Dr. Jack Dekkers and Dr. Bob Rowland - Introduction of the Program
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Introduction of the Program - Dr. Jack Dekkers,Iowa State University, and Dr. Bob Rowland, Kansas State University, from the 2017 North American PRRS/National Swine Improvement Federation Joint Meeting, December 1‐3, 2017, Chicago, Illinois, USA. More presentations at http://www.swinecast.com/2017-north-american-prrs-nsif-joint-meeting
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Dr. Jack Dekkers and Dr. Bob Rowland - Introduction of the Program
- 1. Sponsored by NIFA grant # 2013-68004-20362 Genetically Improving Resistance of Pigs to PRRS Virus Infection 2013-2017 Dr. Margo Holland, NIFA Program Leader Translational Research: The Genetics of Disease Resistance; Updates on Current Progress and a Vision for the Future
- 2. Use genomics to identify genes / genomic regions associated with resistance / susceptibility to PRRS virus infection Led by Joan Lunney – USDA – ARS Beltsville Bob Rowland – Kansas State University Jim Reecy – Iowa State University Jack Dekkers – Iowa State University Strong Industry Participation PHGC Breeding Companies Fast Genetics, Genesus, Choice Genetics PIC/Genus, TOPIGS, PigGen Canada 60 k SNP chip Illumina GeneSeek 2007
- 3. Evolution of PRRS Host Genetics Research 1. Experimental infection of nursery pigs with NVSL 1. Experimental infection of nursery pigs with KS06 1. Experimental co-infection of nursery pigs: PRRS + PCV2 (incl. PRRS vaccination) 1. Field trials 2. Natural Challenge Model NIFA NIFA NIFA Trial Number n Breed PRRSv Isolate 1-3 530 LW x LR NVSL 4 195 Duroc x LW/LR 5 184 Duroc x LR/LW 6 123 LR x LR 7 194 Pietran x LW/LR 8 188 Duroc x LW/LR 15 184 LR x LW 10 176 LR x LW KS06 11 176 LW x LR 12 174 LR x LW 14 180 Duroc x LR/LW
- 4. Funding Industry Partners NIFA Thanks to all Partners Iowa State University Nick Serao Jim Reecy Chris Tuggle Susan Carpenter Kansas State University Bob Rowland PRRS group USDA-ARS Joan Lunney group University of Alberta Graham Plastow group Univ. Saskatchewan John Harding group Roslin Institute Steve Bishop Andrea Doeschl-Wilson Univ. Minnesota Monserat Torremorrell Scientific Collaborators
- 5. PRRS overview and description of disease phenotypes and models for genetic research Bob Rowland – Kansas State University Probing mechanisms of PRRS resistance Joan Lunney – USDA ARS BARC New PRRS disease phenotypes as vaccine and genetic improvement targets Andrea Wilson – Roslin Institute Resilience and PRRS in a natural disease challenge model Graham Plastow – University of Alberta Viral genetics and application to vaccine development Jay Calvert – Zoetis PRRS genetic resistance: an online class at destination of swine experts and professionals Perle Boyer – University of Minnesota Using genetic selection and genomics to combat infectious disease Jack Dekkers – Iowa State University The Genetics of Disease Resistance Updates on Current Progress and a Vision for the Future
- 6. PRRS overview and description of disease phenotypes and models for genetic research Raymond (Bob) Rowland December 1-3, 2017 Intercontinental Chicago Magnificent Mile
- 7. Guest Editors: Raymond (Bob) Rowland Joan Lunney Editor: X.J. Meng Alternative Strategies for the Control of PRRS
- 8. Rodarterivirus Arterivirus LDV PRRSV SHFV EAV Suid Muroid Reorganization and expansion of the nidoviral family Arteriviridae. Kuhn JH, et al. 2016. Arch Virol. 161:755-68 PRRSV-1 PRRSV-2
- 9. Changes in Weight Distribution after Infection “Reproductive Failure of Unknown Etiology” Kerry K. Keffaber, 1989, AASP 1. Influenza-like clinical signs 2. Mid- to late-term abortions 3. Pre-weaning mortality 4. Poor growth performance Porcine reproductive and respiratory syndrome (PRRS) $14 billion in losses ($600million/year)
