Dissertation (M. Appl. Sc.) -- Lincoln University, 2008. / Also available via the World Wide Web.

Thèse--Sc. nat.--Paris 6, 1971. / C.N.R.S. A.O. 5995. Extrait des "Annales de parasitologie humaine et comparée", T. 49, 1974. Résumé en anglais. Bibliogr. p. 282-290.

Made available in DSpace on 2016-04-04T12:45:14Z (GMT). No. of bitstreams: 2 joao_neto_ioc_mest_2013.pdf: 1253667 bytes, checksum: d031c93a4aceeafda254d56260f64a77 (MD5) license.txt: 1748 bytes, checksum: 8a4605be74aa9ea9d79846c1fba20a33 (MD5) Previous issue date: 2013 / Fundação Oswaldo Cruz. Instituto Oswaldo Cruz. Rio de Janeiro, RJ, Brasil / Toxoplasma gondii é o agente causador da toxoplasmose, doença distribuída em grande parte do globo infectando vertebrados endotérmicos. T. gondii é um parasita intracelular obrigatório e não sintetiza colesterol, recrutando-o da célula hospedeira. Esse parasito é dividido em grupos, de acordo com a virulência, sendo a cepa RH e a cepa ME-49 representantes de parasitos de maior e de moderada virulência, respectivamente. Semelhante a outros parasitos protozoários, T. gondii expõem fosfatidilserina (PS) na membrana externa, sendo um exemplo do mimetismo apoptótico, o que tem sido sugerido como mecanismo de evasão. Corpúsculos lipídicos (CL) são organelas lipídicas comumente encontradas em uma gama de diferentes células, de plantas, parasitos e animais. CL são modulados positivamente por diferentes fatores, incluindo células apoptóticas, parasitos como Trypanosoma cruzi, Leishmania spp. e Plasmodium falciparum e citocinas, como o fator de transformação do crescimento (TGF-\03B2). Nosso trabalho visou investigar se a PS exposta na membrana plasmática de T. gondii pode induzir CL em macrófagos. Independente da virulência da cepa utilizada, T gondii foi capaz de induzir CL em macrófagos infectados. A indução foi maior com aumento da relação parasito/macrófago. Subpopulações de T. gondii, PS positivos (PS+) e PS negativo (PS-) foram obtidas e a subpopulação de PS + induziu mais CL em relação a subpopulação de PS-. A neutralização da PS dos parasitos PS+ ou da população total com anexina-V foi capaz de reduzir o número de CL nas células hospedeiras. Lipossomos contendo PS induziram mais CL em macrófagos quando comparados aos lipossomos sem PS Curiosamente, parasitos PS- também foram capazes de aumentar o número de CL em macrófagos, sugerindo a participação de outros fatores nesse processo indutor. Após a interação de macrófagos com T. gondii, observamos aumento no número de CL tanto em células que continham o parasita quanto em células vizinhas não parasitadas. Esse achado sugere uma sinalização 18 parácrina entre as células. O TGF-\03B2 é descrito como sendo secretado após infecção por T. gondii, e sua neutralização causou inibição da formação de CL induzido por T. gondii, revelando sua participação nesse processo. Concluímos que T. gondii é capaz de induzir CL em macrófagos, e essa indução tem a participação da PS exposta em sua membrana. A indução dessas organelas ocorreu independente da virulência da cepa. Esse processo é potencializado pela ação parácrina do TGF-\03B2 / Toxoplasma gondii is the causative agent of toxoplasmosis, a disease largely spread across the globe infecting almost any endothermic vertebrates. T. gondii is an obligatory intracellular par asite that can not synthesize cholesterol thus relying on recruiting it from the host cell. This parasite is divided into groups according to the virulence, and the RH strain and the ME - 49 strain representing highly and moderate virulen t parasites, respect ively. Similar to other protozoan parasites, T. gondii exposes phosphatidylserine (PS) on the outer membrane , an example of the apoptotic mimicry , which has been suggested as a mechanism of evasion ,. Lipid bodies (LB) are lipid organelles commonly found in a range of different cells of plants, animals and parasites. LB are positively modulated by different factors, including apoptotic cells, parasites such as Trypanosoma cruzi , Leishmania spp. and Plasmodium falciparum and cytokines such as transforming gro wth factor (TGF - β) . Our study aimed to investigate whether the PS exposed on the plasma membrane of T. gondii in macrophages can induce LB . T gondii was capable of inducing LB in infected macrophages, i ndependently of the virulence of the strain used . T he LB induction was greater with increase in parasite / macrophage ratio. Subpopulations of T gondii posi tive PS (PS +) and negative (PS - ) were obtained and PS+ subpopulation induced significantly greater LB formation compared to PS - subpopulation. The neutra lization of PS with annexin - V was able to reduce the number of LB o n PS+ parasites or on the total parasite population , but not on PS - parasites. Liposomes containing PS induced more LB than liposomes without PS. Interestingly, PS - parasites induced LB f ormation in macrophages, suggesting the involvement of other factors. After interaction of T. gondii with macrophages , we observed increased LB formation both in cells containing internalized parasite s as well as in bystander cells without internalized par asites , suggestive of a paracrine signaling. TGF - β is described as being secreted after infection with T. gondii and its neutralization caused inhibition of T. gondii - induced LB formation , pointing to TGF - β participation in this process. We conclude that T. gondii is able to induce LB formation in macrophages . The induction of these organelles was independent of strain virulence. T. gondii - induced LB formation occurs through signaling triggered by PS exposed on the parasite membrane. This process is also amplified by TGF - β a cting in a paracrine fashion

