Orientador: Carlos Magno Castelo Branco Fortaleza / Resumo: Illicit drug users (IDUs) are recognized as a group at particular risk for staphylococci. In this population the first outbreak of community-associated Staphylococcus aureus (CAMRSA) was described. However, most studies relating S. aureus and MRSA to drug abuse address users of intravenous substances. In Brazil, most dependents are users of inhaled drugs, especially crack. A proportion of these IDUs have multiple passages through recovery clinics. Another institutionalized group still relevant in Brazil are patients with long-term hospitalizations in psychiatric hospitals. In spite of the psychiatric reform and its emphasis on deinstitutionalization, aspects of social vulnerability and abandonment by relatives also determine the residence of patients in psychiatric hospitals. The two groups described above are exposed to different epidemiological pressures and are both at higher risk of invasive staphylococcal disease and in a strategic position for maintenance and dissemination of isolates. The objective of this study was to identify the prevalence and predictors of the carrying of S. aureus and MRSA in IDUs admitted to the Hospital of Reference for Alcohol and Drugs (SARAD) and patients residing in the Psychiatric Hospital Cantídio de Moura Campos, both located in Botucatu , São Paulo State, Brazil. Molecular characterization of MRSA isolates was performed. A total of 220 subjects were enrolled, 138 from the SARAD and 82 from the psychiatric hospital. The prevalences of S. ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Usuários de drogas ilícitas (UDI) são reconhecidos como grupo de especial risco para estafilococcias. Nessa população foi descrito o primeiro surto de Staphylococcus aureus associados à comunidade (CA-MRSA). No entanto, a maior parte dos estudos relacionando S. aureus e MRSA a drogadição aborda usuários de substâncias endovenosas. No Brasil, a maior parte de dependentes é usuário de drogas inalatórias, especialmente o crack. Uma parcela destes dependentes tem múltiplas passagens por clínicas de recuperação. Um outro grupo institucionalizado ainda relevante no Brasil são os pacientes com internações de longa permanência em hospitais psiquiátricos. A despeito da reforma psiquiátrica e sua ênfase na desinstitucionalização, aspectos de vulnerabilidade social e abandono por familiares determinam ainda a residência de pacientes em hospitais psiquiátricos. Os dois grupos descritos acima são expostos a diferentes pressões epidemiológicas e estão tanto em maior risco de doença estafilocóccica invasiva quanto em posição estratégica para manutenção e disseminação de isolados. Este estudo teve por por objetivo identificar a prevalência e fatores preditores do carreamento de S. aureus e MRSA em UDI internados no Serviço Hospitalar de Referência de Álcool e Drogas (SARAD) e pacientes residentes no Hospital Psiquiátrico Cantídio de Moura Campos, ambos localizados em Botucatu, SP. Caracterização molecular de isolados de MRSA foi realizada. Foram estuados 220 sujeitos, 138 do SARAD e 82 do ho... (Complete abstract click electronic access below) / Mestre

Submitted by LIVIA MARIA GONÇALVES ROSSI null (liv.rossi@yahoo.com) on 2016-01-25T14:04:25Z No. of bitstreams: 1 TESE_HCV Multi-region_LIVIA ROSSI.pdf: 6853283 bytes, checksum: d2ce231bc1292f9fa0469f971cf0856b (MD5) / Approved for entry into archive by Juliano Benedito Ferreira (julianoferreira@reitoria.unesp.br) on 2016-01-25T17:23:43Z (GMT) No. of bitstreams: 1 rossi_lmg_dr_sjrp.pdf: 6853283 bytes, checksum: d2ce231bc1292f9fa0469f971cf0856b (MD5) / Made available in DSpace on 2016-01-25T17:23:43Z (GMT). No. of bitstreams: 1 rossi_lmg_dr_sjrp.pdf: 6853283 bytes, checksum: d2ce231bc1292f9fa0469f971cf0856b (MD5) Previous issue date: 2016-01-18 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O vírus da Hepatite C (HCV) afeta cerca de 3% da população mundial. A cada ano, 3-4 milhões de novos casos são diagnosticados. A identificação de redes transmissão é complexa devido ao longo período de incubação, à falta de sintomas na fase aguda da doença e à heterogeneidade do HCV, que dificulta o estabelecimento de vínculos entre casos relacionados. Uma ampla caracterização das populações intra-hospedeiros pode ser realizada de forma eficiente através do sequenciamento de nova geração (NGS). Com base neste contexto, o sequenciamento de múltiplas regiões subgenômicas é uma solução às limitações impostas pela rápida evolução molecular do HCV. Variantes virais das regiões HVR1 e NS5A de 16 pacientes cronicamente infectados com o HCV, genótipos 1a e 1b, foram sequenciadas com a técnica de NGS. Os pacientes 1-7 compartilhavam fatores de risco, pertencendo ao mesmo grupo de usuários de drogas injetáveis, porém o parentesco genético desses casos não pode ser estabelecido com base apenas no sequenciamento da HVR1 (distância nucleotídica mínima entre 16-23). A amplificação de um fragmento maior (~450 pb), correspondente a um segmento da região NS5A, aprimorou a relação epidemiológica entre os pacientes 1-5, onde as distancias genéticas mínimas foram consideravelmente menores (9-13). Os pacientes 6 e 7 não compartilharam sequências com os outros cinco pacientes dessa rede, apresentando