Development of nested polymerase chain reaction for the detection of<i>Mycoplasma hyopneumoniae</i>in formalin‐fixed paraffin‐embedded lung tissue

Sk, Ha; Choi, Changsun; O, Kim; Hc, Song; Lim, ES; Sh, Kim; Kk, Hwang; Chae, Chanhee

doi:10.1111/j.1751-0813.2005.tb13088.x

Cited by 1 publication

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“…The Mycoplasma hyopneumoniae genome presents a high A+T content of about 70% (Vasconcelos et al 2005) thereby the effects of ϐixatives produce greater damages on its DNA, making its extraction and manipulation a major challenge. Ha et al (2005), found a 100% agreement between seminested PCR on formalin-ϐixed and fresh lung tissues. However, their study included only 5 samples of experimentally infected pigs and the amplicons were of considerably smaller sizes (281 bp and 184 bp for the ϐirst and second reactions, respectively).…”

Section: Discussionmentioning

confidence: 99%

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

et al. 2012

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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.

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Section: Discussionmentioning

confidence: 99%