Zugl.: Berlin, Freie Universiẗat, Diss., 2008.

Submitted by Simone Souza (simonecgsouza@hotmail.com) on 2017-10-18T14:04:08Z No. of bitstreams: 1 DISS_2014_Selma Samiko Miyazaki Onuma.pdf: 1498739 bytes, checksum: 80569040824ee0dd609c6d63ad7c4fa7 (MD5) / Approved for entry into archive by Jordan (jordanbiblio@gmail.com) on 2017-11-08T12:33:06Z (GMT) No. of bitstreams: 1 DISS_2014_Selma Samiko Miyazaki Onuma.pdf: 1498739 bytes, checksum: 80569040824ee0dd609c6d63ad7c4fa7 (MD5) / Made available in DSpace on 2017-11-08T12:33:06Z (GMT). No. of bitstreams: 1 DISS_2014_Selma Samiko Miyazaki Onuma.pdf: 1498739 bytes, checksum: 80569040824ee0dd609c6d63ad7c4fa7 (MD5) Previous issue date: 2014-02-27 / Amostras de sangue de 11 onças-pintadas de vida livre (Panthera onca) foram coletadas em duas unidades de conservação federal no Pantanal de Mato Grosso com objetivo de avaliar o perfil sanitário dessas populações. A presença de anticorpos séricos para Ehrlichia spp., Rickettsia spp., Toxoplasma gondii, Neospora caninum e Sarcocystis neurona foi investigada pela Reação de Imunofluorescência Indireta (RIFI); para Leptospira spp. pela Microtécnica de Soroaglutinação Microscópica (SAM) e para Brucella abortus pelo teste do Antígeno Acidificado Tamponado (Rosa Bengala). O DNA genômico em cada amostra de sangue total coletada foi analisada pela Reação em Cadeia pela Polimerase (PCR) com intuito de identificar a presença de DNA de Ehrlichia spp., Rickettsia spp., Hepatozoon spp. e Cytauxzoon spp. Duas fêmeas de carrapato Amblyomma cajennense coletados em um animal foram também testadas pela PCR para detecção de DNA de Ehrlichia spp. e Rickettsia spp. Duas onças foram soropositivas para Ehrlichia spp., com títulos de anticorpos de 80 e 320, respectivamente. Nove animais reagiram positivamente a pelo menos quatro espécies de Rickettsia spp. Dez, 7 e 8 animais amostrados foram soropositivos para T. gondii, N. caninum e S. neurona, respectivamente. Duas onças apresentaram soros reagentes para Leptospira spp. , e o provável sorotipo infectante em ambos os animais foi um isolado brasileiro do sorovar Canicola (L01). Todas as amostras de soro foram negativas para B. abortus. Nenhum DNA da família Anaplasmataceae e do gênero Ehrlichia foi detectado pela PCR ou pela Heminested- PCR nas amostras de sangue, entretanto uma foi positiva na PCR para Rickettsia spp. segundo o gene citrate sintase (gltA), porém negativa para o gene ompA, comum entre riquétsias pertencentes ao grupo da Febre Maculosa. Um A. cajennense apresentou DNA riquetsial, cujo sequenciamento de nucleotídeos demonstrou 99% de similaridade a R. amblyommii. Sete amostras apresentaram DNA de Hepatozoon spp., com 99% de similaridade para H. felis e 10 das 11 amostras foram positivas para DNA de Cytauxzoon spp., com 99% de similaridade para C. felis. O presente estudo mostrou que as onças de vida livre na porção norte do Pantanal foram expostas a inúmeros agentes patogênicos, e que uma abordagem integrada para a conservação da vida selvagem e a integridade da saúde pública é uma questão eminente a fim de determinar intervenções de gestão em áreas protegidas. Este é o primeiro relato da exposição de onças-pintadas de vida livre a N. caninum e S. neurona. / Blood samples from 11 free-living jaguars (Panthera onca) were collected in two federal conservation units in the Pantanal of Mato Grosso state to determine the health profile of these populations . The presence of serum antibodies for Ehrlichia spp., Rickettsia spp., Toxoplasma gondii, Neospora caninum and Sarcocystis neurona was investigated by Immunofluorescence Assay (IFA); for Leptospira spp by the Microscopic Agglutination Test (MAT). and for Brucella abortus by rose Bengal test . Blood genomic DNA in each sample was tested by Polymerase Chain Reaction( PCR ) to identify Ehrlichia spp . , Rickettsia spp . , Hepatozoon spp . and Cytauxzoon spp. Two females of Amblyomma cajennense ticks collected from an animal were tested by PCR for DNA detection of Ehrlichia spp . and Rickettsia spp. Two jaguars were seropositive for Ehrlichia spp ., with antibody titres of 80 and 320, respectively. Nine animals were positive for at least four species of Rickettsia spp. Ten, 7 and 8 of the sampled animals were seropositive for T. gondii, N. caninum and S. neurona, respectively. Two jaguars were seroreactive for Leptospira spp. antigen and the most likely infecting serotype in both animals was a Brazilian isolate of serovar Canicola ( L01 ) . All serum samples were negative for B. abortus. No Anaplasmataceae and Ehrlichia DNA was detected by PCR or by Heminested-PCR. One sample was positive by PCR for Rickettsia spp. according to the citrate synthase gene (gltA), but it was negative for the ompA rickettsial gene of the spotted fever group. An A. cajennense collected showed riquetsial DNA whose sequence showed 99 % similarity to R. amblyommii . Seven samples presented DNA fragments of Hepatozoon spp., with 99% similarity to H. felis and 10 of the 11 samples were positive for DNA fragments of Cytauxzoon spp., with 99 % similarity to C. felis. The present study showed that free-living jaguars in northern Pantanal were exposed to numerous pathogens, and an integrated approach to wildlife conservation and integrity of public health is a prominent issue in order to determine management interventions in protected areas. This is the first report of exposure of free-living jaguars to N. caninum and S. neurona.