populações virais mais homogêneas. Adicionalmente, Median Joining Networks foram construídas para melhor analisar a variabilidade genética intra-hospedeiro. Em geral, observou-se que as sequências derivadas da NS5A formaram comunidades mais homogêneas e menos divergentes geneticamente. Assim, a tecnologia NGS e o sequenciamento das regiões subgenômicas HVR1 e NS5A podem ajudar a restaurar elos perdidos quando somente a região HVR1 é analisada, aprimorando portanto, a resolução de estudos de associação genética entre populações de HCV. / The hepatitis C virus (HCV) affects approximately 3% of the world's population. Each year 3-4 million new cases are diagnosed. The identification of transmission networks is complicated due to the characteristic long incubation period, the lack of symptoms during the acute phase of the disease and the heterogeneity of HCV, making it challenging to link related cases to a common source of infection. Extensive characterization of intra-host populations can be reliably archived using next generation sequencing (NGS) approaches. Sequencing of multiple and longer subgenomic regions has been proposed as an alternative to overcome the limitations imposed by the rapid molecular evolution of the HCV HVR1. Thus, the NS5A and HVR1 regions of 16 chronically infected individuals, genotypes 1a and 1b, were sequenced using a NGS platform. Patients 1-7 shared risk factors and belonged to the same injection drug users network. However, genetic relatedness could not be established based on the HVR1 sequences (minimal nucleotide distance ranging from 16-23). Amplification and sequencing of a larger PCR fragment (~450 bp) targeting the NS5A region reestablished lost epidemiological links between patients 1-5. The minimum genetic distances in those patients were considerable smaller than the HVR1 counterparts (9-13). Patients 6 and 7 displayed a rather homogeneous viral population and were clearly not sharing any sequences with all other five patients in this network. Additionally, Median Joining Networks analysis was carried out to further analyze the intrahost genetic variability of all seven patients. Overall, NS5A sequences were significantly less diverse than their HVR1 equivalents. Thus, NGS technology and use of both HVR1 and NS5A sequences might help restored otherwise lost links when the HVR1 region alone is analyzed, improving the resolution of HCV genetic relatedness studies. / CAPES: 33004153079P9

Orientador: Paula Rahal / Resumo: O vírus da Hepatite C (HCV) afeta cerca de 3% da população mundial. A cada ano, 3-4 milhões de novos casos são diagnosticados. A identificação de redes transmissão é complexa devido ao longo período de incubação, à falta de sintomas na fase aguda da doença e à heterogeneidade do HCV, que dificulta o estabelecimento de vínculos entre casos relacionados. Uma ampla caracterização das populações intra-hospedeiros pode ser realizada de forma eficiente através do sequenciamento de nova geração (NGS). Com base neste contexto, o sequenciamento de múltiplas regiões subgenômicas é uma solução às limitações impostas pela rápida evolução molecular do HCV. Variantes virais das regiões HVR1 e NS5A de 16 pacientes cronicamente infectados com o HCV, genótipos 1a e 1b, foram sequenciadas com a técnica de NGS. Os pacientes 1-7 compartilhavam fatores de risco, pertencendo ao mesmo grupo de usuários de drogas injetáveis, porém o parentesco genético desses casos não pode ser estabelecido com base apenas no sequenciamento da HVR1 (distância nucleotídica mínima entre 16-23). A amplificação de um fragmento maior (~450 pb), correspondente a um segmento da região NS5A, aprimorou a relação epidemiológica entre os pacientes 1-5, onde as distancias genéticas mínimas foram consideravelmente menores (9-13). Os pacientes 6 e 7 não compartilharam sequências com os outros cinco pacientes dessa rede, apresentando populações virais mais homogêneas. Adicionalmente, Median Joining Networks foram construídas para ... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The hepatitis C virus (HCV) affects approximately 3% of the world's population. Each year 3-4 million new cases are diagnosed. The identification of transmission networks is complicated due to the characteristic long incubation period, the lack of symptoms during the acute phase of the disease and the heterogeneity of HCV, making it challenging to link related cases to a common source of infection. Extensive characterization of intra-host populations can be reliably archived using next generation sequencing (NGS) approaches. Sequencing of multiple and longer subgenomic regions has been proposed as an alternative to overcome the limitations imposed by the rapid molecular evolution of the HCV HVR1. Thus, the NS5A and HVR1 regions of 16 chronically infected individuals, genotypes 1a and 1b, were sequenced using a NGS platform. Patients 1-7 shared risk factors and belonged to the same injection drug users network. However, genetic relatedness could not be established based on the HVR1 sequences (minimal nucleotide distance ranging from 16-23). Amplification and sequencing of a larger PCR fragment (~450 bp) targeting the NS5A region reestablished lost epidemiological links between patients 1-5. The minimum genetic distances in those patients were considerable smaller than the HVR1 counterparts (9-13). Patients 6 and 7 displayed a rather homogeneous viral population and were clearly not sharing any sequences with all other five patients in this network. Additionally, Median Joining... (Complete abstract click electronic access below) / Doutor

