Bartonella henselae ist der Erreger der Katzenkratzkrankheit, einer wenig beachteten, weil meist selbstlimitierend verlaufenden Zoonose. Allerdings kann B. henselae bei Risikogruppen (Kindern, immunsupprimierten Personen) Septikämien, Peliosis hepatis, Bazilläre Angiomatose und andere systemische Erkrankungen verursachen. Mit der vorliegenden Arbeit sollte versucht werden, die Prävalenz von B. henselae in unserer Hauskatzenpopulation abzuschätzen. Dazu wurden 930 EDTA-Katzenblutproben aus dem gesamten Bundesgebiet kulturell untersucht. Die Anzüchtung erfolgte auf doppelt dick gegossenem Hammelblutagar (7,5%) unter mikroaerophilen Bedingungen (5%CO2) über 4 Wochen. Es konnten 15 Stämme isoliert werden (1,61%). Ein Zusammenhang zwischen Alter der Katzen und kulturellem Ergebnis konnte nachgewiesen werden. 302 Katzenseren wurden auf Antikörper untersucht. Die Seroprävalenz betrug 37%. Dieses Ergebnis korreliert mit kulturellem Befund, Haltungsform, Flohbefall und geographischer Herkunft der Katzen. Mit den Isolaten wurden weitere Untersuchungen (PCR, PFGE, SDS-PAGE) zur Charakterisierung durchgeführt. Die Virulenz des Erregers wurde anhand verschiedener Untersuchungen des Referenzstammes untersucht. B. henselae kann nicht ohne schützendes Medium auf dem Fell des Wirtstieres überleben, benötigt bluthaltige Medien zur Isolierung und läßt sich auch nach längerer Aufbewahrung im Kühlschrank oder nach Einfrieren anzüchten. Aufgrund der geringen Anzahl kulturell positiver Befunde ist es schwierig, eine sichere Aussage zum Vorkommen von B. henselae zu treffen. Allerdings läßt die Seroprävalenz von 37% den Schluss zu, dass der Erreger zu einem nicht geringen Anteil unter den Katzen verbreitet ist. Eine Gefährdung gesunder, erwachsener Menschen besteht dabei nicht, aber für Kinder und immunschwache Personen bergen insbesondere verwilderte oder junge Katzen ein gewisses Risiko. Die Infektkette kann allerdings durch konsequente Flohbekämpfung unterbrochen werden.

Ctenocephalides felis est une espèce de puce cosmopolite parasitant majoritairement les carnivores domestiques. Elle est vectrice de nombreux agents pathogènes zoonotiques dont les bactéries du genre Bartonella, bactéries intracellulaires facultatives. La compétence vectorielle de cette puce a été investiguée pour B. henselae, B. quintana, B. clarridgeiae, B. tribocorum et B. birtlesii. Dans ces conditions expérimentales, utilisant un système de gorgement sur membrane, ces espèces ont persisté pendant les trois jours d'une première étude, et pour B. henselae durant les 13 jours de survie des puces, dans une seconde étude. Les cinq espèces de bartonelles ont été retouvées dans les fèces. Pour ces cinq espèces, nos résultats montrent une absence de transmission verticale transovarienne chez la puce et suggèrent une possibilité de contamination horizontale. Nous proposons enfin un protocole original d'évaluation de l'efficacité d'un traitement antiparasitaire chez le chat, pour prévenir sa contamination par Bartonella spp. / Ctenocephalides felis is a cosmopolitan flea species mainly parasitizing pets, transmitting several pathogens of veterinary and zoonotic importance including the facultative intracellular bacteria of the genus Bartonella. The vector competence of this flea was investigated for B. henselae, B. quintana, B. clarridgeiae, B. tribocorum and B. birtlesii, using an artificial feeding system. In these experimental conditions, these bartonellae proved to persist for three days, in a first study, while B. henselae persisted for the 13 days of its life span, in a second study. All five bartonellae were excreted in the flea's faeces. On the whole, these five species were not transmitted transovarially in the fleas, though horizontal transmission was suggested. Furthermore, we propose an original protocol allowing the evaluation of the efficacy of ectoparasiticidal products against Bartonella spp. infection in cats.

