Brief Summary of Minutes of Annual Meeting

8:00 am Dr. Joan K Lunney elcomed everyone to NC1037 annual meeting. As chair, Dr. Lunney presided over the sessions. 8:15 am Business meeting and Election of NC1037 officers. A motion regarding the selection of officers for 2008 was approved. Dr. Jiang (Chair), Dr. Rekaya (Vice-Chair) and Ohio representative (secretary). 8:30 am Dr. Jiang suggested Seattle (Washington State) as location for 2009 meeting. After some discussions, no decision was made. Also there have been some discussions regarding the schedule of the meeting for 2009. Two options were looked at. Regular schedule or at the same time as PAG-2009. 8:50 am Dr. Rothschild , Iowa State University, gave an update on swine sequencing & SNP chip plans. He indicates that a purchase order has to be placed by August 22, 2008 where money will be committed but there is no guaranty on number of chips. He also indicated that an amount of $100 to $110K is already available. After some discussions, it was clear that some policies-rules are needed for the best use of such money. Drs. Rothschild, Hamernik, and Ernst agreed to form a committee to set some rules. 9:30 am Break 10:00 am Updates on USDA bovine genome research - Dr. Zimin gave an update on the assembly of the cow genome o He showed interest for collaboration on the pig genome assembly o Dr. Rothschild and Dr. Cassady suggested being in touch with Dr. Zimin for potential collaboration with the pig genome - Dr. Steven Schroeder from the Bovine Functional Genomics Laboratory (BFGL), BARC, gave an update on the cattle gene atlas - Dr. Matukumalli, BFGL, BARC gave some results on the application of BovineSNP50 for CNVs and genome-wide associations. - Dr. Paul VanRaden from Animal Improvement Programs Laboratory presented an application of Genomic selection in dairy cattle. - 12:00 pm Lunch break 1:00 pm NC1037 Station Reports Chris Tuggle (Iowa state university) Jack Dekkers (Iowa State University) Max Rothschild (Iowa State University) Joe Cassady (North Carolina State University) Rick Barb and Romdhane Rekaya (ARS- University of Georgia) Cathy Ernst (Michigan State University) Le Ann Blomberg (BARC-USDA) Jiang (Washington State University) 3:45 pm Administrative Reports: Deb Hamernik, CSREES, NPL, Animal Physiology; Bert Stromberg, Administrative Advisor; Max Rothschild, National Swine Genome Coordinator Thursday May 15, 2008 8:00 am Review of NC1037 objectives/station reports 1. Discussion of major areas of research in objective 1: Reproduction, growth and development, sow longevity, bone strength and behavior 2. Discussion of major areas of research in objective 2: genetic mechanisms controlling animal health 10:15 am Collaboration and possibility for collaboration - Meat Quality: potential collaboration between Iowa and Michigan. - Growth and Development: Potential collaboration between BARC and Michigan on fetal and neonatal development and between BARC and North Carolina on pre-weaning survival trial with Smithfield, Inc. - Feed efficiency/Intake: Extending the already existing collaboration between Georgia and Iowa on gene expression data analysis, follow up with the MSH/NPY treatment and the used of RFI resources at IA. - Reproduction: Potential collaboration between BARC and Iowa on comparing data for early embryonic development - Behavior: Possibility of joint analysis of chute scores (standardization of scoring systems) 11:15 am Update on PRRS CAP project by J. Lunney - Great potential for collaboration between NC1037 members - Important issue for the industry - Discussion of the potential use of $100K as matching fund 11:45 am Dr. Hamernik suggested adding impact statements to station reports as well as some discussions on CRIS reporting and station reports 11:51 am Meeting adjourned at 11:51 a.m.

