Yaws is a skin debilitating disease caused by Treponema pallidum subspecies pertenue with most cases reported in children. World Health Organization (WHO) aims at total eradication of this disease through mass treatment of suspected cases followed by an intensive follow-up program. However, effective diagnosis is pivotal in the successful implementation of this control program. Recombinase polymerase amplification (RPA), an isothermal nucleic acid amplification technique offers a wider range of differentiation of pathogens including those isolated from chronic skin ulcers with similar characteristics such as Haemophilus ducreyi (H. ducreyi). We have developed a RPA assay for the simultaneous detection of Treponema pallidum (T. pallidum) and H. ducreyi (TPHD-RPA). The assay demonstrated no cross-reaction with other pathogens and enable detection of T. pallidum and H. ducreyi within 15 min at 42 °C. The RPA assay was validated with 49 clinical samples from individuals confirmed to have yaws by serological tests. Comparing the developed assay with commercial multiplex real-time PCR, the assay demonstrated 94% and 95% sensitivity for T. pallidum and H. ducreyi, respectively and 100% specificity. This simple novel TPHD-RPA assay enables the rapid detection of both T. pallidum and H. ducreyi in yaws-like lesions. This test could support the yaws eradication efforts by ensuring reliable diagnosis, to enable monitoring of program success and planning of follow-up interventions at the community level.

Submitted by Cleide Dantas (cleidedantas@ufpa.br) on 2014-02-13T12:08:13Z No. of bitstreams: 2 license_rdf: 23898 bytes, checksum: e363e809996cf46ada20da1accfcd9c7 (MD5) Dissertacao_SoroepidemiologiaChlamydiaTrachomatis.pdf: 714575 bytes, checksum: 150b98376e7d5179deaf87cc15123665 (MD5) / Approved for entry into archive by Ana Rosa Silva (arosa@ufpa.br) on 2014-04-16T14:00:20Z (GMT) No. of bitstreams: 2 license_rdf: 23898 bytes, checksum: e363e809996cf46ada20da1accfcd9c7 (MD5) Dissertacao_SoroepidemiologiaChlamydiaTrachomatis.pdf: 714575 bytes, checksum: 150b98376e7d5179deaf87cc15123665 (MD5) / Made available in DSpace on 2014-04-16T14:00:20Z (GMT). No. of bitstreams: 2 license_rdf: 23898 bytes, checksum: e363e809996cf46ada20da1accfcd9c7 (MD5) Dissertacao_SoroepidemiologiaChlamydiaTrachomatis.pdf: 714575 bytes, checksum: 150b98376e7d5179deaf87cc15123665 (MD5) Previous issue date: 2010 / As bactérias do gênero Chlamydia estão associadas à diversas doenças, como cegueira, infecções genitais e pneumonia. Existem poucos dados sobre como a Chlamydia e o Treponema pallidum afetam indígenas na Amazônia brasileira. Este estudo objetivou determinar a soroprevalência das infecções pela Chlamydia trachomatis, Chlamydia pneumoniae e Treponema pallidum nas aldeias indígenas Bakajá, Apyterewa, Xingu e Mrotdidjãm, no município de Altamira, Pará, Brasil. O estudo incluiu 270 amostras de sangue coletadas no ano de 2007. A detecção de anticorpos das classes IgM e IgG anti-Chlamydia foi realizada empregando-se o ensaio imunoenzimático (ELISA), e selecionada de forma aleatória amostragem de 36, entre os positivos, para determinar a sorotipagem pela microimunofluorescência. Para detecção de anticorpos anti-T. pallidum foi utilizado um teste treponêmico (ELISA) e as amostras positivas foram submetidas a um teste não treponêmico (RPR). A prevalência geral de anticorpos anti-Chlamydia foi de 26,7%, com prevalência de 100% para C. trachomatis entre as amostras testadas pela MIF. Para a C. pneumoniae a prevalência foi de 61,1% e a prevalência de anticorpos contra Treponema pallidum foi baixa. As bactérias do estudo circulam nas comunidades indígenas da Amazônia brasileira estudada, o que requer uma resposta urgente das autoridades de saúde pública, pois estas bactérias podem causar doenças graves, mas são sensíveis a tratamento