PCR multiplex para detecção de patógenos de Apis mellifera L. (Hymenoptera, Apidae) em mel

• Ano de defesa: 2011
• Autor(a) principal: Puker, Anderson
• Orientador(a): Message, Dejair
• Banca de defesa: Moreira, Maria Aparecida Scatamburlo
• Tipo de documento: Dissertação
• Tipo de acesso: Acesso aberto
• Idioma: por
• Instituição de defesa: Universidade Federal de Viçosa
• Programa de Pós-Graduação: Mestrado em Entomologia
• Departamento: Ciência entomológica; Tecnologia entomológica
• País: BR
• Palavras-chave em Português: Multiplex-PCR; Sanidade apícola; Apis mellifera; Abelhas africanizadas
• Palavras-chave em Inglês: Multiplex-PCR; Health beekeeping; Apis mellifera; Africanized bees
• Área do conhecimento CNPq: CNPQ::CIENCIAS BIOLOGICAS::PARASITOLOGIA::ENTOMOLOGIA E MALACOLOGIA DE PARASITOS E VETORES
• Link de acesso: http://locus.ufv.br/handle/123456789/3954

Recently, a decline of pollinators, especially the bees Apis mellifera L. (Hymenoptera: Apidae), has affected beekeeping and consequently agricultural harvests in some countries, although the damage to ecosystems has not been effectively accounted. Among the causes listed for this unexplained phenomenon, pathogens are among the possible factors responsible. There are several pathogens that attack the bees A. mellifera around the world, as bacteria Paenibacillus larvae (White) and the fungi Ascosphaera apis (Maassen ex Claussen) Olive & Spiltoir, Nosema apis Zander and Nosema ceranae Fries et al. Their distributions in some parts of the world, such as Brazil, are not exactly known, not only because of the difficulty of collecting samples in such a large country, and also because of the time and cost of diagnoses involved. At this point, it is important to standardize techniques for rapid diagnosis and to facilitate the safe conduct of epidemiological surveys, and therefore assist in controlling the spread of these microorganisms. The aim of this work was standardize a multiplex PCR for simultaneous detection of the spores of A. apis, N. ceranae and P. larvae present in honey and use it in the analysis of honey samples from some regions. The multiplex PCR was standardized using specific primers and DNA was extracted from honey samples positive for each of the pathogens. The recommendations of existing national legislation were used for the preparation of the solutions of honey to be submitted to the technique developed. The standard technique in this study was effective for diagnosing three pathogens to A. mellifera in honey, A. apis, N. ceranae and P. larvae. The detection threshold of monospecific PCR was of 10 CFU/mL of honey, and of 10 and 100 spores/mL of honey for A. apis and N. ceranae, respectively. The detection sensitivity of multiplex PCR was of 10 CFU/mL of honey for P. larvae, and of 100 spores/mL of honey for A. apis and for N. ceranae. Did not match any of those pathogens in 120 honey samples analyzed with standardized multiplex PCR. Thus this method was suitable for simultaneous detection of pathogens to A. mellifera in honey, but can probably be used in other bee products with minor modifications.