In situ hybridization for the detection and localization of porcine epidemic diarrhea virus in the intestinal tissues from naturally infected piglets

Detection and localization of porcine epidemic diarrhea virus (PEDV) was studied by in situ hybridization with a nonradioactive digoxigenin-labeled probe in formalin-fixed, paraffin-embedded tissues from 10 naturally infected piglets. A 377-base pair cDNA probe for viral RNA encoding the membrane proteins of PEDV cell-culture-adapted strain V215/78 was generated by the reverse transcription polymerase chain reaction. In the retrospective study of pigs from herds with diarrhea, the 10 piglets naturally infected with PEDV had positive signals for PEDV by in situ hybridization. When intestinal tissues were hybridized with the PEDV probe, a strong signal was seen in the villus enterocytes of jejunum and ileum but not in the cecum and colon. Positive cells typically had dark brown reaction products in the cytoplasm. Scattered epithelial cells along the ileal Peyer's patches dome areas contained viral RNA. In one piglet, hybridization signal was also found in the duodenum. PEDV was not demonstrated in tissues outside of the intestinal tract. These findings indicate that jejunal and ileal villus enterocytes are the main target of PEDV replication during epizootic outbreaks of the disease.