específico, quando diagnosticadas adequadamente. / Bacteria from the genius Chlamydia are associated with several diseases including blindness, reproductive tract infections and pneumonias. There is little data about Chlamydia and Treponema pallidum affects on native Indian population in the Brazilian Amazon region. This study determined the prevalence of infections by Chlamydia trachomatis, Chlamydia pneumoniae and Treponema pallidum in Altamira, Para- Brazil among the native Indian populations Bakaja, Apyretewa, Mrotidjam, Xingu and included 270 plasma samples collected in 2007; they were tested using an enzyme immunoassay (ELISA) for the detection of IgM and IgG antibodies to Chlamydia and random samples (36) was chosen from the positives for serotyping through a microimmunofluorescence assay (MIF). Antibodies anti-Treponema were detected using a treponemal (ELISA) test and the positive samples were subjected to a non-treponemal test (RPR). The overall prevalence of antibodies to Chlamydia was 26.7% with a 100% prevalence of C. trachomatis tested through MIF. Prevalence of antibodies to C.pneumoniae was 61.1% and the prevalence of antibodies to T. pallidum was low. This study of bacteria circulating in the Brazilian Amazon indigenous communities confirms that it is necessary for an urgent response from public health authorities, because these bacteria’s can cause serious illness. With adequate diagnoses these bacteria’s can be treated because they are sensitive to specific treatment.

Submitted by Cleide Dantas (cleidedantas@ufpa.br) on 2014-02-13T12:18:42Z No. of bitstreams: 2 license_rdf: 23898 bytes, checksum: e363e809996cf46ada20da1accfcd9c7 (MD5) Dissertacao_SoroepidemiologiaChlamydiaTrachomatisChlamydia.pdf: 2065401 bytes, checksum: 1da33c9028340a826d8e01d04fe2f763 (MD5) / Approved for entry into archive by Ana Rosa Silva (arosa@ufpa.br) on 2014-04-22T13:32:53Z (GMT) No. of bitstreams: 2 license_rdf: 23898 bytes, checksum: e363e809996cf46ada20da1accfcd9c7 (MD5) Dissertacao_SoroepidemiologiaChlamydiaTrachomatisChlamydia.pdf: 2065401 bytes, checksum: 1da33c9028340a826d8e01d04fe2f763 (MD5) / Made available in DSpace on 2014-04-22T13:32:54Z (GMT). No. of bitstreams: 2 license_rdf: 23898 bytes, checksum: e363e809996cf46ada20da1accfcd9c7 (MD5) Dissertacao_SoroepidemiologiaChlamydiaTrachomatisChlamydia.pdf: 2065401 bytes, checksum: 1da33c9028340a826d8e01d04fe2f763 (MD5) Previous issue date: 2009 / CAPES - Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / A Chlamydia trachomatis e o Treponema pallidum compartilham com o HIV uma importante forma de transmissão: a via sexual. Por conta do comprometimento imunológico dos portadores de HIV, a C. pneumoniae pode apresentar um papel potencial em infecções respiratórias. Este trabalho objetivou a descrição da soroprevalência destes três agentes em portadores de HIV do Estado do Pará, Brasil. Entre setembro de 2007 a junho de 2008, foram coletadas 430 amostras de portadores de HIV em Belém, Pará. Estas foram submetidas a um ELISA para detecção de anticorpo IgG e IgM anti-Chlamydia e, dentre os positivos, uma amostragem aleatória foi escolhida e submetida à microimunofluorescência para sorotipagem. Para a detecção de anticorpos anti-Treponema pallidum foi feito um teste não treponêmico (RPR) e um teste treponêmico (ELISA). Os resultados obtidos foram analisados pelo teste do χ2. A prevalência geral de anticorpos anti-Chlamydia foi 64,2% (51,6% para IgG e 4% para IgM). A sorotipagem mostrou uma alta prevalência de C. trachomatis (100% tanto para IgG como IgM), e C. pneumoniae (73,5% IgG e 70,5% IgM), sendo que houve uma larga disseminação dos sorotipos que causam infecções genitais da Chlamydia trachomatis. A prevalência geral de anticorpos contra o Treponema pallidum foi de 34,9%, sendo que 7,3% apresentaram resultado laboratorial indicativo de