Lyme borreliosis, more commonly referred to as Lyme disease, is the fastest growing zoonotic disease in North America with approximately 30,000 confirmed cases and 300,000 estimated infections per year. In nature, the causative agent of Lyme disease, the bacterium Borrelia burgdorferi, cycles between Ixodes sp. ticks and small mammals. Humans become infected with Lyme disease after being bitten by an infected tick. The primary indicator of a Borrelia burgdorferi infection is a bull’s eye rash typically followed by flu-like symptoms with treatment consisting of a 2-4 week course of antibiotics. If not treated, later stages of the disease can result in arthritis, cardiovascular and neurological symptoms. Diagnosis of Lyme disease is challenging and currently requires a complex laboratory diagnostic using indirect detection of host-generated antibodies by a two-tiered approach consisting of an enzyme linked immunosorbent assay (ELISA) followed by IgM and IgG immunoblots. Although two-tier testing has provided an adequate approach for Lyme disease diagnosis, it has weaknesses including subjective analysis, complex protocols and lack of reagent standardization. Immuno-PCR (iPCR) is a method that combines ELISA-based detection specificity with the sensitivity of PCR signal amplification and has demonstrated increased sensitivity for many applications such as detection of disease biomarkers but has yet to be applied for diagnosis of Lyme disease. Herein, using iPCR and recombinant B. burgdorferi antigens, an assay for both the direct and the indirect detection of Lyme disease was developed iv and demonstrated improved sensitivity for detection of B. burgdorferi antibodies using a murine model. Moreover, we present evidence using human Lyme disease patient serum samples that iPCR using both multiple antigens and a unique single hybrid antigen is capable of achieving increased sensitivity and specificity compared to existing methodology. These data represent the first demonstration of iPCR for Lyme disease diagnosis and support the replacement of two-tier testing with a more simplified and objective approach.

The Lyme disease spirochete Borrelia burgdorferi is the only known human pathogen that directly activates invariant natural killer T (iNKT) cells. The number and activation kinetics of iNKT cells vary greatly among different strains of mice. Here, we report the role of the iNKT cell response in the pathogenesis of Lyme disease using C57BL/6 (B6) mice, a strain with optimal iNKT cell activation that is resistant to the development of spirochetal-induced inflammation. During experimental infection of B6 mice with B. burgdorferi , iNKT cells localize to the inflamed heart where they are activated by CD1d-expressing macrophages. Activation of iNKT cells in vivo results in the production of IFNγ, which we demonstrate controls the severity of murine Lyme carditis by at least two mechanisms. First, IFNγ greatly enhances the recognition of B. burgdorferi by macrophages, leading to increased phagocytosis of the spirochete. Secondly, IFNγ activation of macrophages increases the surface expression of CD1d, thereby facilitating further iNKT activation. Collectively, our data demonstrate that in the resistant background, B6, iNKT cells modulate acute murine Lyme carditis through the action of IFNγ, which appears to self-renew through a positive feedback loop during infection. Inflammation during infection with B. burgdorferi is dependent on the ability of the spirochete to evade local mechanisms of clearance. Even though macrophages are the main infiltrating cell during Lyme carditis, the identification of a receptor capable of mediating phagocytosis of B. burgdorferi has been elusive. Here, we demonstrate that the integrin CR3 is able to mediate binding to the spirochete and facilitate phagocytosis in a complement-dependent and independent manner. Expression of CR3, but not CR4, in CHO cells markedly enhanced their capacity to interact with B. burgdorferi , in the absence and presence of complement opsonization. Furthermore, the interaction between CR3 and B. burgdorferi is dependent on the metal-ion-dependent adhesion site (MIDAS) and could be blocked with EDTA. Inhibition of CR3 with blocking antibody was able to completely abrogate phagocytosis of B. burgdorferi by the macrophage-like RAW264.7 cells and partially block uptake by bone marrow-derived macrophages (BMMs), a finding that was recapitulated with CD11b-deficient BMMs. We further show that activation with recombinant IFNγ increases the transcription of CD11b and CD18, which correlates with increased surface expression of CR3, and that the effect of IFNγ on the phagocytosis of B. burgdorferi is circumscribed to CR3 activity, because inhibition of CR3 is able to completely diminish the effect of IFNγ on the phagocytosis of the B. burgdorferi . Lastly, our results demonstrate that CR3 is a negative regulator of proinflammatory cytokine induction in macrophages responding to B. burgdorferi . Overall, our data demonstrate roles for CR3 in the binding, phagocytosis and proinflammatory cytokine elicited by B. burgdorferi and shed light on the role of IFNγ in mediating the clearance of the spirochete during Lyme disease.

No description available.