Objective 1. Further understand the dynamic genetic mechanisms that influence production efficiency and quality of pork. Genetic improvement of production efficiency and pork quality using traditional selection programs. Iowa State University reported a six-generation selection experiment for intramuscular fat (IMF) based on the average EBV estimated using real-time ultrasound. Of the 149 pigs harvested in generation six, the station found that line LS means for tenth rib backfat and loin muscle area were 16.63 mm and 45.45 cm2 in the control line, and 24.22 mm and 38.02 cm2 in the select line (P < 0.01), respectively. Analysis of STAGES data evaluated on all 621 pigs in generation six revealed significant differences between lines for days to 114 kg, backfat, and loin muscle area. Results through generation six indicate that selection for IMF resulted in slower growth, more tenth-rib backfat, and less LMA. Chemical analysis of a loin sample from pigs harvested revealed a significant phenotypic response in IMF (2.41% in the control line vs. 4.53% in the select line) similar to the difference between lines for IMF EBV. Line LS means for pigs harvested in generation six for 24 h Hunter L and Minolta were 47.00 and 22.17 in the control line, and 49.42 and 24.49 in the select line (P < 0.001), respectively. Subjective measures of marbling were significantly different between lines (2.50 in the control line vs. 4.89 in the select line). Evaluation of correlated response in litter traits after selection for IMF showed that dams in the control line farrowed litters that were 1.22 kg heavier at birth. Individual piglets in control line litters were 0.11 kg (P < 0.05) heavier at birth when compared to piglets farrowed by select line dams. Iowa State University also carried out a selection experiment for residual feed intake (RFI) in Yorkshire pigs, which produced generation 6 in summer 2008. Analysis of feed intake and growth curves showed that differences in RFI between the two lines occur after ~60 kg body weight. The slightly lower ADG observed in the select line also occured primarily after ~60 kg. Selection for litter size continues at the Nebraska State University. Two selection lines (Lines 2 and 45) and two contemporary control lines (Lines 1 and 6) have been maintained in the Station for 27 generations. Line 2 was derived from an Index line that had undergone 11 generations of selection for ovulation rate and embryo survival. This selection was followed by three generations of selection for total born per litter, six generations of selection for number of live pigs per litter, and seven generations of number of live pigs per litter and within litter selection for growth rate and backfat thickness. Line 1 has been randomly selected. The Station report indicated that selection lines 2 and 45 farrowed 5 additional fully formed pigs per litter than their respective controls, but also had greater incidence of stillborn piglets so that the difference in number of live pigs per litter is approximately 4 pigs per litter. Recent selection for growth is causing lines to differentiate in weight and backfat, but not in loin eye area. Improvement of growth appears to be greater in Line 45 than in Line 2. North Carolina State University has started a divergent selection for response to the backtest to study pig behavior, the extent to which it is genetically controlled, and its relationship with economically important traits. During their previous work the Station estimated the heritability of the backtest to be approximately 0.40. The long term goal will be to monitor correlated responses to selection for high and low total time spent struggling during the backtest. Between 7 and 21 days of age each pig will be evaluated using the backtest. A pig will be placed on a padded table in a supine position for 60 s and gently restrained. The experimenters right hand will be placed loosely on the pigs thorax with the right foreleg between the index and middle finger. The hind legs will be held with experimenters left hand and stretched and moved downward to begin the test and the hand will remain loosely on the hind legs throughout the test. Each bout of struggling with at least one hind leg will be recorded as an escape attempt. The number of escape attempts, duration of escape behavior, and latency to the first escape attempt will be recorded. It is expected that this test will lead to a better understanding of the relationship between pig behavior, growth, reproduction, and health. North Carolina Agricultural And Technical State University has recently received a SARE Grant entitled A Multi-Disciplinary Approach to Improve the Environmental Performance of Niche Pork Production Systems and Marketability of Heritage Swine Breeds. This project will investigate niche pork production systems that address market demands and natural resource conservation concerns, with a specific focus on maximizing vegetative ground cover and nutrient distribution in pastures and understanding marketability of heritage breeds produced in alternative production systems. Consumer interest in