Effects of fixative type and fixation time on the detection of Maedi Visna virus by PCR and immunohistochemistry in paraffin-embedded ovine lung samples

In doubtful cases, the histopathological diagnosis of lesions induced by Maedi Visna virus (MVV), a chronic multisystemic lentiviral disease of sheep, needs to be confirmed by the demonstration of MVV in the tissues. The influence of fixatives and the duration of fixation on the detection of MVV by immunohistochemistry (IHC) and PCR in paraffin-embedded tissues was assessed in lung samples with lesions in different degree, from five sheep serologically positive. Samples were fixed in 10% neutral buffered formalin (NBF), Bouin's solution (BS) and a zinc salts-based fixative (ZSF), for different periods of time between 24 h and 30 days. The three fixatives preserved the morphology of the tissues, although in ZSF-fixed samples an increase in the number of desquamated cells was seen in the alveoli. Tissues showed a similar degree of immunolabelling, irrespective of the duration of fixation using ZSF and NBF fixatives. MVV nucleic acids could be detected in samples fixed up to 14 days in NBF and 30 days in ZSF. However, in BS fixed tissues, immunostaining was weak and non-specific signals were observed after 4 days of fixation. Amplification of proviral DNA could not be obtained by PCR in these samples. IHC detected viral antigens in all sheep whereas one sheep with mild lesions was always negative by PCR.