Rabies virus quasispecies: Implications for pathogenesis

Abstract

Passage of the mouse-adapted rabies virus strain CVS-24 (where CVS is challenge virus standard) in BHK cells results in the rapid selection of a dominant variant designated CVS-B2c that differs genotypically and phenotypically from the dominant variant CVS-N2c present in mouse-brain-or neuroblastoma-cell-passaged CVS-24. The glycoprotein of CVSB2c has 10 amino acid substitutions compared with that of CVS-N2c. Because CVS-B2c can be reproducibly selected in BHK cells, it is likely to be a conserved minor subpopulation of CVS-24. CVS-N2c is more neurotropic in vitro and in vivo than CVS-B2c, which replicates more readily in nonneuronal cells in vitro and in vivo. These characteristics appear to be relevant to the pathogenicity of the two variants. CVS-N2c is more pathogenic for adult mice than CVS-B2c. In contrast, CVS-B2c is more pathogenic for neonatal mice. These differences in pathogenicity are reflected in the selection pattern when mixtures of CVS-N2c and CVS-B2c were used to infect neonatal and adult mice. Although CVS-N2c was highly selected in adult mice, no selection for either variant was seen in neonates, suggesting that certain aspects of development, such as maturation of the nervous and immune systems, may contribute to the selection process. We speculate that the existence of different variants within a rabies virus strain may facilitate the virus in overcoming barriers to its spread, both within the host and between species.By far the predominant target of rabies virus infection in vivo is the neuron (1). Nevertheless, it has been shown that rabies virus can infect myocytes before the virus infection reaches neurons in the spinal cord or dorsal root ganglia (1, 2). In addition, high titers of rabies virus can be produced in mucogenic acinary cells of the salivary glands during the final phase of infection. In vitro experimentation has shown that street rabies virus can infect macrophages (3) and fixed rabies virus strains can infect murine lymphocytes and human lymphoblast T cells (4). However, it is not clear whether virus replication in nonneuronal tissue, with the exception of the salivary glands, actually represents an integral part of the pathogenesis of rabies nor it is known whether a single virus population or different subpopulations of a particular strain are responsible for the infection of neuronal and nonneuronal cells. In this context, it has been shown, in mice persistently infected with lymphocytic choriomeningitis virus (LCMV), that different variants of the same LCMV strain are associated with infection of neuronal cells, liver cells, or spleen cells (5, 6). Because of the high mutation rates resulting from the lack of proofreading by RNA polymerases RNA virus strains exist as quasispecies (7). It has been demonstrated that in a constant environment quasispecies contain a stable dominant virus species within a pool of viruses with related but often extremely heterogeneous genomes (7-9). Such complexity endows RNA viruses with an enormous capacity to adapt to changing host...

Differentiation of an adult neuron cell line increases susceptibility to rabies infection.

“…No morphological damage or cytolytic effect was observed in cells 24 hours post-infection with CVS rabies virus strain, agreeing with previous data in which it has been proposed that highly neurotropic strains do not cause neuron death (13).…”

“…Preliminary results from our group indicate that the three molecules postulated as rabies virus receptors (p75 NTR , NCAM and NAChR) are found on CAD-R1 cells, but cell number and staining intensity is greater in differentiated cells (23). In spite of it being known that the rabies virus CVS strain acquires different genotypical and phenotypical characteristics, depending on substrate in which it is maintained (13,14), no infection differences between CVS-BHK and CVS-MB strains were found in the present study.…”

“…The main disadvantage regarding primary cultures is non-neuronal cell presence, making it difficult to study the virus interaction with neurons and the need to supplement them with exogenous neurotrophic factors for neuron survival and maintenance, which could affect RV replication (12). On the other hand, the most frequently used cell lines in virus experiments are non-neuronal and those which are neuronal are embryonic, behaving differently to adult neurons (13). There is currently no CNS adult neuron culture model available for studying RV infection.…”

“…It is well known that the fixed Challenge Virus Standard (CVS) rabies virus strain preferentially infects a certain type of cells depending on the type of cells in which they have been adapted (13,14). If it has been maintained in adult mouse brain (CVS-MB) it becomes more pathogenic and neurotropic than if it has been adapted to fibroblastic cells (CVS-BHK), in which case (in spite of continuing to be neurotropic) infecting a greater percentage of non-neuronal cells in primary cultures (14,15).…”

Expressão gênica empregando pseudopartículas em células de mamíferos (HEK 293T e Huh 7.0) cultivadas em diferentes meios de cultura livres de soro.

“…A RVGP apresenta três estados conformacionais possíveis e intercambiáveis: um estado inativo necessário para seu transporte pelo complexo de Golgi, um estado nativo hidrofóbico, presente no vírus circulante e um estado ativo presente no pH ligeiramente ácido a que o vírus é exposto logo após a adesão à célula hospedeira e internalização mediada por endocitose, o que leva à fusão do vírus com a membrana do endossomo (GAUDIN et al, 1991(GAUDIN et al, , 1999LENTZ et al, 1982LENTZ et al, , 1984MORIMOTO et al, 1998;THOULOUZE et al, 1998).…”