heritage breed pork continues to rise; howeve,r little is known about taste characteristics and production potential in alternative systems. The Station will also collaborate with an animal nutritionist in the department to develop a collaborative team that will integrate quantitative and molecular technologies for genetic improvement of pigs. The approach will utilize genomic science and bioinformatics tools to aid in the selection of pigs best fitted for outdoor swine production systems, by evaluating production characteristics such as, nutrient utilization, disease resistance and increased litter weights. Quantitative genomics of production efficiency and pork quality using genome scans and candidate gene approaches. Iowa State University has nearly completed a QTL study using the Berkshire x Yorkshire population. The group found QTL on many chromosomes with special emphasis on chromosomes 1, 17 and 18. From there, the station started to find the underlying genes responsible. Fine mapping of QTL on chromosomes 17 and 18 has continued with the addition of >45 genes in the relevant region on chromosome 17 and >10 on chromosome 18 and increasing the marker density to the linkage map. The Station also used comparative studies to examine candidate genes affecting longevity through reproduction and leg soundness. These include genes in the IGF pathway and genes associated with stress and muscle and bone pathways. Over 100 new genes have been mapped and analyses revealed that some of these genes have effects varying from 0.1 to nearly 1 parity more on average and also affect growth, reproduction and longevity. A NPB funded project is underway to look at 2000 sows (1000 old, 1000 young) to examine the effects of genes on leg traits. The group has identified over 100 genes, mapped 75 new ones and identified several genes that have large effects on many leg action and conformation traits. Using commercial populations they are also examining genes on chromosomes 2 and 12 for effects of scrotal hernia and have found several interesting associations. At Michigan State University, a Duroc x Pietrain resource population has been developed to map quantitative trait loci (QTL) affecting carcass composition and pork quality. In the report, they focused their work on confirmation of QTL on pig chromosome 6 by incorporation of marker genotype data for an additional 452 F2 pigs into the QTL analysis. A total of 962 F2 pigs were genotyped for the SSC6 markers S0087, SW122, SW1881 and SW322. Other SSC6 markers genotyped for the original genome scan (510 F2 pigs) included S0099, SW2406, SW2525, S0220 and SW2419. Data were analyzed with line cross least squares regression interval mapping methods using sex and litter as fixed effects and carcass weight or harvest age as covariates. QTL significant at the 5% chromosome-wise level were found for 24 h carcass temperature, Boston shoulder weight and marbling score. These results confirmed previously identified QTL and included four new QTL (carcass weight, first rib backfat, Boston shoulder weight and belly weight). Michigan State University has also targeted porcine insulin-like growth factor binding protein 2 (IGFBP2) for its association with economically important traits in pigs. For this study, 417 F2 pigs from their Duroc x Pietrain resource population were genotyped and the IGFBP2 was placed on SSC15 at 78.0 cM in a region previously found to contain significant QTL affecting meat color and tenderness. Pigs from litters segregating both alleles (N=226 F2 pigs) were used to determine potential associations between IGFBP2 genotype and growth, carcass and meat quality traits. Genotype effects (P<0.05) were found for loin muscle area (LMA) at 10, 19 and 22 wk of age, ADG, carcass length, ham wt, loin wt, 10th rib carcass LMA, 45 min pH, pH decline from 45 min to 24 h postmortem, CIE L*, CIE a*, subjective color score, Warner-Bratzler shear force, sensory panel muscle fiber tenderness and sensory panel overall tenderness. At Nebraska State University, fine mapping of QTL identified in previous scans was accomplished by using phenotypic and genotypic data from pigs of several generations. Phenotypic data were collected for birth weight (BWT, n = 1422), weaning weight (WWT, n = 1311), age at puberty (AP, n = 669), ovulation rate (OR, n = 797), number of fully formed pigs (FF, n = 841), number of pigs born alive (BA, n = 841), number of mummified pigs (MUM, n = 841), number of nipples (NN, n = 1434), incidence of pigs with splaylegs (n = 458), and number of stillborn pigs (SB, n = 841). Age at puberty was recorded in gilts of Lines 1 and 2 from Generation 2 through Generation 15 and in gilts of Lines 4, 5, and 6 through Generation 16. Ovulation rate was recorded in gilts of Lines 1 and 2 through Generation 11, and in gilts of Lines 4, 5, and 6 through Generation 16. Number of fully formed pigs, MUM, SB, BA were recorded in gilts within 24 h of parturition each generation. The Ohio State University has maintained both purebred Berkshire and Landrace populations of swine, which are available to evaluate the influence of pure and reciprocal crossbred lines on growth, carcass, and fresh and cooked pork quality traits of the loin, belly and ham. Data collected in previous years, including full pedigree and well defined contemporary groups are in place and DNA samples from all parent and progeny are available for use in collaborative gene, QTL discovery and or verification studies. The Station encouraged all interested collaborators within NC-1037 to inquire about potential collaboration opportunities. Litters of purebred and crossbred animals are produced in four farrowing groups annually with the ability to produce up to 60 litters per group. Specific aims of future generations will be discovery of mechanisms influencing variation in loin tenderness, a trait likely to have the greatest impact on consumer assessment of pork palatability and overall desirability. Washington State University has worked on the porcine mitochondrial transcription factor A (TFAM). The group determined both full-length cDNA and promoter sequences of the porcine TFAM gene, which were then used for comparative characterization of the TFAM gene with other species. Using Radiation Hybrid mapping the Station assigned this gene to porcine chromosome 14 (SSC14). A G557A substitution in intron 1 of porcine TFAM gene was detected and genotyped on a total of 252 animals, including 165 from seven Chinese and 87 from five Western pig breeds. Bayesian analysis via MCMC (Markov chain Monte Carlo) revealed that these two groups of pigs were well separated at this locus during the breed history; 95% of the posterior difference of TFAM allelic frequency between these two pig groups was greater than zero. As there are marked differences in fatness and lean meat production between Western and Chinese pig breeds, the TFAM gene deserves further investigation in order to evaluate its phenotypic effect on fat deposition and carcass traits in commercial pig populations. Washington Station has also worked on comparative assembly of Sus scrofa chromosome. The Station assigned more than 1,200 clones to their orthologous regions on human chromosomes 6, 9, 14, 15 and 18, which are consistent with the current comparative relationship between these two species. On the other hand, porcine chromosome 1 might have some small regions orthologous to human chromosomes 1, 10, 11 and 20. The comparative analysis provides information on candidate genes for QTL identified on the pig chromosome. Transcriptomics of production efficiency and pork quality using microarray analysis and real time PCR. University of Georgia has used transcriptional profiling to identify genetic mechanisms that respond to alpha-MSH, a MC3/4-R agonist. Three MC4R genotypes (2 homozygous and the heterozygous for MC4R) were selected from the UGA swine herd (PIC composite). Thirty-six pigs (6 per genotype per treatment) were randomly assigned to one of the following treatments: ICV administration of 150 ul 0.9% saline, or 10 µg NDP-MSH (agonist) in 150 ul of 0.9% saline. Feed intake was measure at 12 and 24 hous after treatment (time 0). All pigs were sacrificed 24 hours post-injection and hypothalamus, liver and middle layer of back fat was collected and mRNA was hybridized to 24,123 probe set Affymetrix Porcine Genome Arrays. The group found that MSH suppressed (P< 0.04) feed intake in all animals at 12 and 24 hr after treatment regardless of genotype with no treatment x genotype interaction detected (P > 0.8). There were 5070, 253 and 282 genes that were differentially expressed (P<0.07) in adipose, liver and hypothalamic tissue, respectively after central administration of MSH. The report concluded that the MC4R genotype did not effect the feeding behavior reponse to MSH but the greatest response to MSH was observed in adipose tissue gene expression. Iowa State University has