We tried to develop detection system of porcine reproductive and respiratory syndrome virus(PRRSV) by in situ hybridization(ISH) in the piglets experimentally infected with KPRRS-2, the Korean isolate(12 piglets) or Mn-lb, the American isolate(4 piglets), and in the natural infection suspected 6 piglets.
Twelve 30-days-old piglets(two pigs per each inoculated group) were inoculated by nasal instillation of KPRRS-2 virus(total dose l0^(4.5)TCID_(50)), Six piglets(one pig per each group) were induced contact infection with inoculated piglets, during the experiment, and two piglets were used as control. Inoculated or contacted piglets were euthanized at 1, 3, 5, 7, 14 and 21 days postinoculation(DPI).
The respiratory signs such as coughing and nasal discharge were observed on day 3 DPI, and ear cyanosis were on day 5 DPI, including contacted piglets. Through the necropsy, purple discolorization of dorsal part of lung, and hypertrophy of local lymph nodes were observed. The histopathological lesions of lung were interstitial pneumonia characterized by type 2 pneumocyte hyperplasia.
We prepared the probe for ISH by RNA isolation from KPRRS-2, RT-PCR, and biotin labeling.
We performed the ISH within only 1∼2 hours using Microprobe^(TM) capillary action system. As the results, the strong red specific positive signals, means PRRSV distribution, was mainly observed in the cytoplasm of alveolar macrophages. And also signals were detected in some type 2 pneumocytes and bronchiolar epithelium of lung, myocardium, liver, kidney, tonsil, spleen, gastrointestinal mucosa, testis and lymph nodes.
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