The Role of CD163 in the Sensitivity to Porcine Reproductive and Respiratory Syndrome Virus.

RS Prather¹, KD Wells¹, KM Whitworth¹, AJ Mileham²

¹University of Missouri, Columbia, MO, ²Genus plc, DeForest, WI

A recent model of porcine reproductive and respiratory syndrome virus (PRRSV) infection involves the virus entering and infecting porcine alveolar macrophages (PAMs). The first contact between PRRSV and the PAMs had been thought to be via heparan sulfate, and this binding was followed by binding to SIGLEC1. The SIGLEC1-virus complex was then thought to be endocytosed. Within the endosome the virus then was uncoated by CD163. This uncoating released the viral genome to infect the cell. To determine which molecules are involved it is necessary to genetically modify the candidate genes. Initially, we used classical homologous recombination to knockout SIGLEC1 in pigs. The SIGLEC1-/- piglets had no resistance to a PRRSV challenge. To modify the next candidate gene we used a meganuclease (CRISPR/Cas9). In our hands the CRISPR/Cas9 injection into zygotes has proven to be very effective. We have to date edited 5 different genes in piglets by zygote injection. Of the 31 piglets born, 29 were edited. This editing technology is not only easy to use, but highly efficient. We used the CRISPR/Cas9 system to both modify somatic cells prior to somatic cell nuclear transfer and to edit CD163 in zygotes. The response of CD163-/- piglets to a PRRSV challenge will validate or invalidate the original model of the method of PRRSV infectivity.

Funding from Genus plc, NIH (U42OD011140), and Food for the 21^st Century.