Rabies pathogenesis. Brief review.

This chapter will emphasize recently derived knowledge concerning the nature of defective interfering (DI) particles of RNA animal viruses, their biological origins and functions, and their involvement in long-term persistent infections. We will not attempt to review all of the DI literature, and we will confine ourselves to DI particles of RNA viruses. The previous review by Huang and Baltimore (1977) amply documents the occurrence and behavior of DI particles in a wide variety of DNA and RNA viruses and discusses their biological effects, and a very thorough recent review of rhabdovirus DI particles by Reichmann and Schnitzlein (1978) provides excellent in-depth coverage of many areas not covered by the present chapter as well as some alternate viewpoints of areas which are considered here. We will omit DNA virus DI particles from extensive consideration because of space limitations and because they are generally less well characterized at present.

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Defective Interfering RNA Viruses and the Host-Cell Response

Characterization of rabies viruses recovered from persistently infected BHK cells.

Summary
BHK 21 carrier cells persistently infected with VSV Indiana for over 2 years have been shedding generally very low levels of mature infectious virus or mature T particles (averaging less than one-hundredth p.f.u./cell/day) yet most cells are producing virus antigens and are resistant to homologous superinfection. However, large amounts of biologically active T particle RNP can be recovered from cytoplasmic extracts of these carrier cells even at times when they are shedding no detectable infectious virus. This recovered cytoplasmic RNP replicates (with helper B virions) to produce mature T particles, interferes strongly after DEAE dextranfacilitated uptake and, together with B virions, allows the establishment of a persistent carrier state in exposed cells.
No ‘provirus’ DNA copies of the VSV RNA genome are detectable (less than 1/40 copy/cell or 1 copy per 40 cells) in carrier cells after more than 2 years of persistent infection, and all transfection attempts have failed using DNA from these VSV carriers or DNA from carrier cells persistently infected with some other negative strand RNA viruses (measles, mumps, LCM, influenza, rabies).
Infectious viruses shed after more than 1 year from carrier cells originally infected with wild-type B virions are small plaque mutants showing a slight temperature sensitivity. Cured cell populations can be obtained from the long term VSV carrier culture by cloning in the presence or absence of antiviral antibody.

Long-Term Persistent Vesicular Stomatitis Virus and Rabies Virus Infection of Cells In Vitro

Previous indications that cloned B virions might be genetically predisposed to generate a particular defective T particle are shown to be inaccurate. T particle generation was found to be a much more random process than was previously believed. We show that the previously observed generation of particular sizes of T particles by B virion pools is due to the random generation of T particles during preparation of first-passage pools of cloned B virions, and these breed true during the additional passages needed to produce visible quantities of T particles. It is also shown that different host cell lines selectively amplify different T particles, suggesting a strong role of host cell factors in T particle replication. Surprisingly, our line of HeLa cells did not generate or replicate detectable T particles of vesicular stomatitis virus (VSV) Indiana after either serial undiluted passage or direct addition of T particles, even though the added T particles strongly interfered with B virion replication. In contrast to VSV, rabies virus generates large amounts of T particles during the first passage of cloned B virions, and every rabies-infected baby hamster kidney-21 cell culture evolves into a persistent carrier state. We find that T particle RNA is biologically inactive although T particle nucleocapsid ribonucleoprotein replicates and interferes in cells coinfected with B virions. Efforts to study the mechanism of T particle generation by in vitro attempts to generate T particles or modify their size (using sheared ribonucleoprotein or chemical or UV mutagenesis) were unsuccessful. The kinetics of UV and nitrous acid inactivation of T particles indicate a smaller target size relative to B virions, even after correcting for lengths of RNA molecules. The intercalating dye proflavine does not photosensitize VSV B virions or T particles when present during replication, indicating that there is little or no RNA base pairing in the helical nucleocapsids of either.

Factors involved in the generation and replication of rhabdovirus defective T particles.

Exposure of both mammalian and reptilian cells in tissue culture to different strains of fixed rabies virus resulted in a carrier type of infection. No cytopathic effect was observed in either type of culture; infected cultures could be maintained by cell transfer for unlimited numbers of passages. A consistent pattern of cyclically rising and falling levels of viral infection was observed by fluorescent-antibody staining techniques and by titration of released infectious virus. Resistance to super-infection by vesicular stomatis virus and the production of an interferon-like substance by infected cells indicated that the maintenance of a carrier type of infection may be interferon-mediated. The degree of susceptibility of rabies-infected cells to immunolysis by antirabies antibody in the presence of complement was found to be correlated with the amount of virus maturation occurring by budding through the cell membrane and not with the presence of immunofluorescent antigen in the cytoplasm of infected cells.

https://iai.asm.org/content/iai/6/6/988.full.pdf

T. J. Wiktor

Papers

Chronic Rabies Virus Infection of Cell Cultures