Bacteria of the Neisseria are predominately commensal, though N. gonorrhoeae and N. meningitidis are capable of causing disease. Both of these species often asymptomatically colonize humans, a trait reminiscent of their commensal cousins. The factors that shift the balance between asymptomatic carriage and disease are unknown. Pathogenic Neisseria use retractile surface structures called Type IV pili to coordinate community behavior and to initiate and sustain infection. Previously, the contributions of pilus retraction have been studied by deleting the pilus retraction motor, PilT. Recent findings suggest the speed and force exerted by pilus retraction is responsive to environmental cues. By examining several PilT mutants that maintain the ability to retract pili, I show retraction, per se, is not required for N. gonorrhoeae social interactions with bacteria or with human cells. Furthermore, Type IV pilus retraction by the commensal N. elongata affects the host cell differently than retraction by N. gonorrhoeae. These observations collectively suggest pilus retraction properties shape the host cell response to Neisseria colonization and could tip the balance of asymptomatic colonization to symptomatic disease.

Pseudomonas aeruginosa (PA) infections in Cystic Fibrosis (CF) patients are not easily cleared due to the conversion from a nonmucoid to a mucoid phenotype. Alginate is an acetylated exopolysaccharide produced by mucoid PA that is responsible for increased resistance to antibiotics, host phagocytic killing, and propagating biofilm formation. Understanding the interaction between PA and host cells is critical to understanding chronic infection and inflammation in CF. In order to investigate this, we used A549 pulmonary epithelial cells and murine alveolar macrophages (MH-S) to examine host response to nonmucoid versus mucoid PA infection. Adhesion assays in A549 pulmonary epithelial cells revealed that mucoid PA mutants adhere poorly compared to their nonmucoid counterparts. Similarly, phagocytosis assays using MH-S infected with PA revealed that mucoid PA are increasingly resistant to phagocytosis. The alginate acetylation mutant FRD1175 is more susceptible to phagocytic killing than alginate+ FRD1. Adherence and phagocytosis of mucoid FRD1 was increased by increasing the multiplicity of infection (MOI) from 50:1 to 500:1. Furthermore, confocal microscopy revealed that mucoid PA are inherently less inflammatory than nonmucoid strains in both A549 and MH-S. Increasing the MOI of mucoid FRD1 from 50:1 to 500:1 significantly increased caspase-1 activation in MH-S but not in A549, revealing that intensity of inflammatory signaling by epithelial cells is likely independent of increased adherence. FRD1175 infection in both A549 and MH-S revealed that alginate acetylation plays a significant role in reducing inflammasome activation. Western analysis revealed that PA does not actively induce TGF-β secretion by A549 epithelial cells. Similarly, NF-κB expression was reduced in both A549 and MH-S when infected with mucoid FRD strains, but not PA from the PAO background, suggesting FRD strains have accumulated additional mutations facilitating escape of inflammation. MH-S treated with cytochalasin D to block phagocytosis were still able to activate NF-κB signaling, suggesting NF-κB activation is adherence but not phagocytosis dependent. These data increase our understanding of the various mechanisms in which mucoid PA is able to evade host immune defenses and provides insight into potential therapies to treat PA infections.

