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Vliv teploty a času na excystaci Cryptosporidium muris izolát CAT21
DUDÁKOVÁ, Hana
The main aim of this master's thesis was to investigate the impact of environmental pH and the host's body temperature on the excystation of Cryptosporidium muris oocysts. The experiment utilized the CAT21 isolate obtained from a naturally infected domestic cat (Felis catus f. domestica), although rodents are the primary hosts of Cryptosporidium muris. The isolate was continuously maintained in immunodefi-cient SCID mice, from which fecal samples were regularly collected for individual experiments. The first factor examined for its influence on oocyst excystation was the host's body temperature. Oocysts stored in dH2O were subjected to water baths set to temperatures ranging from 21 to 44°C for durations of 5, 15 and 30 minutes. According to the results, the optimal temperature for excystation of C. muris CAT21 oocysts was 37°C for a duration of 30 minutes, resulting in up to 92% excystation. Additionally, the study included an examination of the pH impact on excystation. Oocysts were exposed to PBS solutions with varying pH values (ranging from 2.00 to 9.00) and heated in a water bath at 37°C. The optimal pH for excystation was found to be 6.00, with up to 93% excystation observed. The viability of the oocysts used was determined by staining with propidium iodide before each measurement.
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Faktory ovlivňující excystaci žaludečních kryptosporidií hlodavců
SCHULZOVÁ, Tereza
The aim of this study was to determine the effect of host body temperature and en-vironmental pH on the excystation and preservation of the ability to infection of rodent gastric cryptosporidia oocysts. The infectivity of unexcysted oocysts was further examined. Immunodeficiency SCID mice (Mus musculus) were infected with C. proliferans oocysts, then their faeces were collected and individual experi-ments were performed. The effect of temperature on excystation, when oocysts were placed in water baths at the appropriate temperature (32-44 °C) and the effect of pH, when oocysts were exposed to solutions of different pH (2-7), was investi-gated. Cell separation was then performed using a flow cytometer and the values obtained were recorded in the tables. The most oocysts excysted at 40 °C and pH 7. An autopsy of infected mice was also performed to determine the infectivity of oo-cysts from various parts of the digestive system. All taken oocyst samples were infectious for the subsequent host. Infection experiments, which were further car-ried out revealed, that unexcysted oocysts exposed to hypochlorite and laboratory temperatures retain their ability to infect. In contrast, the oocyst sporozoites that excysted were susceptible to both hypochlorite and room temperature, thus losing the ability infect the subsequent host.
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Kryptosporidiové infekce volně žijících a zájmově chovaných holubů
ZIKMUNDOVÁ, Veronika
The object of this thesis was an evaluation of occurrence and prevalence of Cryptosporidium infection in domestic and wild pigeons (Columbiformes). A total of 603 samples (389 domestic and 214 wild pigeons) from 14 farms and 11 locations were collected during two consecutive years (from 2019 to 2021) in the Czech Republic, Croatia and Slovakia. Samples were examined for infection with species of Cryptosporidium using microscopic (following aniline-carbol-methyl violet staining) and molecular tools. Overall, 0,5 % (3/603) and 2,8 % (17/603) of animals were positive for Cryptosporidium by microscopy and PCR, respectively. Phylogenetic analyses based on the small subunit of the rRNA gene revealed the presence of six Cryptosporidium species, namely C. meleagridis (n = 6), C. parvum (n = 4), C. baileyi (n = 2), C. muris (n = 2), C. ornithophilus (n = 2), C. galli (n = 1). Subtyping of isolates of C. parvum and C. melagridis based on sequence analysis of the 60-kDa glycoprotein gene revealed that they belong to the IIa and novel IIIk family, respectively. The isolate of Cryptosporidium meleagridis subtype family IIIk obtained from naturally infected pigeons and used for experiments was infectious for chickens (Gallus gallus f. domesticus) but not for SCID mice. The chickens began to shed oocysts 4 days after infection. The infection was detected in the duodenum, ileum and colon. Cryptosporidium spp. infection has been found frequently in domestic pigeons compared to wild life (2 =14,5919; d.f. = 1, P > 0,001). None of the naturally or experimentally infected birds developed clinical signs of cryptosporidiosis in the present study. Results of this study show that animals bred in hobby breeding and wild pigeons are infected by cryptosporidia and they should represent health risk for humans and domestic animals.
