Orientador: José Correa de Lacerda Neto / Banca: Rosangela Zacarias Machado / Banca: Wilson Roberto Fernandes / Resumo: A transferência de imunidade passiva, através da ingestão do colostro é um fator de grande importância para a sobrevivência de potros neonatos. O colostro é rico em imunoglobulinas, que constitui para esses animais a única fonte de proteção contra os agentes infecciosos durante o período neonatal, fase em que os animais apresentam maior susceptibilidade às doenças. Dentre estas, destaca-se a infecção ocasionada pelo Rhodococcus equi, responsável por elevadas taxas de mortalidade e grandes perdas econômicas. A infecção possui distribuição mundial e causa pneumonia supurativa e enterite associada a linfadenite em potros com menos de seis meses de idade. Apesar da sua grande importância no Brasil, a rodococose ainda é pouco estudada em nossas condições. O objetivo deste estudo foi comparar a resposta imune humoral entre éguas das raças Brasileiro de Hipismo (BH) e Bretã previamente imunizadas para R. equi, e avaliar o efeito da imunoprofilaxia ativa materna na transferência de anticorpos específicos protetores, via colostro, para neonatos eqüinos, utilizando na detecção de anticorpos o ensaio imunoenzimático indireto (ELISA teste), comparando dois diferentes antígenos (APTX e comercial). Os resultados demonstraram que ocorreu aumento na titulação de anticorpos anti-Rhodococcus equi nos grupos de éguas após a vacinação e transferência de anticorpos através do colostro para os potros neonatos. Não houve efeito da raça na produção de anticorpos. O antígeno comercial detectou títulos de anticorpos maiores e mais persistentes do que o antígeno APTX... (Resumo completo, clicar acesso eletrônico abaixo). / Abstract: The passive immunity transfer through colostrum is very important to new-born foals survival. The colostrum is immunoglobulin full, wich is responsable to protect against infectious agents during new-born period, stage that animals present higher susceptibility to diseases. Among these, the infection caused by Rhodococcus equi has the great importance, responsable for large economics damage and high mortality rates in animals illness. The infection has a world-wide distribuition and it causes suppurative pneumonia and enterits associated to limphadenitis in foals under 6 months old. Although its great importance in Brazil, this disease is not well studied. The aims of this study was to compare the humoral immune respost between the breeds "Brasileiro de hipismo" (BH) and Breton, previously vaccinated against Rhodococcus equi, and to evaluate the matern immunoprofilaxy effect in the specific antibodies transfer through colostrum to new-born foals, using the Enzyme linked immunosorbent assay (ELISA-test) comparing two differents antigens ("APTX" and comercial"). The results demonstrated that there were an increase in mares antibodies after vaccination and also antibodies transfer through colostrum to new-born foals. There was no breed effect in the antibodies production. The Comercial antigen demonstrated antibodies titulation higher and more persistents than APTX antigen. The ELISA-test demonstrated sensible R. equi antibodies determination. / Mestre

Made available in DSpace on 2014-06-11T19:29:31Z (GMT). No. of bitstreams: 0 Previous issue date: 2009-06-25Bitstream added on 2014-06-13T19:17:49Z : No. of bitstreams: 1 guazzelli_a_me_botfmvz.pdf: 415224 bytes, checksum: 21f876feebee52f17c73bdef5ce2ef0c (MD5) / Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) / A rodococose suína compreende doença infecciosa caracterizada por linfadenites piogranulomatosas. Diferentes fatores de virulência são reconhecidos na patogenicidade de Rhodococcus equi. A estrutura da parede celular bacteriana, a viabilidade no interior de fagócitos e na ausência de ferro, a produção de citotoxinas, a resistência aos antimicrobianos convencionais e, recentemente, a presença de proteínas associadas à virulência (Vap) reguladas por plasmídios, são considerados os principais mecanismos de virulência do microrganismo. Diferentes fatores de virulência foram avaliados em 23 (6,1%) linhagens de R. equi isoladas de 378 linfonodos submandibulares e mesentéricos de suínos e javalis (Sus scrofa). Foram realizados exames microbiológicos em 129 linfonodos apresentando lesões (linfadenite) e 129 sem lesões (controle) de suínos, e 60 linfonodos com lesões e 60 sem lesões de javalis. Dentre as 23 linhagens de R. equi, 19 (7,4%) foram isoladas de suínos, das quais 17 obtidas de linfonodos com lesões e duas sem lesões. Das 19 linhagens de suínos, 18 (94,7%) foram obtidos de linfonodos submandibulares e um (5,3%) de mesentérico. As quatro (3.3%) linhagens de R. equi isoladas de javalis foram obtidas exclusivamente de linfonodos com lesões. Destes, três foram obtidos de linfonodos submandibulares e um de mesentérico. Dentre nove antimicrobianos testados, azitromicina (100,0%), gentamicina (100,0%), levofloxacina (100,0%), vancomicina (100,0%), amoxicillina/ácido clavulânico (94,7%), eritromicina (94,7%) e rifampicina (94,7%) foram os fármacos mais efetivos. Baixa ocorrência de resistência aos antimicrobianos nos isolados de suínos foi observada contra os fármacos testados. A concentração inibitória mínima (MIC90) da azitromicina, eritromicina e rifampicina foi observada, respectivamente, em ≤2 µg/mL, ≤0,5 µg/mL and ≤1 µg/mL... / The rhodococcosis in swine comprise an infectious disease characterized by pyogranulomatous lymphadenitis. Different virulence factors are recognized in pathogenicity of the Rhodococcus equi. The structure of bacterial cell wall, the viability inside of phagocytes and in absence of iron, the production of cytotoxins, the resistance to conventional antimicrobials and recently, the presence of proteins associated to virulence (Vap) regulated by plasmids, are considered the most important virulence mechanisms of microorganism. Different virulence factors were evaluated in 23 (6.1%) R. equi strains isolated from 378 submandibular and mesenteric lymph nodes of swine and wild boars (Sus scrofa). Microbiological exams were performed in 129 lymph nodes presenting lesions (lymphadenitis) and 129 without lesions (controls) from swine, and 60 lymph nodes with lesions and 60 without lesions from wild boars. Among 23 R. equi strains, 19 (7.4%) were isolated from swine and, from these, 17 were obtained from lymph nodes with lesions and two without lesions. From 19 strains isolated from swine, 18 (94.7%) were obtained of submandibular lymph nodes and one (5.3%) from mesenteric. The four (3.3%) R. equi strains isolated from wild boars were obtained exclusively of lymph nodes presenting lesions. From these, three were obtained from submandibular lymph nodes and one of mesenteric. Among nine antimicrobials tested, azithromycin (100.0%), gentamicin (100.0%), levofloxacin (100.0%), vancomycin (100.0%), amoxicillin/clavulanic acid (94.7%), erythromycin (94.7%) and rifampin (94.7%) were the most–effective drugs. Low rates of resistance to antimicrobials in swine isolates were observed against drugs tested. The inhibitory minimal concentration of 90% of isolates (MIC90) with use of azithromycin, erythromycin and rifampin were observed respectively in 2 μg/mL, 0.5 μg/mL and 1 μg /mL... (Complete abstract click electronic access below)

