African swine fever (ASF) leads to high mortality in domestic pigs and wild boar and is caused by the African swine fever virus (ASFV). Currently, no vaccine is commercially available for prevention, and the epidemic is still spreading. Here, we constructed a recombinant pseudorabies virus (PRV) (PRV-ΔgE/ΔgI/ΔTK-(CD2v)) that expresses the CD2v protein of ASFV and evaluated its effectiveness and safety as a vaccine candidate in mice. A homologous recombination fragment containing ASFV CD2v was synthesized and co-transfected into HEK 293 T cells, a knockout vector targeting the PRV TK gene. The transfected cells were infected with PRV-ΔgE/ΔgI, and the recombinant strain (PRV-ΔgE/ΔgI/ΔTK-(CD2v)) was obtained by plaque purification in Vero cells. The expression of ASFV CD2v in the recombinant virus was confirmed by sequencing, Western blotting, and immunofluorescence analysis, and the genetic stability was tested in Vero cells over 20 passages. The virulence, immunogenicity and protective ability of the recombinant virus were further tested in a mouse model. The PRV-ΔgE/ΔgI/ΔTK-(CD2v) recombinant strain is stable in Vero cells, and the processing of CD2v does not depend on ASFV infection. The vaccination of PRV-ΔgE/ΔgI/ΔTK-(CD2v) causes neither pruritus, not a systemic infection and inflammation (with the high expression of interleukin-6 (IL6)). Besides, the virus vaccination can produce anti-CD2v specific antibody and activate a specific cellular immune response, and 100% protect mice from the challenge of the virulent strain (PRV-Fa). The detoxification occurs much earlier upon the recombinant virus vaccination and the amount of detoxification is much lower as well. The PRV-ΔgE/ΔgI/ΔTK-(CD2v) recombinant strain has strong immunogenicity, is safe and effective, and maybe a potential vaccine candidate for the prevention of ASF and Pseudorabies.

中文翻译：

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表达非洲猪瘟病毒CD2v蛋白的重组伪狂犬病病毒对小鼠安全有效

非洲猪瘟 (ASF) 导致家猪和野猪的高死亡率，由非洲猪瘟病毒 (ASFV) 引起。目前，尚无商业化预防疫苗，且疫情仍在蔓延。在这里，我们构建了一种表达 ASFV 的 CD2v 蛋白的重组伪狂犬病病毒 (PRV) (PRV-ΔgE/ΔgI/ΔTK-(CD2v))，并评估了其作为小鼠候选疫苗的有效性和安全性。合成含有 ASFV CD2v 的同源重组片段并共转染到 HEK 293 T 细胞中，这是一种靶向 PRV TK 基因的敲除载体。转染细胞用PRV-ΔgE/ΔgI感染，在Vero细胞中通过噬菌斑纯化获得重组菌株(PRV-ΔgE/ΔgI/ΔTK-(CD2v))。ASFV CD2v在重组病毒中的表达通过测序、Western blotting、和免疫荧光分析，并在 Vero 细胞中测试了 20 代以上的遗传稳定性。在小鼠模型中进一步测试了重组病毒的毒力、免疫原性和保护能力。PRV-ΔgE/ΔgI/ΔTK-(CD2v)重组株在Vero细胞中稳定，CD2v的加工不依赖ASFV感染。PRV-ΔgE/ΔgI/ΔTK-(CD2v) 的疫苗接种既不会引起瘙痒，也不会引起全身感染和炎症（白细胞介素 6 (IL6) 的高表达）。此外，病毒疫苗接种可产生抗CD2v特异性抗体并激活特异性细胞免疫反应，100%保护小鼠免受毒株（PRV-Fa）的攻击。解毒发生在重组病毒疫苗接种时更早，解毒量也低得多。