Submitted by Claudia Rocha (claudia.rocha@udesc.br) on 2018-02-19T14:47:21Z No. of bitstreams: 1 PGCA16MA200.pdf: 906352 bytes, checksum: a8a793c2fb4ce9135d1ad04376522869 (MD5) / Made available in DSpace on 2018-02-19T14:47:21Z (GMT). No. of bitstreams: 1 PGCA16MA200.pdf: 906352 bytes, checksum: a8a793c2fb4ce9135d1ad04376522869 (MD5) Previous issue date: 2016-02-25 / Among the pathogens foodborne are thermophilic bacteria of genus Campylobacter, gastroenteritis important agents of food-borne, in this case called campylobacteriosis. Campylobacter are defined as Gram-negative, curved or spiral, capnófilos with characteristic movement of corkscrew or comes and goes. They stand out as considerable thermophilic Campylobacter transmission sources for man meat and chicken giblets contaminated during handling and misguided slaughter operations, and cross-contamination of these with other foods that will be eaten raw. Due to the emphasis that the western region of Santa Catarina has the production and marketing of chicken, it was necessary to verify the occurrence of the pathogen in chilled carcasses of chickens slaughtered in this region, and its behavior to the seasons. For a period of 26 months, from January 2013, chicken carcass samples were weekly collected after the cooling process water in slaughterhouses under Federal Inspection of the three largest micro-regions in broiler slaughter number of the western region of Santa Catarina, totaling 808 samples. Campylobacter thermophiles was performed according to the methodology recommended by ISO 10272-1: 2006. Thermophilic Campylobacter were isolated in 1.82% (8/440) of the micro-region 1 of the samples, 4.95% (10/202) of samples of themicro-region 2 and 13.86% (23/166) samples of the micro-region 3 totaling 41 positive samples (5.07%) of the total samples collected. The index reported for the microregion 3 was statistically significant (P <0.05) when compared to the ratios found for the other two microregions. Although the prevalences have solid lower than expected when compared to the already published for this region, still provide risk to consumers, mainly through cross-contamination with food that will be consumed raw state, requiring greater controls on the production chain and slaughter of chickens. With respect to the seasons, the Campylobacter rates thermophilic were not statistically significant (P <0.05), however, it is necessary to evaluate for a longer period of time, since there was a numerically positive trend with respect to summer / Dentre os patógenos veiculados por alimentos, estão as bactérias termofílicas do gênero Campylobacter, importantes agentes de gastrenterite de origem alimentar, neste caso denominada de campilobacteriose. Campylobacter são definidos como bastonetes Gram-negativos, curvos ou espiralados, capnófilos, com movimento característico de saca-rolha ou em vai e vem. Destacam-se como consideráveis fontes de transmissão de Campylobacter termofílicos para o homem a carne e miúdos de frango, contaminadas durante a manipulação e operações de abate mal conduzidas, e a contaminação cruzada destes com outros alimentos que serão ingeridos crus. Devido ao destaque que a região oeste de Santa Catarina possui na produção e comercialização da carne de frango, fez-se necessário verificar a ocorrência do patógeno em carcaças resfriadas de frangos abatidos nesta região, e seu comportamento frente às estações do ano. Por um período de 26 meses, a partir de janeiro de 2013, foram semanalmente coletadas amostras de carcaça de frango, após o processo de resfriamento em água, em abatedouros sob Inspeção Federal das três maiores microrregiões em número de abate de frangos da região oeste de Santa Catarina, totalizando 808 amostras. Foi realizada a pesquisa de Campylobacter termofílicos conforme a metodologia recomendada pela ISO 10272-1:2006. Campylobacter termofílicos foram isolados em 1,82% (8/440) das amostras da microrregião 1, em 4,95% (10/202) das amostras da microrregião 2 e em 13,86% (23/166) amostras da microrregião 3, totalizando 41 amostras positivas (5,07%) do total de amostras coletadas. O índice relatado para a microrregião 3 foi estatisticamente significativo (P<0,05) quando comparado aos índices encontrados para as outras duas microrregiões. Embora as taxas encontradas tenham sidas abaixo das esperadas quando comparadas com as já publicadas para esta região, ainda fornecem riscos aos consumidores, principalmente por meio da contaminação cruzada com alimentos que serão consumidos crús, sendo necessários controles maiores na cadeia produtiva e abate dos frangos. Com relação às estações do ano, os índices de Campylobacter termofílicos não foram estatisticamente significativos (P<0,05), porém, faz-se necessário uma avaliação por um período maior de tempo, já que houve uma tendência numericamente positiva com relação ao verão

Submitted by Geandra Rodrigues (geandrar@gmail.com) on 2018-01-08T17:55:42Z No. of bitstreams: 1 letíciascafuttodefaria.pdf: 1095599 bytes, checksum: 1c63329ef1875b3086d3531350f36ccc (MD5) / Approved for entry into archive by Adriana Oliveira (adriana.oliveira@ufjf.edu.br) on 2018-01-23T11:12:07Z (GMT) No. of bitstreams: 1 letíciascafuttodefaria.pdf: 1095599 bytes, checksum: 1c63329ef1875b3086d3531350f36ccc (MD5) / Made available in DSpace on 2018-01-23T11:12:07Z (GMT). No. of bitstreams: 1 letíciascafuttodefaria.pdf: 1095599 bytes, checksum: 1c63329ef1875b3086d3531350f36ccc (MD5) Previous issue date: 2017-08-22 / O objetivo desse trabalho foi estimar a prevalência de Coxiella burnetii em Queijo Minas Artesanal da microrregião do Serro e identificar os fatores associados à presença desse microrganismo nas propriedades estudadas. Foram sorteados aleatoriamente 55 produtores cadastrados no Instituto Mineiro de Agropecuária (IMA) para participar da pesquisa. Os produtores responderam a um questionário e foi coletada uma amostra de queijo de cada propriedade para representá-la. Nas amostras de queijo coletadas, foram realizadas as análises de Reação de Cadeia da Polimerase (PCR) para C. burnetii e sequenciamento dos produtos de DNA amplificados. Os dados provenientes das entrevistas e resultados das análises laboratoriais, uma vez sistematizados, formaram a base de dados do trabalho. A presença de DNA de C. burnetii foi considerada a variável dependente e as variáveis explicativas foram divididas em três grupos hierárquicos: distal (socioeconômicas e demográficas), intermediário (produção