Orientador : Vicente Amato Neto / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-07-17T05:26:56Z (GMT). No. of bitstreams: 1 Pedro_RogeriodeJesus_D.pdf: 1876469 bytes, checksum: ec8d2fc68c1ee8168368c225705b27e7 (MD5) Previous issue date: 1976 / Resumo: Não informado / Abstract: Not informed / Doutorado / Doutor em Ciências Médicas

Indiana University-Purdue University Indianapolis (IUPUI) / Toxoplasma gondii is an obligate intracellular parasite of the phylum Apicomplexa. Toxoplasma infection is a serious threat to immunocompromised individuals such as AIDS patients and organ transplant recipients. Side effects associated with current drug treatment calls for identification of new drug targets. DNA repair is essential for cell viability and proliferation. In addition to reactive oxygen species produced as a byproduct of their own metabolism, intracellular parasites also have to manage oxidative stress generated as a defense mechanism by the host immune response. Most of the oxidative DNA damage is repaired through the base excision repair (BER) pathway, of which, the apurinic /apyrimidinic (AP) endonucleases are the rate limiting enzymes. Toxoplasma possesses two different AP endonucleases. The first, TgAPE, is a magnesium-dependent homologue of the human APE1 (hAPE1), but considerably divergent from hAPE1. The second, TgAPN, is a magnesium-independent homologue of yeast (Saccharomyces cerevisiae) APN1 and is not present in mammals. We have expressed and purified recombinant versions of TgAPE and TgAPN in E. coli and shown AP endonuclease activity. Our data shows that TgAPN is the more abundant AP endonuclease and confers protection against a DNA damaging agent when over-expressed in Toxoplasma tachyzoites. We also generated TgAPN knockdown Toxoplasma tachyzoites to establish that TgAPN is important for parasite protection against DNA damage. We have also identified pharmacological inhibitors of TgAPN in a high-throughput screen. The lead compound inhibits Toxoplasma replication at concentrations that do not have overt toxicity to the host cells. The importance of TgAPN in parasite physiology and the fact that humans lack APN1 makes TgAPN a promising candidate for drug development to treat toxoplasmosis.

No description available.