Introdução e Objetivo: A doença mixomatosa da valva mitral leva ao comprometimento de sua matriz devido à alteração em sua composição tecidual provocada pelo desequilíbrio na quantidade de ácidos mucopolissacarídeos ou glicosaminoglicanos. Sua etiologia ainda não está totalmente esclarecida, podendo ocorrer em formas familiares com transmissão autossômica dominante de penetrância variável, que pode ser dependente do tempo ou de prováveis fatores ambientais, situações em que a interação de agentes infecciosos necessita de maiores esclarecimentos. O objetivo deste estudo é a análise dos produtos dos patógenos da Chlamydophila pneumoniae, Mycoplasma pneumoniae e Borrelia burgdorferi em segmentos de cúspide retirados da valva mitral com degeneração mixomatosa, comparada ao grupo controle e a relação dos produtos bacterianos com aumento de marcadores inflamatórios (CD20, CD48, CD68) e de metaloproteinase (MMP9) na etiopatogenia da degeneração mixomatosa da valva mitral. Método: Estudo observacional, analítico, tipo caso-controle, que analisou 2 grupos contendo 20 pacientes cada e divididos em grupo 1, composto por fragmentos de tecido valvar mitral com diagnóstico de degeneração mixomatosa extraídos em procedimentos de troca ou plásticas valvares mitrais; e grupo 2, formado por segmentos de valvas mitrais sem valvopatia retirados no serviço de verificação de óbito. Foram realizadas colorações de hematoxilina e eosina e Movat para diagnóstico histológico da degeneração mixomatosa e técnica de imunohistoquímica para detecção de antígenos da Borrelia burgdorferi, Mycoplasma pneumoniae, mediadores inflamatórios (CD20, CD45, CD68) e marcadores de metaloproteinase (MMP9). A presença de antígenos da Chlamydophila pneumonia e foi pesquisada pela técnica de hibridização in situ. A análise quantitativa dos aspectos microscópicos foi realizada com o analisador de imagens Aperio. A pesquisa de elementos bacterianos foi feita através de microscopia eletrônica de transmissão. Resultados: No grupo 1, 14 (70%) pacientes são do gênero masculino e 6 (30%) do gênero feminino. A idade média é de 67,4 anos (51 a 79 anos, dp = 9,2). No grupo 2, 11 (55%) pacientes são do gênero masculino e 9 (45%) do gênero feminino. A idade média é de 67,6 anos (42 a 84 anos, dp= 12,0). Na análise da porcentagem de degeneração mixomatosa pela coloração Movat, houve diferença com significância estatística entre os grupos DM (G1), com média de 54,6 % ± 23,7 e grupo controle (G2) com média de 35,5 % ± 22,5 com valor de p = 0,013. Houve um maior número de células CD20 positivas/mm2 no grupo com DM com mediana igual a 17,8 (6,7 - 27,9) x 4,6 (3,6 - 9,8) com p = 0,007 para a área 1. Houve maior número de células CD45 positivas/mm2 no grupo com DM com mediana igual a 17,3 (3,4 - 92,5) x 2,8 (1,4 - 10,1) com p = 0,008 para a área 1. Houve maior número de células CD68 positivas/mm2 no grupo controle (G2), porém sem significância estatística com mediana igual a 38,7 (26,6 - 81,8) x 70 (42,7 - 120,4) com p = 0,098 para a área 1. Em relação à presença de antígenos de Mycoplasma pneumoniae, houve uma maior área (?m2) de antígenos detectados no grupo 1, quando comparadas com o grupo 2 com diferença estatisticamente significante para as duas áreas. Na área 1, mediana de 180.993 (24.856 - 387.477) x 7.970 (2.736 - 15.992) com p < 0,001 e na área 2, mediana igual a 105.968 (2.503 - 416.585) x 7.190 (3.314 - 17.833) com p = 0,02 A análise da presença de antígenos de Chlamydophila pneumoniae demonstrou que em ambas as áreas, houve uma maior área (?m2) de antígenos detectados no grupo de valvas com degeneração mixomatosa, quando comparadas com o grupo controle, porém sem diferença estatística com mediana para o G1 de 9.905 (4.716 - 16.912) x 5.864 (2.382 - 8.692) com p = 0,2 e para o G2, mediana de 3.199 (1.791 - 10.746) x 2.536 (683 - 6.125) com p = 0,3. Em relação à presença de antígenos de Borrelia burgdorferi, houve uma maior área (um2) de antígenos detectados no grupo 2 em relação ao grupo 1, em ambas as áreas. Na área 1, mediana de 7.596 (3.203 - 13.519) x 10.584 (7.223 - 15.974) com p = 0,14 e na área 2, mediana igual a 5.991 (3.009 - 8.475) x 8.403 (1.626 - 27.887) com p = 0,47. Em relação à presença da metaloproteinase MMP9, observamos maior área (um2) de antígeno marcado de MMP9 no grupo com degeneração mixomatosa tanto na área 1 quanto na área 2, com diferença estatística significante. Na área 1, mediana de 503.894 (202.428 - 938.072) x 269.244 (111.953 - 354.022) com p = 0,03. Na área 2, houve diferença estatística representada pela mediana de 389.844 (214.459 - 679.711) x 144.397 (29.894 - 247.453) com p < 0,001. No grupo DM houve correlação positiva entre Borrelia burgdorferi e porcentagem de DM com R = 0,52 e p = 0,018. Em relação às células inflamatórias, houve correlação positiva entre CD45 e Mycoplasma pneumoniae com R = 0,51 e p = 0,02. A presença de MMP9 se correlacionou positivamente com a presença de Mycoplasma pneumoniae com R = 0,45 e p = 0,04. Estas correlações estiveram ausentes no grupo controle. Conclusões: Houve associação de agentes infecciosos Mycoplasma