worked on a USDA-NRICGP funded project to determine the genes expressed in both the endometrium and embryo/conceptus during the elongation and implantation phase of reproduction in both the Yorkshire and the Meishan breeds. using Affymetrix Genechip transcript profiling analysis. The endometrial results were analyzed and 47 genes were tested by Q-PCR across several contrasts; approximately 85% of expression patterns across > 200 pairwise comparisons were confirmed. Conceptus data analyzed in conjunction with similar data collected at Oklahoma State University determined that a joint analysis improves the power to detect differentially expressed genes. An analysis of the OSU data on embryonic gene expression during the critical time of elongation has been submitted for publication. In addition to the selection experiment for residual feed intake described above, Iowa State University also performed gene expression profiling for this project and initial analysis of gene expression data has been completed. Michigan State University has evaluated the new Swine Protein-Annotated Oligonucleotide Microarray (http://www.pigoligoarray.org) by analyzing transcriptional profiles for longissimus dorsi muscle (LD), liver, brain and uterine endothelium (UE). The extent of non-specific hybridization was evaluated by comparing intensities of 60 negative control oligonucleotides to background intensity and by estimating the distribution of intensities of non-control oligonucleotides. The study indicated that increasing temperature appreciably improved overall specificity while hybridizations performed at 40 °C showed unacceptable levels of non-specific hybridization. Diagnostics also utilized 60 perfect match/mismatch (PM/MM) oligonucleotide sets designed as part of the array elements (1, 2, 3, 5, 7 and 10 mismatched bases). A small number of negative controls showed consistently high signals suggesting they may not be suitable for use as negative controls. This work was supported in part by the USDA Pig Genome Coordination program. Michigan State University also used the Swine Protein-Annotated Long Oligonucleotide Microarray with pig longissimus dorsi (LD) samples for an expression QTL (eQTL) study. Microarrays have been completed for 176 LD samples, quality assessments of the arrays have been performed and data analysis is in progress. This work was supported by the USDA CSREES National Research Initiative. In collaboration with Dr. Chens Animal Genomics Laboratory at Nanjing Agricultural University, China, Washington State University has worked on porcine nuclear receptor coactivators 1 (NCOA1), 2 (NCOA2) and 3 (NCOA3) for their roles in adipogenesis. The group cloned these three porcine genes, identified their transcript variants and analyzed their expression level in relation to intramuscular fat (IMF) content in Longissimus Dorsi (LD) muscle. The station found that both NCOA1 transcript variant 2 (r=-0.554, p<0.01) and total NCOA1 (r=-0.516, p<0.01) expression levels were negatively correlated with IMF contents, while NCOA2 transcript variant 1 (r=0.605, p<0.01) and NCOA3 (r=0.435, p<0.05) were positively associated with IMF content in LD muscle. A similar study was also performed on porcine sterol regulatory element binding transcription factor 1 (SREBF1) for its role in intramuscular fat (IMF) deposition in pigs. Objective 2: Discover genetic mechanisms controlling animal health in pork production. Nomenclature and identification of new alleles for the SLA complex. USDA ARS BARC has focused their major efforts on updating the nomenclature for the SLA complex in collaboration with Michigan State University and other members of the ISAG SLA Nomenclature Committee. New SLA allele sequences and haplotypes have been designated: 74 new SLA alleles, including 18 SLA-1 alleles, 11 SLA-2 alleles, six SLA-3 alleles, two SLA-6 alleles, one SLA-DRA allele, 20 SLA-DRB1 alleles, three SLA-DQA alleles and 13 SLA-DQB1 alleles; in addition, 12 new SLA class I and 4 new class II haplotypes. These updates are now posted on the Immuno Polymorphism Database-Major Histocompatibility Complex (IPD-MHC) website (http://www.ebi.ac.uk/ipd/mhc/sla) which serves as the repository for maintaining a list of all recognized SLA genes and their allelic sequences. BARC, Japanese, and French scientists have worked on developing new methods for optimizing SLA allele genotyping. The analyses covered genetic polymorphisms in 23 SLA homozygous/heterozygous