Avaliação soro-epidemiológica e molecular de cães assintomáticos para leishmaniose tegumentar americana em área endêmica

• Autor(a) principal: Passos, Gabriela Porfirio
• Data de Publicação: 2013
• Tipo de documento: Dissertação
• Idioma: por
• Título da fonte: Repositório Institucional da Universidade Federal do Espírito Santo (riUfes)
• Texto Completo: http://repositorio.ufes.br/handle/10/7725
• topic: Extração de DNA; ELISA; Western Blot; PCR; Leishmania brazil; Ciências Agrárias; Cão; Leishmaniose; Diagnóstico; Teste imunoenzimático

The aim to make the diagnosis of American tegumentary leishmaniasis (ATL) in dogs, it was used isolation and culture methods, ELISA and Western Blot (WB) serological tests and parasite DNA research for two animals groups, one group composed of animals without clinical suggestive lesions of ATL, but residents around confirmed human clinical cases of ATL, and another group of animals with suggestive lesions of ATL which served as control of the performed protocols. The study was conducted in the municipality of Iúna, ES, Brazil, an endemic disease region. In the first group, were analyzed serum samples from 109 animals with no history or indicative of ATL injuries, they were subjected to ELISA and WB techniques, which resulted in 20 animals serologically positive for both techniques. The ELISA test presented sensitivity of 100.00% (CI95% - 0.83 to 1.00) and specificity of 77.53% (CI95%- 0.67 to 0.86), compared tothe WB technique. The WB test was more accurate, and proved to be more suitable for diagnosis of asymptomatic animals, while the ELISA technique for screening. For parasite DNA research in the 20 asymptomatic animals and positive by WB technique it was used whole blood and intacttissue biopsy of pinna by the technique of polymerase chain reaction (PCR). The results of intact tissue biopsy PCR and tissue blood PCR showed a sensitivity of 30.0% (CI95%- 0.12 to 0.54) and 20.0% (CI95%- 0.06 to 0.44), respectively. The specificity, 99.0% (CI95%- 0.93 to 0.99) and 100% (CI95%- 0.96 to 1.00), for intact tissue biopsy PCR and tissue blood PCR when compared the WB test. The results show that the pinna intact tissue and blood of asymptomatic animals subjected to PCR, showed a low sensitivity and high specificity, thereby intact tissue biopsy PCR is a better indicator for asymptomatic animals compared to the blood tissue PCR. For the second group, three animals were identified with ATL suggestive lesions and serologically positive. From a sample of an ATL suggestive lesion biopsy present in pinna, part of this was destined to culture and isolation and another portion was destined for comparison of three protocols for DNA extraction from animal tissue for canine ATL diagnosis. The used protocols were based on Phenol-Chloroform, Potassium Acetate and an association of the two methodologies. Compared to molecular standards of DNA concentration, it was concluded that the protocol with potassium acetate was the most suitable for the used tissue. Finally, in possession of data presented for the studied asymptomatic and seropositive animals, it was concluded that these did not represent potential parasitereservoirs, which also shows that for evaluation of this disease profile must be given the combination of diagnosis methods