Bovine respiratory syncytial virus in-situ hybridization from sheep lungs at different times postinfection

Abstract

We studied the distribution of bovine respiratory syncytial virus (BRSV) RNA in lungs of experimentally infected sheep by in situhybridization at different times postinfection. The probe used for in-situ hybridization was prepared by reverse transcription of BRSV RNA, followed by PCR amplification of the cDNA. Twenty five Merino sheeps of both sexes with a live weight of 55± 10 Kg, received a intratracheal inoculation of 40 ml saline solution containing 1.26 x 10⁶ TCID₅₀ BRSV (strain NMK7) per ml. Sheep were slaughtered 1,3,7,11 and 15 postinoculation days (PID). Bronchial and bronchiolar epithelial cells were positive for BRSV nucleic acid by ISH at 1, 3, 7 and 11 PID. However, alveolar epithelial cells contained positive hybridization signals cells at 1,3 and 7 PID. Cells containing viral RNA were detected from 1 to 11 PID, in exudate within bronchial and bronchiolar lumina; and from 3 to 7 PID in alveolar exudates. Positive hybridization signals were identified in interstitial mononuclear cells and in bronchi associated lymphoid tissue from 3 to 11 PID. The highest signal intensity of positive cells were observed at 3 and 7 PID, coinciding with high virus antibodies titres and with the most important histopathological findings.