Biological and biochemical properties of defective interfering particles of equine herpesvirus type 1

Abstract

Defective interfering (DI) particles of equine herpesvirus type 1(EHV-1) were purposely generated in an in vitro system of L-M cells by repeated high-multiplicity, serial, undiluted passage. Quantitation of infectious virus revealed a definite cyclic pattern which increased in magnitude with continued passage; additional experiments indicated that these fluctuations in virus titer were due to the presence of a population of DI particles as judged by interference assays. Attempts to separate standard and defective EHV-1 were unsuccessful. Analysis of DNA isolated from mixed populations of these virions revealed the presence of a high-density (H) variant DNA (ϱ = 1.724 g/cm 3) in addition to standard EHV-1 DNA (ϱ = 1.716 g/cm 3). Furthermore, it was found that the relative amount of this H-DNA in each passage corresponded very closely to the fluctuations in infectious virus titer. Sedimentation velocity studies of DNA isolated from populations of virions rich in H-DNA (>99%) indicated that the variant genomes were the same size as the standard EHV-1 genome (50–55 S). Comparisons of purified virion populations from 17 high-multiplicity passages with regard to particle counts, relative amount of H-DNA, and infectious virus titer indicated that the relative interference capacity of EHV-1 DI particles increased significantly with continued passage. Although the factor(s) responsible for the increased interference activity is unknown, DNA-DNA hybridization analyses of selected passages rich in H-DNA indicated that the genomes of EHV-1 DI particles became genetically less complex with passage and contained significant amounts of reiterated sequences. The possible mechanisms of the evolution of EHV-1 DI particles and their role in the interference process are discussed.