Enhancement of plaque formation and cell fusion of an enteropathogenic coronavirus by trypsin treatment

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Abstract
Plaque formation, replication and related cytopathic functions of the enteropathogenic bovine coronavirus strain L9 in bovine fetal thyroid (BFT) and bovine fetal brain (BFB) cells were investigated in the presence and absence of trypsin. Plaque formation was enhanced in both cell types. Plaques reached a size with an average diameter of 5 mm within 4 days with trypsin in the overlay; their diameter remained < 1 mm at this time after plating without trypsin in the overlay. Fusion of both cell types was observed 12-18 h after infection when trypsin was present in the medium. Fusion was not observed in infected BFB cell cultures and was rarely observed 48 h after infection of BFT cells maintained with the trypsin-free medium. The largest polycaryons formed had 15-22 nuclei. They then lysed and detached. Cell fusion depended on de novo synthesis of hemagglutinin and infectivity. Fusion from without was not observed. Virus produced under trypsin-enhancing conditions accompanied by cell fusion did not lyse mouse erythrocytes that reacted with L9 coronavirus hemagglutinin. Trypsin-treated, infected BFT cultures produced coronaviral particles that excluded stain from the envelope confinement. These virions had uniformly shorter surface projections than did the viral forms generated by trypsin-free cell cultures.