de queijo) e proximal (características do rebanho, produção de leite e ordenha). Os fatores associados à presença de C. burnetii nos queijos foram identificados por análises de regressão logística univariada e multivariada. Do total de amostras analisadas (n = 53), cinco (9,43%) apresentaram DNA de C. burnetii confirmado por sequenciamento. Os fatores associados com a presença de DNA de C. burnetii nos queijos foram: uso do pingo (soro- fermento) para a fabricação dos queijos (OR= 12,09), tipo de ordenha (OR= (16,45) e número de vacas em lactação (OR= 1,05). As variáveis que permaneceram no modelo final como explicativas para presença de C. burnetii nos queijos foram: número de vacas em lactação, tipo de ordenha e uso do pingo. A presença de DNA de C. burnetii nos queijos artesanais enfatiza, por um lado, a necessidade de medidas de controle mais rigorosas para que essa bactéria não esteja presente nos rebanhos de propriedades produtoras de queijos artesanais. Por outro lado, o conhecimento dos fatores associados à presença de C. burnetii nos queijos artesanais poderá auxiliar no estabelecimento de medidas preventivas e na criação de novas normas e legislação para a produção de queijos artesanais, já que essa lacuna na legislação seria algo para reflexão, dada a importância da febre Q para a saúde pública. / The objective of this study was to estimate the prevalence of Coxiella burnetii in Artisan Minas cheese of region of Serro and identify the factors associated with the presence of this organism in the properties studied. Were drawn at random 55 registered producers in the Instituto Mineiro de Agropecuária (IMA) to participate in the research. The producers responded to a questionnaire and was collected a sample of cheese at a property to represent her. The cheese samples collected, the analysis of reaction of polymerase chain reaction (PCR) to C. burnetii and sequencing of amplified DNA products. Data from the interviews and results of laboratory tests, once organized, form the basis of work data. The presence of DNA of C. burnetii was considered as the dependent variable and the explanatory variables were divided into three groups: hierarchical distal (socioeconomic and demographic), intermediate (cheese production) and proximally (characteristics of the herd, milk production and milk). The factors associated with the presence of C. burnetii in cheeses were identified by analysis of univariate and multivariate logistic regression. Of the total samples analyzed (n = 53), five (9.43%) presented DNA of C. burnetii confirmed by sequencing. The factors associated with the presence of DNA of C. burnetii in cheeses were: use of the pingo (serum- baking) for the manufacture of the cheeses (OR = 12.09), type of milking (OR = (16.45) and number of lactating cows (OR = 1.05). The variables that remained in the final model as explanatory for the presence of C. burnetii in cheeses were: number of lactating cows, milking and type use the pingo. The presence of DNA of C. burnetii in artisanal cheeses emphasizes, on the one hand, the need for more stringent control measures so that this bacteria is not present in flocks of artisanal cheese-producing properties. On the other hand, the knowledge of the factors associated with the presence of C. burnetii in artisanal cheeses may assist in establishing preventive measures and the creation of new standards and legislation for the production of artisanal cheeses, since this gap in the legislation would be something for reflection, given the importance of Q fever to public health.

Submitted by Cássia Santos (cassia.bcufg@gmail.com) on 2017-03-31T11:35:53Z No. of bitstreams: 2 Dissertação - Fernanda Martins de Aquino - 2017.pdf: 2825789 bytes, checksum: 21dd668e99af981f702f5b0d0823a01c (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2017-04-03T12:12:17Z (GMT) No. of bitstreams: 2 Dissertação - Fernanda Martins de Aquino - 2017.pdf: 2825789 bytes, checksum: 21dd668e99af981f702f5b0d0823a01c (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-04-03T12:12:17Z (GMT). No. of bitstreams: 2 Dissertação - Fernanda Martins de Aquino - 2017.pdf: 2825789 bytes, checksum: 21dd668e99af981f702f5b0d0823a01c (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2017-03-08 / Amongst the several parasite-diseases that may be found on the bovine species slaughter line, cysticercosis surely is the most frequent one. Besides being a zoonosis and a public health issue, it also causes great economic losses on the Brazilian meat productive chain. On the other hand, fasciolosis, also considered a zoonosis, has a lower occurrence when compared to cysticercosis, fact that may be due to its epidemiologic dynamics. Aiming to learn the level of infection of these zoonosis in the bovine herd of the state of Goiás, this project was undertaken with the scope of determining the prevalence and spatial distribution, as well to evaluate the association with some epidemiologic variables with the occurrence of these diseases and also to estimate economic losses inflicted on the producers and industries. A retrospective study was carried out on a total of 23.255.979 animals slaughtered per year, mesoregions and microregions. The data were used to create one epidemiologic map for bovine cysticercosis and one for bovine fasciolosis, gathering all cities of the state of Goiás. A prevalence of bovine cysticercosis on the state of Goiás of 0,53% (CI 95% 0,5295 – 0,5354), where the percentage of viable cysticercosis was 42,31%, non-viable cysticercosis 57,69% and the presence of bovine fasciolosis was de 0,0026% (CI 95% 0,0024 – 0,0028). The mesoregion Centro presented a greater chance (OR = 4,44) of finding positive cattle for cysticercosis when compared to the mesoregions Norte (OR = 1) and Nordeste (OR = 1,02), whilst the mesoregions diagnosed with greater chances of finding animals positive for fasciolosis, due to having a greater effective herd (p ≤ 0,05), were Noroeste, Sul, part of the region Centro and two cities of the region Leste of the state. The losses estimated for the evaluated period, from 2007 to 2014, ranged from R$ 64.809.817,50 (US$ 20.574.545,24) due to the presence of cysticercosis, and around R$ 15.072,75 (US$ 4.785) due to the presence of Fasciola hepatica in bovine liver. Such results outline the importance of developing strategic measures and action policies to try and control the spreading of these relevant zoonosis. / Dentre os vários parasitos que podem