Submitted by Marlene Santos (marlene.bc.ufg@gmail.com) on 2016-05-04T18:41:09Z No. of bitstreams: 2 Dissertação - Denisy Marques Mendanha de Oliveira - 2013.pdf: 1154542 bytes, checksum: e32fe195aaa52077c2e1d030caeadea3 (MD5) license_rdf: 19874 bytes, checksum: 38cb62ef53e6f513db2fb7e337df6485 (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2016-05-05T13:05:48Z (GMT) No. of bitstreams: 2 Dissertação - Denisy Marques Mendanha de Oliveira - 2013.pdf: 1154542 bytes, checksum: e32fe195aaa52077c2e1d030caeadea3 (MD5) license_rdf: 19874 bytes, checksum: 38cb62ef53e6f513db2fb7e337df6485 (MD5) / Made available in DSpace on 2016-05-05T13:05:48Z (GMT). No. of bitstreams: 2 Dissertação - Denisy Marques Mendanha de Oliveira - 2013.pdf: 1154542 bytes, checksum: e32fe195aaa52077c2e1d030caeadea3 (MD5) license_rdf: 19874 bytes, checksum: 38cb62ef53e6f513db2fb7e337df6485 (MD5) Previous issue date: 2013-04-17 / The calicivirus (CV) are important etiologic agents of acute gastroenteritis, affecting people of different ages and in various parts of the world. The present study aimed at the detection and molecular characterization of caliciviruses - norovirus (NoV) and sapovirus (SaV) - from fecal samples from children attending a daycare in Goiânia, Goiás. Were collected and analyzed 539 fecal samples 56 children from October 2009 to October 2011. The viral detection and genotyping was done by polymerase chain reaction post-transcription (RT-PCR) and by genomic sequencing using primers designed for partial regions of the viral capsid. From the 56 children participants of the study, 29 (51.8%) had at least one positive sample for CV during the study period. Positive children, 21 (37.5%) had at least one sample for NoV, 16 (28.6%) for SaV and eight (14.3%) positivity to both the virus. The CV were detected in children of both genders, and a higher positivity rate was detected among children aged 13 to 24 months, when compared to the other age groups. A significantly higher positivity for CV was observed in samples from asymptomatic children, and a defined seasonality pattern was not detected for these agents. In terms of molecular characterization for NoV the circulation of samples GII.6, GII.2, GII.1, GII.4, GI.1 and GI.7 was observed, whereas for SaV samples were detected GI.1 and GI.3. During the study period five outbreaks took place, being three of NoV, GII.6, GII.2 and GII.1, two SaV GI.1 and GI.3. Two children presented by prolonged NoV excretion, one for two months and another for 23 days. Twelve (20.4%) children had more than infection for NoV and / or SaV. This study showed circulation of CV at the daycare, a higher rate of infection in asymptomatic children, and prolonged de NoV excretion, which may have contributed to the occurrence of outbreaks and to the dissemination of these agents in such environment. / Os calicivírus (CV) são importantes agentes etiológicos da gastroenterite aguda, acometendo pessoas de diversas idades e em várias partes do mundo. O presente estudo teve como objetivo a detecção e caracterização molecular de amostras de calicivírus - norovírus (NoV) e sapovírus (SaV) - provenientes de amostras fecais de crianças frequentadoras de uma creche em Goiânia, Goiás. Foram coletadas e analisadas 539 amostras fecais de 56 crianças no período de outubro de 2009 a outubro de 2011. A detecção viral e genotipagem foi feita pela reação em cadeia pela polimerase pós-transcrição (RT-PCR) e por sequenciamento genômico, utilizando iniciadores desenhados para regiões parciais do capsídeo viral. Das 56 crianças participantes do estudo, 29 (51,8%) apresentaram pelo menos uma amostra positiva para CV durante o período do estudo. Das crianças positivas, 21 (37,5%) apresentaram pelo menos uma amostra para NoV e 16 (28,6%), para SaV, sendo que 8 (14,3%) apresentaram positividade para ambos os vírus. Os CV foram detectados em crianças de ambos os gêneros, sendo observada uma maior positividade entre as crianças com idade entre 13 a 24 meses. Foi observada ainda detecção significativamente maior nas crianças assintomáticas não tendo sido observado padrão definido de sazonalidade para esses agentes. Em termos da caracterização molecular para NoV, foi observado circulação de amostras GII.6, GII.2, GII.1, GII.4, GI.1 e GI.7 enquanto que para SaV as amostras detectadas foram GI.1 e GI.3. Durante o período do estudo foi observado a ocorrência de cinco surtos sendo três deles por NoV, amostras, GII.6, GII.2 e GII.1, e dois por SaV, GI.1 e GI.3. Duas crianças apresentaram excreção prolongada por NoV, uma por dois meses e outra por 23 dias. Doze (21,4%) crianças apresentaram mais de uma infecção por NoV e/ou SaV. Este estudo mostrou circulação de CV no ambiente de creche, maior índice de infecção em crianças assintomáticas, além de excreção prolongada de NoV, o que pode ter contribuído para a ocorrência de surtos e a disseminação destes agentes naquele ambiente.