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OCCURRENCE AND GENOTYPISATION OF CRYPTOSPORIDIES IN OSTRICHES IN FARM BREED
HEJZLAROVÁ, Adéla
Avian cryptosporidiosis is a common parasitic disease caused by unicellular parasites belonging to the genus Cryptosporidium spp. These parasites have been described as a cause of various respiratory and gastrointestinal diseases of humans, farm and wild animals. Total of 204 faecal samples of common ostriches (Struthio camelus) were collected on four ostrich farms in the Czech Republic. Samples were screened for Cryptosporidium presence by microscopy and PCR/sequencing. Overall, 2.5 % (5/204) and 5.9 % (12/204) of animals were positive for Cryptosporidium by microscopy and PCR, respectively. Phylogenetic analysis of small subunit rRNA, actin and gp60 gene sequences showed the presence of Cryptosporidium avian genotype II (n=7) and C. ubiquitum IXa (n=5). Only ostriches infected with Cryptosporidium avian genotype II shed oocysts that were detectable by microscopy. Oocysts were purified from a pooled sample of four birds, characterised morphometrically and used in experimental infections to determine biological characteristics. Oocysts of Cryptosporidium avian genotype II measure 6.13 × 5.15 m, which is indistinguishable from C. baileyi and C. avium. Cryptosporidium avian genotype II was experimentally infectious for geese (Anser anser f. domestica), chickens (Gallus gallus f. domestica) and cockatiels (Nymphicus hollandicus), with a prepatent period of four, seven and eight days post infection, respectively and not infectious for mice (Mus musculus) and rats (Rattus norvegicus). The infection intensity ranged from 1,000 to 16,000 oocysts per gram. None of the naturally or experimentally infected birds developed clinical signs in the present study.
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Cryptosporidial infections of pets
MAROUŠOVÁ, Michaela
This thesis was focused on the incidence and prevalence of Cryptosporidium spp. in selected zoo boutiques and companion animals in České Budějovice. A total of 104 fecal samples of companion animals (Mus musculus, Rattus norvegicus, Cricetinae ssp., Cavia aperea porcellus, Oryctolagus cuniculus) were examined during 2011 to 2012. Samples were investigated by microscopic methods; positive samples were investigated by PCR. Cryptosporidium infections were observed only in mice (Mus musculus) and rats (Rattus norvegicus). There were detected two species of zoonotic cryptosporidia, C. tyzzeri and C. muris. The results show that animals bred in hobby breeding are infected by cryptosporidia and they should represent health risk for human.
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Intraspecies variability of Cryptosporidium parvum infecting calves
HOLUBOVÁ, Nikola
Faecal samples for parasitologic examination were obtained from ten farms. A total of 161 faecal specimens were fixed on a slide by methanol and stained with aniline-carbol-methyl violet. Of the 161 specimens, 47 (29,2 %) were positive for Cryptosporidium oocysts presence, namely Cryptosporidium parvum. The prevalence of C. parvum infection was highest in calves around 2 weeks of age, the erder ones were also recorded to be positive but in a weeker intensity. As the most risky management system was evaluated to be technology group housing and housing in ``Staimanové boudy{\crqq}, which were in close proximity and contact between neighboring calves was not prevented. In each breeding, only one type of subtype was detected.
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The occurrence of cryptosporidia in children
MACHOVCOVÁ, Petra
The aim of this thesis was to determine the incidence of unicellular parasites of the genus Cryptosporidium, which is the causative agent of cryptosporidiosis in children. 67 stool samples of children at the age of 1-15 were analyzed in this study. Out of it, 4 children were admitted to the infectious ward of the České Budějovice Hospital, Corp. Other samples were taken due to the suspicion of pathogene presence which causes these disorders, mainly abdominal pain, diarrhoea and fever. The samples were analyzed microscopically with the sedimentation method M.I.F.C., which serves for the intestinal parasite diagnosis, along with the staining method according to Miláček ?Vítovec, which is used to prove oocyst crystosporidia in the stool. They were also analyzed molecularly with the help of PCR. The cryptosporidia oocysts were not detected in any sample.
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