Recovery of a Far-Eastern Strain of Tick-Borne Encephalitis Virus with a Full-Length Infectious cDNA Clone

et al. 2022

Preprint

The intron-based stabilization approach is a very useful strategy for construction of stable flavivirus infectious clones. SA14-14-2 is a highly attenuated Japanese encephalitis live vaccine and widely used in China. To develop safe and effective recombinant vaccines with SA14-14-2 as a backbone vector, we constructed the DNA-based infectious clone pCMW-JEV of the vaccine strain using the intron-based stabilization approach, and acquired the rescued virus rDJEV which retained the identical biological properties of the parental virus. Unexpectedly, a rescued virus strain, designated rHV-DJEV, with altered virulence was acquired in one of the transfection experiments. rHV-DJEV showed up to 105-fold increased neurovirulence compared with SA14-14-2 parental strain. Genome sequencing found that the inserted introns still existed in the genome of rHV-DJEV. Therefore, we think that the intron-based stabilization approach should be used carefully in vaccine development and direct iDNA immunization.

Section: Resultssupporting

confidence: 80%

Accidental acquisition of a JEV rescued virus with introns unspliced in the viral genome using intron-based stabilization approach

Huang

et al. 2022

Preprint

“…Upon transfection of the cDNA to Vero cells, the added introns were spliced from the precursor nuclear messenger RNA (pre-mRNA) in the nucleus, resulting in a true copy of the viral RNA which transported to the cytoplasm. Like other rescued viruses which has been reported elsewhere [10][11][12][13][14], our rescued virus rDJEV retained the biological properties of the parental virus well. However, we also accidentally found a rescued virus, termed rHV-DJEV, with altered virulence identi ed in the neurovirulence test.…”

Section: Discussionsupporting

confidence: 80%

Accidental acquisition of a JEV rescued virus with introns unspliced in the viral genome using intron-based stabilization approach

Huang

et al. 2022

Preprint

The intron-based stabilization approach is a very useful strategy for construction of stable flavivirus infectious clones. SA14-14-2 is a highly attenuated Japanese encephalitis (JE) live vaccine strain and widely used since 1989 in China. To develop safe and effective recombinant vaccines with SA14-14-2 as a backbone vector, we constructed the DNA-based infectious clone pCMW-JEV of SA14-14-2 using the intron-based stabilization approach, and acquired the rescued virus rDJEV which retained the identical biological properties of the parental virus. Unexpectedly, a rescued virus strain, designated rHV-DJEV, with altered virulence was accidentally acquired in one of the transfection experiments. rHV-DJEV showed up to 105-fold increased neurovirulence compared with SA14-14-2 parental strain. Genome sequencing found that the inserted introns still existed in the genome of rHV-DJEV. Therefore, we think that the intron-based stabilization approach should be used prudently in vaccine development and direct iDNA immunization.

“…Upon transfection of Vero cells with cDNA, the added introns were spliced from the precursor nuclear messenger RNA (pre-mRNA) in the nucleus, resulting in a true copy of the viral RNA, which was transported to the cytoplasm. Like other rescued viruses that have been reported elsewhere [10][11][12][13][14], our rescued virus rDJEV retained the biological properties of the parental virus. However, we also accidentally found a rescued virus, termed rHV-DJEV, that was found to have altered virulence in the neurovirulence test.…”

Section: Discussionsupporting

confidence: 80%

Accidental acquisition of a rescued Japanese encephalitis virus with unspliced introns in the viral genome when using an intron-based stabilization approach

Huang¹,

et al. 2023

Arch Virol

The intron-based stabilization approach is a very useful strategy for construction of stable flavivirus infectious clones. SA14-14-2 is a highly attenuated Japanese encephalitis (JE) live vaccine strain that has been widely used in China since 1989. To develop safe and effective recombinant vaccines with SA14-14-2 as a backbone vector, we constructed the DNA-based infectious clone pCMW-JEV of SA14-14-2 using the intron-based stabilization approach and acquired the rescued virus rDJEV, which retained the biological properties of the parental virus. Unexpectedly, a rescued virus strain with altered virulence, designated rHV-DJEV, was accidentally acquired in one of the transfection experiments. rHV-DJEV showed up to 105-fold increased neurovirulence compared with the SA14-14-2 parental strain. Genome sequencing showed that the inserted introns were still present in the genome of rHV-DJEV. Therefore, we think that the intron-based stabilization approach should be used with caution in vaccine development and direct iDNA immunization.