pneumoniae e Borrelia burgdorferi na etiopatiopatogenia da degeneração mixomatosa da valva mitral. Na análise da relação dos produtos bacterianos com os marcadores inflamatórios e com a metaloproteinase, houve relação positiva entre o marcador inflamatório CD45 e a metaloproteinase (MMP9) apenas com a Mycoplasma pneumoniae, nas valvas com degeneração mixomatosa. O marcador inflamatório CD68 foi encontrado em maior número no grupo controle / Background: The myxomatous mitral valve disease leads to impairment due to changes in their tissue composition caused by the imbalance in the amount of acid mucopolysaccharides or glycosaminoglycans. Its etiology is not yet fully understood and may occur in familial forms of autosomal dominant trait with variable penetrance that can be time-dependent or probable environmental factors, where the interaction of infectious agents requires further elucidation. The purpose of this study is the analysis of the pathogens products of Chlamydophila pneumoniae, Mycoplasma pneumoniae and Borrelia burgdorferi in removed cusp segments of the mitral valve with myxoid degeneration, compared to the control group and the ratio of bacterial products with increased inflammatory markers (CD20, CD48, CD68) and metalloproteinase (MMP9) in the pathogenesis of myxomatous degeneration of the mitral valve. Method: Observational, analytical, case-control study which analyzed 2 groups of 20 patients each and divided in group 1, consisting of fragments of mitral valve tissue with diagnosis of myxomatous degeneration extracted in replacement procedures or mitral valve repair; group 2, formed by segments of mitral valves without valvolpaty clinial disease removed in the coroner service. Hematoxylin and eosin and Movat stains were done for histological diagnosis of myxoid degeneration and immunohistochemical technique for the detection of Borrelia burgdorferi, Mycoplasma pneumonia antigens, inflammatory mediators (CD20, CD45, CD68) and markers of metalloproteinase (MMP9). The presence of Chlamydophila pneumonia antigens was verified through an in situ hybridization technique. The quantitative analysis of the microscopic aspects was performed with the Aperio image analyzer. The research of bacterial elements was performed by a transmission electron microscopy. Results: In group 1, 14 (70%) patients were male and 6 (30%) were female. The mean age was (51 to 79 years, sd = 9.2). In group 2, 11 (55%) patients were male gender and 9 (45%) were female. The mean age was 67,6 years (42 to 84 years, sd= 12). In the analysis percentage of myxomatous tissue by Movat staining, there was a significant difference between the DM (G1) groups, with a media of 54.6 % ± 23,7 and control group (G2) with a media of 35.5 % ± 22.5 with p = 0.013. There was an increased number of CD20 cells/mm2 in myxomatous degeneration group (G1) with a median of 17.8 (6.7 - 27.9) x 4.6 (3.6 - 9.8) with p = 0.007 for the area 1. There was a higher number of CD45 cells/mm2 in myxomatous degeneration group (G1) with a median of 17.3 (3.4 - 92.5) x 2.8 (1.4 - 10.1) with p = 0.008 for the area 1. There was a higher number of CD68 cells/mm2 in control group (G2) without a statistically significant difference, with a median of 38.7 (26.6 - 81.8) x 70 (42.7 - 120.4) with p = 0,098 for the area 1. In quantifying Mycoplasma pneumoniae we observed a higher area (um2) antigen marked by, there was a higher amount of antigen detected in myxomatous degeneration group. In area 1, a median of 180,993 (24,856 - 387,477) x 7,970 (2,736 - 15,992) with p < 0.001 and in area 2, a median of 105,968 (2,503 - 416,585) x 7,190 (3,314 - 17,833) with p = 0.02. The analysis of the presence of Chlamydophila pneumoniae antigens showed that in both area, there was a larger area (?m2) antigens detected in the group of valves with MD when compared with the control group, but without significant differences with median for the G1 of 9,905 (4,716 - 16,912) x 5,864 (2,382 - 8,692), with p = 0.2 and the G2, median 3,199 (1,791 - 10,746) x 2,536 (683 - 6,125) with p = 0.3. Regarding the presence of Borrelia burgdorferi antigens, there was a greater area (?m2) antigens detected in group 2 than in group 1, in both areas. In one area, median 7,596 (3,203 - 13,519) x 10,584 (7,223 - 15,974), with p = 0.14 and in area 2, a median of 5,991 (3,009 - 8,475) x 8,403 (1,626 - 27,887) with p = 0.47. Regarding the presence of metalloproteinase MMP9, we observed a higher area (um2) antigen marked by MMP9 in the group with MD both in area 1and area 2, with statistically significant difference. In area 1, median of 503,894 (202,428 - 938,072) x 269,244 (111,953 - 354,022), p = 0.03. In area 2, median 389,844 (214,459 - 679,711) x 144,397 (29,894 - 247,453) with p < 0.001. In the DM group there was a positive correlation between Borrelia burgdorferi and the percentage of MD with R = 0.52 and p = 0.018. Regarding inflammatory cells, there was a positive correlation between CD45 and Mycoplasma pneumoniae with R = 0.51 and p = 0.02. The presence of MMP9 was positively correlated with the presence of Mycoplasma pneumoniae with R = 0.45 and p = 0.04. These correlations were absent in the control group. Conclusions: There was an association of infectious agents Mycoplasma pneumoniae and Borrelia burgdorferi in etiopathogeny of myxoid degeneration of the mitral valve. In the analysis of the relationship of bacterial products with the inflammatory markers and the metalloproteinase, there was a positive relationship between the inflammatory marker CD45 and metalloproteinase (MMP9) only with Mycoplasma pneumoniae. The inflammatory marker CD68 was found in greater numbers in the control group