- 10. PRRS is a production system disease Endemic phase with outbreaks of severe disease 2003-Eric Neumann Stealthy Easily transmitted Persistent Participates in polymicrobial diseases Viremia Day after infection Persistence in a production system Persistence in a population and within a pig
- 11. Corn Prices The greatest cost of PRRSV is wasted feed Sick and dead pigs, Slow growing pigs Secondary infections Nutritional, Environmental and Social Impacts
- 12. Vaccines Detection Ecology Epidemiology Biosecurity Sociology Resistance Tolerance Resilience Vaccine readiness Genome editing Feed efficiency Feed formulation Microbiome Nutritional Program Disease Control Pig Genetics Nutrition Integrated approach to PRRS control
- 13. Rodarterivirus genome organization-PRRSV 1a 1b 2a 2b 3 4 5 6 7 5’ UTR pA 1/5 of the genome 3’ UTR 2b GP2 GP3 GP4 GP5 M N pA pA pA pA pA pA 3 4 5 M N 15 kb genome nsp2 5 a 2
- 14. PRRS vaccines • Modified live virus (MLV) vaccine introduced in the U.S.1994- approved for use in PRRSV-infected herds • MLV limitations-virus shedding, persistent infection, incomplete immune protection, inability to differentiate infected from vaccinated animals (DIVA), potential for reversion to virulence • Killed vaccines are not effective • Subversion of host immunity and antigenic variation have made further advances in vaccines difficult to achieve
- 15. The application of genetics for improving animal health •Marker selected breeding to improve response to infection •Genetic modification Insertion of genes or alleles that promote resistance Deletion of genes involved in virus susceptibility
- 16. 2007-PRRS Host Genetics Consortium (PHGC) Joan Lunney, Bob Rowland, Jack Dekkers, NPB “Integrating state of the art genomics with state of the art virology” • Develop marker-assisted breeding tools PRRS-resistant pig PRRS-tolerant or resilient pig Vaccine-ready pig • Identify new host response pathways to aid vaccine development (RNA-Seq) • Create a community resource of data and sample repositories for future research
- 17. Models for the study of genetics (PHGC) • Nursery pig model (PRRSV infection) -3200 pigs-16 trials • Field models- 1,000 pigs • Dual-infection disease model- 1,000 pigs All pigs and parents are genotyped for >60,000 SNPs using the Porcine SNP60 BeadChip (Illumina Corp) Well-characterized PRRSV-2 isolates, NVSL, KS06, KS62 Genome-wide association studies (GWAS)
- 18. PHGC data/sample collection timeline Nursery model -7 0 7 11 14 21 35 424 28 Acclimation Weight Blood, Tempus (RNA) Weight Blood Tempus Weight Blood Tempus Weight Blood Tempus Weight Blood Tempus Weight Blood Tempus Weight Blood Tempus TonsilsBlood Tempus Blood Tempus Infection Acute Infection Rebound Persistence Oral fluid samples Fecal samples
- 19. Phenotypic data-sample types • Serum- virus, cytokines, antibody, etc. • Whole blood- PBMC • Whole blood RNA preserved in Tempus tubes- RNA-Seq • Oral fluid – virus, antibody • Feces- caloric retention, microbiome • Tonsil- persistent infection • Other tissues
- 20. Phenotypic data • Reproducible (commercial kits or standard diagnostic tests) Tetracore PCR • Transferable (pool data from different studies) • Cost-effective Weighing a pigs is cheap, but commercial tests may be expensive • Collection is non-terminal and “non-invasive”
- 21. Phenotypic data-types • Binary- live versus dead • Qualitative (subjective) – clinical scores • Quantitative- weight gain, virus load, antibody levels