ser encontrados na linha de abate da espécie bovina, é notório que a cisticercose é a de maior frequência. Além de ser uma zoonose e um problema de saúde pública, ainda causa grandes prejuízos econômicos na cadeia produtiva de carnes do Brasil. Por outro lado a fasciolose, também considerada uma zoonose, possui uma menor ocorrência quando comparada à cisticercose, fato que pode estar atribuído a sua dinâmica epidemiológica. Visando conhecer o nível de infecção destas zoonoses no rebanho bovino goiano, realizou-se este trabalho com o objetivo de determinar a prevalência e a distribuição espacial, além de avaliar a associação de algumas variáveis epidemiológicas com a ocorrência destas enfermidades nos animais e estimar as perdas econômicas causadas aos produtores e indústrias. Foi realizado um estudo do tipo retrospectivo referente a um total de 23.255.979 animais abatidos agrupados por ano, mesorregiões e microrregiões de Goiás. Os dados foram utilizados para criar um mapa epidemiológico da cisticercose bovina e um mapa epidemiológico da fasciolose bovina abrangendo todos os municípios goianos. A prevalência de cisticercose bovina no estado de Goiás foi de 0,53% (IC 95% 0,5295 – 0,5354), sendo a porcentagem de cisticercose viável 42,31% e cisticercose inviável 57,69% e a prevalência de fasciolose bovina de 0,0026% (IC 95% 0,0024 – 0,0028). A mesorregião Centro apresentou maior chance (OR = 4,44) de encontrar bovinos positivos para a cisticercose quando comparada as mesorregiões Norte (OR = 1) e Nordeste (OR = 1,02), enquanto as mesorregiões diagnosticadas com chances mais elevadas de se encontrar animais com fasciolose, porque continham maior rebanho efetivo (p ≤ 0,05) foram a Noroeste, Sul, parte da região Centro e dois municípios da região Leste do estado. As perdas estimadas para o período analisado, de 2007 a 2014, foram de R$ 64.809.817,50 reais (US$ 20.574.545,24 dólares) devido à presença da cisticercose no rebanho e cerca de R$ 15.072,75 reais (US$ 4.785 dólares) devido à presença de Fasciola hepatica no fígado de bovinos. Os resultados encontrados neste trabalho destacam a importância de se realizar medidas e ações políticas estratégicas na tentativa de controlar a disseminação destas importantes zoonoses.

Made available in DSpace on 2014-08-20T14:37:53Z (GMT). No. of bitstreams: 1 tese_samuel_felix.pdf: 483417 bytes, checksum: 782ad0a9939b35b83a255622221d018c (MD5) Previous issue date: 2013-02-01 / Leptospirosis is a disease caused by pathogenic spirochetes of the Leptospira genus. This zoonosis of worldwide distribution causes veterinarian and public health issues, especially in underdeveloped and developing countries with tropical and subtropical climates. In veterinary medicine, leptospirosis is important both as a clinical problem, causing illness in domestic animals, and an economic problem, causing productive and reproductive losses in commercial herds. Vaccination of these animals is applied, however protection conferred by these conventional vaccines is limited, and the carrier status is not always avoided. Recombinant outer membrane proteins seem to be the most promising antigens to replace the traditional bacterins (whole cell inactivated preparations), but thus far none of the tested proteins have turned satisfactory results. The goal of this study was to assess recombinant antigens and vaccine preparations, regarding their capability of producing protective immunity in hamsters, against lethal leptospirosis. Moreover, heterologous protection was sought, and assessed. The prevalence anti-Leptospira antibodies in stray dogs from the city of Pelotas was assessed using serogroups Icterohaemorrhagie and Canicola antigens. Several experiments were conducted to assess the protective potential of previously described leptospiral proteins. Twenty seven proteins were used to immunize hamsters which were then challenged with virulent Leptospira. Furthermore, leading vaccine candidates, LipL32 and LigB, were assessed regarding their protective potential when co-administered with traditional bacterins in previously established heterologous challenge experiments. A total of 28.96% of the animals tested were seropositive for the disease in the prevalence assay. Of the 27 antigens tried, two were shown to have some protective potential. Although no protection was demonstrated in the coadministration experiment, leptospiral bacterins seem to have some immunestimulating activity. / A leptospirose é uma doença causada por espiroquetas do gênero Leptospira. É uma zoonose de ampla distribuição geográfica, sendo um problema de saúde pública e veterinária, principalmente em países subdesenvolvidos e em desenvolvimento de clima tropical e subtropical. Em medicina veterinária, a leptospirose é uma doença importante tanto para a clínica quanto para a produção, devido ao risco à saúde pública, perdas reprodutivas e óbitos. A vacinação animal é realizada como medida de prevenção da enfermidade, entretanto, a proteção conferida pelas vacinas comerciais é limitada e não evita a condição de portador. Os antígenos protéicos da membrana externa parecem ser a melhor alternativa para substituir as vacinas atualmente disponíveis, porém, após diversos estudos, nenhum apresentou resultados satisfatórios. O objetivo deste trabalho foi avaliar antígenos recombinantes e preparações vacinais, capazes de conferir proteção de amplo espectro, em hamsters, contra leptospirose letal. A prevalência da leptospirose em cães da cidade de Pelotas foi aferida em um ensaio de diagnóstico sorológico usando como antígeno cepas dos sorogrupos Icterohaemorrhagie e Canicola. Em uma série de experimentos de prospecção de alvos, grupos de hamsters foram vacinados com diferentes proteínas recombinantes e posteriormente desafiados com cepa virulenta de Leptospira sp. Após o desenvolvimento de modelo para avaliação de proteção contra desafios heterólogos, as proteínas rLipL32 e rLigBNI foram avaliadas quando coadministradas com bacterinas (preparações de células inteiras inativadas por calor) em hamsters, sofrendo posterior desafio com quatro cepas de sorogrupos diferentes. Uma soroprevalência de 28,96% foi encontrada nos ensaios de prevalência. Duas de 27 proteínas recombinantes triadas foram identificadas como possíveis imunógenos. Apesar da falta de proteção demonstrada no experimento de coadministração de proteína e bacterina contra desafio homólogo ou heterólogo, a bacterina parece ter ação imunoestimulante.