La résistance bactérienne aux antibiotiques est devenue un problème majeur de santé publique impliquant des actions de surveillance et de lutte contre sa diffusion. L’épidémiologie de la résistance aux antibiotiques au sein des pathogènes cliniques est indispensable notamment à la prise en charge thérapeutique. Cependant, elle est également pertinente au sein des bactéries animales et environnementales afin d’en apprécier l’ampleur et d’en appréhender la diffusion. Alors que de nombreux travaux ont été conduits sur le microbiote des animaux de compagnie, les études portant sur la faune sauvage restent rares. Or, il apparaît opportun de l’intégrer dans l’étude de la dynamique des bactéries antibiorésistantes afin d’apprécier son rôle épidémiologique dans leur dissémination et d’évaluer les risques zoonotiques qui en découlent.Sur la base de notre revue de la littérature, nous avons mis en évidence un lien étroit existant entre les activités humaines et la présence de bactéries antibiorésistantes dans la faune sauvage. Ceci nous a conduits à discuter de l’existence probable de voies d’échanges entre le compartiment humain et animal.Sur la base d’une étude ayant démontré la présence dans le sud de la France d’un réservoir aviaire d’Escherichia coli producteurs de béta-lactamases à spectre élargi, nous avons exploré le microbiote cloacal de deux espèces de goélands, différentes par leurs niches écologiques et leur mode d’alimentation, en tant que réservoir potentiel de bactéries multirésistantes aux antibiotiques.Dans un premier temps nous nous sommes intéressés à la présence de Proteus mirabilis producteurs d'AmpC acquises dans le microbiote des goélands au cours des deux années d’étude. Ces isolats étaient producteurs de céphalosporinases de type CMY-2 dont le support génétique était un élément intégratif et conjugatif (ICE) de la famille SXT/R391-like. Deux souches cliniques humaines avaient les mêmes enzymes, supports et fond génétiques que des souches aviaires. Ceci permet de supposer que ces goélands constituent un réservoir de P. mirabilis porteurs du gène blaCMY-2, et que les structures de type ICE SXT/R391-like joueraient un rôle important dans la dissémination et la persistance de ce gène de résistance.Nous avons également isolé des souches d’Escherichia coli productrices de carbapénèmases acquises, qui constituent actuellement l'une des menaces les plus préoccupantes pour la santé publique en termes d'antibiorésistance. Ces souches proviennent uniquement de goélands leucophées et sont porteuses du gène blaVIM-1. L’analyse de leur patrimoine génétique montre qu’elles sont liées à des souches humaines sensibles. Nous n’avons en effet pas isolé de souches humaines productrices de carbapénèmases de type VIM dans le même temps. Cette découverte pose la question d’un réservoir aviaire potentiel et d’une menace de diffusion.Lors du screening nous avons identifié une souche de Vibrio cholerae non-O1/non-O139 résistante aux carbapénèmes et provenant d’un goéland leucophée. Elle possédait des gènes blaVIM-1 et blaVIM-4 qui faisaient partie d’un integron de classe 1, situé sur un plasmide IncA/C. Il s’agit de la première description d’une souche de V. cholerae productrice de ce type de carbapénèmases. Ce travail démontre la complexité de la circulation de l’antibiorésistance au sein du microbiote étudié. Il ouvre de nombreuses perspectives d’un point de vue épidémiologique mais également fondamental sur les mécanismes et les supports génétique de cette antibiorésistance. En effet, il illustre bien les apports importants des outils d’épidémiologie moléculaire dans la surveillance de l’émergence et la compréhension de la dynamique de transmission et de diffusion des bactéries multirésistantes dans la faune sauvage. / Bacterial resistance has become a major public health problem leading to a strengthening of spread surveillance and control. The epidemiology of antimicrobial resistance (AMR) in clinical pathogens is essential for therapeutic management. It is also relevant in animal and environmental bacteria to determine and understand AMR existence and diffusion. While much work has been done on the microbiota of companion animals, studies involving wildlife are scarce. It is essential to consider wildlife when studying AMR dynamics to assess its epidemiological role in AMR spread and understand the zoonotic risk which ensues from it.With our literature review, we highlight the close link between human activities and the presence of AMR in wildlife. It led us to discuss the pathways between the human and animal compartments.A previous study reported the presence of an avian reservoir of extended spectrum beta-lactamases-producing Escherichia coli in the South of France. Based on this finding, we explored the microbiota of two gull species, differentiated by their ecological niches and diet, as a potential reservoir of AMR.First, we investigated the presence of acquired AmpC-producing Proteus mirabilis in the gulls’ microbiota over two years. The isolates produced CMY-2 cephalosporinases with the genetic support of an integrative and conjugative element (ICE) which belongs to the SXT/R391-like family. Two human strains had the same enzymes, genetic support and genetic background as the avian isolates. This suggests that these gulls may act as a reservoir of blaCMY-2-carrying P. mirabilis, and the SXT/R391-like ICEs may play an important role in this gene’s dissemination and persistence.We also isolated acquired carbapenemases-producing E. coli, which is currently one of the most serious AMR threats to public health. These strains, which carried the blaVIM-1 gene, were recovered from yellow-legged gulls. The phylogenetic analyses showed that the gulls are significantly linked with human susceptible isolates. However, VIM carbapenemase producing-human isolate was not isolated in the same time period. This discovery raises the question of a potential avian reservoir and the threat of diffusion.During the screening, we identified a carbapenem resistant non-O1/non-O139 Vibrio cholerae strain, recovered from a yellow-legged gull. It carried both blaVIM-1 and blaVIM-4 genes which were part of a class 1 integron structure located in an IncA/C plasmid. This is the first description of a V. cholera strain producing this type of carbapenemase.This work demonstrates the complexity of the AMR circulation in the microbiota studied. It opens many perspectives from an epidemiological and fundamental point of view on the mechanisms and genetic supports of AMR. It further illustrates the contribution of molecular epidemiology tools in the understanding of the dynamics of transmission and diffusion and the surveillance of the emergence of AMR in wildlife.