sífilis. As variáveis que apresentaram associação com a infecção por Chlamydia e Treponema pallidum foram: o gênero masculino, maior idade, baixa escolaridade, número de parceiros por semana, a prática de sexo anal, homossexualismo/bissexualismo, uso de droga não-endovenosa, histórico de IST. Faz-se necessário tanto a conscientização como o monitoramento da população, para impedir a transmissão destes agentes e para a melhoria da qualidade de vida dos indivíduos portadores de HIV. / Chlamydia trachomatis and Treponema pallidum share the sexual route of transmission with HIV-1. In consequence of the compromise of the immune response among HIV-1 carriers, C. pneumoniae is a potential harassment in respiratory infections. The present study intended the description of the seroprevalence of those three agents among 430 HIV-1 infected persons residing in the State of Para, Brazil, attended at the State Reference Unit (URE-DIPE), between September 2007 to June 2008. Plasma samples were tested using an enzyme immuno assay for the detection of IgM and IgG antibodies to Chlamydia and those which elicited positive results were ramdomly selected for serotyping through a microimmunofluorescence assay. Antibodies to T. pallidum were detected using a flocculation reaction (RPR) and an enzyme immunoassay. Results were compared statistically using the Chi square test (χ2). The general prevalence to Chlamydia was 64.2% (51.6% IgG reactivity and 4% to IgM). Serotyping showed 100% reactivity to C. trachomatis (for both IgG and IgM), a high prevalence to C. pneumoniae (73.5% IgG and 70.5% to IgM) and a large distribution of reactivity to strains of C. trachomatis which cause genital infections. Prevalence of antibodies to T. pallidum was 34.9% and 7.3% showed laboratory evidence of syphilis. Infection with both pathogens were associated to several characteristics which included: higher prevalence among males, high age, low number of study years, high number of sexual partners, anal sexual relations, homosexual/bisexual habits, use of non injecting drugs and the history of sti. It is necessary not only the individual attention for prevention, but also the continuous monitoring to block transmission and the improvement of the well being of HIV-1 infected persons.

FundaÃÃo Cearense de Apoio ao Desenvolvimento Cientifico e TecnolÃgico / CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / Over the past few years, there has been increasing evidence of the effect of the oral health over the general health of individuals, supported by a series of biological and epidemiological studies that show a relation between the mouth and many diseases, including cardiovascular diseases. Structural deficiencies and functional abnormalities of heart valves represent an important cause of cardiovascular morbidity and mortality in Brazil, and a few defects have been recently associated with infectious agents. The aim of this study was to identify cariogenic and periodontopathogenic bacteria in dental plaque, saliva and heart valves, without clinical endocarditis, of patients with heart valve diseases, and correlate these findings with the oral health status of the patients. Oral exams using the DMTF (decayed, missing and filled teeth) and PSR (Periodontal Screening and Recording) indexes to evaluate caries and periodontal disease, respectively, were performed. Samples of supragingival and subgingival dental plaque, saliva and cardiac valves were evaluated, through Real Time Polymerase Chain Reaction, for the presence of DNA of Streptococcus mutans (S. mutans), Prevotella intermedia (P. intermedia), Porphyromonas gingivalis (P. gingivalis) and Treponema denticola (T. denticola). A total of 114 samples were collected from 42 patients with a mean age of 55.6  13.8 years. The average number of missing teeth due to caries was 23.52  