- 22. Nursery model-Deep phenotyping for analysis of disease traits • Morbidity and mortality • Viremia, qRT-PCR log PRRSV RNA templates/rxn • Virus Load in serum, area under the virus PCR curve (AUC) for the first 21 days • Weight (weekly) – average weight gain (WG) • Total antibody and virus neutralizing activity (42 dpi) • Circulating cytokine levels • Virus level in tonsil (persistence) • “Ultra-deep” phenotyping-Transcriptome analysis (whole blood and tonsil)
- 23. Nursery pig model NVSL 97-7895 (1,499 pigs) KS 2006 (845 pigs)
- 24. Viral load-area under the curve (graph) (AUC) Quantified as area under the curve from day 0 to 21
- 25. Criteria for pigs • 200 pigs per trial • ~6 pigs/litter from a limited number of sires • Mated with 2-3 dams/sire • Pig sources provide pedigree information and DNA from the parents for later genotyping analyses • Weaned pigs are of a “high-health” status and free of PRRSV, Mycoplasma hyopneumoniae and swine influenza virus (SIV) • No pre-selection for a PRRS-associated trait
- 26. viremia weight rebound Nursery pig model Phenotypic variation predicts genetic variation 200 pigs per trial
- 27. Nursery pig model-The host responses, virus load and weight gain, are moderately inherited traits • Approximately 40% of how a pig responds to PRRSV infection is inherited • The remaining 60% is dependent on Maternal effects Environment Virus • Impact- breed pigs for improved disease resistance
- 28. Nursery pig model-Regions on the swine genome associated with weight gain and virus load ProportionofGenetic Variance ProportionofGenetic Variance 5-SNP window ordered by chromosome5-SNP window ordered by chromosome GWAS Viral Load (AUG) GWAS Weight Gain (WG) SSC 4 SSC 4 SSC X SSC 17 SSC 1 Boddicker, et al. 2012. J Anim Sci. 90:1733-1746.
- 29. Weight gain and virus load locate to the same marker on chromosome 4 WUR10000125
- 30. 5 10 15 20 25 0 1 2 3 4 5 6 7 0 5 10 15 20 25 30 35 40 Weight(kg) Viremia(Log10Templates/ml,qPCR) Days Post Infec on AA AB WUR SNP GenotypeViremia Weight The favorable SSC4 marker, WUR, results in a 10% increase in weight and a decrease in viremia Boddicker, et al. 2012. Anim Sci. 90:1733-1746
- 31. EXON 9 EXON 10 281 bp The unfavorable genotype is composed of exon 9/10 splice variations that create premature stop codons Globular Domain Helical Domain CAAX Stop Guanylate binding protein 5 (GBP5)
- 32. Anti-PRRSV activities of GBP5
- 33. Serum Days post 0 7 14 21 28 42 60 90 Market arrival Serum Serum Serum Serum Field model 200 pigs placed in a “health-challenged” barn NIFA Serum Serum Serum Serum weight 50 pigs200 pigs
- 34. Arriv eVacci- nate Days post vaccination -1 0 4 7 11 14 21 28 32 35 39 42 49 56 63 70 Days post infection 0 4 7 11 14 21 28 35 42 Infect Vaccination response Vaccination + infection response Early response Later response Body weight and Blood samples Dual infection (PCVAD) model 50 pigs AA Non-vaccinated 50 AB 50 pigs AA Vaccinated (PRRS MLV) 50 AB GBP5 (-) GBP5 (WT) = 200 pigs
- 35. Clinical signs, survival, viremia PRRS PCVAD %Survival No MLV MLV
- 37. December 7, 2015 Virus Antibody No CD163 No PRRSV CD163-Positive No CD163
- 38. Oct. 2107
- 39. Reproductive PRRS- Infection of dam at 90 days gestation 109 days gestation
- 41. Prevention of reproductive PRRS Dam Fetus ++ ++ -- +/- -- -/-
- 42. The end of a disease • The absence of CD163 in the dam surrounds the developing fetus with a protective barrier, preventing infection with PRRSV • The piglets are born with normal CD163 levels and are susceptible to PRRSV • Still need that miracle vaccine
- 43. Co-Directors USDA ARS BARC Joan Lunney Kansas State University RRR (Bob) Rowland Iowa State University Jack Dekkers • NIFA award #2013-68004-20362 • National Pork Board • Genome Canada • LLNL • Genus PIC The Rowland Lab