Toxoplasma gondii est un parasite intracellulaire obligatoire potentiellement capable d’infecter tous les animaux homéothermes. L’Amérique du Sud, et plus particulièrement le Brésil occupe une place particulière dans l’épidémiologie de ce parasite cosmopolite en raison d’une part de formes cliniques sévères observées chez l’Homme et d’autre part d’une diversité génétique du parasite, sans équivalent jusqu’à présent sur d’autres continents. L’Etat de Rio de Janeiro présente des environnements très contrastés allant d’une ville capitale comportant différents degrés d’urbanisation à des zones isolées de haute altitude, habitat d’une faune très riche.Nous avons cherché à étudier l’influence de facteurs anthropiques et physiques sur l’épidémiologie du parasite dans différents environnements de l’Etat de Rio de Janeiro. Pour cela, nous avons d’une part étudié la séroprévalence chez les chats dans deux environnements avec différents degrés d’urbanisation de la ville de Rio, d’autre part mené des études d’épidémiologie moléculaire à l’aide de prélèvements sur des animaux de la ville de Rio, d’un parc national situé dans les zones de hautes altitudes et des zones rurales avoisinantes.L’étude de séroprévalence chez les chats dans la ville de Rio a permis de montrer l’intérêt de l’utilisation des prélèvements sur papier-filtre pour la réalisation de la technique de Modified-Agglutination-Test (MAT). Elle a révélé une différence significative de prévalence entre les chats errants du quartier résidentiel privé (4/107- 3,74%) et ceux du refuge municipal (32/265-12,08%). La densité des animaux et d’autres facteurs écologiques peuvent expliquer cette différence.La détection d’ADN toxoplasmique a été positive chez 8/16 chats de la ville de Rio, 14/18 animaux domestiques et 23/33 animaux sauvages de la zone d’amortissement du parc, 31/38 petits rongeurs ou marsupiaux piégés dans les zones de haute altitude. La détection a été positive sur les tissus de 3 félidés sauvages de la zone d’amortissement et sur les fèces d’un Puma yagouaroundi retrouvées dans le parc témoignant du rôle de ces félidés sauvages dans la contamination du sol aussi bien en zone préservée que dans les zones rurales proches du parc.Le génotypage par 15 marqueurs microsatellites a été possible pour 6 échantillons (dont une souche viable). Il a révélé une grande diversité génétique, mais aussi la présence aussi bien dans le parc qu’au centre-ville de Rio de souches appartenant aux lignées brésiliennes majeures BRII et BRIII. / Toxoplasma gondii is a protozoan parasite, potentially infecting all warm-blooded animals. In South America, and especially in Brazil, this cosmopolitan parasite presents some peculiarities due to an exceptional genetic diversity and the existence of severe clinical forms in Human, probably linked to the genetic background of some strains. In Brazil, the state of Rio de Janeiro presents highly contrasted environments from the city of Rio de Janeiro with different degrees of urbanizaton to isolated “sky islands”, hosting a rich and endemic fauna.We looked for physical and anthropic factors that may have an impact on the epidemiology of T. gondii in different environments of the state of Rio de Janeiro. We performed a seroprevalence study in cat populations living in 2 districts of Rio de Janeiro city with different degrees of urbanization, and molecular epidemiology studies on tissues of animals collected in Rio city, in a National Park characterized by “sky islands” and in intermediate and rural areas surrounding this park..The seroprevalence study on cats in Rio de Janeiro city allowed to show the value of blood sampling on filter-paper for performing the Modified-Agglutination-Test (MAT) for antibody detection in cats. There was a significant difference between seroprevalence observed in stray-cats from a residential district area with a relatively well-preserved natural environment (4/107- 3.74%) and in cats from a downtown public shelter (32/265-12.08%). Cat population density and other ecological factors may be involved in these differences.T. gondii DNA was detected in tissues of 8/16 cats from Rio city, 14/18 domestic and 23/33 wild animals in the area surrounding the park, and 31/38 small rodents or marsupials captured inside the National Park. DNA detection was positive in 3 tissues from wild felids found in the intermediate area surrounding the park and in feces from a Puma yagouaroundi found inside the park. This showed the role of wild felids in soil contamination in both environments.Genetic characterization using 15 microsatellite markers was successful for 6 samples, including one live strain (full genotyping for 5, and incomplete genotyping for one sample). This small sampling of strains was characterized by a large genetic diversity and revealed the presence of the main brazilian lineages, BRII and BRIII in isolated sky islands as well as in the city center of Rio de Janeiro.

Ce travail a analysé tous les cas de HFMD déclarés à Hai Phong pendant l’épidémie de 2011 et 2012 qui a été la plus importante au Vietnam et la première enregistrée dans le nord du pays. Hai Phong a connu la plus forte incidence au Nord Vietnam. C’était donc un bon modèle pour étudier la dynamique de cette maladie sans interférence et reliquats de précédentes épidémies ou de patients immunoadaptés.La première section est consacrée à une revue de la littérature sur EV-A71 et les entérovirus. La seconde section est divisée en trois chapitres, chacun abordant un aspect spécifique du projet.Le premier chapitre aborde la dynamique de la maladie et le rôle des directives officielles pour la gestion de l’épidémie de 2011-2012. Outre les éléments de base, cette étude apporte des résultats sur l’influence des directives HFMD durant l’épidémie, ce qui n’avait pas encore été fait. La publication des directives a conduit à un accroissement du score de sévérité et d’une réduction du délai entre le premier pic de fièvre et l’admission. Cet effet est associé à un accroissement de la sensibilisation qui a conduit à déclarer la plupart des patients avec des symptômes sévères pour assurer de meilleurs traitements et suivis. Le travail décrit dans ce chapitre a aussi démontré que trois vagues avec des caractéristiques différentes et causées par trois