Cette thèse présentait comme objectif mettre en évidence le potentiel apport de l’immuno-PCR dans le diagnostic des maladies infectieuses à travers 3 exemples. L’application de l’iPCR dans le diagnostic précoce de la fièvre Q aiguë via la détection des IgM Phase II anti Coxiella burnetii a permis de détecter 90% des sérums prélevés dans les 2 premières semaines après l’apparition des symptômes contre 55% détectés par PCR, 38% par ELISA et 35% par l’IF la technique de référence. De plus une spécificité de 92% a été retrouvée par iPCR, 100% par PCR et l’IF et 90% par ELISA. L’application de l’iPCR à la fièvre Q aiguë constitue un exemple d’application de la technique plus globalement pour tous types d’infections aiguës. D’autre part, dans le cadre de la mise au point d’un modèle expérimental murin d’infection à Tropheryma whipplei, l’iPCR a servi à mesurer la réponse immunitaire mucosale via la détection des IgA anti T. whipplei dans les selles de souris. Les résultats obtenus ont permis de confirmer le rôle de la bactérie comme agent de gastroentérite via entre autre la détection d’IgA anti T. whipplei dans les selles lorsque des atteintes intestinales étaient provoquées. Enfin l’’utilisation de l’iPCR pour la détection de l’antigène Y. pestis dans des dents anciennes a permis de confirmer Y. pestis comme agent étiologique de la peste noire dans 5 charniers à travers la France et l’Italie. Une sensibilité de 41% a été retrouvée par iPCR contre 32% par PCR et 10% par ELISA. Nos résultats suggèrent que la détection des antigènes et la détection de l’ADN du pathogène semblent être 2 approches complémentaires permettant de confirmer le rôle de Y. pestis dans les différentes pandémies de peste et de mettre fin aux controverses suscitées par la seule utilisation des techniques de biologie moléculaire.Globalement, les résultats que nous avons obtenus au cours de cette thèse démontrent le potentiel énorme de l’iPCR comme technique de détection des anticorps et d’antigènes dans le domaine des maladies infectieuses tant au niveau de la sensibilité de détection qu’au niveau de son adaptabilité à différentes applications. / The objective of this thesis was to highlight the potential contribution of immuno-PCR in the diagnosis of infectious disease through 3 examples of infection. The iPCR was adapted for the early diagnosis of acute Q fever by the detection of IgM anti phase II Coxiella burnetii in patient’s sera. The results that we obtained show that iPCR could allow an early diagnosis of acute Q fever since 90% of early sera of patients with acute Q fever collected during the 2 first weeks after the onset of symptoms are detected against 55% by PCR, 38% by ELISA and 35% by IFA the gold standard. In addition, a specificity of 92% was found by iPCR, 90% by ELISA and 100% by PCR and IF. Application of iPCR to the diagnosis of acute Q fever is an example of application of the technique more generally for all types of acute infections. In a second time, the use of iPCR for the detection of Yersinia pestis antigen in ancient teeth allowed a confirmation of its role as the etiologic agent of plague in five mass graves across France and Italy. A sensitivity of 41% was recovered by IPCR against 32% by PCR and 10% by ELISA. Our results suggest that antigen and DNA detection of pathogen in ancient samples are 2 complementary approaches allowing the confirmation of the role of Y. pestis in different plague pandemic. Finally, as part of the development of an experimental murine infection with Tropheryma whippleii, the iPCR was used to measure the mucosal immune response via the detection of IgA anti T. whippleii in mouse stools. The results have confirmed the role of the bacterium as an agent of gastroenteritis via the detection of IgA anti T. whipleii in the stool when intestinal damages were caused.Overall, the results that we obtained during my thesis demonstrate the enormous potential of iPCR as a diagnostic tool of infectious disease by the ultrasensitive detection of antigens and antibodies.

Orientador: Mariangela Ribeiro Resende / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-09T21:35:03Z (GMT). No. of bitstreams: 1 Barao_SandraCristina_D.pdf: 3775784 bytes, checksum: ffb11ed337f2da69883370a4bf0d4b32 (MD5) Previous issue date: 2007 / Resumo: As informações que existem acerca da leishmaniose visceral humana, provêm em sua maioria das notificações realizadas nas áreas de alta endemicidade. Por isso, ainda há muitos aspectos a respeito da transmissão urbana e dos quadros de infecção assintomática que precisam ser elucidados. O dimensionamento real da prevalência da infecção por Leishmania chagasi pode contribuir para a definição e avaliação do impacto das medidas de controle. Com o objetivo de determinar a prevalência da infecção por L. chagasi em área de autoctonia recente, município de Araçatuba e, avaliar os fatores associados em relação aos casos humanos de leishmaniose visceral notificados, foi realizado um estudo transversal, com amostra estratificada de fase única, realizada em duas áreas urbanas de níveis sócio-econômicos distintos, designadas A1 (periférica, menor nível sócio-econômico) e A2 (central, melhor nível sócio-econômico). A soroprevalência foi avaliada com a utilização do teste imunocromatográfico com antígeno recombinante K39 (Ag-RK39). A prevalência observada foi de 18,4% (23/125) em A1 e 4,8% (6/125) em A2. A proporção entre indivíduos assintomáticos e casos de doença ativa nas áreas 1 e 2 foram respectivamente 1,35:1 e 2:1. Não houve diferença significativa da soropositividade na distribuição por idade, nem por sexo, entre as áreas. Contudo, foi observada diferença na proporção de casos assintomáticos entre as áreas, possivelmente associada aos níveis sócio-econômicos e intensidade de transmissão. Também houve relação com a presença canina nos últimos dois anos e a soropositividade para o Ag-rK39. As informações obtidas sugerem a associação da soroprevalência à presença canina nos dois últimos anos e reforça a estratégia de controle adotada / Abstract: Many information exist about human visceral leishmaniasis are origin to thepontificated cases, moreover, almost all data substantiating derive high levels transmission. So, there are many aspects about the urban transmission and asymptomatic infection to need to elucidated. The real comprehensive measurements about the Leishmania chagasi infection to be able to contribute to improve the assessment impact about the measures control. The objective to this study was determining the prevalence of asymptomatic visceral leishmaniasis infection in Araçatuba city, a recent autoctone area. This was a cross-sectional survey on a random sample of the population in two urban different areas, called A1 (outskirts, low social-economic condition) and A2 (central, good social-economic condition). The seroprevalence was assessing by the Immunochromatographic test with recombinant antibody K39 dipstick. The prevalence was 18.4% (23/125) in A1 and 4.8% (6/125) in A2. And the proportion between the asymptomatic and active disease in areas 1 and 2 was 1.35:1 and 2:1, respectively. There was no significant difference in age, nor gender, distribution of seropositivity between the areas. But we observed a difference in asymptomatic infection rates between the two areas, possibly associated with socioeconomic levels and transmission intensity. The data from this study suggest an associate between the human symptomatic seroprevalence and the presence of dogs in last two years old / Doutorado / Ciencias Basicas / Doutor em Clínica Médica