Made available in DSpace on 2014-06-11T19:23:47Z (GMT). No. of bitstreams: 0 Previous issue date: 2003-02-21Bitstream added on 2014-06-13T19:50:56Z : No. of bitstreams: 1 martins_cb_me_jabo.pdf: 234153 bytes, checksum: a0a5965509f8505fb5b72f7bfe8b4650 (MD5) / 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... . / 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.

No description available.

Nitrile biocatalysts are of use in the chemical and pharmaceutical industries for the synthesis of carboxyamides and carboxylic acids. In particular, the application of biocatalysts in the synthesis of single enantiomer compounds is of increasing interest, but requires novel substrate specific highly stereoselective biocatalysts. Addition to the limited toolbox of known nitrile biocatalysts requires definitive characterisation of the biocatalysts through accurate determination of the substrate profiles and quantification of activity. The accurate quantification of stereoisomers chiral mixtures to determine biocatalyst stereoselectivity remains a significant challenge due to the difficulty in separating stereoisomers by physical methods. The known nitrile metabolising organism, Rhodococcus rhodochrous ATCC BAA-870, was grown in a defined medium and harvested, providing whole cell biocatalyst. Additional biomass was disrupted to provide a cell free enzyme extract, which was put through an enzyme purification protocol to provide a solution with specific activity of 351 U.mg⁻¹. A portion of the enzyme was self immobilised using the SphereZyme™ technique. The nitrile hydratase SphereZymes™ (1.2 U.mg⁻¹ initial activity) that were prepared had pH and temperature optima of 6 and 30°C respectively, and could be recovered by repeated washing. The particles retained activity in the presence of the organic solvents isooctane and n-hexadecane saturated with 50 mM phosphate buffer (pH 7.5). An initial analytical system was devised for quantification of the nitrile hydratase activity using the non-chiral substrate benzonitrile. An improved reversed phase high performance liquid chromatography method was developed to separate and quantify benzamide, benzoic acid and benzonitrile. The mobile phase consisting of 0.1% trifluoroacetic acid in H₂O and acetonitrile (70:30, %v/v), at a flow rate of 0.5 ml.ml⁻¹, 25°C, resolved all three analytes in 3.5 minutes on a Waters X-Terra MS C18 3.5μm column. UV detection was carried out at 210 nm. Analytical methods to determine activity and enantioselectivity of the whole cell biocatalyst were subsequently developed for both β-amino nitriles and β-hydroxy nitrile substrates and hydrolysis products.

Nitrilases are enzymes which catalyze the hydrolysis of nitriles to corresponding carboxylic acids. These enzymes have a great potential in biocatalysis, for example in the synthesis of mandelic acid and mandelamide, because of their chemo- and enantioselectivity. As bioremediation agents they are also applicable to sites contaminated with organic nitriles. In this work, activities of recombinant strains of E. coli expressing hypothetical nitrilases from fungi Giberella moniliformis and Nectria haematococca mpVI 77-13-4 were studied, as well as the biodegradation potential of bacteria from Rhodococcus and Nocardia genera towards benzonitrile herbicides dichlobenil (2,6-dichlorobenzonitrile), ioxynil (3,5-diiodo-4- hydroxybenzonitrile) and bromoxynil (3,5-dibromo-4-hydroxybenzonitrile). The hypothetical fungal nitrilases were expressed as functional enzymes. Nitrilase from G. moniliformis showed highest activity towards benzonitrile (30.9 U/mg protein), total activity yield was 2,560 U/l cell culture. The preferred substrate of the nitrilase from N. haematococca was phenylacetonitrile (12.3 U/mg prot.), total activity yield was 28,050 U/l cell culture. Nitrilase from N. haematococca was also able to hydrolyze mandelonitrile (5.9 U/mg prot.). Soil bacteria Rhodococcus rhodochrous PA-34, Nocardia globerula...