Endocardite infecciosa é uma doença associada à elevada morbidade e letalidade. O diagnóstico precoce e o reconhecimento de sua etiologia podem contribuir para o sucesso do tratamento antibiótico; entretanto, cerca de um quarto das endocardites permanece sem diagnóstico etiológico. Este estudo teve como objetivo principal identificar a frequência de endocardite por Bartonella spp. e Coxiella burnetii dentre as endocardites com culturas negativas comunitárias e avaliar os fatores preditores dessas infecções. Como objetivo secundário compararam-se as características clínicolaboratoriais e prognósticas entre as endocardites comunitárias com culturas negativas e positivas. Foram avaliados também os fatores associados à letalidade intra-hospitalar das endocardites com culturas negativas. Entre janeiro de 2004 e janeiro de 2009, foram investigados 369 episódios consecutivos de endocardite em pacientes atendidos no Instituto do Coração do Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo - InCor HC-FMUSP. Foram estudados os casos que ocorreram em adultos, classificados pelos critérios de Duke modificados como \"endocardite definida\" e de origem comunitária. Assim, foram incluídos 221 episódios de endocardite, 170 com culturas positivas e 51 com culturas negativas. Neste último grupo, foram feitas as pesquisas sorológicas (reação de imunofluorescência indireta) e histopatológica de Bartonella spp. e Coxiella burnetii. Consideraram-se positivos títulos de imunoglobulina G (IgG) >= 800 para Bartonella henselae e ou Bartonella quintana, e IgG antifase I para C. burnetii > 800. O estudo histopatológico das valvas cardíacas foi capaz de identificar morfologicamente a etiologia de 87% das endocardites com culturas negativas, enquanto que o método de Gram do tecido a fresco o fez em somente 10% dos casos. As endocardites com culturas negativas apresentaram maior frequência de dispneia à admissão (p=0,001), menor valor de proteína C reativa (p=0,009), menor Fração de Ejeção do Ventrículo Esquerdo (Feve) (p=0,022) e necessitaram de mais tempo para o início do tratamento antibiótico para endocardite (p < 0,001) quando comparadas àquelas com culturas positivas. Não houve diferença estatisticamente significante entre os grupos na letalidade intra-hospitalar e na sobrevida após alta hospitalar. Verificou-se que a presença de diabetes mellitus (p=0,009) ou sepse grave na admissão (p=0,01) esteve independentemente associada ao óbito intra-hospitalar entre as endocardites com culturas negativas. Dez casos de endocardite por Bartonella spp. (frequência 19,6% [IC95%: 9,8 - 33,1]) e quatro casos de endocardite por Coxiella burnetii (frequência 7,8% [IC95%: 2,2 - 18,9]) foram diagnosticados dentre os 51 episódios de endocardite com culturas negativas. As endocardites por Bartonella spp. apresentavam menor Feve (p=0,025), associação com a identificação de cocobacilo Gram-negativo no exame histológico da valva cardíaca (p=0,001) e presença de gato no domicílio (p=0,001). Conclusões: Bartonella spp. e Coxiella burnetii foram as etiologias de quase um terço (27,5%) das endocardites comunitárias com culturas negativas. A presença de gato no domicílio, Feve <= 45%, e a identificação de cocobacilo Gramnegativo no exame histológico da valva cardíaca em pacientes com endocardite com culturas negativas parecem estar associadas à infecção por Bartonella spp. O exame histológico da valva cardíaca permitiu a identificação morfológica do micro-organismo na maioria dos casos, mesmo quando as hemoculturas estavam negativas. Não se observou diferença na letalidade intra-hospitalar e na sobrevida em longo prazo entre os dois grupos. A presença de diabetes mellitus ou sepse grave à admissão associou-se ao óbito hospitalar nas endocardites com culturas negativas / Infective endocarditis is associated with high morbidity and lethality. Early diagnosis and recognition of the specific etiology can contribute to successful antibiotic treatment. However, approximately one-fourth of endocarditis cases remain without an etiologic diagnosis. This study aimed to identify the frequency of endocarditis caused by Bartonella spp. and Coxiella burnetii among cases of community-acquired culture-negative endocarditis and to also assess risk factors for such infections. As a secondary objective, the clinical, laboratory and prognostic features of community-acquired endocarditis were compared. Factors related to the in-hospital lethality of culture-negative endocarditis were also assessed. Between January 2004 and January 2009, 369 consecutive cases of endocarditis were investigated in patients attending the no Instituto do