A Doença de Lyme-símile brasileira ou Síndrome Baggio-Yoshinari (SBY) é uma zoonose emergente, transmitida por carrapatos e até o momento, de descrição restrita ao território brasileiro. O agente etiológico da SBY era desconhecido até o presente trabalho. O objetivo principal do estudo foi identificar a etiologia da SBY. Foi selecionado 2 grupos de pacientes: grupo A (n=68) composto por pacientes com suspeita diagnóstica de SBY, a maioria na fase latente da doença; grupo B (n=10), composto por pacientes com diagnóstico de SBY, que apresentaram obrigatoriamente eritema migratório e que encontravam-se sintomáticos no momento da coleta. Foi utilizado também um grupo controle composto por indivíduos saudáveis e com epidemiologia negativa (n=50). Amostras de sangue foram coletadas para a realização de sorologias, culturas, análises microscópicas (óptica e eletrônica) e reação de cadeia da polimerase (PCR) para diferentes micro-organismos (Mycoplasma spp, Chlamydia spp e Borrelia spp). Além disso, foi realizado um estudo preliminar, através da PCR para Borrelia spp em 47 amostras de carrapatos oriundos de áreas de risco do Espírito Santo (sendo 17 Rhipicephalus microplus e 30 Rhipicephalus sanguineus), e amostras de sangue total de 27 bovinos e 26 equinos, animais estes oriundos da Universidade Federal Rural do Rio de Janeiro. Os resultados mostraram que a SBY não se trata de uma zoonose causada por um conjunto de micro-organismos como pensado inicialmente e sim pela Borrelia burgdorferi sensu lato. Descoberta essa que foi possível empregando-se novos primers amplificadores do principal gene envolvido na síntese do gancho flagelar da Borrelia, chamado flgE. A positividade para flgE foi confirmada em 6 pacientes do grupo B, 2 carrapatos, 1 bovino e 1 equino, os quais apresentaram homologia de 99% com o gene da proteína do gancho flagelar da Borrelia burgdorferi (flgE) depositado no GenBank (L43849). Esta importante descoberta, associada às pesquisas anteriores, permitiu definir a SBY como zoonose emergente e própria do país, causada pela bactéria B. burgdorferi na apresentação morfológica atípica, transmitida por carrapatos não pertencentes ao complexo Ixodes ricinus, responsável por manifestações clínicas semelhantes à Doença de Lyme, exceto pela grande frequência de sintomas recorrentes / Brazilian Lyme disease-like illness (BLDL) or Baggio-Yoshinari Syndrome (BYS) is an emerging zoonosis, transmitted by ticks and so far, restricted to the description of the Brazilian territory. The causative agent of BYS was unknown until now. The main objective of this study was to identify the etiology of BYS. We have selected two groups of patients: group A (n = 68) consisting of patients suspected of BYS, mostly in the latent stage of disease; group B (n = 10), composed of patients diagnosed with BYS, who had compulsorily erythema migrans and that were symptomatic at the time of blood collection. We also used a control group composed of healthy individuals with negative epidemiology (n = 50). Blood samples were collected, in which we performed serology, cultures, microscopic analysis (optical and electron) and polymerase chain reaction (PCR) for different microorganisms (Mycoplasma spp, Chlamydia spp and Borrelia spp). In addition, a preliminary study was conducted by PCR for Borrelia spp in 47 samples of ticks from risk areas at Espirito Santo State (being 17 Rhipicephalus microplus and 30 Rhipicephalus sanguineus), 27 cattle and 26 horses, being these animals from the Universidade Federal Rural do Rio de Janeiro. The results showed that BYS is not a zoonosis caused by a set of microorganisms as initially thought, but by Borrelia burgdorferi sensu lato. These findings were possible after employing new primers that are able to amplify portions of the main genes involved in the synthesis of the Borrelia flagellar hook protein, called flgE. We confirmed positivity for the flgE in 6 patients from group B, 2 ticks, a cow, and a horse, which showed 99% homology with the gene of Borrelia burgdorferi flagellar hook protein (flgE) deposited in GenBank (L43849). This important discovery, coupled with previous research, helped to define BYS as an emerging zoonosis particular from Brazil, caused by B. burgdorferi of atypical morphologic presentation, transmitted by ticks outside the Ixodes ricinus complex, responsible for clinical signs similar to Lyme disease, except for the high frequency of relapsing symptoms