Many important antibiotics have become nearly obsolete due to the rise of antibiotic resistant pathogens. Rhodococcus, an actinomycete related to the prolific antibiotic producing genus Streptomyces, harbors over 30 genes for secondary metabolism that could be involved in antibiotic production. Several antibiotics have already been reported for Rhodococcus, suggesting the genus may be a good source for new inhibitory compounds. Fifty four soil bacteria were isolated using enrichment culture techniques (including 37 Rhodococcus) and screened for antibiotic producers. BTHX2, a species of Bacillus was found to have activity against Micrococcus luteus and Rhodococcus erythropolis. BTHX2 has a 16S rDNA sequence 97% homologous to Bacillus licheniformis, and may be a new strain of B. licheniformis. The inhibitory substance produced by BTHX2 was and found to have a spectrum of activity against a broad Gram-positive bacteria and some fungus, and may have cytolytic activity.

Iron is an essential nutrient for most bacteria because enzymes like nitrate reductase and cytochromes use it as a cofactor. However, in most aerobic, neutral pH environments, iron is essentially insoluble and not easily available for bacteria to use. Many bacteria respond to this problem by releasing small organic compounds called siderophores that bind and effectively solubilize iron so that it can be transported into the cell for growth. The focus of this study was to learn more about the iron acquisition and especially the transport of iron by the soil bacterium Rhodococcus erythropolis. To fulfill this aim, mutant strains of the bacteria were screened for those that overproduce siderophore. Often, a bacterium will over produce siderophore to compensate for a defect in transport. One such mutant, R187-12, was further analyzed by cloning the region of the chromosome containing the defective gene responsible for over production of siderophore into a plasmid vector. The DNA sequence of this region was determined and analyzed for the presence of similar genes encoding transport proteins.

Many drugs used in the treatment of various cancers are derived from or influenced by compounds from nature. The soil bacterium Rhodococcus is of interest because of its identified secondary metabolic pathways and the production of novel natural antibiotics from several strains. In this study, a solid agar extraction method was used to collect compounds from strains of Rhodococcus. These bacterial compound extracts were then tested using a MTT assay in order to evaluate their effectiveness in augmenting MCF7 breast cancer cell death. The results of two way ANOVA analyses revealed 18 compound extracts from 15 strains of Rhodococcus that showed significant p-values when assayed with MCF7 breast cancer cells but nonsignificant interaction p-values when assayed with the healthy cell control. These results prompt further identification of specific compounds present in the bacterial extract that caused cell death as well as a mechanism of interaction with the breast cancer cells.

The objective of this study was to purify IgG from commercially available hyperimmune Rhodococcus equi plasma and to assess the delivery of IgG as an aerosol to the equine lung. IgG was purified from plasma, and the IgG concentration of both the plasma and the purified IgG was determined by ELISA. The purified IgG was aerosolized using a vibrating mesh nebulizer and aerosol characterization was performed using cascade impaction. The purified IgG was nebulized to six healthy adult horses in order to assess the efficacy of pulmonary delivery and safety of administration. Bronchoalveolar fluid was retrieved endoscopically using a low volume technique prior to aerosolization (time 0) and at 0.5, 4 and 24 hours post aerosolization. The BAL fluid IgG concentration was determined and cytologic analysis was performed. The IgG concentrations of the plasma and purified IgG were 2,175 mg/dL and 1,145 mg/dL, respectively. The MMAD of the purified IgG aerosol was 4.7 microns. The mean BAL fluid IgG concentration increased 61% from 19.33 µg/dL at time 0 to 31.5 µg/dL at 0.5 hours, but this increase was not significant (P=0.603). No significant change was observed in inflammatory cell numbers over time or at any time point during the study. This study demonstrated that IgG antibodies were purified at a concentration acceptable for nebulization, and that the nebulization unit generated aerosol particles from the IgG solution of appropriate size for pulmonary delivery. Nebulization of purified IgG to adult horses was well tolerated and caused no local or systemic adverse effects. / Master of Science