Les maladies émergentes représentent actuellement 65% de toutes les pathologies infectieuses récentes. Récemment, un nombre croissant de nouveaux virus a été associé à de petits mammifères terrestres, plus particulièrement à des rongeurs, désignant ce groupe comme étant l'une des possibles sources de dangereuses pathologies émergentes et ré-émergentes. Actuellement, la réaction en chaîne par polymérase (PCR) est l'outil principal utilisé pour la détection d'agents pathogènes dans la diagnostique de routine et dans la recherche. Or, plusieurs recherches ont montré que certaines substances inhibent la PCR, causant de faux résultats. Aussi, nous avons lancé un programme de capture de rongeurs pour le dépistage de virus connus et non identifiés. Au total 1441 rongeurs ont été capturés pendant des campagnes organisées en Europe et Afrique entre 2002 et 2011. Tout d'abord, nous avons examiné l'inhibition de la PCR et étudié les différentes techniques de traitement d'échantillons qui favorisent la réduction de la quantité d'inhibiteurs dans les échantillons de rongeurs. Parmi les techniques d'extraction évaluées, l'EZ1 virus mini kit et le réactif d'extraction RNAnow se sont avéré plus efficaces que le NucleoSpin virus kit ou le réactif d'extraction TRIzol. De même, l'utilisation des poumons et de reins était préférable à l'utilisation du foie et de la rate. Aucune différence significative n'a été observée entre le stockage à -80°C et le stockage dans le réactif RNAlater. Nous avons conduit le dépistage des virus, en utilisant les tests moléculaires et la culture cellulaire. Deux nouvelles souches de virus ont été isolées, séquencées et caractérisées. / Emerging diseases currently represent 65% of recent major disease outbreaks. Of them, 75% are associated with wildlife. Recently, an increasing number of newly discovered viruses have been associated with small terrestrial mammals, particularly with rodents, pointing at this group as one of the most dangerous potential sources of emerging or re-emerging diseases. To meet these challenges for public health, a proper surveillance becomes necessary, which passes by detection of pathogens in human and risky groups of animals, including field investigations. Yet this can be achieved only by using proper techniques of samples treatment and pathogen detection. Currently, polymerase chain reaction (PCR) is the main tool used for the detection of pathogens in routine diagnostic and research. Yet, several researches showed that some substances can inhibit PCR, causing false-negative results. Therefore, we initiated a screening program targeting rodents for the presence of known and unidentified viruses. A total of 1441 rodents were trapped during field campaigns organized in Europe and Africa, between 2002 and 2011. At first we investigated on PCR inhibitors and discussed techniques of treatment of samples allowing reducing the influence of inhibitors in rodent samples. Among the extraction techniques tested, EZ1 virus mini kit and RNAnow extraction reagent were more effective than NucleoSpin virus kit or TRIzol extraction reagent. Also, the use of lungs and kidneys was preferable to the use of liver and spleen, the quantity of inhibitors being higher in the last two organs. No significant difference was observed between storage at -80°C, or in RNAlater RNA stabilization reagent.

L’encéphalopathie spongiforme bovine (ESB) est une maladie neuro-dégénérative fatale affectant les bovins ; elle est également une zoonose à l’origine du variant de la maladie de Creutzfeldt-Jakob. Identifiée pour la première fois au Royaume-Uni en 1986, cette maladie s’est rapidement étendue en Europe, malgré la mise en place de mesures de contrôle. En raison des particularités épidémiologiques de l’ESB (longue période d’incubation, âge précoce à l’infection et diagnostic post-mortem possible uniquement en fin d’incubation), l’évolution temporelle de l’exposition des bovins à l’ESB ne peut être appréhendée qu’à partir de la modélisation. Nous avons utilisé le modèle Age-Période-Cohorte afin de (ré)évaluer, en relation avec les principales mesures de contrôle, l’évolution de l’épizootie d’ESB à la lumière des données de surveillance les plus récente, en France, et dans six autres pays européens : Allemagne, Irlande, Italie, Pays-Bas, Pologne et Royaume-Uni. / Bovine spongiform encephalopathy is a fatal neurodegenerative disease affecting cattle and transmissible to humans as the cause of variant Creutzfeldt-Jakob disease. BSE was first identified in 1986 in United Kingdom, before spreading to European countries despite the implementation of control measures. Due to BSE epidemiological characteristics (long incubation period, early age at infection and post-mortem diagnostic at end stage of incubation period), time trend of BSE cattle exposure can only be estimated by modeling. We used age-period-cohort model in order to (re)evaluate, in relation to the main control measures, the trend of BSE epidemic, using the most recent surveillance data in France and six other European countries: Germany, Ireland, Italy, the Netherlands, Poland and United Kingdom.