O Toxoplasma gondii é um parasito coccídio que se localiza intracelularmente em vários órgãos e tecidos de uma ampla gama de hospedeiros. O estudo da soroepidemiologia, deste parasito, na espécie felina é de grande relevância, pois o estreito convívio de seres humanos com esses animais pode acarretar na transmissão de algumas doenças como a toxoplasmose. Com o objetivo de contribuir com dados sobre a freqüência de anticorpos para Toxoplasma gondii em felinos domiciliados da cidade de Porto Alegre, os soros desses animais foram avaliados pelas técnicas de Hemaglutinação Indireta (HAI) e Reação de Imunofluorescência Indireta (RIFI). A freqüência de anticorpos de T. gondii para a amostragem em 245 soros felinos foi de 26,94% pela técnica de HAI e 37,96% pela técnica de RIFI. Dados epidemiológicos foram incluídos no trabalho, como gênero, raça, idade, acesso ou não à rua e tipo de alimentação. Estes parâmetros foram analisados estatisticamente para mensurar suas influências nos resultados obtidos com os testes. A percentagem de co-positividade e co-negatividade nas duas técnicas foi de 56% e 90%, respectivamente, e uma percentagem de concordância total de 77,5%, enquanto que o valor Kappa foi de 0.49. Este estudo mostra que os valores encontrados são relativamente altos, levando-nos a crer, que estes felinos, em algum momento de sua existência poderiam ser fonte de contaminação ambiental, como potenciais eliminadores de oocistos, principalmente aqueles que têm livre acesso à rua. / Toxoplasma gondii is an intracellular coccidian parasite that infects several organs and tissues in a large variety of hosts. Its seroepidemiology in feline species is of great value since the close relationship between human beings and cats may serve as a vector for the transmission of some diseases such as toxoplasmosis. The sera of 245 cats from Porto Alegre, southern Brazil, were submitted to indirect hemagglutination antibody (IHA) test and to indirect immunofluorescence (IIF) assay in order to determine the frequency of antibodies against Toxoplasma gondii. The IHA test showed that 26.94% of the cats had antibodies against Toxoplasma gondii compared to 37.96% in the IIF assay. Epidemiological data such as gender, race, age, access or not to the street and eating behavior were assessed. These parameters were statistically analyzed to measure the influence on test results. The co-positive and co-negative values amounted to 56 and 90% for the IHA test and IIF assay, respectively, yielding an overall agreement of 77.5% and a kappa coefficient of 0.49. The rates obtained by this study are relatively high, leading us to the assumption that these cats, mainly those with free access to the street, could be a source of environmental contamination, due to oocyst shedding, at some time over the course of their lifetime.

