Utilização da PCR em tempo real na detecção e quantificação de PCV2 em diferentes órgãos de suínos clinicamente sadios e apresentando PMWS

• Ano de defesa: 2008
• Autor(a) principal: Silva, Fernanda Miquelitto Figueira da
• Orientador(a): Lamêgo, Márcia Rogéria de Almeida
• Banca de defesa: Viloria, Marlene Isabel Vargas , Fietto, Luciano Gomes , Zerbini Júnior, Francisco Murilo
• Tipo de documento: Dissertação
• Tipo de acesso: Acesso aberto
• Idioma: por
• Instituição de defesa: Universidade Federal de Viçosa
• Programa de Pós-Graduação: Mestrado em Bioquímica Agrícola
• Departamento: Bioquímica e Biologia molecular de plantas; Bioquímica e Biologia molecular animal
• País: BR
• Palavras-chave em Português: PCR em tempo real; Circovírus suíno 2; PMWS
• Palavras-chave em Inglês: Real time PCR; Swine circovirus 2; PMWS
• Área do conhecimento CNPq: CNPQ::CIENCIAS BIOLOGICAS::BIOQUIMICA
• Link de acesso: http://locus.ufv.br/handle/123456789/2398

Porcine circovirus 2 (PCV2) is a non-enveloped virus containing a single stranded circular DNA of approximately 1.76 Kb. PCV2 is the primary etiologic agent of porcine circovirus-associated disease (PCVAD) responsible for large economic losses in the swine industry. The objectives of this work were to use molecular methods to determine the most suitable organs for quantification of PCV2 viral load and compare the genotype and viral load in organs with different levels of tissue injury. A plasmid containing the complete viral genome was quantified by spectrophotometry and used to create a standard curve for PCV2 quantification. DNA was extracted using the Wizard SV Genomic Purification System (Promega). Reactions of nested PCR were performed using two pairs of primers and analyzed in agarose gel. Real-time PCR was carried out using TaqMan. Tissue samples were fixed in 10% formalin, embedded in paraffin blocks and processed for histopathological investigation. Comparison of sensitivity of real time PCR with nested PCR showed 91% of positive samples by real time PCR, whereas nested PCR detected only 66.5% of positive samples. Results showed that real time PCR is more effective to detect PCV2 in tissue samples than nested PCR. In the tested swine samples, lymph nodes showed mean viral load significantly higher than other types of organs (P <0.01) and thus it is recommended for PCV2 diagnosis and pathogenesis studies. Comparisons among viral load and the four injury levels did not show any significant results, it is believed, therefore, that the different levels of lymphoid lesions may be associated with other infectious or non-infectious co-factors.