A sensitive fluorescence in situ hybridization technique for detection of porcine reproductive and respiratory syndrome virus

A sensitive fluorescence in situ hybridization (ISH) for detecting porcine reproductive and respiratory syndrome virus (PRRSV) RNA in viral infected t issue was developed using digoxigenin-labeled RNA probes targeted on the nu cleocapsid gene of PRRSV. In situ RNA/RNA hybrids were detected with an ant i-digoxigenin antibody-alkaline phosphatase conjugate and further revealed with Fast Red TR salt;naphthol AS-MX phosphate using a fluorescent microsco pe. Viral nucleic acid was readily demonstrated within macrophages, known t o be the major target of PRRSV. In addition, positively stained cells were found in the salivary gland and skin tissues which have not been reported t o contain PRRSV infected cells before. In conclusion, the fluorescence ISH used in this study provides a fast and sensitive means for screening virus- infected tissues in which relatively few cells are affected. This advantage will be especially beneficial for studying viral persistence and for routi ne diagnosis of PRRSV infection. (C) 1999 Elsevier Science B.V. All rights reserved.