O Toxoplasma gondii é um parasito coccídio que se localiza intracelularmente em vários órgãos e tecidos de uma ampla gama de hospedeiros. O estudo da soroepidemiologia, deste parasito, na espécie felina é de grande relevância, pois o estreito convívio de seres humanos com esses animais pode acarretar na transmissão de algumas doenças como a toxoplasmose. Com o objetivo de contribuir com dados sobre a freqüência de anticorpos para Toxoplasma gondii em felinos domiciliados da cidade de Porto Alegre, os soros desses animais foram avaliados pelas técnicas de Hemaglutinação Indireta (HAI) e Reação de Imunofluorescência Indireta (RIFI). A freqüência de anticorpos de T. gondii para a amostragem em 245 soros felinos foi de 26,94% pela técnica de HAI e 37,96% pela técnica de RIFI. Dados epidemiológicos foram incluídos no trabalho, como gênero, raça, idade, acesso ou não à rua e tipo de alimentação. Estes parâmetros foram analisados estatisticamente para mensurar suas influências nos resultados obtidos com os testes. A percentagem de co-positividade e co-negatividade nas duas técnicas foi de 56% e 90%, respectivamente, e uma percentagem de concordância total de 77,5%, enquanto que o valor Kappa foi de 0.49. Este estudo mostra que os valores encontrados são relativamente altos, levando-nos a crer, que estes felinos, em algum momento de sua existência poderiam ser fonte de contaminação ambiental, como potenciais eliminadores de oocistos, principalmente aqueles que têm livre acesso à rua. / Toxoplasma gondii is an intracellular coccidian parasite that infects several organs and tissues in a large variety of hosts. Its seroepidemiology in feline species is of great value since the close relationship between human beings and cats may serve as a vector for the transmission of some diseases such as toxoplasmosis. The sera of 245 cats from Porto Alegre, southern Brazil, were submitted to indirect hemagglutination antibody (IHA) test and to indirect immunofluorescence (IIF) assay in order to determine the frequency of antibodies against Toxoplasma gondii. The IHA test showed that 26.94% of the cats had antibodies against Toxoplasma gondii compared to 37.96% in the IIF assay. Epidemiological data such as gender, race, age, access or not to the street and eating behavior were assessed. These parameters were statistically analyzed to measure the influence on test results. The co-positive and co-negative values amounted to 56 and 90% for the IHA test and IIF assay, respectively, yielding an overall agreement of 77.5% and a kappa coefficient of 0.49. The rates obtained by this study are relatively high, leading us to the assumption that these cats, mainly those with free access to the street, could be a source of environmental contamination, due to oocyst shedding, at some time over the course of their lifetime.

O Toxoplasma gondii é um parasito coccídio que se localiza intracelularmente em vários órgãos e tecidos de uma ampla gama de hospedeiros. O estudo da soroepidemiologia, deste parasito, na espécie felina é de grande relevância, pois o estreito convívio de seres humanos com esses animais pode acarretar na transmissão de algumas doenças como a toxoplasmose. Com o objetivo de contribuir com dados sobre a freqüência de anticorpos para Toxoplasma gondii em felinos domiciliados da cidade de Porto Alegre, os soros desses animais foram avaliados pelas técnicas de Hemaglutinação Indireta (HAI) e Reação de Imunofluorescência Indireta (RIFI). A freqüência de anticorpos de T. gondii para a amostragem em 245 soros felinos foi de 26,94% pela técnica de HAI e 37,96% pela técnica de RIFI. Dados epidemiológicos foram incluídos no trabalho, como gênero, raça, idade, acesso ou não à rua e tipo de alimentação. Estes parâmetros foram analisados estatisticamente para mensurar suas influências nos resultados obtidos com os testes. A percentagem de co-positividade e co-negatividade nas duas técnicas foi de 56% e 90%, respectivamente, e uma percentagem de concordância total de 77,5%, enquanto que o valor Kappa foi de 0.49. Este estudo mostra que os valores encontrados são relativamente altos, levando-nos a crer, que estes felinos, em algum momento de sua existência poderiam ser fonte de contaminação ambiental, como potenciais eliminadores de oocistos, principalmente aqueles que têm livre acesso à rua. / Toxoplasma gondii is an intracellular coccidian parasite that infects several organs and tissues in a large variety of hosts. Its seroepidemiology in feline species is of great value since the close relationship between human beings and cats may serve as a vector for the transmission of some diseases such as toxoplasmosis. The sera of 245 cats from Porto Alegre, southern Brazil, were submitted to indirect hemagglutination antibody (IHA) test and to indirect immunofluorescence (IIF) assay in order to determine the frequency of antibodies against Toxoplasma gondii. The IHA test showed that 26.94% of the cats had antibodies against Toxoplasma gondii compared to 37.96% in the IIF assay. Epidemiological data such as gender, race, age, access or not to the street and eating behavior were assessed. These parameters were statistically analyzed to measure the influence on test results. The co-positive and co-negative values amounted to 56 and 90% for the IHA test and IIF assay, respectively, yielding an overall agreement of 77.5% and a kappa coefficient of 0.49. The rates obtained by this study are relatively high, leading us to the assumption that these cats, mainly those with free access to the street, could be a source of environmental contamination, due to oocyst shedding, at some time over the course of their lifetime.

