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The development, constitution and functions of the gland apparatus in schistosome cercariae and endopeptidases in its contents
Titlová, Lucie ; Mikeš, Libor (advisor) ; Konečný, Lukáš (referee)
Flukes of the family Schistosomatidae are blood parasites with two-host life cycles involving aquatic snails as intermediate hosts and avian or mammalian definitive hosts. Cercariae, as invasive aquatic stages of schistosomes, enter the lumen of vessels by penetrating the skin of the definitive host. During their short life, cercariae possess a glandular apparatus consisting of three types of glands with external secretion: penetration glands, escape glands, and a head gland. These glands secrete granules containing, among other things, proteolytic enzymes, which play an important role in the process of penetrating the host's skin, but at the same time they occupy many other functions during the life of the cercaria and other life stages of schistosomes. This thesis summarizes basic knowledge about the anatomy, function and development of the gland apparatus of schistosome cercariae and further focuses on the representation of proteolytic enzymes in these glands, specifically endopeptidases. At the end, it briefly compares the different representation of endopeptidases in some members of the family.
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Glycocalyx shedding by cercariae of bird schistosomes
Chaloupecká, Jana ; Mikeš, Libor (advisor) ; Štěrba, Ján (referee)
Trichobilharzia spp. are avian schistosomes related to medically important human parasites of the genus Schistosoma. Penetrating cercariae are well known as causative agent of cercarial dermatitis in humans. Cercariae actively penetrate the skin of definitive hosts and transform into schistosomula. This process is preceded by cercarial tail detachment and includes emptying of penetration glands and extensive surface changes. One of these changes is the loss of highly immunogenic glycocalyx which represents a protective coat in the aquatic environment. The glycocalyx has specific composition of saccharide molecules which are bound to lipids or proteins on the membrane of cercarial tegument. There is only limited information about the mechanism of shedding. Hypotheses based on indirect evidences suggest that peptidases or (phospho)lipases from penetration glands could be involved. This work describes the changes in surface glycosylation during transformation of cercariae into schistosomula by fluorescently labelled lectins and monoclonal antibodies against Lewis X antigen. Lectins UEA-I, LTA and PNA have been chosen as markers of transformation of T. regenti. Further, our experiments have been focused on shedding of cercarial glycocalyx. During in vitro induction of penetration gland emptying and...
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Features and functions of glycocalyx of trematode cercariae
Chaloupecká, Jana ; Mikeš, Libor (advisor) ; Kašný, Martin (referee)
Trematodes are parasites from phylum Platyhelminthes which have compex life cycles involving two to four hosts. This work focuses especially on trematodes of the family Schistosomatidae. Their cercariae which leave the snail intermediate host, actively penetrate the skin of definitive hosts and transform into schistosomula. This is accompanied by detachment of cercarial tail and emptying of penetration glands. During transformation, cercarial bodies undergo extensive ultrastructural and molecular changes. One of these changes is the loss of surface glycocalyx which represents a protective coat in the aquatic environment. In glycocalyx shedding, participation of proteolytic enzymes from cercarial penetration glands is expected during invasion of the host. Glycocalyx has specific composition of saccharide molecules which are bound to lipids or proteins on the membrane of cercarial tegument. This work describes the origin, ultrastructure, saccharide composition, function and shedding mechanism of cercarial glycocalyx.
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Cathepsins L of Diplostomum pseudospathaceum cercariae
Perháčová, Terézia ; Mikeš, Libor (advisor) ; Hartmann, David (referee)
This study is focused on cercarial cysteine peptidases of the trematode Diplostomum pseudospathaceum. It follows previous research which confirmed the presence of a 24kDa cysteine peptidase in cercariae biochemically and by mass spectrometry. It was postulated, that the function of this peptidase is histolytic, when cercariae penetrate the tissues. During an attempt to purify this peptidase and characterize its peptidolytic activity, it was found out that the cercarial homogenate containsmore different peptidases varying in their pI. Tests of peptidolytic activity and inhibition have shown that these peptidases are cathepsin L-like. They are active over a broad spectrum of pH with optima of activities in weakly acidicor neutral pH. Using degenerate primers based on conserved motifs of cysteine pepridases, partial sequences of three genes for cathepsin L of D. pseudospataceum (DpCL1, 2 a 3) were obtained. Then the complete sequences of DpCL2 and 3 genes and partial sequence (without 5'end) of DpCL1 were obtained by RACE PCR. To confirm function of these peptidases we tried to immunolocalize them. We assumed that they are localized in penetration glands. Preliminary results suggested that some of the cathepsins could be also localized in the gut of cercariae. For more detailed biochemical...
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Morphological and functional variability of secretory glands in cercariae of chosen trematode groups
Krčmářová, Veronika ; Bulantová, Jana (advisor) ; Mikeš, Libor (referee)
Trematodes are characterized by their complex life cycles that include definitive hosts and variable number of intermediate hosts. Transfer of the parasite from the first intermediate host to the other is usually realized by larval stage called cercaria. After finishing of their development within the first intermediate host, morphology and fate of these larval stages vary according to the way of infection of the next host. Some cercariae actively penetrate directly to their definitive hosts trough their body surface, other encystate in the outer environment where they wait in a form of metacercariae to be ingested by definitive host. Both of these ways can be combinated and cercariae encystate inside second intermediate host after they actively penetrate them. Exceptionally, cercariae do not leave the sporocyst in which they were developing inside the first intermediate host. They encyst there waiting for ingestion by the definitive host. Various types of secretory glands have been developed in cercariae for successful direct infection of next hosts, survival of parasite in adverse conditions of outer environment or for transformation of one larval stage to subsequent one. Variability in morphology and function of these secretory glands in cercariae is closely connected with differences in life...
