Summary auto-generated
This study analyzed the envelope (E) protein sequences of six West Nile virus strains with different neuroinvasiveness properties in mice. Researchers compared a neuroinvasive parental strain (WNI) with attenuated derivatives obtained through cell culture passage or selection. Prolonged passage in mosquito cells led to loss of neuroinvasiveness and acquisition of an N-linked glycosylation site at the conserved flavivirus position. However, glycosylation alone does not fully explain attenuation—a leucine-to-proline substitution at position 68 also appeared associated with loss of neuroinvasiveness. A monoclonal antibody escape mutant with a lysine-to-glutamate substitution at residue 307 was also attenuated. Significantly, a partially neuroinvasive revertant recovered the parental E protein sequence despite remaining attenuated, suggesting that mutations outside the E protein region contribute to the attenuation phenotype. The Israeli strain (WNI) differed substantially from the Nigerian strain at both nucleotide and amino acid levels. The data support the hypothesis that West Nile virus neuroinvasiveness determinants are multifactorial and may involve mechanisms similar to other flaviviruses.
Key findings
- Prolonged passage in mosquito cells causes loss of neuroinvasiveness and acquisition of an N-linked glycosylation site, but glycosylation may not directly cause attenuation
- A leucine-to-proline substitution at position 68 of the envelope protein is associated with loss of neuroinvasiveness in cell culture-passaged strains
- A monoclonal antibody escape mutant with a lysine-to-glutamate substitution at residue 307 is attenuated, similar to escape mutants in other flaviviruses
- A neuroinvasive revertant virus regained parental E protein sequence but remained partially attenuated, indicating that determinants outside the E protein region contribute to attenuation
- West Nile virus Israeli and Nigerian strains diverge substantially, differing by approximately 20% at the nucleotide level and 5% at the amino acid level
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Abstract
Several neuroinvasive and non-neuroinvasive West Nile (WN) viruses were characterized by nucleotide sequencing of their envelope (E) protein regions. Prolonged passage in mosquito cells caused loss of neuroinvasiveness and acquisition of an N-linked glycosylation site, which is utilized. Limited passage in cell culture also caused glycosylation but not attenuation, suggesting that glycosylation may not be directly responsible for attenuation and that a second mutation (L68 --> P) may also be involved. A monoclonal antibody-neutralization escape mutant with a substitution at residue 307, a site common to other flavivirus escape mutants, was also attenuated. A partially neuroinvasive revertant regained the parental E sequence, implying that determinants outside of the E region may also influence attenuation. Data suggest that the neuroinvasive determinants may be similar to those for other flaviviruses. Also, sequence comparison with the WN virus (Nigeria) strain revealed considerable divergence of the E protein at the nucleotide and amino acid levels.