Coração do Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo - InCor HC-FMUSP. Cases occurring in adults, those classified by the modified Duke criteria as \"defined endocarditis\" and community-acquired cases were studied. In total, 221 cases of endocarditis comprising 170 culture-positive and 51 culturenegative cases were included. For the culture-negative cases, serology (indirect immunofluorescence reaction) and histopathological analyses for Bartonella spp. and Coxiella burnetii were performed. Cases were considered positive for Bartonella henselae or Bartonella quintana with IgG titers >= 800 and for Coxiella burnetii with antiphase I IgG titers > 800. Histopathological studies of the cardiac valves were capable of morphologically identifying the etiology in 87% of the culture-negative endocarditis cases, whereas the Gram stain was only positive in 10% of cases using fresh tissue. Culture-negative endocarditis patients presented a greater frequency of dyspnea on admission (p=0.001), lower C-reactive protein levels (p=0.009), and a lower left ventricular ejection fraction (LVEF) (p=0.022), and they required more time to start antibiotic therapy (p < 0.001) when compared with culture-positive patients. There was no statistically significant difference between the two groups regarding in-hospital lethality or survival after hospital discharge. Diabetes mellitus (p=0.01) or severe sepsis on admission (p=0.01) were independently associated with in-hospital death for culture-negative endocarditis. Ten cases of endocarditis caused by Bartonella spp. (frequency 19.6% [IC95%: 9.8 - 33.1]) and 4 caused by Coxiella burnetii (frequency 7.8% [IC95%: 2.2 - 18.9]) were diagnosed among the 51 cases of culture-negative endocarditis. Endocarditis caused by Bartonella spp. was associated with lower LVEF values (p=0.025), the identification of Gram-negative coccobacilli in cardiac valve histology (p=0.001) and the presence of a cat in the patient\'s residence (p=0.001). Conclusions: Bartonella spp. and Coxiella burnetii were the causative etiology of almost one-third (27.5%) of the community-acquired cases of culture-negative endocarditis. The presence of a cat in the patient\'s residence, a LVEF <= 45% and the identification of Gram-negative coccobacilli in the histological examination of the cardiac valve in patients with culturenegative endocarditis appear to be associated with Bartonella spp. as the causative etiology. Histological examination of the cardiac valves allowed for morphological identification of the causative microorganism in the majority of cases, even when blood cultures were negative. There was no difference in in-hospital lethality or long-term survival between the two groups. The presence of diabetes mellitus or severe sepsis at admission was associated with in-hospital death in cases of culture-negative endocarditis

Bartoneloses são infecções causadas por Bartonella spp e constituem zoonoses de importância crescente devido à gravidade da infecção em pacientes imunodeficientes. Existem muitas síndromes clínicas associadas às bartoneloses, desde quadros típicos de doença da arranhadura do gato até lesões cutâneas linfoproliferativas (angiomatose bacilar), ou granulomatosas em órgãos (peliose bacilar), passando por quadros neurológicos, oculares, endocardites e febre prolongada. O diagnóstico laboratorial deveria ser baseado em isolamento da bactéria em cultura, resultados de sorologias e exames imunohistoquímicos, porém estas técnicas não se encontram disponíveis em nosso meio. A Reação da Cadeia da Polimerase (PCR) tem sido utilizada para aprimorar o diagnóstico das bartoneloses, e o presente estudo teve como objetivo padronizar três sistemas de dupla amplificação para detecção de DNA de Bartonella henselae em amostras biológicas de pacientes com suspeita de bartonelose e determinar dentre os sistemas de dupla amplificação (nested-PCR), aquele com melhor desempenho. Os nested-PCR empregaram oligonucleotídeosiniciadores das sequências ITS, HSP e FtsZ da Bartonella spp. Foram incluídos 19 pacientes, sendo 14 crianças/adolescentes e cinco adultos, tendo oito deles referido contato com gatos. Dos pacientes, 10 não tinham doenças de base, e nove apresentavam doença de base (três evoluíram a óbito). Foram coletadas 25 amostras: 14 de sangue periférico, sete biópsias de gânglios, duas punções de gânglios e duas biópsias de pele. Do total de 19 pacientes, 14 tiveram resultados positivos por PCR, e considerando as 25 amostras, 18 foram positivas por PCR (nove amostras de sangue, cinco biópsias de gânglios, duas punções de gânglios e duas biópsias de pele). Dez