Nested-PCR for the detection of Mycoplasma hyopneumoniae in bronchial alveolar swabs, frozen tissues and formalin-fixed paraffin-embedded swine lung samples: comparative evaluation with immunohistochemical findings and histological features

Abstract

Pesq. Vet. Bras. 32(8) The diagnosis of Mycoplasma hyopneumoniae infection is often performed through histopathology, immunohistochemistry (IHC) and polymerase chain reaction (PCR) or a combination of these techniques. PCR can be performed on samples using several conservation methods, including swabs, frozen tissue or formalin-ϐixed and parafϐin-embedded (FFPE) tissue. However, the formalin ϐixation process often inhibits DNA ampliϐication. To evaluate whether M. hyopneumoniae DNA could be recovered from FFPE tissues, 15 lungs with cranioventral consolidation lesions were collected in a slaughterhouse from swine bred in herds with respiratory disease. Bronchial swabs and fresh lung tissue were collected, and a fragment of the corresponding lung section was placed in neutral buffered formalin for 48 hours. A PCR assay was performed to compare FFPE tissue samples with samples that were only refrigerated (bronchial swabs) or frozen (tissue pieces). M. hyopneumoniae was detected by PCR in all 15 samples of the swab and frozen tissue, while it was detected in only 11 of the 15 FFPE samples. Histological features of M. hyopneumoniae infection were presented in 11 cases and 7 of these samples stained positive in IHC. Concordance between the histological features and detection results was observed in 13 of the FFPE tissue samples. PCR was the most sensitive technique. Comparison of different sample conservation methods indicated that it is possible to detect M. hyopneumoniae from FFPE tissue. It is important to conduct further research using archived material because the efϐiciency of PCR could be compromised under these conditions.

Diagnóstico imuno-histoquímico e caracterização anatomopatológica de Mycoplasma gallisepticum em galinhas de subsistência

“…compararam IHQ, PCR e cultura bacteriana como método de diagnóstico de micoplasmose em frangos de corte e encontraram concordância de 96,3% para MG e 95,7% para MS entre essas três técnicas. Em mamíferos existem muitos estudos utilizando IHQ para detecção da infecção por micoplasma (Almeida et al 2012), no entanto em aves o número de trabalhos é limitado (Nunoya et al 1995, Nunoya et al 1997, Radi et al 2000.…”

Infecção pelo vírus influenza A em suínos no sul de Moçambique

“…As amostras de pulmão foram igualmente submetidas a exame de IHQ para Circovírus suíno tipo 2 (PCV2) empregando um anticorpo policlonal anti-PCV2 (Iowa State University, USA) na diluição de 1:1000 (Zlotowski et al 2008) e para Mycoplasma hyopneumoniae usando um antisoro autoimune anti-Mycoplasma hyopneumoniae, gentilmente cedido por Nelson Morés (CNPSA/Embrapa), na diluição de 1:200 (Almeida et al 2012).…”