virus différents s’étaient succédées. La vague 1 et la vague 3 ont été causées respectivement par EV-A71 et par une combinaison de CV-A6 et CV-A16 alors que la vague 2 a été causée par un virus inconnu. Ce travail est aussi une analyse intégrative incluant une analyse spatiotemporelle. La maladie semble s’être étendue vers l’est en suivant les rivières pour atteindre les des zones plus peuplées à partir desquelles elle s’est répandue par les routes secondaires locales. Etant donné l’âge moyen des patients, environ 2 ans, la source de contamination doit être cherchée chez les adultes asymptomatiques contaminés lors de leurs activités professionnelles et des mobilités locales.Le deuxième chapitre aborde la phylogénie et la distribution spatiotemporelle de EV-A71 dans le nord du Vietnam et apporte un éclairage sur l’évolution et la dynamique de cet entérovirus. La protéine de capside VP1 a été ciblée. La première conclusion de ce chapitre est que l’épidémie de 2011 et 2012 n’a pas été causée par une souche exogène mais par des souches d’EV-A71 déjà présentes au Nord Vietnam. Ceci indique qu’elles peuvent se maintenir à faible niveau, asymptomatique, en stase génomique et avec une structuration géographique. La cause de l’épidémie devrait donc être recherchée dans le tissu socio-économique plutôt que dans une émergence extérieure. Une autre conclusion de ce chapitre est la corrélation observée ente les groupes de variants I/V et phylogénie, pathogénicité et groupe ethnique. Les profils des mutations I/V aux positions 249, 262 et 284 sur la protéine VP1 pourraient jouer un rôle dans la pathogénicité, ce qui est appuyé par la corrélation entre variants I/V et sévérité/ethnicité.Le dernier chapitre aborde la modélisation mathématique d’une maladie multiphases telle que HFMD. Il est essentiel de détecter aussi tôt que possible une nouvelle vague associée à un nouvel agent. Grace à la grande taille de la cohorte disponible pour ce travail (environ 9000 patients), nous avons pu développer un système d’équations différentielles apportant une forte correspondance avec les données observées. Le modèle a confirmé l’existence de trois vagues en 2011-2012, ayant des niveaux de virulence différents. Il permet aussi de caractériser chaque vague, de détecter l’apparition d’une nouvelle vague et d’associer des groupes patients à un tableau clinique.En conclusion, ce travail de thèse a permis de souligner plusieurs éléments clés à aborder de façon coordonnée afin de faciliter une surveillance efficace de l’HFMD au Vietnam. / This work analyzed all HFMD cases reported in Hai Phong in 2011 and 2012 outbreak which was the largest to have ever occurred in Vietnam and the first recorded in the northern part of the country. Hai Phong city experienced the highest HFMD incidence in North Vietnam. It was thus a good model for investigating the dynamic of the disease without interference and potential remains from previous outbreaks or patient immunological adaptation.The first section is dedicated to a review of the literature on EV-A71 and enteroviruses. The second section is divided in three chapters, each one addressing a specific issue of the project.The first chapter addresses the dynamic of the disease and the role of official guidelines in the handling of the 2011-2012 epidemic. Beside basic epidemiological features, the study also provides findings relating to the influence of HFMD guidelines during the outbreak period that has never been described before. The guideline release led to a significant increase of the severity score and reduced delay between onset and admission. This effect is linked to an increased awareness leading to patients being mostly declared with severe symptoms in order to ensure a better treatment and surveillance. The work presented in this chapter also demonstrated that three waves occurred with different characteristics and caused by three different viruses. Wave 1 and wave 3 were caused by EV-A71 and a combination of CV-A6 and CV-A16, respectively while Wave 2 was caused by an unknown virus. This work is also an integrative analysis including a spatiotemporal analysis. The disease seems to have expanded following the eastbound river system to reach densely populated settlements from where it secondarily expanded through local roads. Owing to the average age of the patients, around 2, the source of contamination must be sought for within asymptomatic adults being contaminated during their occupational activities and in local movements.The second chapter addresses the phylogeny and spatiotemporal distribution of EV-A71 in North Vietnam and provides an insight on the evolution and dynamic of the EV-A71 enterovirus. The VP1 capsid protein was used as target. The first conclusion of this chapter is that the 2011-2012 outbreak was not caused by an incoming strain but by EV-A71 strains which were already present in North Vietnam. This indicates that they can remain in a low level, asymptomatic state, in genomic stasis and with a geographic structuration. The cause for outbreaks should thus be sought for in the socio-economic patterns rather than in exogenous emergence. Another outcome of this chapter is the observed correlation between I/V variant groups and phylogeny, pathogenicity and ethnicity. The I/V pattern at positions 249, 262 and 284 on the VP1 protein might play a role in pathogenicity. The observed correlation of the I/V variant populations with severity and ethnicity strengthen this hypothesis.The last chapter addresses the mathematical modelling of a multiphase disease such as HFMD. It is essential to detect as soon as possible the emergence of new wave, associated to a novel agent. Owing to the large size of the cohort available for this work (ca. 9000 patients), we have been able to develop a differential equation model providing a very high fit with the observed data. The model confirmed that three waves were present in 2011-2012 with differing virulence. It also allows to characterize each wave, detect the start of a new one and associate groups of patients with specific patterns of symptoms.As a conclusion, this PhD work as underlined some key issues to be addressed in a coordinated way in order help developing an efficient surveillance and monitoring system for HFMD in Vietnam.