Pathogenic Listeria are the causative agents of listeriosis, a severe food-borne infection. They are able to invade various non-phagocytic cell types including epithelial cells. The life cycle of these intracellular parasites involves penetrating into host cells, rupturing of the phagocytic vacuole, rapidly proliferating in the cytosol, and directly spreading cell to cell. Each step of the listerial intracellular infection involves activation of virulence factors dependent on PrfA, the master regulator of Listeria virulence. PrfAmediated virulence gene activation occurs within host cells by mechanisms that remain unknown. This thesis explores several aspects of PrfA regulation and its impact in the host-pathogen interaction. Methods for assessing PrfA-dependent gene expression were first developed and standardized, including a highly sensitive and accurate quantitative reverse-transcription real-time PCR (RT-QPCR), as well as procedures to investigate the correlation with virulence using cell culture-based assays. These techniques were applied in an investigation into the structure-function of PrfA. We studied the role of a solvent-accessible pocket identified in the N-terminal domain of PrfA, homologous to the cyclic nucleotide-binding (CNB) domain of Crp and other cAMP-regulated proteins, in intracellular virulence gene activation. Site-directed PrfA mutants were constructed. Our data support the notion that PrfA activity is allosterically regulated and are consistent with a role for the pocket as putative binding site for the PrfA-activating allosteric effector. The characterization of spontaneously occurring PrfA mutations that identified in our laboratory as PrfA*- suppressor or attenuator mutations, A129T, E173G and C229Y, allowed us to gain additional insight into PrfA structure-function. The role of the C229Y in sugar-mediated repression was investigated and found to explain the anomalous phenotype of strain NCTC 7973, a prfA* (G145S) mutant that carries this second mutation and is repressed by cellobiose but not glucose. We also carried out experiments to address the intriguing activation of PrfAdependent virulence genes upon addition of an adsorbent to the culture medium, the socalled "charcoal effect". Using a chemically defined culture medium and resin, Amberlite™ XAD-4, we provided evidence that the virulence gene activation may involve the sequestration of a medium component rather than a bacteria-derived autorepressor, as initially thought. We also explored the role of PrfA and the sigma factor σB in L. monocytogenes entry into host cells. ΔsigB mutants in different prfA regulation backgrounds were constructed. We showed that σB has no major effect on host cell invasion, and that L. monocytogenes invasiveness is a strictly PrfA-dependent trait. Our results also demonstrate a differential role of σB in L. monocytogenes serotypes. σB apparently plays no role in stress tolerance in serotype 4b, whereas it is important in serotype 1/2a for maintenance of bacterial fitness in stress conditions. Finally, we investigated the occurrence of apoptosis in Listeria-infected cells and developed normalized methods to accurately determine and quantify this cellular response in infected cell monolayers.

Pas de résumé français / Each Bartonella species appears to be highly adapted to one or a limited number of reservoir hosts, in which it establishes a long-lasting intraerythrocytic bacteremia as the hallmark of infection. Although the course of Bartonella infection has been precisely described, the molecular mechanisms of host specific erythrocyte infection and the stages of precedent the arrival in the bloodstream are poorly understood. In this thesis we purposed to identify the mechanisms of erythrocyte infection by Bartonella and characterize the possible locations of Bartonella during the days before the intraerythrocytic stage.By the establishment of an in vitro model of adhesion and invasion of erythrocytes by Bartonella spp., we demonstrated that host specificity was determined by the interaction between bacteria and erythrocytes. By screening signature-tagged mutagenesis (STM) library of B. birtlesii in vivo and in vitro and ectopic expression, we revealed that type IV Trw locus was required for host-restricted adhesion to erythrocytes in a wide range of mammals. After that, we further characterized that only TrwJ1 and TrwJ2 were expressed and present on the surface of the bacteria and had the ability to bind to mouse erythrocytes, and the receptor of them was erythrocyte band3 by different technology (phage display, electron microscopy, far western blot and adherence and invasion inhibition assay). By the model of experimental infection of laboratory normal Balb/C mice and splenectomized mice with B. birtlesii, we showed that during the first 7 days, no bacteria were recovered from lymph nodes, bone marrow and brain, but in the spleen, transient in the liver, And bacteremia was the same in both infection models during the first 7 days, thereafter, bacteremia was 10 fold higher in splenectomized mice than in normal mice and lasted 2 weeks longer. This suggested that the spleen was able to retain Bartonella.In conclusion, the host specific adhesion between Bartonella and erythrocyte was mediated by Trw and erythrocytic band 3, and spleen had a role in retention Bartonella.