Introdução e Objetivo: A doença mixomatosa da valva mitral leva ao comprometimento de sua matriz devido à alteração em sua composição tecidual provocada pelo desequilíbrio na quantidade de ácidos mucopolissacarídeos ou glicosaminoglicanos. Sua etiologia ainda não está totalmente esclarecida, podendo ocorrer em formas familiares com transmissão autossômica dominante de penetrância variável, que pode ser dependente do tempo ou de prováveis fatores ambientais, situações em que a interação de agentes infecciosos necessita de maiores esclarecimentos. O objetivo deste estudo é a análise dos produtos dos patógenos da Chlamydophila pneumoniae, Mycoplasma pneumoniae e Borrelia burgdorferi em segmentos de cúspide retirados da valva mitral com degeneração mixomatosa, comparada ao grupo controle e a relação dos produtos bacterianos com aumento de marcadores inflamatórios (CD20, CD48, CD68) e de metaloproteinase (MMP9) na etiopatogenia da degeneração mixomatosa da valva mitral. Método: Estudo observacional, analítico, tipo caso-controle, que analisou 2 grupos contendo 20 pacientes cada e divididos em grupo 1, composto por fragmentos de tecido valvar mitral com diagnóstico de degeneração mixomatosa extraídos em procedimentos de troca ou plásticas valvares mitrais; e grupo 2, formado por segmentos de valvas mitrais sem valvopatia retirados no serviço de verificação de óbito. Foram realizadas colorações de hematoxilina e eosina e Movat para diagnóstico histológico da degeneração mixomatosa e técnica de imunohistoquímica para detecção de antígenos da Borrelia burgdorferi, Mycoplasma pneumoniae, mediadores inflamatórios (CD20, CD45, CD68) e marcadores de metaloproteinase (MMP9). A presença de antígenos da Chlamydophila pneumonia e foi pesquisada pela técnica de hibridização in situ. A análise quantitativa dos aspectos microscópicos foi realizada com o analisador de imagens Aperio. A pesquisa de elementos bacterianos foi feita através de microscopia eletrônica de transmissão. Resultados: No grupo 1, 14 (70%) pacientes são do gênero masculino e 6 (30%) do gênero feminino. A idade média é de 67,4 anos (51 a 79 anos, dp = 9,2). No grupo 2, 11 (55%) pacientes são do gênero masculino e 9 (45%) do gênero feminino. A idade média é de 67,6 anos (42 a 84 anos, dp= 12,0). Na análise da porcentagem de degeneração mixomatosa pela coloração Movat, houve diferença com significância estatística entre os grupos DM (G1), com média de 54,6 % ± 23,7 e grupo controle (G2) com média de 35,5 % ± 22,5 com valor de p = 0,013. Houve um maior número de células CD20 positivas/mm2 no grupo com DM com mediana igual a 17,8 (6,7 - 27,9) x 4,6 (3,6 - 9,8) com p = 0,007 para a área 1. Houve maior número de células CD45 positivas/mm2 no grupo com DM com mediana igual a 17,3 (3,4 - 92,5) x 2,8 (1,4 - 10,1) com p = 0,008 para a área 1. Houve maior número de células CD68 positivas/mm2 no grupo controle (G2), porém sem significância estatística com mediana igual a 38,7 (26,6 - 81,8) x 70 (42,7 - 120,4) com p = 0,098 para a área 1. Em relação à presença de antígenos de Mycoplasma pneumoniae, houve uma maior área (?m2) de antígenos detectados no grupo 1, quando comparadas com o grupo 2 com diferença estatisticamente significante para as duas áreas. Na área 1, mediana de 180.993 (24.856 - 387.477) x 7.970 (2.736 - 15.992) com p < 0,001 e na área 2, mediana igual a 105.968 (2.503 - 416.585) x 7.190 (3.314 - 17.833) com p = 0,02 A análise da presença de antígenos de Chlamydophila pneumoniae demonstrou que em ambas as áreas, houve uma maior área (?m2) de antígenos detectados no grupo de valvas com degeneração mixomatosa, quando comparadas com o grupo controle, porém sem diferença estatística com mediana para o G1 de 9.905 (4.716 - 16.912) x 5.864 (2.382 - 8.692) com p = 0,2 e para o G2, mediana de 3.199 (1.791 - 10.746) x 2.536 (683 - 6.125) com p = 0,3. Em relação à presença de antígenos de Borrelia burgdorferi, houve uma maior área (um2) de antígenos detectados no grupo 2 em relação ao grupo 1, em ambas as áreas. Na área 1, mediana de 7.596 (3.203 - 13.519) x 10.584 (7.223 - 15.974) com p = 0,14 e na área 2, mediana igual a 5.991 (3.009 - 8.475) x 8.403 (1.626 - 27.887) com p = 0,47. Em relação à presença da metaloproteinase MMP9, observamos maior área (um2) de antígeno marcado de MMP9 no grupo com degeneração mixomatosa tanto na área 1 quanto na área 2, com diferença estatística significante. Na área 1, mediana de 503.894 (202.428 - 938.072) x 269.244 (111.953 - 354.022) com p = 0,03. Na área 2, houve diferença estatística representada pela mediana de 389.844 (214.459 - 679.711) x 144.397 (29.894 - 247.453) com p < 0,001. No grupo DM houve correlação positiva entre Borrelia burgdorferi e porcentagem de DM com R = 0,52 e p = 0,018. Em relação às células inflamatórias, houve correlação positiva entre CD45 e Mycoplasma pneumoniae com R = 0,51 e p = 0,02. A presença de MMP9 se correlacionou positivamente com a presença de Mycoplasma pneumoniae com R = 0,45 e p = 0,04. Estas correlações estiveram ausentes no grupo controle. Conclusões: Houve associação de agentes infecciosos Mycoplasma pneumoniae e Borrelia burgdorferi na etiopatiopatogenia da degeneração mixomatosa