das 18 amostras positivas por PCR só foram detectadas pelo nested-FtsZ (seis amostras de sangue periférico, duas punções de gânglios, uma biópsia de gânglio e uma biópsia de pele). A primeira amplificação do sistema HSP-PCR detectou uma amostra positiva em 25 (1/25 ou 4%), e após o nested-HSP este percentual foi de 24% (6/25). Na primeira amplificação do sistema ITS-PCR a positividade foi de 20% (5/25), e no nested-ITS 32% (8/25). Após a primeira amplificação do sistema FtsZ-PCR a positividade foi de 16% (4/25), e no nested-FtsZ foi de 72% (18/25). O teste de McNemar testou a concordância ou discordância dos resultados obtidos pelos três sistemas de nested-PCR e revelou que os mesmos foram discordantes, com superioridade do sistema nested-FtsZ em relação aos outros dois nested-PCR, sendo a diferença estatisticamente significante (p<0,05). Os resultados do presente estudo permitiram concluir que o nested-FtsZ apresentou vantagens para o diagnóstico das bartoneloses em relação aos outros sistemas testados, em materiais biológicos previamentedescritos na literatura (biópsias e punções de gânglios), e até mesmo em sangue periférico de pacientes sem doença de base. Além disso, foi possível determinar, no presente estudo, que as infecções foram causadas por B. henselae, uma vez que não houve nenhuma PCR positiva pelos outros dois sistemas que não tenha sido detectada pelo nested-FstZ, e este amplifica apenas DNA de Bartonella henselae após a segunda amplificação. / Bartonellosis are infections caused by Bartonella spp and this zoonosis is of growing importance due to the severity of infection in immunodeficient patients. Many clinical syndromes are associated with bartonellosis, varying from typical manifestations of cat scratch disease, to lymphoproliferative skin lesions (bacillary angiomatosis) or granulomatous ones in organs (bacillary peliosis), and also neurologic and ocular manifestations, endocarditis and prolonged fever. The laboratory diagnosis should be based on bacteria isolation in culture, serological tests and immunohistochemical examination, but these techniques are not available in our country. The Polymerase Chain Reaction (PCR) has been used to improve the diagnosis of bartonellosis, and this study has aimed at standardizing three nested-amplifications for detection of Bartonella henselae DNA in biological samples of patients with suspected bartonellosis, and establishing among the three nested-PCR, the one presenting with the best performance. Nested-PCR amplifications were performed using the ITS, the HSP and the FtsZ gene sequences of Bartonella spp. We included 19 patients, being 14 children / adolescents andfive adults, and eight reported contact with cats. From these patients, 10 had no underlying disease, and nine patients had underlying conditions (3 evolved to death). We collected 25 samples: 14 peripheral blood samples, seven lymph nodes biopsies, two lymph nodes punctures and two skin biopsies. From the total of 19 patients, 14 had positive results by PCR, and considering the 25 samples, 18 were positive by PCR (nine peripheral blood samples, five lymph nodes biopsies, two lymph nodes punctures and two skin biopsies). Ten of the 18 PCR-positive samples were only detected by the nested-FtsZ (six peripheral blood samples, two lymph nodes punctures, one lymph node biopsy and one skin biopsy). The first HSP-PCR amplification detected one positive sample in 25 (1 / 25 or 4%), and after the nested-HSP this percentage was 24% (6 / 25). In the first ITS-PCR amplification the positivity was 20% (5 / 25), and after the nested-ITS 32% (8 / 25). The first FtsZ-PCR amplification positivity was 16% (4 / 25), and after the nested-FtsZ it was 72% (18/25). The McNemar test was used to verify the concordance or discordance of the results obtained by the three nested-PCR systems and showed that they were discordant, with superiority of the nested-FtsZ in relation to the other two nested-PCR, being the difference statistically significant (p<0.05). The results of this study indicated that the nested-FtsZ showed advantages for the diagnosis of bartonellosis with respect to the other two systems when biological materials previously described in literature were tested (punctures and lymph nodes biopsies), and also in peripheral blood of patients without underlying conditions. Furthermore, it was possible to determine that the infections were caused byB. henselae, since there was no positive PCR results obtained by the other two systems that were not detected by the nested-FstZ, that only amplifies DNA from Bartonella henselae after the second round of amplification.