Orientador: Daniel Júnior de Andrade / Resumo: O ácaro-da-ferrugem-dos-citros Phyllocoptruta oleivora é uma das principais pragas dos citros no Brasil. Os problemas atribuídos a P. oleivora têm se intensificado e suspeita-se que os danos estejam relacionados a uma nova espécie de eriofiídeo descrita recentemente, o ácaro-marrom-dos-citros Tegolophus brunneus. No entanto, não há estudos com essa espécie e informações sobre sua distribuição, características bioecológicas, danos, suscetibilidade a acaricidas e inimigos naturais associados são ausentes. O objetivo principal do projeto foi estudar a distribuição de T. brunneus na principal região citrícola brasileira e sobre as principais espécies e variedades cítricas. Além disso, estudou-se a biologia de T. brunneus em laboratório, determinou-se a suscetibilidade desse ácaro aos principais acaricidas, caracterizando seus danos em plantas cítricas, bem como o potencial de predação das principais espécies de predadores associadas ao ácaro e seu potencial para o uso no controle biológico. As coletas realizadas em vários municípios do estado São Paulo e Triângulo Mineiro demonstraram que T. brunneus infestou apenas lima ácida ‘Tahiti’, enquanto P. oleivora infestou todas as outras espécies e variedades cítricas. O ácaro completa o desenvolvimento em 7 dias, com período de incubação de 3 dias, duração de larva de 2,1 dias e de ninfa 2,8 dias e as fêmeas apresentam fecundidade de 8,5 ovos a 25°C. Os danos da espécie caracterizam-se pelo prateamento dos frutos e formação de manchas... (Resumo completo, clicar acesso eletrônico abaixo) / Doutor

Volunteers, governments, agencies and organizations from the North have too often defined the benefits and limitations of international volunteer programs without incorporating the perspectives of the organizations they seek to help. In fact, scholars and practitioners have relied on Northern experiences to develop a critical analysis of this development practice. As the experiences of the South are often absent from the conversations about international volunteerism, the goal of this thesis is to leverage the voices and the stories of Southern hosts to further understand the impact of international volunteerism. The research draws on the experiences of host organizations in Uganda. The interviewed participants are Ugandans who have worked with international volunteers to address various development issues. A review of the existing literature on international volunteerism, combined with the field research, support the analysis of the benefits and limitations of international volunteerism from the perspectives of host organizations. It also enables an exploration of the agency of volunteerism and determines key principles to empower host organizations and their employees. Thus, the analysis establishes the following conclusions: international volunteers are valuable actors for Southern hosts; volunteer programs must consider the impact, the challenges and the recommendations identified by host organizations and their communities; international volunteers enable alternative voices to be heard; and volunteerism fosters cooperation and partnerships within the Global South.

This thesis explores perspectives of 30 women hosting international students in a rural, coastal town in Costa Rica through an International Studies lens - interdisciplinary, critical, and bridging theory and practice. Analysis of 30 semi-structured interview sessions, which included 2 questionnaires, conducted over 10 weeks living with 3 host mothers contributes to understanding the impact of study abroad on host families. Hosting is discussed as a preferred form of paid care work in that it is flexible and enjoyable. Women host for the income as well as for the joy of mothering students. Host perspectives are shared regarding benefits and challenges of and lessons learned from hosting. Recommendations are made for homestay program administrators and international educators, including recommendations for addressing power dynamics to ensure reciprocal exchanges.