da valva mitral. Na análise da relação dos produtos bacterianos com os marcadores inflamatórios e com a metaloproteinase, houve relação positiva entre o marcador inflamatório CD45 e a metaloproteinase (MMP9) apenas com a Mycoplasma pneumoniae, nas valvas com degeneração mixomatosa. O marcador inflamatório CD68 foi encontrado em maior número no grupo controle / Background: The myxomatous mitral valve disease leads to impairment due to changes in their tissue composition caused by the imbalance in the amount of acid mucopolysaccharides or glycosaminoglycans. Its etiology is not yet fully understood and may occur in familial forms of autosomal dominant trait with variable penetrance that can be time-dependent or probable environmental factors, where the interaction of infectious agents requires further elucidation. The purpose of this study is the analysis of the pathogens products of Chlamydophila pneumoniae, Mycoplasma pneumoniae and Borrelia burgdorferi in removed cusp segments of the mitral valve with myxoid degeneration, compared to the control group and the ratio of bacterial products with increased inflammatory markers (CD20, CD48, CD68) and metalloproteinase (MMP9) in the pathogenesis of myxomatous degeneration of the mitral valve. Method: Observational, analytical, case-control study which analyzed 2 groups of 20 patients each and divided in group 1, consisting of fragments of mitral valve tissue with diagnosis of myxomatous degeneration extracted in replacement procedures or mitral valve repair; group 2, formed by segments of mitral valves without valvolpaty clinial disease removed in the coroner service. Hematoxylin and eosin and Movat stains were done for histological diagnosis of myxoid degeneration and immunohistochemical technique for the detection of Borrelia burgdorferi, Mycoplasma pneumonia antigens, inflammatory mediators (CD20, CD45, CD68) and markers of metalloproteinase (MMP9). The presence of Chlamydophila pneumonia antigens was verified through an in situ hybridization technique. The quantitative analysis of the microscopic aspects was performed with the Aperio image analyzer. The research of bacterial elements was performed by a transmission electron microscopy. Results: In group 1, 14 (70%) patients were male and 6 (30%) were female. The mean age was (51 to 79 years, sd = 9.2). In group 2, 11 (55%) patients were male gender and 9 (45%) were female. The mean age was 67,6 years (42 to 84 years, sd= 12). In the analysis percentage of myxomatous tissue by Movat staining, there was a significant difference between the DM (G1) groups, with a media of 54.6 % ± 23,7 and control group (G2) with a media of 35.5 % ± 22.5 with p = 0.013. There was an increased number of CD20 cells/mm2 in myxomatous degeneration group (G1) with a median of 17.8 (6.7 - 27.9) x 4.6 (3.6 - 9.8) with p = 0.007 for the area 1. There was a higher number of CD45 cells/mm2 in myxomatous degeneration group (G1) with a median of 17.3 (3.4 - 92.5) x 2.8 (1.4 - 10.1) with p = 0.008 for the area 1. There was a higher number of CD68 cells/mm2 in control group (G2) without a statistically significant difference, with a median of 38.7 (26.6 - 81.8) x 70 (42.7 - 120.4) with p = 0,098 for the area 1. In quantifying Mycoplasma pneumoniae we observed a higher area (um2) antigen marked by, there was a higher amount of antigen detected in myxomatous degeneration group. In area 1, a median of 180,993 (24,856 - 387,477) x 7,970 (2,736 - 15,992) with p < 0.001 and in area 2, a median of 105,968 (2,503 - 416,585) x 7,190 (3,314 - 17,833) with p = 0.02. The analysis of the presence of Chlamydophila pneumoniae antigens showed that in both area, there was a larger area (?m2) antigens detected in the group of valves with MD when compared with the control group, but without significant differences with median for the G1 of 9,905 (4,716 - 16,912) x 5,864 (2,382 - 8,692), with p = 0.2 and the G2, median 3,199 (1,791 - 10,746) x 2,536 (683 - 6,125) with p = 0.3. Regarding the presence of Borrelia burgdorferi antigens, there was a greater area (?m2) antigens detected in group 2 than in group 1, in both areas. In one area, median 7,596 (3,203 - 13,519) x 10,584 (7,223 - 15,974), with p = 0.14 and in area 2, a median of 5,991 (3,009 - 8,475) x 8,403 (1,626 - 27,887) with p = 0.47. Regarding the presence of metalloproteinase MMP9, we observed a higher area (um2) antigen marked by MMP9 in the group with MD both in area 1and area 2, with statistically significant difference. In area 1, median of 503,894 (202,428 - 938,072) x 269,244 (111,953 - 354,022), p = 0.03. In area 2, median 389,844 (214,459 - 679,711) x 144,397 (29,894 - 247,453) with p < 0.001. In the DM group there was a positive correlation between Borrelia burgdorferi and the percentage of MD with R = 0.52 and p = 0.018. Regarding inflammatory cells, there was a positive correlation between CD45 and Mycoplasma pneumoniae with R = 0.51 and p = 0.02. The presence of MMP9 was positively correlated with the presence of Mycoplasma pneumoniae with R = 0.45 and p = 0.04. These correlations were absent in the control group. Conclusions: There was an association of infectious agents Mycoplasma pneumoniae and Borrelia burgdorferi in etiopathogeny of myxoid degeneration of the mitral valve. In the analysis of the relationship of bacterial products with the inflammatory markers and the metalloproteinase, there was a positive relationship between the inflammatory marker CD45 and metalloproteinase (MMP9) only with Mycoplasma pneumoniae. The inflammatory marker CD68 was found in greater numbers in the control group

