Some Properties of the Transmissible Interfering Component of Vesicular Stomatitis Virus Preparations

Abstract

SUMMARY:In vesicular stomatitis virus inocula containing the transmissible interfering component (T) an exponential relation, a t low doses, between inoculum concentration and virus yield suggested that adsorption of one T particle was enough to exclude infective virus. This relation was not maintained a t high doses of inoculum, possibly because maximal interference required several hours between adsorption of T and virus particles. An assay method for T, based on the dose which gave 37% of the yield from T-free inocula, showed that the T content of inocula was usually related to passage history and yield of virus on undiluted passage. Treatment with immune serum did not appear to neutralize T, which sedimented in the centrifuge more slowly than did virus. T was much less rapidly inactivated at 56O and by U.V.irradiation than was the infectivity. Despite resemblances between the agent T, incomplete influenza virus and interferon, direct evidence is lacking that T is an incomplete form of the vesicular stomatitis virus or an interferon-like substance.In a previous paper (Cooper & Bellett, 1959) we described a phenomenon with vesicular stomatitis virus which resembled the interference due to 'incomplete "virus in influenza (von Magnus, 1951). It was concluded that the low yields from serial undiluted passage of virus were due to a transmissible interfering component (T). The present paper attempts a further definition of the properties of the agent T, and a comparison of T with incomplete influenza virus and influenza; interferon . METHODSThe preparation'of media, virus stocks and tissue cultures, and the assay of virus by plaque count were described previously (Cooper, 1957; Cooper, 1958a; Cooper & Bellett, 1959). Virus titres are expressed as plaque-forming units (pfu)/ml., dilutions being made in phosphate-buffered saline (PBS).Assay of the inteyfering agent T. Samples (0.5 ml.) of the virus stock to be assayed in twofold serial dilutions in the medium composed of Earle's saline + 5 mg. lactalbumin hydrolysatelml. + 2 mg. yeast extractlml. + 5 % (v/v) horse serum were added to washed confluent (20 hr.) chick embryo cell monolayers. After 1 hr. at 87" 5'ml. of 'conditioned' medium (fluid from 20 hr. monolayers) were added, and 20 hr. later the supernatant fluids were removed and plaque-assayed immediately, or after 2-4 days at -2 O O . The derivation of the content of T agent from these values is discussed below.

Assay of Defective-interfering Semliki Forest Virus by the Inhibition of Synthesis of Virus-specified RNAs

“…At present, assays measure the reduction in yield of standard virus or the inhibition of infectious centre formation. The former was originally described for DI VSV (Bellett & Cooper, 1959) and adapted recently to assay DI Sindbis virus (Kowal & Stollar, 1980;Fuller & Marcus, 1980) while the latter has been used for DI lymphocytic choriomeningitis (LCM) virus (Welsh et al, 1972), DI Sindbis virus (Johnston et al, 1975) and DI influenza virus (Janda et al, 1979). A focus-forming assay based on the ability of some DI viruses to inhibit cytopathic effects has been used to measure DI LCM virus (Popescu et aL, 1976) and DI VSV (Winship & Thacore, 1980).…”

Review Article: An Analytical Review of Defective Infections of Vesicular Stomatitis Virus

“…Since the key comparisons (equations 4 and 8) are formulated as ratios, the resulting errors will be minimal provided that the m.o.i, of ND and DI particles are modest (~< 10) and near equal. Interestingly, both the mathematics and the experimental observations of Bellett & Cooper (1959) break down when the m.o.i, of ND or DI particles is too high. In so far as the differential rate equations (2), (3) and (5) reflect the properties of viral RNA synthesis, the reciprocal exponential relationship that arises naturally by integration reflects the properties of virus replication.…”

“…This may be the case where superinfection with too great a multiplicity of DI particles leads to repression of both DI and ND progeny (Bellett & Cooper, 1959;Stampfer et al, 1969;Wild, 1972;Holland & Villareal, 1975;Khan & Lazzarini, 1977). This defective autointerference now might be attributed to a surfeit of easily suppressed DI terminators.…”

“…Under conditions of mixed virus infections where the input ratio of Dis to ND virus is varied, and samples are taken at a fixed time when the harvest is maximal, as in the study of Bellett & Cooper (1959), then equation (10) becomes the reciprocal exponential relationship described in Fig. 1 of these authors:…”

“…Another test of the model would be the derivation of a reciprocal exponential relationship like that of Bellett & Cooper (1959) by including terms for DI replication in the analysis. The rate equation for DI genome synthesis can be written:…”