A novel Pseudorabies virus vaccine developed using HDR-CRISPR/Cas9 induces strong humoral and cellular immune response in mice

Chen Luo and Qianqian Wang and Ruhai Guo and Jingnan Zhang and Jingya Zhang and Riteng Zhang and Xin Ma and Peixin Wang and Fathalrhman Eisa Addoma Adam and Basit Zeshan and Zengqi Yang and Yefei Zhou and Xinglong Wang (2022) A novel Pseudorabies virus vaccine developed using HDR-CRISPR/Cas9 induces strong humoral and cellular immune response in mice. Virus Research, 322. pp. 1-35. ISSN 0168-1702

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Outbreaks of Pseudorabies (PR) by numerous highly virulent and antigenic variant Pseudorabies virus (PRV) strains have been causing severe economic losses to the pig industry in China since 2011. However, current commercial vaccines are often unable to induce thorough protective immunity. In this study, a TK/gI/gE deleted recombinant PRV expressing GM-CSF was developed by using the HDR-CRISPR/Cas9 system. Here, a four-sgRNA along with the Cas9D10A targeting system was utilized for TK/gI/gE gene deletion and GM-CSF insertion. Our study showed that the four-sgRNA targeting system appeared to have higher knock-in efficiency for PRVs editing. The replication of the recombinant PRVs were slightly lower than that of the parental strain, but they appeared to have similar properties in terms of growth curves and plaque morphology. The mice vaccinated with the recombinant PRV expressing GM-CSF via intramuscular injection showed no obvious clinical symptoms, milder pathological lesions, and were completely protected against wild-type PRV challenge. When compared to the triple gene-deleted PRV, the gB antibodies and neutralizing antibody titers were improved and the immunized mice appeared to have lower viral load and higher mRNA levels of IL-2, IL-4, IL-6, and IFN-γ in spleens. Our study offers a novel approach for recombinant PRV construction, and the triple gene-deleted PRV expressing GM-CSF could serve as a promising vaccine candidate for PR control.

Item Type: Article
Keyword: Pseudorabies virus , GM-CSF, CRISPR/Cas9 , Vaccine
Subjects: R Medicine > RA Public aspects of medicine > RA1-1270 Public aspects of medicine > RA421-790.95 Public health. Hygiene. Preventive medicine > RA638 Immunity and immunization in relation to public health
S Agriculture > SF Animal culture > SF1-1100 Animal culture
Department: FACULTY > Faculty of Sustainable Agriculture
Date Deposited: 10 Nov 2022 20:19
Last Modified: 10 Nov 2022 20:19
URI: https://eprints.ums.edu.my/id/eprint/34792

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