Uropathogenic E. coli (UPEC) is the most frequent cause of urinary tract infection (UTI), being responsible for up to 85% of community acquired and 40% of nosocomial cases. UPEC strains harbour various virulence factors that contribute to their ability to cause disease. The high prevalence across the globe of multidrug resistant UPEC is a significant threat to therapy. Virulent and resistant UPEC strains have been recognised as belonging to major lineages and we have only recently begun to understand the factors contributing to their successful global dissemination. Work in this thesis was carried out to identify the population structure of E. coli isolates recovered from urosepsis and biliary sepsis, to reveal any differences in genetic background. A total of 100 isolates from the blood and urine of 50 patients presenting with urosepsis and 27 isolates from cases of biliary sepsis were subjected to genotypic and phenotypic analysis, including MLST, virulence gene detection and antibiogram and metabolic profiling. Urosepsis paired isolates showed identical genotypes and antimicrobial resistance profiles. However, several pairs of isolates showed discrepant metabolic activity profiles suggesting niche specific regulation of metabolism. Members of the ST131 clone were significantly associated with antibiotic resistance and ST38 isolates were associated with the highest level of metabolic activity. An in vivo infection model was used to investigate the virulence potential of isolates from the major UPEC lineages. Galleria mellonella larvae inoculated with ST69 and ST127 isolates showed significantly higher mortality rates than those infected with other strains. However, one isolate of ST127 (strain EC18) was avirulent and comparative genomic analyses with a single virulent ST127 strain revealed an IS1 mediated deletion in the O-antigen cluster in strain EC18, which is likely to explain the lack of virulence in the larvae and demonstrates the importance of this cell surface molecule in the model system. Finally, a total of 202 UPEC isolates were recovered from community and hospital urine samples from a tertiary care hospital in Riyadh, Saudi Arabia. Molecular epidemiological investigation of the strains was carried out to examine the overall UPEC population structure, for the first time in any part of Saudi Arabia. The most common lineages were ST131 (17.3%), ST73 (11.4%), ST38 (7.4%), ST69 (7.4%) and ST10 (6.4%). The findings highlight the successful spread of multidrug resistant, CTX-M positive ST38, ST131 and ST405 UPEC in Saudi Arabia. The high proportion (35%) of ESBL producing E. coli isolates is a particular concern and is driving frequent prescription of carbapenem antibiotics. A total of four isolates of ST38 were positive for aggR, which is a virulence marker of enteroaggregative E. coli (EAEC); ST38 strains that cause UTI but have an EAEC genetic background are becoming recognised as novel UPEC and this clonal group warrants further study.

Orientador: Maria de Lourdes Ribeiro de Souza da Cunha / Resumo: A epidemiologia das infecções estafilocócicas tem sofrido importante modificação nas últimas décadas. A emergência de linhagens de Staphylococcus aureus resistentes à meticilina associados à comunidade (CA-MRSA) representa risco especial para populações reconhecidamente vulneráveis como os idosos. Neste âmbito, duas situações distintas são de especial interesse: os indivíduos institucionalizados, vivendo em casas de repouso, que representam um espaço intermediário entre a comunidade e o hospital; e os dependentes (acamados) cuidados em domicílio, expostos de forma intermitente aos serviços de saúde. Nosso objetivo foi identificar a prevalência e fatores associados ao carreamento nasal, oral e retal de S. aureus e MRSA em indivíduos acamados ou residindo em instituições de longa permanência para idosos (ILPIs) no município de Botucatu, SP, bem como a identificação de clones importantes de S. aureus e MRSA nessa população. Estudo com delineamento transversal, em que swabs da nasofaringe, orofaringe e reto de 226 indivíduos (150 residentes em nove ILPIs e 76 acamados em domicílio) foram coletados juntamente com um questionário que, através de entrevista com o próprio indivíduo ou responsável legal, levantou informações como dados demográficos (gênero e idade), tempo de institucionalização ou restrição ao leito, dados clínicos (comorbidades), dispositivos invasivos, internações recentes, doenças infeccionas e uso de antimicrobianos, para identificação dos fatores de risco. O isol... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The epidemiology of staphylococcal infections has undergone important changes in recent decades. The emergence of community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) strains represents a special risk to populations recognized as vulnerable, such as the elderly. Within this context, two different situations are of special interest: institutionalized individuals living in nursing homes, which represent an intermediary space between the community and the hospital, and dependent (bedridden) patients cared for at home, who are intermittently exposed to health services. Our objective was to determine the prevalence and factors associated with nasal, oral and rectal carriage of S. aureus and MRSA in bedridden patients and residents of long-term care facilities (LTCF) for the elderly in the city of Botucatu, SP, and to identify important S. aureus and MRSA clones in this population. In a cross-sectional study, nasopharyngeal, oropharyngeal and rectal swab samples were collected from 226 individuals (150 individuals from nine LTCF and 76 bedridden patients living at home). In addition, a questionnaire was applied by interview with the subject himself or the legal representative for the collection of demographic data (gender and age), length of institutionalization or bedridden period, clinical data (comorbidities), invasive devices, recent hospitalizations, infectious diseases, and antimicrobial use in order to identify risk factors. Staphylococcus aureus was is... (Complete abstract click electronic access below) / Mestre