A Doença de Lyme-símile brasileira ou Síndrome Baggio-Yoshinari (SBY) é uma zoonose emergente, transmitida por carrapatos e até o momento, de descrição restrita ao território brasileiro. O agente etiológico da SBY era desconhecido até o presente trabalho. O objetivo principal do estudo foi identificar a etiologia da SBY. Foi selecionado 2 grupos de pacientes: grupo A (n=68) composto por pacientes com suspeita diagnóstica de SBY, a maioria na fase latente da doença; grupo B (n=10), composto por pacientes com diagnóstico de SBY, que apresentaram obrigatoriamente eritema migratório e que encontravam-se sintomáticos no momento da coleta. Foi utilizado também um grupo controle composto por indivíduos saudáveis e com epidemiologia negativa (n=50). Amostras de sangue foram coletadas para a realização de sorologias, culturas, análises microscópicas (óptica e eletrônica) e reação de cadeia da polimerase (PCR) para diferentes micro-organismos (Mycoplasma spp, Chlamydia spp e Borrelia spp). Além disso, foi realizado um estudo preliminar, através da PCR para Borrelia spp em 47 amostras de carrapatos oriundos de áreas de risco do Espírito Santo (sendo 17 Rhipicephalus microplus e 30 Rhipicephalus sanguineus), e amostras de sangue total de 27 bovinos e 26 equinos, animais estes oriundos da Universidade Federal Rural do Rio de Janeiro. Os resultados mostraram que a SBY não se trata de uma zoonose causada por um conjunto de micro-organismos como pensado inicialmente e sim pela Borrelia burgdorferi sensu lato. Descoberta essa que foi possível empregando-se novos primers amplificadores do principal gene envolvido na síntese do gancho flagelar da Borrelia, chamado flgE. A positividade para flgE foi confirmada em 6 pacientes do grupo B, 2 carrapatos, 1 bovino e 1 equino, os quais apresentaram homologia de 99% com o gene da proteína do gancho flagelar da Borrelia burgdorferi (flgE) depositado no GenBank (L43849). Esta importante descoberta, associada às pesquisas anteriores, permitiu definir a SBY como zoonose emergente e própria do país, causada pela bactéria B. burgdorferi na apresentação morfológica atípica, transmitida por carrapatos não pertencentes ao complexo Ixodes ricinus, responsável por manifestações clínicas semelhantes à Doença de Lyme, exceto pela grande frequência de sintomas recorrentes / Brazilian Lyme disease-like illness (BLDL) or Baggio-Yoshinari Syndrome (BYS) is an emerging zoonosis, transmitted by ticks and so far, restricted to the description of the Brazilian territory. The causative agent of BYS was unknown until now. The main objective of this study was to identify the etiology of BYS. We have selected two groups of patients: group A (n = 68) consisting of patients suspected of BYS, mostly in the latent stage of disease; group B (n = 10), composed of patients diagnosed with BYS, who had compulsorily erythema migrans and that were symptomatic at the time of blood collection. We also used a control group composed of healthy individuals with negative epidemiology (n = 50). Blood samples were collected, in which we performed serology, cultures, microscopic analysis (optical and electron) and polymerase chain reaction (PCR) for different microorganisms (Mycoplasma spp, Chlamydia spp and Borrelia spp). In addition, a preliminary study was conducted by PCR for Borrelia spp in 47 samples of ticks from risk areas at Espirito Santo State (being 17 Rhipicephalus microplus and 30 Rhipicephalus sanguineus), 27 cattle and 26 horses, being these animals from the Universidade Federal Rural do Rio de Janeiro. The results showed that BYS is not a zoonosis caused by a set of microorganisms as initially thought, but by Borrelia burgdorferi sensu lato. These findings were possible after employing new primers that are able to amplify portions of the main genes involved in the synthesis of the Borrelia flagellar hook protein, called flgE. We confirmed positivity for the flgE in 6 patients from group B, 2 ticks, a cow, and a horse, which showed 99% homology with the gene of Borrelia burgdorferi flagellar hook protein (flgE) deposited in GenBank (L43849). This important discovery, coupled with previous research, helped to define BYS as an emerging zoonosis particular from Brazil, caused by B. burgdorferi of atypical morphologic presentation, transmitted by ticks outside the Ixodes ricinus complex, responsible for clinical signs similar to Lyme disease, except for the high frequency of relapsing symptoms

