Molecular Therapy: Nucleic Acids (Sep 2020)

Prevention of Avian Retrovirus Infection in Chickens Using CRISPR-Cas9 Delivered by Marek’s Disease Virus

  • Kai Li,
  • Yongzhen Liu,
  • Zengkun Xu,
  • Yu Zhang,
  • Yongxiu Yao,
  • Venugopal Nair,
  • Changjun Liu,
  • Yanping Zhang,
  • Yulong Gao,
  • Xiaole Qi,
  • Hongyu Cui,
  • Li Gao,
  • Xiaomei Wang

DOI
https://doi.org/10.1016/j.omtn.2020.06.009
Journal volume & issue
Vol. 21
pp. 343 – 353

Abstract

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Reticuloendotheliosis virus (REV) is an avian retrovirus that causes an oncogenic, immunosuppressive, and runting-stunting syndrome in avian hosts. The co-infection of REV and Marek’s disease virus (MDV), an oncogenic herpesvirus in chickens, further increases disease severity and reduces MDV vaccine efficacy. The clustered regularly interspaced short palindromic repeats (CRISPR)-Cas9 system has successfully been used against pathogens in mammalian cells. However, the large size of the CRISPR-Cas9 coding sequences makes its in vivo delivery challenging. Here, following the design of a panel of single-guided RNAs targeting REV, we demonstrate that CRISPR/Cas9 can efficiently mediate the editing of the long terminal repeats of REV, resulting in the inhibition of viral protein expression. The CRISPR-Cas9 system disrupts the integrated proviral genome and provides defense against new viral infection and replication in chicken cells. Moreover, by constructing recombinant MDV carrying CRISPR-Cas9 components using an attenuated MDV vaccine strain as the vector, we efficiently delivered the CRISPR-Cas9 system into chickens, and the MDV-delivered CRISPR-Cas9 drastically reduced REV viral load and significantly diminished REV-associated symptoms. To our knowledge, this is the first study establishing avian retrovirus resistance in chickens utilizing herpesvirus-delivered CRISPR-Cas9, which provides a novel and effective strategy against viral infections.

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