A sensitive bioassay system for detecting defective interfering particles of rabies virus

Abstract

Rabies virus has been known to produce CPE after several days incubation. By increasing the incubation temperature to 39-41° after an initial period at 34-36°, we found a rapid CPE accompanied by a cessation of viral replication. When this procedure was utilized with a mixed infection of infectious and DI rabies virions, the surviving cells infected with DI particles stood out against a background of lysed cells which had been infected with infectious virions alone. With this finding, an interference focus-forming assay of rabies virus DI particles was devised. The critical m.o.i. of helper infectious virus was determined to be 0.2 PFU/cell. When a sample had many infectious virions which interfered with this narrow limit of the optimal m.o.i., that problem was overcome by UV irradiation, which inactivated infectious virions at a much higher rate than DI particles. Numbers of interference focus-forming units (IFU) were in linear proportion to the input dose of DI particles. Under optimal conditions, this temperature increase procedure was sensitive enough to measure as few as 20 IFU/ml, almost equal in sensitivity to the plaque assay. © 1982.