for particular species.
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Characterization of excretory-secretory proteins of liver fluke Fascioloides magna.
Beránková, Kateřina ; Kašný, Martin (advisor) ; Horn, Martin (referee)
Fascioloides magna (the giant liver fluke) originated from North America, is known in the Czech Republic since 1930s. This pathogenic fluke invades mostly cervids, but livestock too. Excretory-secretory products (ES products) contain number of esential biomolecules which are produced by excretory and secretory system of the fluke. These molecules play key role in many biological process during the life cycle not only of fascioloid flukes (e.g. migration in the host tissues, immune evasion and digestion). Due to their antigenic properties they could be also used in immunodiagnostics. Excretory-secretory proteins from adult Fascioloides magna and comparative related species Fasciola hepatica were purified and separated by the basic biochemical methods (1D, 2D electrophoresis, ion-exchange chromatography) and their activity was confirmed by specific (fluorogenic peptide) and nonspecific (gelatine) substrates. By using the mass spectrometry methods (MALDI TOF/TOF), the most abundant peptidolytically active proteins from ES products of F. magna were identified as cathepsin L (FmCL). Recombinant analog of FmCL was expressed in Pichia pastoris expression system. The peptidolytic activity was again confirmed using the synthetic fluorogenic substrates; the specifity of recombinant FmCL active site was...
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The occurrence of Fascioloides magna in deers in South Bohemia
VERNEROVÁ, Kateřina
Giant liver fluke Fascioloides magna is the biggest represenative of flukes in the Czech Republic. This originally North American parasite was brought to Europe with wapiti deers in 1960s. Since that time Fascioloides magna has extended the area of its appearance and its host spectrum a lot. The first appearance of this parasite in the Czech Republic was noted for the first time at a fallow deer which was hunted down in Middle Bohemia in 1930. However in the early 1960s the number of positive findings started to increase at hoofed. In the last decade was observed the expansion of areas of appearance of this fluke in South Bohemia. That is why I decided to make examinations to evaluate the recent appearance of F. magna at deer family in the South Bohemia from March 2009 to January 2010. 74 samples of dropping of a deer from three different areas (Českobudějovicko, Třeboňsko and Šumava national park) were collected and examined and only in the sample from Šumava national park were proved the ovulums of F. magna. Then I examined 11 livers of deers which were hunted down in Šumava national park. In all these livers were found macroscopic changes. In all these examined livers was also finding of fluke F. magna in number from 1 to 72. From the reached results follows that the problems of appearance of F. magna is in the Czech Republic still actual and it is necessery to count with the extending area of appearance of this parasite in the future. Edification among hunting community and the use of information from trained people who make the primary examination of animals after hunting down are possible ways to catch up appearance and expansion of this serious parasite at hoofed.
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