Bartoneloses são infecções causadas por Bartonella spp e constituem zoonoses de importância crescente devido à gravidade da infecção em pacientes imunodeficientes. Existem muitas síndromes clínicas associadas às bartoneloses, desde quadros típicos de doença da arranhadura do gato até lesões cutâneas linfoproliferativas (angiomatose bacilar), ou granulomatosas em órgãos (peliose bacilar), passando por quadros neurológicos, oculares, endocardites e febre prolongada. O diagnóstico laboratorial deveria ser baseado em isolamento da bactéria em cultura, resultados de sorologias e exames imunohistoquímicos, porém estas técnicas não se encontram disponíveis em nosso meio. A Reação da Cadeia da Polimerase (PCR) tem sido utilizada para aprimorar o diagnóstico das bartoneloses, e o presente estudo teve como objetivo padronizar três sistemas de dupla amplificação para detecção de DNA de Bartonella henselae em amostras biológicas de pacientes com suspeita de bartonelose e determinar dentre os sistemas de dupla amplificação (nested-PCR), aquele com melhor desempenho. Os nested-PCR empregaram oligonucleotídeosiniciadores das sequências ITS, HSP e FtsZ da Bartonella spp. Foram incluídos 19 pacientes, sendo 14 crianças/adolescentes e cinco adultos, tendo oito deles referido contato com gatos. Dos pacientes, 10 não tinham doenças de base, e nove apresentavam doença de base (três evoluíram a óbito). Foram coletadas 25 amostras: 14 de sangue periférico, sete biópsias de gânglios, duas punções de gânglios e duas biópsias de pele. Do total de 19 pacientes, 14 tiveram resultados positivos por PCR, e considerando as 25 amostras, 18 foram positivas por PCR (nove amostras de sangue, cinco biópsias de gânglios, duas punções de gânglios e duas biópsias de pele). Dez das 18 amostras positivas por PCR só foram detectadas pelo nested-FtsZ (seis amostras de sangue periférico, duas punções de gânglios, uma biópsia de gânglio e uma biópsia de pele). A primeira amplificação do sistema HSP-PCR detectou uma amostra positiva em 25 (1/25 ou 4%), e após o nested-HSP este percentual foi de 24% (6/25). Na primeira amplificação do sistema ITS-PCR a positividade foi de 20% (5/25), e no nested-ITS 32% (8/25). Após a primeira amplificação do sistema FtsZ-PCR a positividade foi de 16% (4/25), e no nested-FtsZ foi de 72% (18/25). O teste de McNemar testou a concordância ou discordância dos resultados obtidos pelos três sistemas de nested-PCR e revelou que os mesmos foram discordantes, com superioridade do sistema nested-FtsZ em relação aos outros dois nested-PCR, sendo a diferença estatisticamente significante (p<0,05). Os resultados do presente estudo permitiram concluir que o nested-FtsZ apresentou vantagens para o diagnóstico das bartoneloses em relação aos outros sistemas testados, em materiais biológicos previamentedescritos na literatura (biópsias e punções de gânglios), e até mesmo em sangue periférico de pacientes sem doença de base. Além disso, foi possível determinar, no presente estudo, que as infecções foram causadas por B. henselae, uma vez que não houve nenhuma PCR positiva pelos outros dois sistemas que não tenha sido detectada pelo nested-FstZ, e este amplifica apenas DNA de Bartonella henselae após a segunda amplificação. / Bartonellosis are infections caused by Bartonella spp and this zoonosis is of growing importance due to the severity of infection in immunodeficient patients. Many clinical syndromes are associated with bartonellosis, varying from typical manifestations of cat scratch disease, to lymphoproliferative skin lesions (bacillary angiomatosis) or granulomatous ones in organs (bacillary peliosis), and also neurologic and ocular manifestations, endocarditis and prolonged fever. The laboratory diagnosis should be based on bacteria isolation in culture, serological tests and immunohistochemical examination, but these techniques are not available in our country. The Polymerase Chain Reaction (PCR) has been used to improve the diagnosis of bartonellosis, and this study has aimed at standardizing three nested-amplifications for detection of Bartonella henselae DNA in biological samples of patients with suspected bartonellosis, and establishing among the three nested-PCR, the one presenting with the best performance. Nested-PCR amplifications were performed using the ITS, the HSP and the FtsZ gene sequences of Bartonella spp. We included 19 patients, being 14 children / adolescents andfive adults, and eight reported contact with cats. From these patients, 10 had no underlying disease, and nine patients had underlying conditions (3 evolved to death). We collected 25 samples: 14 peripheral blood samples, seven lymph nodes biopsies, two lymph nodes punctures and two skin biopsies. From the total of 19 patients, 14 had positive results by PCR, and considering the 25 samples, 18 were positive by PCR (nine peripheral blood samples, five lymph nodes biopsies, two lymph nodes punctures and two skin biopsies). Ten of the 18 PCR-positive samples were only detected by the nested-FtsZ (six peripheral blood samples, two lymph nodes punctures, one lymph node biopsy and one skin biopsy). The first HSP-PCR amplification detected one positive sample in 25 (1 / 25 or 4%), and after the nested-HSP this percentage was 24% (6 / 25). In the first ITS-PCR amplification the positivity was 20% (5 / 25), and after the nested-ITS 32% (8 / 25). The first FtsZ-PCR amplification positivity was 16% (4 / 25), and after the nested-FtsZ it was 72% (18/25). The McNemar test was used to verify the concordance or discordance of the results obtained by the three nested-PCR systems and showed that they were discordant, with superiority of the nested-FtsZ in relation to the other two nested-PCR, being the difference statistically significant (p<0.05). The results of this study indicated that the nested-FtsZ showed advantages for the diagnosis of bartonellosis with respect to the other two systems when biological materials previously described in literature were tested (punctures and lymph nodes biopsies), and also in peripheral blood of patients without underlying conditions. Furthermore, it was possible to determine that the infections were caused byB. henselae, since there was no positive PCR results obtained by the other two systems that were not detected by the nested-FstZ, that only amplifies DNA from Bartonella henselae after the second round of amplification.