9.41 teeth per patient, and according to the highest score of periodontal disease observed for each patient, excluding edentulous patients (44.0%), periodontal pockets over 4mm (43.4%) and dental calculus (34.7%) were detected in a higher number of patients. The molecular analysis of the oral samples revealed high frequency of S. mutans and P. intermedia in supragingival dental plaques, subgingival dental plaques and saliva of dentate and edentulous patients (variation 60.0% - 100.0%), while P. gingivalis and T. denticola were detected in a smaller number of oral samples (variation 17.6% - 64.0%). The microorganism most frequently detected in heart valve samples was the S. mutans (89.3%), followed by P. intermedia (19.1%), P. gingivalis (4.2%) e T. denticola (2.1%). Significant difference was observed between the frequency of P. intermedia, P. gingivalis and T. denticola in the heart valve and dental plaque, as oposed to S. mutans. The identification of oral bacteria, especially S. mutans, in heart valves of patients with a previous history of dental caries and gingivitis/periodontitis suggests the possible involvement of these pathogens in the etiopathogenesis of heart valve diseases. / Atualmente, cada vez mais se tem evidÃncias do efeito da condiÃÃo oral na saÃde geral dos indivÃduos, atravÃs de uma sÃrie de estudos epidemiolÃgicos e biolÃgicos que mostram uma relaÃÃo entre a boca e diversas doenÃas, incluindo as doenÃas cardiovasculares. Desordens estruturais e nas funÃÃes das vÃlvulas cardÃacas representam uma importante causa de morbidade e mortalidade cardiovascular no Brasil, sendo alguns processos, como a estenose aÃrtica degenerativa, mais recentemente associados a agentes infecciosos. O objetivo desta pesquisa foi identificar bactÃrias cariogÃnicas e periodontopatogÃnicas na placa dental, saliva e vÃlvulas cardÃacas, sem endocardite clÃnica, de pacientes com doenÃa valvar, correlacionando esses achados à condiÃÃo bucal dos indivÃduos. AvaliaÃÃo, quanto Ãs doenÃas cÃrie e periodontal, foi realizada, atravÃs dos Ãndices CPO-D (Dentes Permanentes Cariados, Perdidos e Obturados) e PSR (Registro Periodontal Simplificado), respectivamente. Amostras de placa dental supragengival, subgengival, saliva e vÃlvula cardÃaca foram coletadas para investigaÃÃo da presenÃa de DNA, atravÃs de PCR (ReaÃÃo em Cadeia de Polimerase) em tempo real, de Streptococcus mutans (S. mutans), Prevotella intermedia (P. intermedia), Porphyromonas gingivalis (P. gingivalis) e Treponema denticola (T. denticola). Um total de 114 amostras foi coletado de 42 pacientes com mÃdia de idade de 55.6  13.8 anos. A mÃdia de dentes perdidos devido à cÃrie, por paciente, foi em torno de 23.52  9.41 e, segundo o maior grau de doenÃa periodontal observado no indivÃduo, excluindo-se os pacientes desdentados totais (44.0%), bolsa superior a 4 mm (43.4%) e o cÃlculo dental (34.7%) esteve presente em um maior nÃmero de pacientes. A anÃlise molecular das amostras bucais revelou alta frequÃncia de S. mutans e P. intermedia nas placas supragengival, subgengival e saliva de pacientes dentados e desdentados (variando entre 60.0% e 100.0%), enquanto que P. gingivalis e T. denticola estiveram presentes em menor nÃmero de amostras bucais (variando entre 17.6% e 64.0%). O micro-organismo mais frequentemente encontrado nas amostras valvares foi o S. mutans (89.3%), seguido da P. intermedia (19.1%), P. gingivalis (4.2%) e T. denticola (2.1%). DiferenÃa significativa foi encontrada entre a presenÃa de P. intermedia, P. gingivalis e T. denticola na vÃlvula e na placa dental, diferentemente do S. mutans. A identificaÃÃo de bactÃrias orais, principalmente S. mutans, em vÃlvulas cardÃacas de pacientes com elevada experiÃncia prÃvia de cÃrie e ocorrÃncia de gengivite/periodontite, sugere o possÃvel envolvimento desses patÃgenos nas doenÃas valvares.