Les interactions entre les membres d'une communauté microbienne peuvent être un moyen d'adaptation dans l'environnement. Parmi les nombreuses interactions qui ont lieu dans un écosystème et qui joue un rôle majeur sur la diversité et la dynamique des populations microbiennes est celui des virus procaryotes et leurs hôtes. Les virus peuvent également arbitrer le transfert de matériel génétique entre les procaryotes (transduction), qui pourrait être un mécanisme d'adaptation rapide. Afin de déterminer l'impact potentiel des virus et la transduction, nous avons besoin d'une meilleure compréhension de la dynamique des interactions entre virus et leurs hôtes dans l'environnement. Les données sur les virus de l'environnement sont rares, et les méthodes pour le suivi de leurs interactions avec les procaryotes sont nécessaires. Clustered regularly interspaced short palindromic repeats (CRISPRs), qui contiennent des séquences virales dans les génomes bactériens, pourraient aider à documenter l'histoire des interactions virus-hôte dans l'environnement. Ainsi, cette thèse vise à explorer les interactions virus-hôte dans un environnement donné à travers du séquences CRISPR.Les virus de la cryosphère sont considérés comme abondantes, très actif et avec de larges gammes d'hôtes. Ces caractéristiques pourraient faire de la transduction virale, un facteur clé pour l’adaptation microbienne dans ces environnements. Des métagénomes publics créés à partir des environnements avec une gamme de températures différents ont été examinés. De cette manière, certaines dynamiques d'interactions virus-hôte se sont révélées comme ayant une corrélation avec la température. Un flux de travail a ensuite été développé pour créer un réseau reliant les virus et leurs hôtes en utilisant des séquences CRISPR obtenus à partir de données métagénomiques de la glace des glaciers et du sol de l'Arctique. La création de réseaux d'infection à traves du CRISPRs a fourni une nouvelle perspective sur les interactions virus-hôte. En outre, nous avons cherché des événements de transduction dans les données métagénomiques par la recherche de séquences virales contenant de l'ADN microbien. L’analyse indiquée que les bactériophages du Ralstonia pourraient être des agents de transduction dans la glace des glaciers de l'Arctique. / Interactions between the members of a microbial community can be a means of adaptation in the environment. Among the many interactions that take place in an ecosystem and have been seen to play a major role on microbial diversity and population dynamics is that of prokaryotic viruses and their hosts. Viruses can also mediate the transfer of genetic material between prokaryotes (transduction), which could be a mechanism for rapid adaptation. In order to determine the potential impact of viruses and transduction, we need a better understanding of the dynamics of interactions between viruses and their hosts in the environment. Data on environmental viruses are scarce, and methods for tracking their interactions with prokaryotes are needed. Clustered regularly interspaced short palindromic repeats (CRISPRs), which contain viral sequences in bacterial genomes, might help document the history of virus-host interactions in the environment. Thus, this thesis aimed to explore virus-host interactions in a given environment through CRISPRs. Viruses in the cryosphere have been seen to be abundant, highly active and with broad host ranges. These characteristics could make viral transduction a key driver of adaptation in these environments. Public metagenomes created from environments over a range of temperatures were examined through sequence and CRISPR analysis. In this fashion, certain virus-host interaction dynamics were found to have a correlation with temperature. A workflow was then developed to create a network linking viruses and their hosts using CRISPR sequences obtained from metagenomic data from Arctic glacial ice and soil. The creation of CRISPR-based infection networks provided a new perspective on virus-host interactions in glacial ice. Moreover, we searched for transduction events in metagenomic data by looking for viral sequences containing microbial DNA. Further analysis of the viral sequences in the CRISPRs indicated that Ralstonia phages might be agents of transduction in Arctic glacial ice.

