Abstract
Porcine reproductive and respiratory syndrome virus (PRRSV) 1 and 2 differ in their recognition of CD163. Substitution of porcine CD163 SRCR5 domain with a human CD163-like SRCR8 confers resistance to PRRSV 1 but not PRRSV 2. The deletion of CD163 SRCR5 has been shown to confer resistance to PRRSV 1 in vivo and both PRRSV 1 and 2 in vitro. However, the anti-PRRSV 2 activity of modifying the CD163 SRCR5 domain has not yet been reported. Here, we describe the highly efficient generation of two pig breeds (Liang Guang Small Spotted and Large White pigs) lacking a short region of CD163 SRCR5, including the ligand-binding pocket. We generated a large number of gene-edited Large White pigs of the F0 generation for use in viral challenge studies. The results of this study show that these pigs are completely resistant to infection by species 2 PRRSV, JXA1, and MY strains. There were no clinical symptoms, pathological abnormalities, viremia, or anti-PRRSV antibodies in the CD163 SRCR5-edited pigs compared to wild-type controls after viral challenge. Porcine alveolar macrophages (PAMs) isolated from CD163 SRCR5-edited Large White pigs also displayed resistance to PRRSV in vitro. In addition, CD163 SRCR5-edited PAMs still exhibited a cytokine response to PRRSV infection, and no significant difference was observed in cytokine expression compared to wild-type PAMs. Taken together, these data suggest that CD163 SRCR5-edited pigs are resistant to PRRSV 2, providing a basis for the establishment of PRRSV-resistant pig lines for commercial application and further investigation of the essential region of SRCR5 involved in virus infection.
Highlights
Porcine reproductive and respiratory syndrome (PRRS), caused by PRRS virus (PRRSV), is one of the most prevalent and serious infectious diseases in the global swine industry [1]
SCD163 levels were found to be equal in supernatants of mock infected CD163 SRCR5-edited Porcine alveolar macrophages (PAMs) and WT PAMs, and were not significantly different between virus infected Porcine reproductive and respiratory syndrome virus (PRRSV)-infected WT PAMs and CD163 SRCR5-edited PAMs. These results suggest that PAMs from CD163 SRCR5-edited pigs are completely resistant to PRRSV infection
Porcine embryonic fibroblasts (PEFs) with targeted deletion of the SRCR5 region in CD163 were enriched using our previously developed dual fluorescence selection strategy [30]. Using these enriched cells as donors for Somatic Cell Nuclear Transfer (SCNT), we generated a high number of pigs harboring a 41-aa deletion of CD163 SRCR5
Summary
Porcine reproductive and respiratory syndrome (PRRS), caused by PRRS virus (PRRSV), is one of the most prevalent and serious infectious diseases in the global swine industry [1]. Since it was first described in 1987 in the United States, PRRS has caused huge economic losses worldwide, especially in China [2,3,4,5]. Cellular molecules including heparan sulfate, CD163, sialoadhesin (CD169), CD151, vimentin, and DC-SIGN (CD209) have been described as potential receptors for PRRSV [11, 12] Among these cellular receptors, CD163 has been identified as the essential receptor mediating both species 1 and 2 PRRSV infection [13,14,15,16]. PRRSV species 1 and 2 differ in their recognition of CD163 [17]
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