The protozoan parasite Toxoplasma gondii (T. gondii) is an important zoonotic pathogen, which has the ability to infect all warm blooded mammals including humans, with approximately one third of the human population predicted to be infected. Transmission of the parasite to the foetus during pregnancy can result in miscarriage, however, a child infected during pregnancy may go on to develop clinical symptoms such as retinochoroiditis (ocular toxoplasmosis), hydrocephalus or learning difficulties in later life. Post-natally acquired infection in humans is generally asymptomatic, however, individuals who are immunocompromised may develop ocular toxoplasmosis or toxoplasmic encephalitis. T. gondii type II is reported to be the predominant genotype in Europe and the United States, but currently very little information exists about the prevalence and genotypes present within Great Britain. Consumption of T. gondii tissue cysts from raw or undercooked meat is a main source of infection for humans, with infected pork being considered a high risk. Currently the “gold standard” for assessing the viability of infective T. gondii tissue cysts is by an in vivo mouse bioassay. However, more recent ethical requirements to reduce, refine or replace experimental animals raises the question as to whether molecular technologies could be incorporated into these studies to reduce mouse numbers. The main aims of this PhD were to: (i) determine the prevalence and genotypes of T. gondii within different wildlife populations and humans in Great Britain; (ii) determine whether vaccination of pigs with a live attenuated strain of T. gondii would reduce the load of viable T. gondii tissue cysts within this species; (iii) study the viability and dissemination of tissue cysts from oocyst and bradyzoite infected pigs and (iv) to compare mouse bioassay with molecular detection of T. gondii DNA from experimentally infected pigs. The main findings of this work show that the prevalence of T. gondii within carnivorous wildlife varied from 6.0% to 44.4% depending on the host species with type II being the predominant lineage identified, however, type III and two alleles for type I were also present. In humans, serological detection of the parasite from a group of Scottish blood donors from Glasgow and Dundee (n=1403) was determined at 13.0%, molecular detection of T. gondii in human brains (n=151) from the Sudden Death Brain Bank show a prevalence of 17.9%. A correlation between increasing age and an increase in the detection of parasite was identified from both study groups. T. gondii strain genotyping using DNA extracted from human brains identified alleles for type I and III, however, no direct link between cause of death and detection of parasite DNA could be made. Live vaccination and subsequent oocyst challenge of pigs showed a significant reduction in the establishment of viable T. gondii tissue cysts. Mouse bioassay clearly demonstrates this result, where 100% of mice that were inoculated with homogenised tissues from vaccinated/challenged pigs survived, compared to the survival of only 51% of mice, which received homogenised tissues from non-vaccinated/oocyst challenged animals. In addition, porcine tissues from pigs challenged with either oocysts or bradyzoites did not show a significant difference in mouse survival following bioassay of these tissues. Challenge with either stage of the parasite (oocysts or bradyzoites) showed a preference to form tissue cysts in brains and highly vascular muscles (tongue, diaphragm, heart or masseter) of pigs. The findings, comparing mouse bioassay with molecular detection of parasite DNA from homogenised porcine tissue (prior to inoculation into mice), showed similar levels of detection. However, mouse bioassay was more sensitive and also provides evidence of parasite viability. In conclusion, this research not only provides current figures for prevalence and genotypes of T. gondii in both wildlife and humans in Great Britain, it also successfully answers the question as to whether live vaccination of pigs with the S48 strain can reduce the tissue cyst burden. These promising results show the potential of a vaccine against T. gondii in producing safer pork for human consumption. Although the mouse bioassay still remains the most sensitive method for the detection and viability assessment of tissue cysts, further research should be carried out in this area, perhaps incorporating a technique such as magnetic capture qPCR, to enable an effective in vitro technique to be developed.