Noctiliostrebla Wenzel, 1966 pertence a Streblidae, uma família de moscas parasitas exclusivas de morcegos, e compreende atualmente quatro espécies divididas em dois grupos, grupo A: Noctiliostrebla dúbia (Rudow, 1871) e N. traubi Wenzel, 1966; e grupo B: N. aitkeni Wenzel, 1966 e N. maai Wenzel, 1966. O gênero está incluído dentro da subfamília Trichobiinae, junto com Paradyschiria Speiser, 1900, sugerido como seu grupo-irmão hipotético. Com uma classificação sistemática complicada e poucos estudos taxonômicos, Noctiliostrebla caracteriza-se como um grupo muito homogêneo e de difícil distinção entre suas espécies. Restrito ao continente americano, exibe um alto grau de especificidade em relação às duas únicas espécies de morcegos do gênero Noctilio Linnaeus, 1766, N. albiventris Desmarest, 1818 e N. leporinus (Linnaeus, 1758). Não existem trabalhos filogenéticos para Noctiliostrebla, sendo que, os únicos estudos envolvendo Streblidae foram feitos para propor uma hipótese de relacionamentos entre as famílias de Hippoboscoidea. O projeto tem como objetivos realizar a revisão taxonômica do gênero Noctilostrebla e reconstruir uma hipótese de relacionamento entre as espécies do gênero, utilizando caracteres morfológicos e um conjunto de dados moleculares composto por três genes mitocondriais (12S, COI e cytB) e um gene nuclear (CAD). Dois métodos de análise foram empregados: análise de parcimônia, com pesagem igual dos caracteres, e análise de máxima verossimillhança. As análises foram realizadas com dados morfológicos e moleculares separados, enquanto os dados moleculares foram analisados com genes concatenados e separados em mitocondriais e nucleares. Como resultados, as quatro espécies já descritas foram consideradas válidas, sendo aqui redescritas, e outras seis novas espécies foram descritas. Do total de onze espécies, cinco são parasitas restritos de N. albiventris e seis de N. leporinus. As estruturas abdominais forneceram os únicos caracteres diagnósticos encontrados para separação entre as espécies. Tanto a análise morfológica como a molecular recuperaram Noctiliostrebla como monofilético e corroboraram a divisão do gênero em dois clados morfologicamente distintos. As análises com genes mitocondriais e nucleares foram incongruentes com relação a alguns clados, o que pode ser um indicativo de histórias filogenéticas diferentes. Os resultados obtidos para Noctiliostrebla apresentaram semelhanças com as hipóteses consideradas para os hospedeiros / Noctiliostrebla Wenzel 1966 belongs to Streblida, a family of batflies, and currently comprises four species divided into two groups, group A: Noctiliostrebla dubia (Rudow, 1871) and N. traubi Wenzel, 1966; Group B: N. aitkeni. Wenzel, 1966 and N. maai Wenzel, 1966. The genus is included within the Trichobiinae subfamily, along with Paradyschiria Speiser, 1900, suggested as your hypothetical sister group. With a complicated systematic classification and a few taxonomic studies, Noctiliostrebla is characterized as a very homogeneous group and difficult to distinguish between their species. Restricted to the American continent, exhibits a high degree of specificity with the only two bats species of the genus Noctilio Linnaeus, 1766, N. albiventris Desmarest, 1818 and N. leporinus (Linnaeus, 1758). There are no phylogenetic works with Noctiliostrebla and the only studies involving Streblidae were made to propose a relationship hypothesis among Hippoboscoidea families. The project aims to conduct a taxonomic revision of the Noctilostrebla genus and to rebuild a relationship hypothesis among Noctilostrebla species using morphological and molecular data set, with three mitochondrial genes (12S, COI and cytB) and one nuclear gene (CAD). Two analysis methods were used: parsimony with equal weighting of characters and lielihood. The analyses were performed using morphological and molecular data separate, while the molecular data were analyzed with concatenated genes and separated into mitochondrial and nuclear genes. As a result, the four described species were considered valid being here redescribed, and six new species have been described. Of all eleven species, five are restricted parasites of N. albiventris and six are restricted parasites of N. leporinus. The abdominal structures provided the only diagnostic characters found for separation of the species. Both morphological and molecular analysis recovered a monophyletic Noctiliostrebla and supported the division of the genus into two distinct clades. The analyses of mitochondrial and nuclear genes showed inconsistent clades, which may be indicative of different phylogenetic histories. The results obtained for Noctiliostrebla showed similarities with the assumptions for the hosts