Bovine Dermatitis digitalis (DD) ist eine weltweit verbreitete Infektionskrankheit bei Rindern, die primär die plantare Haut über dem Kronenrand nahe des Zwischenzehenspalts der Hinterklauen betrifft. Schmerzhafte ulzero-proliferative Läsionen mit akuten und chronischen Erscheinungsformen führen zu Verhaltensänderungen und Lahmheit der Tiere. DD hat damit einen erheblichen Einfluss auf deren Wohl und ihre Leistungen. Zahlreiche Untersuchungen zur Ätiologie der Krankheit ergaben, dass es sich um das Zusammenspiel verschiedener Ursachen handelt. Einer synergistischen multifaktoriellen Infektion mit starker Beteiligung von Bakterien der Gattung Treponema kommt dabei besondere Bedeutung zu. Aspekte wie Tierhaltung, Hygienestandards und genetische Prädispositionen wurden ebenfalls intensiv untersucht. Nichtsdestotrotz bleiben Infektionsherde, Transmissionsrouten und Pathomechanismen weitgehend unklar. Zum besseren Verständnis der Ereignisse, die zu DD-Läsionen führen, sollte im Zuge dieser Arbeit ein organotypisches Zellkulturmodell der bovinen Haut erstellt werden, welches in späteren Versuchen mit dem Krankheitserreger zum Einsatz kommen soll. Verlässliche und reproduzierbare Techniken zur Isolation und Kultur von bovinen primären Keratinozyten und Fibroblasten wurden etabliert; geeignete Zellkulturmedien für die Langzeitkultivierung und –aufbewahrung der Hautzellen wurden identifiziert. Zur Erstellung des Hautmodells wurden zwei verschiedene Ansätze miteinander verglichen. Der zweite Ansatz, bei dem Keratinozyten direkt auf ein dermales Äquivalent, d.h. ein Pad aus bovinem Kollagen I mit eingesäten post-mitotischen Fibroblasten, gesät wurden, brachte ein vielversprechendes Hautmodell hervor. Die inkorporierten post-mitotischen Fibroblasten wiesen eine charakteristische Zellmorphologie mit intakten Nuklei auf. Die terminale Differenzierung der Keratinozyten auf dem dermalen Äquivalent wurde mittels Immunfluoreszenzfärbungen mit Antikörpern gegen die Markerproteine Keratin 14 und Desmoglein 1 gezeigt. Die Ergebnisse erster Experimente mit Treponema spp. verdeutlichen, dass das Hautäquivalent ein geeignetes Modell zur Untersuchung der Pathogenese der DD darstellt. / Bovine digital dermatitis (DD) is a worldwide occurring, infectious disease in cattle primarily affecting the plantar skin above the coronary band near the interdigital cleft on hind feet. Painful ulceroproliferative lesions with acute and chronic appearances lead to behavioral changes and lameness. Hence, DD has a major impact on animal welfare and performance. Substantial efforts in investigating the etiology of the disease revealed a synergistic origin with evidence for a multibacterial infection and the strong involvement of bacteria from the genus Treponema. As the interaction between host, pathogen and environment is not negligible, surrounding circumstances such as housing, general hygiene and genetic predispositions have been investigated intensively. Nevertheless, infection reservoirs, transmission routes and pathomechanisms remain widely unclear. To better understand the cellular and molecular events during Treponema-infection of bovine skin, it was the specific aim of this study to establish an organotypic in vitro skin model, which could be challenged with the causative agent of the disease. A technique to reliably and reproducibly isolate primary keratinocytes and fibroblasts from the site of infection was established. Appropriate cell culture media for the long-term cultivation and storage of bovine skin cells were identified. Two different methods to develop the skin model were compared. The second strategy in which keratinocytes were directly seeded on top of a dermal equivalent, i.e. a bovine collagen type I pad with embedded post-mitotic fibroblasts, gave rise to a promising organotypic skin equivalent. The incorporated post-mitotic fibroblasts showed a characteristic cell morphology with intact nuclei. The terminal differentiation of the keratinocytes on top of the dermal equivalent was shown with anti-K14 and anti-Dsg1 immunofluorescence stainings. The results of initial Treponema-experiments proved that the skin equivalent is a suitable model to investigate the underlying mechanisms during Treponema-infection of bovine skin and hence, the pathogenesis of DD.

