Abstract
Classical swine fever (CSF) caused by classical swine fever virus (CSFV) is one of the most detrimental diseases, and leads to significant economic losses in the swine industry. Despite efforts by many government authorities to stamp out the disease from national pig populations, the disease remains widespread. Here, antiviral small hairpin RNAs (shRNAs) were selected and then inserted at the porcine Rosa26 (pRosa26) locus via a CRISPR/Cas9-mediated knock-in strategy. Finally, anti-CSFV transgenic (TG) pigs were produced by somatic nuclear transfer (SCNT). Notably, in vitro and in vivo viral challenge assays further demonstrated that these TG pigs could effectively limit the replication of CSFV and reduce CSFV-associated clinical signs and mortality, and disease resistance could be stably transmitted to the F1-generation. Altogether, our work demonstrated that RNA interference (RNAi) technology combining CRISPR/Cas9 technology offered the possibility to produce TG animal with improved resistance to viral infection. The use of these TG pigs can reduce CSF-related economic losses and this antiviral strategy may be useful for future antiviral research.
Highlights
Classical swine fever virus (CSFV) belongs to the genus Pestivirus within the family Flaviviridae [1]
The results of this study suggested that these TG pigs offered potential benefits over commercial vaccination and reduced CSFVrelated economic losses
At 72 h post-infection, the viral inhibition efficiencies of each small interfering RNAs (siRNAs) were evaluated by qPCR (Fig 1A) and indirect immunofluorescence assay (IFA) (S2A Fig) [32,33,34]
Summary
Classical swine fever virus (CSFV) belongs to the genus Pestivirus within the family Flaviviridae [1]. Congenital infection with CSFV can result in persistently infected animals, which do not develop specific antibodies against the virus [6]. This effect is probably due to immunotolerance developed during foetal lymphocyte differentiation. Infected animals continuously shed the virus and are potential sources of new CSF outbreaks, in addition, this phenomenon leads to difficulties in diagnosis[7]. Infections with highly virulent CSFV strains exhibit low age dependence of clinical courses and may result in 100% mortality in all age classes of animals and severe neurological signs within 7 to 10 days [8]. Infected pork and pork products are dangerous sources of CSFV
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