Les protozooses digestives restent une des premières causes de morbidité, de malnutrition et de mortalité dans le monde. Cependant, la biologie de certains protozoaires entériques comme Blastocystis est mal connue et il reste négligé par les autorités sanitaires. Ce parasite colonise le tractus intestinal de l’Homme et de nombreux animaux. Son principal mode de transmission est la voie oro-fécale et sa prévalence peut dépasser 50% dans les pays en développement. Il présente une large diversité génétique avec 17 sous-types (STs) identifiés à ce jour. Un large faisceau de données récentes suggère que l’infection à Blastocystis est associée à une variété de troubles gastro-intestinaux et de l’urticaire.Dans le cadre de ma thèse, des études épidémiologiques ont été menées dans différents pays (Liban, Sénégal, France) afin de déterminer la prévalence de ce parasite dans la population humaine et identifier des facteurs de risque d’infection. En parallèle, à travers une enquête dans des zoos français, des réservoirs animaux de transmission zoonotique du parasite ont été proposés. D’autre part, les mécanismes impliqués dans la colonisation de l’hôte par Blastocystis ont été étudiés.Dans le cadre des enquêtes épidémiologiques, le parasite a été recherché dans les selles par PCR en temps réel et l’amplicon obtenu séquencé pour le sous-typage. La première étude menée au Liban a montré une prévalence de 19% dans la population générale mais cette prévalence atteint 60% dans une population d’écoliers vivant dans la même région. Une prévalence de 100% a été obtenue dans une cohorte d’enfants sénégalais. Ces fortes prévalences s’expliquent par des conditions d’hygiènes très précaires. Le ST3 était prédominant dans ces deux pays suivi des ST1 et ST2. Dans une étude multicentrique menée en France, une prévalence globale de 18,3% a été obtenue avec une prédominance du ST3 suivi des ST1, ST4 et ST2. Cette distribution est aussi celle observée dans une majorité de pays européens. Dans l’étude française, des variables (voyage récent, âge, saison) ont été identifiés comme des facteurs de risque de transmission du parasite. Le contact avec des animaux peut représenter un autre facteur de risque du fait du potentiel zoonotique du parasite. Dans une large étude épidémiologique réalisée dans deux zoos français sur plus de 160 espèces animales, la prévalence globale de Blastocystis dépasse 30% avec des variations importantes selon les groupes d’animaux. En comparant la distribution des STs entre l’Homme et les différents groupes d’animaux, les primates, les artiodactyles (bovins et cochons) et les oiseaux représenteraient les principaux réservoirs potentiels d’infection pour l’Homme.Une association entre l’infection à Blastocystis et l’appendicite a été mise en évidence chez une enfant au Maroc confirmant la pathogénie et le potentiel invasif et inflammatoire du parasite. De plus, 26 autres membres de sa famille ont présenté des symptômes digestifs suggérant une épidémie de blastocystose d’origine hydrique. L’hypothèse d’une relation entre ST de Blastocystis et pouvoir pathogène a été émise d’où l’intérêt d’une étude de génomique comparative afin d’identifier des facteurs de virulence pouvant être spécifiques d’un ST. A ce jour, aucune différence n’a pu être mise en évidence entre le génome de ST4 séquencé durant ma thèse et celui de ST7 disponible dans les bases de données alors qu’ils présentent une virulence différente in vitro. Enfin, l’impact de la colonisation par Blastocystis sur la composition du microbiote intestinal humain a été évalué. Une approche par séquençage à haut-débit a permis de comparer les compositions des microbiotes de patients infectés ou non par Blastocystis montrant une diversité bactérienne plus élevée chez les patients colonisés par le parasite. Ces données suggèrent que la colonisation par Blastocystis ne serait pas associée à une dysbiose intestinale généralement observée dans les maladies infectieuses intestinales. / Digestive protozoan infections are a major cause of morbidity, malnutrition and mortality worldwide. However, the biology of some enteric protozoa as Blastocystis is not well known and these microorganisms remain still neglected by the health authorities. Briefly, this parasite colonizes the intestinal tract of humans and various animals. Its main mode of transmission is the fecal-oral route and its prevalence can exceed 50% in developing countries. It exhibits a large genetic diversity with 17 subtypes (STs) identified to date. Recent data suggest that infection with Blastocystis is associated with a variety of gastrointestinal disorders and urticaria. As part of my thesis, epidemiological studies have been conducted in different countries (Lebanon, Senegal, France) to determine the prevalence of this parasite in the human population and identify risk factors for infection. In parallel, through a survey in French zoos, animal reservoirs of zoonotic transmission of Blastocystis have been proposed. Moreover, mechanisms involved in the colonization of the host by the parasite were studied.As part of, epidemiological, the parasite was identified in faecal samples by real-time PCR and the resulting amplicon was sequenced for subtyping. The first study conducted in Lebanon in the Tripoli area showed a prevalence of 19% in the general population but this prevalence reached 60% in a population of school children living in the same region. A prevalence of 100% was obtained in a cohort of Senegalese children. The high prevalence observed in these countries can be explained by poor hygiene conditions in connection with the faecal peril. In terms of distribution of STs, the ST3 was predominant in both countries followed by ST1 and ST2. In