Ao processo de envelhecimento pode estar associado o surgimento das doenças crônicas não transmissíveis (DCNT) que, por sua vez, podem contribuir para a ocorrência dos agravos clínicos e levar os idosos ao atendimento de urgência/emergência, expondo tanto o paciente quanto seus familiares a uma situação de sofrimento e angústia pelo agravamento da sua condição de saúde e interrupção inesperada de seu cotidiano. Dessa forma, torna-se relevante conhecer como os profissionais e instituições de saúde estão recebendo e se preparando para acolher, tanto o idoso quanto o familiar que o acompanha. Os objetivos deste estudo são: descrever as características sociais e demográficas dos idosos internados na Unidade Funcional de Atendimento às urgências/emergências da Unidade de Emergência do HCFMRP/USP, bem como do seu familiar acompanhante; identificar os núcleos temáticos extraídos da experiência do familiar que acompanha o idoso nesta situação; analisar o acolhimento prestado a partir da experiência do familiar acompanhante do idoso. Trata-se de um estudo qualitativo, em que a história oral foi utilizada como método de investigação na modalidade história oral temática. Para a análise dos dados, seguiram-se os passos da técnica de Análise Indutiva de Conteúdo. A coleta de dados foi realizada no período de abril e maio de 2010. As entrevistas foram realizadas com 11 familiares acompanhantes do idoso que forneceram subsídios para a construção de três núcleos de sentidos: \"Entre a vida e a morte: a busca pelo atendimento de urgência/emergência\", \"Os desafios enfrentados pelo familiar acompanhante do idoso na situação de urgência/emergência\" e \"Alternativas para o atendimento humanizado e acolhedor em unidades de urgência/emergência sob a ótica do familiar acompanhante do idoso\". A partir da análise e discussão destas categorias, fica evidente a necessidade de que os profissionais da saúde precisam conhecer a realidade e as necessidades das pessoas atendidas. Verificou-se que a clientela está atenta aos aspectos da humanização, e a comunicação se revelou uma ferramenta de extrema importância no manejo das relações entre os profissionais e os usuários do serviço. Os conceitos e especificações sobre a humanização e o acolhimento são, na atualidade, bem fundamentados, formulados e amplamente discutidos, no entanto, é necessário que se tornem ações efetivas no âmbito das instituições de saúde. / The aging process may be associated with the emergence of chronic noncommunicable diseases (CNCD), which in turn can contribute to the occurrence of clinical diseases and the elderly to take care at emergency, exposing both the patient and their families to a situation of suffering and distress by the deterioration of his health condition and unexpected interruption of their daily lives. Therefore, it becomes relevant to know how the professionals and health institutions are receiving and preparing to welcome both the old as the family accompanying him. The objectives of this study are to describe the social and demographic characteristics of elderly inpatients in the Unit Functional assistance to emergency care of the Emergency Unit of HCFMRP/USP, as well as your family companion, to identify the core themes drawn from the experience of relatives in accompanies the elderly in this situation, analyze the reception provided from the experience of family companion for the elderly. This is a qualitative study; \"Oral History\" was used as a method of research in oral story mode. For the data analysis followed the steps of the technique of \"Inductive Analysis of Content\". Data collection was performed during April and May 2010. The interviews were conducted with 11 family companions for the elderly, which provided subsidies for the construction of three groups of meaning: \"Between life and death: the search for care at emergency\", \"The challenges faced by the elderly in family companion urgency/emergency\" and \"alternatives for humanized and friendly units at emergency from the viewpoint of the elderly family companion\". From the analysis and discussion of these categories is an evident necessity for health care professionals needs to know the reality and needs of people served. It was found that the customer is looking to aspects of humanization and communication has proved an extremely important tool in the management of relationships between professionals and service users. The concepts and details on humanization and the reception are nowadays well-founded, formulated and widely discussed; however it is necessary to become effective actions within the health institutions.