Summary auto-generated
Researchers isolated a highly virulent field strain of Sendai virus (M1) from an animal laboratory epidemic and adapted it to LLC-MK2 monkey kidney cells, producing a mutant strain (MVC11) with greatly enhanced replication in these cells. While MVC11 produced 20-fold more infectious virus than M1 in cell culture through enhanced viral gene expression, it became highly attenuated in mice, failing to replicate efficiently in mouse lungs and causing no lethal disease even at high doses. Sequencing the entire genomes of both strains revealed only two amino acid mutations in MVC11: one in the C protein (phenylalanine to serine at position 170) and one in the L protein (glutamate to alanine at position 2050). The C protein mutation appears to reduce its inhibitory effect on RNA polymerase activity, promoting adaptation to monkey cells, while impairing viral fitness in mouse lungs. MVC11-infected epithelial cells showed strong cell death in lung tissue, contrasting with M1's protective cellular response. These findings demonstrate that adaptation to non-natural host cells can be accompanied by dramatic virulence attenuation and that specific viral mutations determine host cell tropism and pathogenicity.
Key findings
- Sendai virus adapted to LLC-MK2 monkey cells (MVC11 mutant) produced 20-fold higher infectious virus titers than the parental M1 strain through enhanced viral gene expression
- MVC11 was highly attenuated in mice despite its superior growth in cell culture, failing to replicate efficiently in mouse lungs and causing no lethal infections
- Only two amino acid substitutions were found between M1 and MVC11: C protein position 170 (Phe→Ser) and L protein position 2050 (Glu→Ala)
- The C170 mutation reduces the inhibitory function of the C protein on viral RNA polymerase, enhancing gene expression in monkey cells but impairing replication in natural mouse host
- MVC11-infected epithelial cells underwent strong cell lysis in mouse lungs, limiting viral spread compared to M1's non-lytic infection pattern
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Abstract
A field strain of Sendai virus (SeV) Ohita-M1 (M1) was isolated from an epidemic in an animal laboratory by passaging in mice. A mutant strain, Ohita-MVC11 (MVC11), was then obtained by passaging M1 in rhesus monkey (LLC-MK2) cells. MVC11 was adapted to LLC-MK2 cells and produced 20 times higher levels of infectious virus than M1. This increased production of infectious virus in LLC-MK2 cells was associated with enhanced viral gene expression. However, MVC11 could not replicate efficiently in mouse lung and was not lethal to mice even when inoculated at a titre of 8 x 10(5) cell-infecting units (CIU) per mouse. On the other hand, with an inoculum of only 4 x 10(1) CIU per mouse, corresponding to 1 LD50, M1 replicated well in mouse lung and was highly virulent to mice. Nucleotide and deduced amino acid sequence analyses of the entire genomes of M1 and MVC11 revealed that adaptation to LLC-MK2 cells and the attenuation of mouse pathogenicity of MVC11 were associated with only two amino acid substitutions; one on the C protein (Phe substituted by Ser at position 170) and the other on the RNA polymerase, the L protein (Glu substituted by Ala at position 2050).