O Rhodococcus equi é uma das principais causas de pneumonia piogranulomatosa e linfadenite em potros de 1-6 meses de idade, resultando em alta morbidade e muitas vezes em óbito. Em propriedades endêmicas, a enfermidade é responsável por gerar altos custos para tratar e prevenir a doença. Com o objetivo de propor uma alternativa para o controle ambiental do Rhodococcus equi, foi aplicado um protocolo para isolamento de bacteriófagos. Foram isoladas 14 amostras de bacteriófagos, obtidas do ambiente de criação de cavalos. Estas apresentaram capacidade de lisar in vitro amostras de campo de Rhodococcus equi com diferentes intensidades. O resultado obtido nos possibilita sugerir que os fagos podem ser uma alternativa natural para a redução da bactéria no ambiente. / Rhodococcus equi is an important cause of pyogranulomatus pneumonia and lymphadenitis in foals 1-6 months of age, resulting in high morbidity and often mortality. In endemic properties, the disease is responsible for generating high costs to treat and prevent disease. In order to propose an alternative to the environmental control of Rhodococcus equi, a protocol for isolation of bacteriophages was applied. Fourteen bacteriophage samples were isolated obtained from horse breeding environment. They showed capacity to lyse in vitro Rhodococcus equi field samples with different sensitivities. The results obtained allows us to suggest the phage as a natural alternative to reducing R. equi population.

Rhodococcus equi (R.equi), uma bactéria gram-positiva intracelular facultativa, é uma causa importante de pneumonia em potros com idade entre 3 semanas e 5 meses. A manifestação clinica mais comum da doença é a broncopneumonia piogranulomatosa com abscedação. Na pneumonia causada por R. equi os primeiros sinais clínicos podem não ser aparentes até que as alterações patológicas estejam bastante avançadas, por esse motivo, o diagnóstico precoce e acurado de potros com pneumonia por R. equi se torna fundamental. O diagnóstico definitivo baseia-se na detecção de R. equi na cultura bacteriológica e identificação molecular a partir da amostra de lavado traqueal; no entanto, essa técnica é invasiva, traz riscos para o animal e é relativamente cara. A ultrassonografia (US) para detecção precoce tem se tornado uma pratica de rotina em muitas fazendas endêmicas para rodococose equina. Com o advento dessa prática de triagem, a forma mais identificada de pneumonia por R. equi tem sido a subclínica, onde os animais apresentam presença de alterações pulmonares mas não apresentam sinais clínicos da doença. Atualmente, vapA é o gene com função demonstrada na virulência. Identificação de R. equi por Reação em Cadeia da Polimerase (PCR) em amostras de fezes tem se mostrado efetivo para o reconhecimento precoce do agente. Ultrassonografia torácica e PCR das amostras de fezes e swab nasal foram realizadas em 22 potros desde as 3 até as 16 semanas de idade (intervalos de 15 dias) de 3 fazendas endêmicas de criação de cavalos no sul do Brasil para identificar a ocorrência de doença subclínica. A associação entre a ultrassonografia torácica e PCR das amostras de fezes possibilitaram a detecção de doença subclínica e identificação de pontos críticos de controle dessa doença. Considerando o fato de que 95.4% dos potros apresentaram doença subclínica e que nenhum deles desenvolveu a doença clínica demonstra que o tratamento desses casos não é justificável para a população analisada. / Rhodococcus equi, a gram-positive facultative intracellular pathogen, is an important cause of pneumonia in foals between 3 weeks and 5 months of age. Pneumonia caused by R. equi is an insidious disease in which clinical signs may not be apparent until pathologic changes are well progressed. Because of the insidious progression of infection to severe clinical signs, early and accurate diagnosis of foals with R. equi pneumonia is important. Definitive diagnosis is based on R. equi detection by bacterial culture and molecular identification from tracheobronchial aspirate (TBA), this procedure is invasive, labor-intensive, requires skill, carries risks to foals, and is relatively expensive. The sequential thoracic ultrasonography (TUS) to early detection of the disease has been adopted as a screening method in many endemic farms; for this reason, subclinical disease has been the most frequently observed form. Nowadays, vapA is the only virulent gene identified. Fecal polymerase chain reaction (PCR) is a noninvasive technique with good diagnostic accuracy. Thoracic ultrasound screening (TUS) and PCR from fecal and nasal swab samples were performed in 22 foals from 3 to 16 weeks of age from 3 endemic farms at south of Brazil to identify the occurrence of R. equi subclinical disease. The association of TUS and fecal PCR detection of virulent R. equi provided a possibility of identification of critical points in disease control. Considering the fact that 95.4% of the foals showed evidence of subclinical disease and none of them developed any signs of clinical disease, the antibiotic treatment was not reasonable for the foals followed.