Construction of pseudorabies virus variant attenuated vaccine: codon deoptimization of US3 and UL56 genes based on PRV gE/TK deletion strain

Since 2011, pseudorabies based on the pseudorabies virus (PRV) variant has emerged as a serious health issue in pig farms in China. The PRV gE/TK or gE/gI/TK deletion strains protect against emerging PRV variants. However, these variants may cause lethal infections in newborn piglets without PRV antibodies. Previous studies have shown that codon deoptimization of a virulence gene causes virus attenuation. Accordingly, we deoptimized US3-S (US3 gene encoding a short isoform that represents approximately 95% of the total US3 transcription) and UL56 genes (first 10 or all codons) of PRV gE/TK deletion strain (PRV Delta TK&gE-AH02) to generate six recombinant PRVs through bacterial artificial chromosome technology. In swine testicular cells, recombinant PRVs with all codon deoptimization of US3-S or UL56 genes were grown to lower titers than the parental virus. Notably, US3-S or UL56 with all codon deoptimization reduced mRNA and protein expressions. Subsequently, the safety and immunogenicity of recombinant PRVs with codon deoptimization of US3-S or UL56 are evaluated as vaccine candidates in mice and piglets. The mice inoculated with recombinant PRVs with codon deoptimization of US3-S or UL56 showed exceptional survival ability without severe clinical signs. All codons deoptimized (US3-S and UL56) significantly decreased virus load and attenuated pathological changes in the brains of the mice. Moreover, the protection efficiency offered by recombinant PRVs with codon deoptimization of US3-S or UL56 showed similar effects to PRV Delta TK&gE-AH02. Remarkably, the 1-day-old PRV antibody-negative piglets inoculated with PRV Delta TK&gE-US3-ST-CD (a recombinant PRV with all codon deoptimization of US3-S) presented no abnormal clinical symptoms, including fever. The piglets inoculated with PRV Delta TK&gE-US3-ST-CD showed a high serum neutralization index against the PRV variant. In conclusion, these results suggest using codon deoptimization to generate innovative live attenuated PRV vaccine candidates.

Automated summary

Pseudorabies based on the pseudorabies virus (PRV) variant has become a serious health issue in pig farms in China since 2011. In this study, codon deoptimization was used to modify PRV Delta TK&gE-AH02, creating recombinant PRVs with improved safety and immunogenicity. The results showed that these modified PRVs provided strong protection against PRV variants in mice and piglets, making them promising vaccine candidates.