Endocardite infecciosa é uma doença associada à elevada morbidade e letalidade. O diagnóstico precoce e o reconhecimento de sua etiologia podem contribuir para o sucesso do tratamento antibiótico; entretanto, cerca de um quarto das endocardites permanece sem diagnóstico etiológico. Este estudo teve como objetivo principal identificar a frequência de endocardite por Bartonella spp. e Coxiella burnetii dentre as endocardites com culturas negativas comunitárias e avaliar os fatores preditores dessas infecções. Como objetivo secundário compararam-se as características clínicolaboratoriais e prognósticas entre as endocardites comunitárias com culturas negativas e positivas. Foram avaliados também os fatores associados à letalidade intra-hospitalar das endocardites com culturas negativas. Entre janeiro de 2004 e janeiro de 2009, foram investigados 369 episódios consecutivos de endocardite em pacientes atendidos no Instituto do Coração do Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo - InCor HC-FMUSP. Foram estudados os casos que ocorreram em adultos, classificados pelos critérios de Duke modificados como \"endocardite definida\" e de origem comunitária. Assim, foram incluídos 221 episódios de endocardite, 170 com culturas positivas e 51 com culturas negativas. Neste último grupo, foram feitas as pesquisas sorológicas (reação de imunofluorescência indireta) e histopatológica de Bartonella spp. e Coxiella burnetii. Consideraram-se positivos títulos de imunoglobulina G (IgG) >= 800 para Bartonella henselae e ou Bartonella quintana, e IgG antifase I para C. burnetii > 800. O estudo histopatológico das valvas cardíacas foi capaz de identificar morfologicamente a etiologia de 87% das endocardites com culturas negativas, enquanto que o método de Gram do tecido a fresco o fez em somente 10% dos casos. As endocardites com culturas negativas apresentaram maior frequência de dispneia à admissão (p=0,001), menor valor de proteína C reativa (p=0,009), menor Fração de Ejeção do Ventrículo Esquerdo (Feve) (p=0,022) e necessitaram de mais tempo para o início do tratamento antibiótico para endocardite (p < 0,001) quando comparadas àquelas com culturas positivas. Não houve diferença estatisticamente significante entre os grupos na letalidade intra-hospitalar e na sobrevida após alta hospitalar. Verificou-se que a presença de diabetes mellitus (p=0,009) ou sepse grave na admissão (p=0,01) esteve independentemente associada ao óbito intra-hospitalar entre as endocardites com culturas negativas. Dez casos de endocardite por Bartonella spp. (frequência 19,6% [IC95%: 9,8 - 33,1]) e quatro casos de endocardite por Coxiella burnetii (frequência 7,8% [IC95%: 2,2 - 18,9]) foram diagnosticados dentre os 51 episódios de endocardite com culturas negativas. As endocardites por Bartonella spp. apresentavam menor Feve (p=0,025), associação com a identificação de cocobacilo Gram-negativo no exame histológico da valva cardíaca (p=0,001) e presença de gato no domicílio (p=0,001). Conclusões: Bartonella spp. e Coxiella burnetii foram as etiologias de quase um terço (27,5%) das endocardites comunitárias com culturas negativas. A presença de gato no domicílio, Feve <= 45%, e a identificação de cocobacilo Gramnegativo no exame histológico da valva cardíaca em pacientes com endocardite com culturas negativas parecem estar associadas à infecção por Bartonella spp. O exame histológico da valva cardíaca permitiu a identificação morfológica do micro-organismo na maioria dos casos, mesmo quando as hemoculturas estavam negativas. Não se observou diferença na letalidade intra-hospitalar e na sobrevida em longo prazo entre os dois grupos. A presença de diabetes mellitus ou sepse grave à admissão associou-se ao óbito hospitalar nas endocardites com culturas negativas / Infective endocarditis is associated with high morbidity and lethality. Early diagnosis and recognition of the specific etiology can contribute to successful antibiotic treatment. However, approximately one-fourth of endocarditis cases remain without an etiologic diagnosis. This study aimed to identify the frequency of endocarditis caused by Bartonella spp. and Coxiella burnetii among cases of community-acquired culture-negative endocarditis and to also assess risk factors for such infections. As a secondary objective, the clinical, laboratory and prognostic features of community-acquired endocarditis were compared. Factors related to the in-hospital lethality of culture-negative endocarditis were also assessed. Between January 2004 and January 2009, 369 consecutive cases of endocarditis were investigated in patients attending the no Instituto do Coração do Hospital das Clínicas da Faculdade de Medicina da Universidade de São Paulo - InCor HC-FMUSP. Cases occurring in adults, those classified by the modified Duke criteria as \"defined endocarditis\" and