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

Abstract

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.