Les acyl stéryl glycosides (ASGs) appartiennent à une famille de glycolipides qui possèdent un caractère amphiphile particulier dû à la présence de deux parties hydrophobes, un stéroïde et une chaine grasse. Dans le cadre de nos études des propriétés d’auto-organisation des glycoamphiphiles, ce travail est dédié à l’étude de deux ASGs, α-CAG et BbGL 1, composés naturels présents respectivement dans les membranes des bactéries Helicobacter pylori et Borrelia burgdorferi., présentant des structures similaires mais des activités biologiques différentes. Notre travail a consisté à déterminer les paramètres structuraux qui gouvernent leurs propriétés d’auto-assemblage. Deux séries de 6-O-acyl cholestéryl glycosides (glucosides et galactosides) variant dans leur configuration anomérique et la longueur et le niveau d’insaturation de leur chaine grasse ont été synthétisées et leur capacité à former des cristaux liquides et à promouvoir une ségrégation lipidique dans des monocouches de Langmuir modèles de membrane ont été étudiées. Les relations structure-propriétés établies montrent que la longueur de la chaine grasse est le paramètre le plus discriminant dans le comportement d’auto-assemblage dans les deux types d’expériences. Pour les cristaux liquides thermotropes, l’autre facteur discriminant est la configuration anomérique, deux phases colonnaires successives rectangulaires puis hexagonales étant observées pour les séries α alors qu’une seule a été observée en séries β Changer de sucre n’induit pas de différence significative dans le comportement LC. Concernant la formation de domaines lipidiques, les modifications de la configuration (α/β) et du sucre influencent significativement leur temps d’apparition, apportant pour la première fois une définition claire des paramètres structuraux et physicochimiques qui gouvernent le comportement de l’α-CAG et ses analogues, en lien avec les données commues sur l’augmentation de pathogénicité d’Helicobacter pylori. Ce travail de thèse donne une illustration de l’importance de la structure des carbohydrates dans les processus biologiques et du concept de glycoamphiphilie. / Acyl steryl glycosides (ASGs) are a family of glycolipids which exhibit a peculiar amphiphilic character based on the presence of two hydrophobic appendages, one steroid moiety and one fatty alkyl chain, attached on a polar carbohydrate backbone. In the frame of our studies on the self-organisation properties of carbohydrate-based amphiphiles, this thesis is an investigation of the behavior of ASGs, in particular α-CAG and BbGL 1, two natural compounds found in bacterial membranes, Helicobacter pylori, Borrelia burgdorferi repectively, who exhibit close structures but different bioactivity. Our work has aimed at determining the key structural parameters governing their self-organization behavior. Two series of acyl cholesteryl glycosides (glucosides or galactosides) have been synthesized, with variations in the anomeric configuration, the 6-O-acyl chain length and level of unsaturation, and investigated with respect to their ability to form liquid crystalline mesophases, and to drive lipid domain segregation in Langmuir monolayers as model membranes. Structure-properties relationships have been established, indicating that the fatty chain length showed the most remarkable influence on the self-organization behavior, in LC and model membrane experiments. For the LC mesophases, the other important parameter is the anomeric configuration, two successive columnar phases, rectangular then hexagonal, being observed for the α-anomers, whereas only one was found for the β-anomers. No significant changes were observed when comparing glucosides and galactosides. With respect the formation of domains, configuration modifications at both C-1 (α or β) and C-4 (gluco or galacto) influenced significantly the domains appearance time, giving the first, clear physicochemical proof of the structural influential factors in the behavior of α-CAG and analogues, in the context of the known increased pathogenicity of Helicobacter pylori. Overall, this thesis provides a nice illustration of the subtlety and the importance of carbohydrate structure in biological processes, and of the concept of glycoamphiphilicity.

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Indiana University-Purdue University Indianapolis (IUPUI) / Borrelia burgdorferi, a bacterium which causes Lyme disease, is maintained in nature through a cycle involving two distinct hosts: a tick vector and a mammalian host. To adapt to these two diverse environments, B. burgdorferi undergoes dramatic alterations in its surface lipoprotein. Two essential lipoproteins, outer surface protein A (OspA) and outer surface protein C (OspC), are reciprocally regulated throughout the B. burgdorferi lifecycle. Very little is known about the regulation of OspA. These studies elucidate the regulatory mechanisms controlling the expression of OspA. Various truncations of the ospA promoter were created and then studied in our novel in vitro model of ospA repression or grown within the host-adapted model. A T-Rich region of the ospA promoter was determined to be a cis-element essential for both the full expression and full repression of ospA.