A Noval Strategy of US3 Codon De-Optimization for Construction of an Attenuated Pseudorabies Virus against High Virulent Chinese Pseudorabies Virus Variant

In this study, we applied bacterial artificial chromosome (BAC) technology with PRVΔTK/gE/gI as the base material to replace first, central, and terminal segments of US3 gene codon-deoptimized fragments via two-step Red-mediated recombination in E. coli GS1783 cells. The three constructed BACs were co-transfected gI part gE carrying homologous sequences (gI+gE’), respectively, swine testicular These recombinant viruses codon de-optimization ((PRVΔTK&gE-US3deop−1, PRVΔTK&gE-US3deop−2, PRVΔTK&gE-US3deop−3) obtained purified. exhibited similar growth kinetics parental AH02LA strain, stably retained deletion TK fragments, inherited recoded US3. Mice inoculated intraperitoneally or control virus PRVΔTK&gEAH02 at a 107.0 TCID50 dose. immunized PRVΔTK&gE-US3deop−1 did not develop clinical signs had decreased load attenuated pathological changes lungs brain compared group. Moreover, mice challenged 100 LD50 provided protection that PRVΔTK&gEAH02. Finally, was injected intramuscularly into 1-day-old PRV-negative piglets dose 106.0 TCID50. Immunized showed only slight temperature reactions mild signs. However, high levels seroneutralizing antibody produced 14 21 days post-immunization. addition, immunization 105.0 complete prevented shedding by 106.5 PRV variant 1 week post immunization. Together, these findings suggest displays great potential vaccine candidate.