Rapid Detection of Pseudorabies Virus by the Shell Vial Technique

Abstract

Conventional 24-well microtiter plates and shell vials were seeded with pig kidney (PK-15) and bovine turbinate (BT) cells. The monolayers were inoculated with 244 clinical specimens from pigs suspected of having pseudorabies virus (PRV) infection. The results of a shell vial assay (SVA) were compared with those obtained in a 24-well plate cell culture assay in terms of sensitivity and speed of virus isolation. All samples were passaged only once in cell cultures in both assays. Samples producing cytopathic effects (cpe) in 1 or both assay systems and showing positive fluorescence in a direct fluorescent antibody assay were considered to be positive for PRV. Of the 244 samples examined, 118 (48.4%) and 121 (49.2%) were positive by the 24-well plate assay and SVA, respectively. Of the 118 samples positive in 24-well plates, 113 (95.8%) were positive in BT cells and 117 (99.2%) were positive in PK-15 cells. The SVA detected 121 positive samples of which 121 (100%) were positive in PK-15 cells and 113 (93.4%) were positive in BT cells. Virus-specific cpe appeared earlier in the SVA than in the 24-well assay. At 24 hours postinoculation, 91 (75.2%) samples were cpe positive by SVA, whereas only 15 (12.7%) were positive in 24-well plates. All but 2 of the 121 (98.3%) SVA-positive samples were positive within 48 hours postinoculation, whereas only 56 of 118 (47.5%) were positive in 24-well plates during the same time period. These results indicate that the SVA is comparable in sensitivity to 24-well plate assay but yields virus isolation results more quickly. Also, PK-15 cells appeared to be more sensitive than BT cells for PRV isolation.