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Složení společenstev larválních stádií motolic (Digenea) u vybraných zástupců plicnatých plžů čeledi Planorbidae
VYHLÍDALOVÁ, Tereza
In the present study the species diversity, seasonal dynamics, composition of larval trematode communities and factors influencing their structure were studied in two species of planorbid gastropods (Gyraulus albus and Segmentina nitida) in typical eutrophic fishponds in South Bohemia, Czech Republic. The study significantly contributed to the present knowledge about the fauna of trematodes in small planorbids, provided evidence for validity of general rules affecting trematode community composition and structure and demonstrated the underestimated role of both snail species as first intermediate hosts for trematodes in the past.
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In vitro cultivation of the trematode species Trichobilharzia regenti
Vrbová, Kristýna ; Kašný, Martin (advisor) ; Skelly, Patrick (referee)
The class Trematoda includes many pathogenic representatives. Main subject of this thesis, avian schistosome Trichobilharzia regenti, is a close relative to the important human pathogen Schistosoma mansoni (family Schistosomatidae). In vitro cultivation of trematodes enables closer understanding of their biology and parasite- host interactions; however, no trematode species has been successfully kept in vitro from the egg stage to the adults producing eggs. Many studies are focused on the problematic of S. mansoni cultivation, but data concerning T. regenti cultivation remain scarce. Only the ability of T. regenti cercariae to transform into schistosomula in vitro was documented, with following survival in a culture medium for a few days. Comparison of eight transformation methods was performed with T. regenti cercariae. Based on the number of tailless cercarial bodies obtained, five transformation methods were selected for further evaluation of the early schistosomula characteristics (glycocalyx shedding, penetration glands emptying and survival in vitro). It was observed that the largest quantity of cercarial bodies can be obtained by using a syringe needle or the BeadBeater cell disrupter. The largest quantity of schistosomula meeting the criteria of early schistosomulum was recorded after...
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Excretory-secretory proteins of schistosome intravertebrate stages in context of their biology
Konečný, Lukáš ; Mikeš, Libor (advisor) ; Kašný, Martin (referee)
Schistosomatidae is a family of blood-dwelling trematodes which cause serious disease in humans and warm blooded animals. Human schistosomiasis affects over 258 million people worldwide. The fundamental pathological manifestations of the disease are not caused by adult worms, but by responses to antigens released from eggs, which remain trapped in the tissues. All stages of these parasites release excretory-secretory products, which mostly comprise of proteins which are, to a certain extent, species- and stage-specific. These products have many functions that play roles in successful transmission of the parasite and its interactions with the host. Various approaches to the collection and analysis of these molecules do not always accurately reflect the secretory processes in vivo, because they always employ in vitro techniques. The research of these secreted molecules discovered many mechanisms, through which the parasite successfully modulates and evades host immune system. Despite the fact, that many of these molecules were identified as potential vaccine candidates, still no sufficiently effective vaccine has been developed. With great technological advances used for the identification of these proteins, this area is becoming more and more topical. This review focuses on composition of...
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Cathepsins L of Diplostomum pseudospathaceum cercariae
Perháčová, Terézia ; Mikeš, Libor (advisor) ; Hartmann, David (referee)
This study is focused on cercarial cysteine peptidases of the trematode Diplostomum pseudospathaceum. It follows previous research which confirmed the presence of a 24kDa cysteine peptidase in cercariae biochemically and by mass spectrometry. It was postulated, that the function of this peptidase is histolytic, when cercariae penetrate the tissues. During an attempt to purify this peptidase and characterize its peptidolytic activity, it was found out that the cercarial homogenate containsmore different peptidases varying in their pI. Tests of peptidolytic activity and inhibition have shown that these peptidases are cathepsin L-like. They are active over a broad spectrum of pH with optima of activities in weakly acidicor neutral pH. Using degenerate primers based on conserved motifs of cysteine pepridases, partial sequences of three genes for cathepsin L of D. pseudospataceum (DpCL1, 2 a 3) were obtained. Then the complete sequences of DpCL2 and 3 genes and partial sequence (without 5'end) of DpCL1 were obtained by RACE PCR. To confirm function of these peptidases we tried to immunolocalize them. We assumed that they are localized in penetration glands. Preliminary results suggested that some of the cathepsins could be also localized in the gut of cercariae. For more detailed biochemical...
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Orientation of helminths during host seeking
Vojtová, Terezie ; Mikeš, Libor (advisor) ; Horák, Petr (referee)
The parasitic helminths are often forced in the environment to search for their host quickly in order to continue the ontogenetic development, or else to finish it. To search for and indentify the host, they use various types of receptors which allow them to provoke a taxes corresponding to the oncoming stimulus and to react regarding the current situation. In respect to the concept relevant to the helminths, this work concerns trematodes, whose freely movable stadia are miracidium and cercacia, and then monogeneans which are viparous or oviparous with oncomiracidium larva, and nematodes during which the third larva stage moves. The representatives of the groups of Myxozoans and Ciliophorea are also mentionned, since they use similar mechanisms to look for hosts as helminths. This thesis includes the summary of the existing knowledge related to the orientation of helminths in the environment. On the basis of the data that is already known, it is possible to compare which mechanisms are used for host-seeking by particular species, and according to which signals from the surroundings they most often orientate themselves. Powered by TCPDF (www.tcpdf.org)
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