a multicenter study conducted in France, an overall prevalence of 18.3% was obtained with a predominance of ST3, followed by ST1, ST2 and ST4. This distribution is quite similar to that observed in most European countries. In the French study, parasite prevalence was significantly higher in summer than in winter. Other variables such as a recent trip and age have been identified as risk factors for transmission of the parasite. The contact with animals may represent another risk factor because of the zoonotic potential of the parasite. In a large epidemiological study conducted in two French zoos and including over 160 animal species, the overall prevalence of Blastocystis exceeds 30% with significant variations between animal groups. By comparing the distribution of STs between humans and different groups of animals, primates, artiodactyls (cattle and pigs) and birds represent major potential reservoirs of infection for humans.An association between infection with Blastocystis and appendicitis was demonstrated in a child in Morocco confirming the pathogenicity and invasive and inflammatory potential of the parasite. In addition, 26 other family members presented digestive symptoms suggesting waterborne outbreak of blastocystosis. The hypothesis of a relationship between Blastocystis ST and pathogenicity was suggested hence the interest of a comparative genomics study to identify virulence factors that may be present or absent for some STs. No difference was found between the ST4 genome sequenced during my thesis and the ST7 genome available in the database while these STs have different virulence in vitro. Finally, the unknown impact of colonization by Blastocystis on the composition of the human intestinal microbiota was evaluated. The compositions of the bacterial microbiota of 96 patients infected or not by Blastocystis were obtained by high-throughput sequencing and compared. A higher bacterial diversity was found in colonized patients compared to non-infected patients. These data suggest that colonization by Blastocystis would not be associated with dysbiosis generally observed in intestinal infectious diseases but rather to a healthy intestinal microbiota.

Introduction Hepatitis E is a worldwide distributed disease, which is caused by the hepatitis E virus (HEV). In addition to humans, domestic pigs, wild boars, rabbits and dromedaries can be subclinically infected as reservoir animals with the zoonotic HEV genotypes 3, 4 and 7. In addition, HEV and HEV-like viruses have been described sporadically in other mammals, as well as in birds and fish, although their distinct role as reservoirs or carriers of the virus is still unclear. Aims The aim of the study was therefore to analyse in more detail the importance of different mammalian species, which do not belong to the known HEV reservoirs, for the epidemiology of HEV infections, thus enabling a better assessment of the risk of virus transmission by these animal species. Material and Methods Fourteen non-human primate species and 66 other mammal species, as well as Norway rats (Rattus norvegicus) and feeder rats (Rattus norvegicus forma domestica) from German zoos were selected for the investigations. In total 259 individual non-human primate sera and 244 individual mammalian sera of clinically healthy zoo animals were analysed for the presence of HEV-specific antibodies (ab) using a species-independent double-antigen sandwich ELISA. The non-human primate sera were additionally examined using a commercial human ELISA. Real-time reverse-transcription (RT)-PCR, nested broad-spectrum RT-PCR and a rat HEV-specific RT-PCR were used to detect the HEV genome in sera of mammals and rat liver samples. A commercial and an in-house method were used for the DNA sequencing. Results HEV-specific ab were detected in 3.9% (10/259) of the non-human primate sera (4 species) and 11.5% (28/244) of the mammalian sera (16 species). The highest detection rates were recorded with 33.3% (9/27) in porcines and with 27.0% (10/37) in carnivores. HEV-RNA was detected in a clinically healthy female Syrian brown bear (Ursus arctos syriacus) and in 8 of the investigated Norway rats. Sequence analysis identified the virus as rat HEV; the viruses from the bear and the free-ranging rats from the same zoo showed a high nucleotide sequence identity (94.6%–97.8%). Because of the small number of samples due to the small populations within the individual zoos, further statistical evaluations were not carried out. Conclusions The results show that non-human primates in zoos may be infected with HEV or HEV-like viruses; however, the low ab detection rates together with the negative genome detection argue against a high risk of virus transmission to humans. The study in other zoo-housed mammalian species was able to significantly increase the number of animal species with indications of HEV infections. In most animal species, only rare evidence and low detection rates were available, which can best be explained by “spillover-infections”. In addition to the expected high detection rate in porcine species, the high percentage of HEV antibody-positive carnivores is remarkable. Their role as possible HEV reservoir animals should therefore be clarified in further investigations. The detection of rat HEV in the serum of the bear and its high nucleotide sequence identity with the HEVs of the pest rodents provides first evidence of transmission of this virus species between rodents and carnivores.:List of content List of figures List of tables List of abbreviations 1 General introduction 1.1 Discovery of HEV 1.2 Taxonomy 1.3 Morphology 1.4 Genomic organisation 1.5 Viral replication 1.6 Hepatitis E in humans 1.7 Tools for HEV diagnosis 1.8 