La chromoblastomycose (CBM) et la sporotrichose (SPT) sont des infections fongiques chroniques des tissus sous-cutanés. Elles touchent surtout les membres, après blessure végétale ou souillure tellurique. Les principaux agents fongiques responsables de CBM appartiennent aux genres Fonsecaea et Cladophialophora tandis que Sporothrix est à l’origine de la SPT. A Madagascar, les études menées entre 1955 et 1994 ont montré une prévalence de la CMB à 0,5/100 000 habitants faisant considérer ce pays comme le premier foyer mondial. Depuis, aucune donnée n’a permis d’actualiser ces observations. Malgré les nombreux cas de SPT observés par les médecins, son épidémiologie à Madagascar n’a jamais été décrite.L'objectif général était d'évaluer l’incidence actuelle de ces mycoses à Madagascar. Les objectifs spécifiques étaient de caractériser les espèces fongiques responsables et de mettre en place un réseau clinico-biologique pérenne permettant une gestion adaptée des patients et l’utilisation de méthodes moléculaires pour l’identification des souches.Une étude prospective conduite entre mars 2013 et juin 2017 a donné lieu à des consultations dans les régions ou dans le service de dermatologie de l’hôpital universitaire d’Antananarivo et a permis d’inclure des patients présentant des lésions sous-cutanées chroniques. Les méthodes conventionnelles de diagnostic mycologique ont été complétées par des méthodes moléculaires (PCR, séquençage, MALDI-TOF-MS). Les cas ont été classés au cours de réunions de concertation clinico-biologique.Au total, 148 patients d’âge moyen de 41 ans avec une prédominance d’hommes (75,0%) ont été inclus : 63 cas de SPT (42,5%) et 50 cas de CBM (33,8%) ont été confirmés. L’incidence annuelle de le CBM a été estimée à 0,38/100 000 habitants dans la région Sava au Nord où les cas prédominent avec au niveau du district Anosibe An’Ala (à l’Est) une incidence maximale de 1,12/100 000. Alors que la CBM était prédominante dans le Nord-Est, l'Est et le Sud de l'île, étonnamment, la SPT était presque exclusivement localisée sur les hautes terres centrales. L’incidence globale moyenne de SPT était de 0,17/100 000 habitants dans les hauts plateaux et de 0,07/100 000 pour l’ensemble de Madagascar. Pour la SPT, le risque est plus élevé chez les jeunes (< 18 ans) et les formes cutanéo-lymphatiques des membres supérieurs sont les plus fréquentes. Pour la CBM, le risque de contamination est élevé chez les agriculteurs et les professions de services et la majorité des lésions (82,9%) était localisée au niveau des membres inférieurs. Sur le plan mycologique, 63 Sporothrix schenckii, 7 Cladophialophora carrionii, 29 Fonsecaea sp dont 22 F. nubica ont été identifiés. Pour le genre Fonseceae, l’identification de l’espèce nubica remplace l‘espèce pedrosoi initialement proposée pour les isolats malgaches et souligne l’importance de l’identification moléculaire pour une classification précise des espèces.Cette première étude sur la SPT humaine à Madagascar a permis d’actualiser les données épidémiologiques mondiales et de classer Madagascar avec un niveau endémique modéré alors qu’il était considéré comme faible jusque-là. La CBM persiste à une incidence élevée et comparable à celle décrite jusqu’en 1994, illustrant l’absence de contrôle de cette mycose dans le pays. La constitution d’un réseau clinico-biologique local stable et les nouveaux outils diagnostics mis en place dans ce travail (PCR et le MALDI-TOF MS), vont faciliter la conduite de programmes de surveillance et de contrôle alors que l’OMS a récemment classé la CBM en maladie tropicale négligée. Une enquête environnementale est en cours pour détecter les sources de contamination dans l’environnement. Le réseau mis en place permettra dans un futur proche de nouvelles études thérapeutiques (nouveaux schémas thérapeutiques) ou génétique (facteurs d’hôtes) sur la CBM et SPT mais également sur d’autres mycoses endémiques et encore négligées à Madagascar / Chromoblastomycosis (CBM) and sporotrichosis (SPT) are chronic subcutaneous or cutanéo-lymphatic infections found mostly in tropical and subtropical regions. Studies carried out by the Institut Pasteur of Madagascar between 1955 and 1994 provided an inventory of the number of cases of CBM and identified this country as the leading focus of this mycosis worldwide. Mean incidence was estimated at about 0.5/100,000 inhabitants at the time. No new data have been obtained to update the epidemiological situation. About SPT, only sporadic cases have been reported in Madagascar, due to the absence of specific surveillance. CBM is usually caused by dematiaceous fungi, principally Fonsecaea spp. and Cladophialophora spp. The causal agent of SPT is Sporothrix schenckii, a dimorphic hyphomycete.The objectives of this study was to update the data on epidemiology and to evaluate the current burden of these two fungal infections. In addition, we aimed to set up a durable local bio-clinical network and to implement molecular tools to ensure reliable species identification (PCR, sequencing, mass spectrometry).A prospective study, involving the recruitment of patients with suspect lesions was undertaken from March 2013 to June 2017. Patients were included in the dermatology department of the univerity hospital of Antananarivo and through field campaigns in rural areas. Clinical samples were collected and analyzed with conventional mycological methods and molecular tools. Classification of the cases was achieved by the confrontation of mycological and clinical features.Among the 148 patients (mean age 41; male 75.0%): 63 SPT cases (42.5%) and 50 CBM cases (33.8%) were diagnosed. The highest annual incidence of CBM was estimated at 0.38/100,000 inhabitants in the Sava region located to the north. At the district level, the peak incidence was 1.12/100,000 at Anosibe An’Ala, eastern part of the country. Whereas CBM predominated at the periphery of the island, SPT was surprinsgly concentrated in the highlands where the mean incidence was 0.17/100,000 inhabitants. The incidence in the whole country was 0.07/100,000. SPT likelihood of infection was higher in young (<18 years) and the cutaneo-lymphatic forms of the upper limbs were the most frequent. For CBM, farmers and service workers were at high risk and the lesions were mostly (82.9%) located to the lower limbs. The mycological analyses revealed 63 strains of Sporothrix schenckii, 7 of Cladophialophora carrionii, 29 of Fonsecaea sp including 22 F. nubica. F. nubica identification corrected the one of F. pedrosoi previously found in Madagascar, highlighting the need for a molecular analysis of the strains.This is the first study describing human SPT in Madagascar. The SPT burden can now be considered as moderate instead of low as it was described before. It reveals an unexpected concentration of the patients in the central highlands. CBM burden was found at a high level, regrettably similar to the one described 20 years ago, showing the lack of control of this infection. The implementation of a durable bio-clinical network and of the new molecular tools (PCR et le MALDI-TOF MS) developed in this work will easy the development of surveillance programs, especially as the WHO recently added CBM in the neglected tropical diseases list. The network that was built for this work will be used for further therapeutic trials on new schedules of treatment, new drugs or new formulations as well as genetic studies about predisposing factors of CBM and SPT and others deep fungal infections that are still neglected in Madagascar. Yet, an environmental survey is ongoing to describe the sources of contamination.