Evolution of mouse hepatitis virus: detection and characterization of spike deletion variants during persistent infection

Abstract

High-frequency RNA recombination has been proposed as an important mechanism for generating viral deletion variants of murine coronavirus. Indeed, a number of variants with deletions in the spike glycoprotein have been isolated from persistently infected animals. However, the significance of generating and potentially accumulating deletion variants in the persisting viral RNA population is unclear. To study this issue, we evaluated the evolution of spike variants by examining the population of spike RNA sequences detected in the brains and spinal cords of mice inoculated with coronavirus and sacrificed at 4, 42, or 100 days postinoculation. We focused on the S1 hypervariable region since previous investigators had shown that this region is subject to recombination and deletion. RNA isolated from the brains or spinal cords of infected mice was rescued by reverse transcription-PCR, and the amplified products were cloned and used in differential colony hybridizations to identify individual isolates with deletions. We found that 11 of 20 persistently infected mice harbored spike deletion variants (SDVs), indicating that deletions are common but not required for persistent infection. To determine if a specific type of SDV accumulated during persistence, we sequenced 106 of the deletion isolates. We identified 23 distinct patterns of SDVs, including 5 double-deletion variants. Furthermore, we found that each mouse harbored distinct variants in its central nervous system (CNS), suggesting that SDVs are generated during viral replication in the CNS. Interestingly, mice with the most severe and persisting neurological disease harbored the most prevalent and diverse quasispecies of SDVs. Overall, these findings illustrate the complexity of the population of persisting viral RNAs which may contribute to chronic disease.

The complete sequence of the bovine torovirus genome

“…In that regard, it is remarkable that most differences occur at isolated nucleotides except for a region of 217 nts in the putative bulbous domain (S1) of the spike glycoprotein. This is highly reminiscent of a well-characterized hypervariable region in the S1 domain of the spike glycoprotein of the murine hepatitis virus (Lai and Cavanagh, 1997;Rowe et al, 1997).…”

Detection of feline coronaviruses by culture and reverse transcriptase-polymerase chain reaction of blood samples from healthy cats and cats with clinical feline infectious peritonitis

“…Studies in mice have been performed that show that antigen-or mitogen-stimulated lymphocytes acquire the capacity to replicate exogenous or endogenous viruses (Hirsch, 1976). Also, in vitro maintenance of PBMCs from infected animals can activate the expression or replication of certain viruses producing persistent infections (Stock and Ferrer, 1972;Rowe et al, 1997). Similarly, PBMCs from FCoVinfected cats may have acquired the capacity to express or replicate virus in vitro, to coinfect other cells, or to release soluble factors (e.g.…”

The Molecular Biology of Coronaviruses

“…The complementarity of the MHV and TGEV RBD loci is further emphasized by the fact that substantial deletions are tolerated in TGEV S1 in the region that corresponds to the MHV RBD . Conversely, substantial deletions are tolerated in MHV S1 in the region that corresponds to the TGEV RBD (Parker et al, 1989;Rowe et al, 1997).…”