No description available.

CONTEXTE La syphilis est une maladie ré-émergente depuis 2000. Son traitement est simple, mais son diagnostic est complexe. La technique d’amplification génique de Treponema pallidum (Tp-PCR) existe depuis 1990 mais le CDC l’a incluse dans sa définition de cas en janvier 2014. OBJECTIFS 1) Evaluer la performance diagnostique de la Tp-PCR à différents stades cliniques et milieux biologiques. 2) Mesurer la sensibilité, spécificité et les valeurs prédictives de la Tp-PCR en fonction de 3 groupes de référence dans des ulcères récents. 3) Comparer les performances des 2 principales cibles de Tp-PCR.MATÉRIEL ET MÉTHODES Premièrement, une revue systématique et méta-analyse des études publiées depuis 1990 ont été menées. Ensuite une étude multicentrique prospective a été conduite dans 5 villes européennes pendant 2 ans chez des patients avec un ulcère oro-génital. Tous ont reçu le test de référence local et 2 Tp-PCRs dans l’ulcère (gène tpp47 vs. polA). Les valeurs de sensibilité, spécificité et valeurs prédictives de la Tp-PCR ont été calculées comparativement au fond noir (FN), à la sérologie et à un gold standard amélioré. La concordance des 2 cibles a été évaluée par un coefficient kappa.RÉSULTATS PRINCIPAUX La méta-analyse conclut que la Tp-PCR a une meilleure performance dans les ulcères récents. L’étude clinique montre que la Tp-PCR décrit une meilleure performance comparativement au gold standard amélioré et a même une meilleure sensibilité que le FN. Les 2 cibles ont la même valeur diagnostique et une concordance quasi parfaite. CONCLUSIONS La Tp-PCR ciblant tpp47 ou polA est cliniquement utile pour diagnostiquer une syphilis primaire et pourrait même remplacer le FN sous certaines conditions. / BACKGROUND Syphilis has re-emerged in at-risk populations since 2000. Although the treatment of syphilis is simple, its diagnosis remains challenging. Treponema pallidum Polymerase Chain Reaction (Tp-PCR) has been used in the diagnosis of syphilis since 1990 but it is included in the case definition of the CDC since January 2014. OBJECTIVES 1) To assess the accuracy of Tp-PCR in various biological specimens and syphilis stages. 2) To measure its diagnostic performance (sensitivity, specificity and predictive values) in ulcers from early syphilis compared to three groups of reference. 3) To compare the accuracy of the two most currently used targets: tpp47 and polA genes.METHODS We conducted a systematic review and meta-analysis of all studies published from 1990. We implemented a multicentre, prospective, observational study in 5 European cities between 09/2011 and 09/2013 among patients with an oral or genital ulcer suggestive of syphilis. All patients were tested with traditional reference tests plus 2 Tp-PCRs (tpp47 and polA). We estimated the sensitivity, specificity and predictive values of Tp-PCR compared to darkfield microscopy (DFM), serology and an enhanced gold standard. We used the kappa coefficient to assess the agreement between the 2 targets.MAIN RESULTST p-PCR had the best accuracy in ulcers from early syphilis. Tp-PCR performed better when compared to the enhanced gold standard and had a higher sensitivity than DFM. The 2 Tp-PCRs had a similar accuracy and an almost perfect agreement.CONCLUSIONS Tp-PCR targeting either tpp47 or polA is clinically useful to confirm an early syphilis in smears and could even replace DFM under specific conditions.