community-acquired cases were studied. In total, 221 cases of endocarditis comprising 170 culture-positive and 51 culturenegative cases were included. For the culture-negative cases, serology (indirect immunofluorescence reaction) and histopathological analyses for Bartonella spp. and Coxiella burnetii were performed. Cases were considered positive for Bartonella henselae or Bartonella quintana with IgG titers >= 800 and for Coxiella burnetii with antiphase I IgG titers > 800. Histopathological studies of the cardiac valves were capable of morphologically identifying the etiology in 87% of the culture-negative endocarditis cases, whereas the Gram stain was only positive in 10% of cases using fresh tissue. Culture-negative endocarditis patients presented a greater frequency of dyspnea on admission (p=0.001), lower C-reactive protein levels (p=0.009), and a lower left ventricular ejection fraction (LVEF) (p=0.022), and they required more time to start antibiotic therapy (p < 0.001) when compared with culture-positive patients. There was no statistically significant difference between the two groups regarding in-hospital lethality or survival after hospital discharge. Diabetes mellitus (p=0.01) or severe sepsis on admission (p=0.01) were independently associated with in-hospital death for culture-negative endocarditis. Ten cases of endocarditis caused by Bartonella spp. (frequency 19.6% [IC95%: 9.8 - 33.1]) and 4 caused by Coxiella burnetii (frequency 7.8% [IC95%: 2.2 - 18.9]) were diagnosed among the 51 cases of culture-negative endocarditis. Endocarditis caused by Bartonella spp. was associated with lower LVEF values (p=0.025), the identification of Gram-negative coccobacilli in cardiac valve histology (p=0.001) and the presence of a cat in the patient\'s residence (p=0.001). Conclusions: Bartonella spp. and Coxiella burnetii were the causative etiology of almost one-third (27.5%) of the community-acquired cases of culture-negative endocarditis. The presence of a cat in the patient\'s residence, a LVEF <= 45% and the identification of Gram-negative coccobacilli in the histological examination of the cardiac valve in patients with culturenegative endocarditis appear to be associated with Bartonella spp. as the causative etiology. Histological examination of the cardiac valves allowed for morphological identification of the causative microorganism in the majority of cases, even when blood cultures were negative. There was no difference in in-hospital lethality or long-term survival between the two groups. The presence of diabetes mellitus or severe sepsis at admission was associated with in-hospital death in cases of culture-negative endocarditis

Memoria para optar al Título Profesional de Médico Veterinario / Con el propósito de conocer si existe asociación entre la presentación de gingivitis con la seropositividad a Bartonella henselae en felinos domésticos, se analizaron 144 gatos en la ciudad de Santiago, provenientes de los Hospitales Clínicos Veterinarios de la Universidad de Chile, de ambos sexos, mayores de un año, negativos a las pruebas de Inmunodeficiencia Viral Felina y Leucemia Viral Felina. Todos los gatos fueron clasificados, según la presencia o ausencia de gingivitis, en dos grupos de 72 gatos cada uno. Para la determinación clínica de gingivitis se utilizó una adaptación del índice descrito por Loe y Silness. Para el diagnóstico serológico de Bartonella henselae se utilizó una prueba comercial de inmunofluorescencia indirecta (SLIDE ®). De los todos gatos analizados, el 73,6% fue seropositivo a Bartonella henselae y de los 72 gatos con gingivitis, el 87,5% de ellos fueron seropositivos. Mediante la prueba de Chi cuadrado, se determinó que no hay independencia entre la presentación de gingivitis y la seropositividad a Bartonella henselae (p<0,05)

Cat scratch's disease caused by Bartonella henselae, is known to be a self-limited benign process in immunocompetent children. The association with neurologic manifestations is very uncommon especially in patient with no immunologic defects and in cases without specific treatment. A 7 years old male patient, without any immunocompromised defect, presented an atypic presentation of the cat scratch disease. The patient came to the hospital in two opportunities in a status epilepticus, in both cases the diagnosis was encephalitis by Bartonella henselae and the evolution with treatment was monitored with PCR (polymerase chain reaction) in cerebrospinal fluid and blood, as well as IFI (IgM, IgG) serology (indirect immunofluorescence). The patient had a favorable clinical and laboratory evolution for 6 months showing no recurrence of the disease. / This work has been partially supported by the Programa Nacional de Innovacion para la Competitividad y Productividad ´ (Innovate Per ´ u), under the contract 116-PNICP-PIAP-2015.