Rhodococcus equi is an important pathogen in foals between one to six months of age and is a major cause of death in in these animals. In addition, R. equi has recently emerged as a significant opportunistic pathogen in immunosuppressed humans, especially those infected with HIV. Despite the ability of the organism to survive stressful growth conditions, for example, exposure to elevated temperature and oxygen radicals, the role of heat shock proteins in the pathogenesis of R. equi has not been well documented. In this project we developed and optimised methods to purify the heat shock protein DnaK from R. equi, using a combination of ion-exchange and affinity chromatography. The effectiveness of the purification protocols were assessed using SDS-PAGE and Western-blotting with anti-DnaK antibodies, and the enzymic activity of the purified DnaK was verified with an ATPase assay. ATPase assays were also used to investigate the roles of other heat shock proteins in enhancing the activity of DnaK.

Although Rhodococcus equi (R. equi), previously known as Corynebacterium equi, was first isolated from pneumonic foals almost a century ago, it remains the most common cause of subacute or chronic granulomatous bronchopneumonia in foals. While the majority of foals exposed to R. equi develop a protective immune response (regressors), others exhibit a unique susceptibility to infection (progressors). The determinants for either outcome are not completely understood. Therefore, current diagnostic and preventive measures are suboptimal and require betterment. In light of this current need, we hypothesized that immunoglobulin G subisotype T [IgG(T)] against the virulence-associated protein A (VapA) of R. equi, and whole blood cytokine expression profile of foals predict the outcome of infection and can be used as diagnostic markers of clinical disease. Further, we hypothesized that the use of R. equi hyperimmune plasma (HIP) decreases severity of disease in naturally infected foals, playing an important role in disease prevention in the field. Lastly, we hypothesized that specific anti-Rhodococcus equi pili antibodies passively acquired by foals via colostrum after immunization of pregnant mares with a Rhodococcus equi pili-based candidate vaccine will confer protection against induced disease, and therefore have an immediate impact on R. equi pneumonia prophylaxis. The objectives of this study were: (1) to describe the humoral immune response of progressor and regressor foals to R. equi following experimental challenge and natural infection, (2) to compare the cytokine and cell-marker expression profile in whole blood of progressor and regressor foals after challenge, (3) to evaluate the Vap-A specific IgG profile of a commercially available HIP product and its value as a prophylactic tool on an endemic farm, and (4) to evaluate the efficacy of a vaccine based on the Rhodococcus equi pili (Rpl). Although the IgG(T) response of progressor foals after challenge or following natural infection tended to be more pronounced than that observed in regressor foals, its performance as a diagnostic test for predicting disease outcome was poor. Likewise, whole blood cell-marker and cytokine expression profiles of progressor and regressor foals were not significantly different, undermining its reliability as a diagnostic tool. Evaluation of the association of HIP VapA specific IgG profile and rhodococcal disease outcome in the field resulted in the conclusion that progressor foals received significantly less VapA specific IgG, suggesting that HIP may have provided some protection to regressor foals. Although HIP appeared to have provided some protection against clinical pneumonia, Rpl maternally-derived IgG failed to confer any advantage to foals born from vaccinated mares. The Rpl candidate vaccine failed to confer protection to foals after challenge, and did not decrease disease severity in comparison to a control group. In summary, the results of this study do not support the use of VapA specific IgG(T) or whole blood cytokine expression profile as predictors of disease outcome. Further, our results suggest a positive effect of HIP on disease outcome. Lastly, the presence of systemic and local Rpl antibodies was not protective in foals.