Therapy 1.9 Animal infections with HEV and HEV-like viruses 1.10 Experimental infections of animals 1.11 Geographical distribution 1.12 Transmission pathways 1.13 Epidemiology 1.14 Prevention 2 Aims of the study 3 Publications 3.1 Publication I 3.1.1 Summary of Publication I 3.1.2 Key messages of Publication I 3.1.3 Own contribution to Publication I 3.2 Publication II 3.2.1 Summary of Publication II 3.2.2 Key messages of Publication II 3.2.3 Own contribution to Publication II 3.3 Publication III 3.3.1 Summary of Publication III 3.3.2 Key messages of Publication III 3.3.3 Own contribution to Publication III 4 General discussion 4.1 HEV infections in various animal species 4.2 Prevalence of natural HEV infections in non-human primates 4.3 Prevalence of natural HEV infections in other zoo-housed mammals 4.4 Transmission pathways of HEV in a zoo-setting 4.5 Risk of virus transmission from zoo animals to humans 5 Conclusion and perspectives 6 Summary 7 Zusammenfassung 8 References List of animals investigated in the study List of publications Acknowledgements / Einleitung Hepatitis E ist eine durch das Hepatitis E-Virus (HEV) verursachte, weltweit verbreitete Erkrankung. Neben dem Menschen können Hausschwein, Wildschwein, Kaninchen und Dromedar als Reservoirtiere subklinisch mit den zoonotischen HEV-Genotypen 3, 4 und 7 infiziert werden. Darüber hinaus wurden HEV und HEV-ähnliche Viren vereinzelt bei weiteren Säugetieren, sowie Vögeln und Fischen beschrieben, wobei deren genaue Rolle als Reservoir oder Überträger des Virus bislang unklar ist. Ziele Ziel der Arbeit war es deshalb, die Bedeutung verschiedener Säugetierarten, die nicht zu den bekannten HEV-Reservoiren gehören, für die Epidemiologie der HEV-Infektionen besser zu erfassen und dadurch das Risiko einer Virusübertragung durch diese Tierarten besser abzuschätzen. Material und Methoden Vierzehn Affenarten und 66 weitere Säugetierarten, sowie Wanderratten (Rattus norvegicus) und Futterratten (Rattus norvegicus forma domestica) aus deutschen Zoos wurden für die Untersuchungen ausgewählt. Insgesamt wurden 259 individuelle Affenseren und 244 individuelle Säugerseren klinisch gesunder Zootiere mittels eines Spezies-unabhängigen Doppel-Antigen-Sandwich-ELISAs auf das Vorhandensein von HEV-spezifischen Antikörpern (AK) untersucht. Die Affenseren wurden zusätzlich mittels eines kommerziellen humanen ELISAs untersucht. Real-time reverse-transcription (RT)-PCR, nested broad-spectrum RT-PCR sowie eine Ratten-HEV-spezifische RT-PCR wurden für den HEV-Genomnachweis in Seren der Säuger und in Ratten-Lebern verwendet. Für die DNA-Sequenzierungen wurden eine kommerzielle und eine In-house-Methode verwendet. Ergebnisse In 3,9% (10/259) der Affenseren (4 Arten) und 11,5% (28/244) der Säugerseren (16 Arten) wurden HEV-spezifische AK nachgewiesen. Die höchsten Nachweisraten wurden mit 33,3% (9/27) in Schweineartigen und 27,0% (10/37) in Fleischfressern ermittelt. HEV-RNA wurde in einer klinisch gesunden Syrischen Braunbärin (Ursus arctos syriacus), sowie in 8 der untersuchten Wanderratten nachgewiesen. Die Sequenzanalyse identifizierte das Virus als Ratten-HEV; die Viren aus der Bärin und aus den wildlebenden Ratten desselben Zoos zeigten eine hohe Nukleotidsequenz-Identität (94,6%–97,8%). Weitergehende statistische Auswertungen wurden wegen der geringen Probenzahlen aufgrund der kleinen Populationen innerhalb der einzelnen Zoos nicht durchgeführt. Schlussfolgerungen Die Ergebnisse zeigen, dass Affen in Zoos mit HEV oder HEV-ähnlichen Viren infiziert sein können, jedoch sprechen die geringen AK-Nachweisraten zusammen mit den negativen Genomnachweisen gegen ein hohes Übertragungsrisiko auf den Menschen. Die Studie an anderen Säugetierarten in Zoos konnte die Zahl der Tierarten mit Hinweisen auf HEV-Infektionen deutlich erhöhen. Bei den meisten Tierarten lagen nur seltene Nachweise und niedrige Detektionsraten vor, die am besten durch „Spillover-Infektionen“ erklärt werden können. Neben der erwarteten hohen Nachweisrate bei Schweineartigen ist der hohe Prozentsatz an HEV AK-positiven Fleischfressern bemerkenswert, weshalb deren Rolle als mögliche HEV-Reservoirtiere in weiteren Untersuchungen geklärt werden sollte. Der Ratten-HEV-Nachweis im Serum der Bärin, sowie dessen hohe Nukleotidsequenz-Identität zu den HEVs der Schadnager geben erstmals Hinweise auf eine Übertragung dieser Virusart zwischen Nagern und Fleischfressern.:List of content List of figures List of tables List of abbreviations 1 General introduction 1.1 Discovery of HEV 1.2 Taxonomy 1.3 Morphology 1.4 Genomic organisation 1.5 Viral replication 1.6 Hepatitis E in humans 1.7 Tools for HEV diagnosis 1.8 Therapy 1.9 Animal infections with HEV and HEV-like viruses 1.10 Experimental infections of animals 1.11 Geographical distribution 1.12 Transmission pathways 1.13 Epidemiology 1.14 Prevention 2 Aims of the study 3 Publications 3.1 Publication I 3.1.1 Summary of Publication I 3.1.2 Key messages of Publication I 3.1.3 Own contribution to Publication I 3.2 Publication II 3.2.1 Summary of Publication II 3.2.2 Key messages of Publication II 3.2.3 Own contribution to Publication II 3.3 Publication III 3.3.1 Summary of Publication III 3.3.2 Key messages of Publication III 3.3.3 Own contribution to Publication III 4 General discussion 4.1 HEV infections in various animal species 4.2 Prevalence of natural HEV infections in non-human primates 4.3 Prevalence of natural HEV infections in other zoo-housed mammals 4.4 Transmission pathways of HEV in a zoo-setting 4.5 Risk of virus transmission from zoo animals to humans 5 Conclusion and perspectives 6 Summary 7 Zusammenfassung 8 References List of animals investigated in the study List of publications Acknowledgements