22 November 2024

4 minute read

By: Dr. Lucina Galina

North America

Disease-resistance genes in livestock provide health benefits to pigs and opportunities for producers to meet the growing demand for affordable, high-quality protein. The porcine reproductive and respiratory syndrome (PRRS) is one of the most persistent and economically impactful swine diseases. Thirty years after discovering the PRRS virus (PRRSV), the swine industry struggles to control it despite significant efforts and implementation of innovative approaches.

Gene-editing and CD163

Recent breakthroughs in gene editing have shown animals resistant to infection and, in so doing, created the potential to prevent epidemics. Using gene-editing with CRISPR-CAS9, a portion of DNA coding for a protein can be deleted. Unlike genetically modified organisms, in the case of the PRRS-resistant pig, DNA from another organism is not introduced to alter the genetic code. Evidence supports that CD163 is an indispensable monocyte and macrophage receptor, and specifically, its Scavenger Receptor Cysteine Rich 5 domain, also called domain 5, is crucial for PRRSV infection and replication.1 CD163's primary function is the clearance of cell-free hemoglobin and participation in inflammatory processes through domains different than domain 5, and no specific role has been associated with domain 5 other than PRRSV infection.2 A list of findings about the relevance of CD163 in PRRSV infections can be found in Table 1.

Commercially-scale founder population

A founder population carrying a 414 base pair deletion in domain 5 of the CD163 gene was used to build commercial breeding populations.3,4 The populations include two dam PIC commercial lines (L02 and L03) and two sire lines (337 and 800). Gene-editing using CRISPR-CAS9 was conducted at the embryonic level in the founder population. After the first set of gene-edited pigs were born, they were bred for multiple generations, where the deleted version of the gene was maintained, and the trait was passed via Mendelian inheritance to the next generations. Today, one important goal is to increase the number of homozygous pigs in this population.

Inheritance of the PRRS-resistant gene

The inheritance of the CD163 edited gene is recessive. This means that pigs must carry two copies of the edited gene version to be resistant (homozygous, aa); pigs carrying only one edited version (heterozygous pigs or Aa) are not resistant to PRRSV. Pigs that inherit the complete CD163 gene that includes domain 5 from both parents are not resistant (null, AA). For implementation postregulatory approvals, the most practical and efficient way to disseminate resistance to PRRS will be via semen of homozygous resistant boars and heterozygous or homozygous gilts.

Phenotypic Evaluations

Challenge studies

Homozygous edited, heterozygous, and null pigs were inoculated with PRRSV isolates, including type I (SD13-15) and II (1-4-4 L1C, 1-8-4 L1H, 1-7-4 L1A, 1-4-2 L1E NVSL97 L8). The studies were conducted in BSL2 facilities at Midwest Veterinary Services in Oakland, NE. Veterinarians and caregivers who oversaw the inoculation and sample collection were blinded to the zygosity of the pigs. Weaned pigs arrived at the BSL2 facilities; pigs from different zygosities were housed together for each of the PRRS inoculates, acclimated for one week, and were intranasally inoculated with 3 mL of PRRSV (± 104 TCID50). Sera were taken before and at 3, 7, 10, 14, and 21 days post-inoculation. Clinical assessments were obtained, including daily temperatures, demeanor, and respiratory scores (0-3). PRRSV PCR and ELISA tests were conducted at the Iowa State University Veterinary Diagnostic Laboratory, and the diagnostician was blinded to the pig identifications. Overall, the type II isolates were more pathogenic than type I. The homozygous pigs were consistently negative by PCR across the bleeding times, while heterozygous and null pigs were positive. The homozygous pigs did not mount an immune response detectable by ELISA. In contrast, heterozygous and null pigs seroconverted.

Key performance indicators

The homozygous edited pigs were also evaluated relative to heterozygous and null pigs to assess key performance production indicators from birth to maturity, reproductive traits, carcass traits, and meat composition. The results of a recent study did not detect statistical differences in meat composition and showed no changes in the growth rate, health, or ability to farrow.5

Conclusions

The outcomes of the studies above suggest that gene-editing offers an unprecedented opportunity to control PRRSV that has plagued the global swine industry for several decades. This technology could also become a platform for disease control for pathogens that have been difficult to control by industry-standard approaches.