The Gram-negative bacterium Bartonella henselae (Bh) is an emerging zoonotic pathogen that has been associated with a variety of human diseases including bacillary angiomatosis which is characterized by vasoproliferative tumor-like lesions on the skin and internal organs of some immunosuppressed individuals. Several virulence factors associated with Bartonella-induced pathogenesis have been characterized. However, the study of those virulence factors has been limited to in vitro cell culture systems due to the lack of a practical animal model. Therefore, we wanted to investigate whether the zebrafish embryo (Danio rerio) could be used to model human infection with Bh. We investigated if Bh can mount an infection in zebrafish embryos during their early stage of development. Our data showed that Tg(fli1:egfp)y1 zebrafish embryos supported a sustained Bh infection for 7 days with >10-fold bacterial replication when inoculated in the yolk sac. This was evident by plating of zebrafish homogenates, quantitative PCR, and confocal microscopy analysis. We assessed the interaction of Bh with EC and the phagocytic cells in live embryos by microscopy. Our data showed that aggregates of Bh interact with the endothelium of the embryo vasculature. Evidence showed that Bh recruited phagocytes to the site of infection in the Tg(mpx:GFP)uwm1 embryos. We also wanted to determine the response to infection with Bh. Infected embryos showed evidence of a Bh-induced angiogenic phenotype as well as an increase in expression of genes encoding pro-inflammatory factors and pro-angiogenic markers. A deletion mutant for the entire VirB type IV secretion system (ΔvirB2-11 supported bacterial replication although to a lesser degree compared to the wild type control. However, infection of zebrafish embryos with a deletion mutant in the major adhesin (BadA) resulted in little or no bacterial replication and a diminished pro-angiogenic and pro-inflammatory host response compared to wild type Bh, providing the first evidence that BadA is critical for in vivo infection. Thus, the zebrafish embryo provides the first practical animal model of Bh infection that will facilitate efforts to identify virulence factors and define molecular mechanisms of Bh pathogenesis.

Dans le cadre de cette thèse, nous avons mis au point un outil de différenciation moléculaire performant, simple et transférable pour Bartonella henselae, basé sur la technique MLVA. 5 VNTR dits " principaux " ont été sélctionnés pour leur polymorphisme (BHVA- E). Ceci nous a permis d'évaluer la diversité des souches et/ou isolats de B. henselae. Avec ces 5 VNTR, et pour 178 isolats et /ou souches testés, un index de diversité de 0.98 et 99 profils ont été obtenus. Ces profils se répartissent en groupes A et B. Le groupe A n'inclut que des soldats félins, alors que le groupe B est constitué d'isolats félins, d'un isolat canin et de la totalité des isolats humains testés. Une étude réalisée sur des isolats de chats et de leurs propriétaires a montré que la technique MLVA est un outil efficace pour la traçabilité. Les VNTR les moins polymorphes semblent pouvoir jouer le rôle de marqueur géographique, alors que pour les BHV les plus polymorphes, certains allèles sont particulièrement associés aux isolats humains. Aucun profil commun aux génotypes I et II n'a été rencontré. Nos observations suggèrent par ailleurs que tous les isolats du groupe B, c'est-à-dire les isolats de génotype I (d'origine humaine et féline) ainsi que tous les isolats humains appartenant à l'un ou l'autre des 2 génotypes, pourraient être dérivés d'isolats félins de génotype II (groupe A). En outre, la technique MLVA s'est avérée capable de typer des " variants " de B. henselae issus de félidés sauvages. La comparaison des performances de la technique MLVA versus les autres techniques déjà développées telles que ECP, MLST et MST, utilisant des souches communes, a montré que la technique MLVA est plus discriminante que l'ensemble de ces techniques et par ailleurs plus stable que l'ECP. Enfin, nos résultats suggèrent que seul le groupe B serait zoonotique, et que parmi les 5 VNTR polymorphes intragéniques que nous avons utilisés, certains au moins joueraient un rôle dans le potentiel zoonotique, la persistance chez le chat et/ou la virulence pour l'Homme.