samples, proved that haplotype-specific patterns and variations of MS existed, and refined recombination points for several SLA haplotypes. These studies affirmed the existence of strong linkage disequilibrium (LD) in the entire SLA region; large haplotype blocks extended across >2-Mb segments. These MS markers constitute an alternative method to characterize the SLA haplotypes. BARC has also developed a simple and rapid method using the polymerase chain reaction (PCR)-sequence-specific primer (PCR-SSP) strategy for direct determination of SLA alleles, specifically, to type alleles at the multiple SLA class I genes and alleles at each of the three classical SLA class I loci (designated SLA-1, SLA-3 and SLA-2). The PCR-SSP typing system was designed with 47 discriminatory PCR primer pairs to amplify the current SLA class I alleles based on groups that have similar sequence motifs. Overall 24 class I allele groups corresponding to 56 class I genotypes were detected; additionally 23 low-resolution SLA class I haplotypes were identified in the 202 pig DNAs tested. This provides a reliable and unambiguous method for detecting SLA class I alleles. The MS and PCR-SSP typing methods provide important alternates to direct determination of the SLA alleles based on sequencing or SNP typing. Identification and mapping of genes responding to PRRS infection. BARC in collaboration with Kansas State and Iowa State has been coordinating the PRRS Host Genomic Consortium. This is a national effort to collect phenotype and genotype of 1000s of commercial pigs for their response to PRRSV infection. The PHGC database is being developed by Iowa State, Kansas State and BARC scientists to house the large amounts of phenotypic and genotypic data that will be collected across several research labs for the PHGC and stored for continued use by the PRRS research community. The schema for the database was originally designed based on data sets generated from the PRRS virus Big Pig project. This included data on pig, sex, birth date, infection details, weights, PRRS viral and serum antibody and cytokine levels. A new PRRS CAP2 grant will support BARC in collaboration with NC229 and PRRS CAP scientists to start SNP genotyping and whole genome association studies with this data. These analyses should reveal associations of PRRS disease resistance and growth traits with SNP alleles. BARC has used the new Pigoligoarray to identify immune regulatory and protective pathways in samples from swine infected with virulent PRRS virus with comparison to samples from vaccinated pigs. In their initial PRRS experiment animals were divided into three groups: (1) pigs infected with a virulent PRRSV isolate MNW2B; (2) pigs vaccinated using a contemporary PRRS ATP vaccine (Ingelvac®); and (3) control pigs. Different tissues were collected between days 3-6 post infection/vaccination. Results obtained for cranial lung tissues affirmed 923, 619 and 747 putatively differentially expressed genes for vaccinated versus control, vaccinated versus infected, and control versus infected, respectively. As expected, pathways involving interferons and other cytokines, as well as chemokines, have been identified as critical for differentiating infected from vaccinated pigs. Final statistical and pathway analyzes are underway to identify the biological functions and regulatory pathways that are most significant. At the University of Nebraska, two replications of a PRRSV challenge experiment with a total of 400 pigs have been conducted. Replication 1 occurred during summer 2002, and Replication 2 in winter of 2003. In each replicate a total of 100 PRRSV-negative, SEW pigs of the NE Index line (Line 2) and 100 pigs of a commercial Duroc-Hampshire (DH) line were used. Line 2, described above, is an inbred Large White-Landrace population that, at the time of the experiment, had been selected for 24 generations for increased litter size. It was expected to have low resistance to disease because of its relatively high inbreeding (~25%). Line DH is a non-inbred, terminal sire line that excels in growth and leanness and was expected to have greater disease resistance than Line 2. Two pigs from each of 200 litters by 163 dams and 83 sires were sampled to ensure genetic diversity within lines to maximize the chance that genes for both resistance and susceptibility to PRRSV existed in the sample. RNA was extracted from lung and lymph tissue of these same pigs and differences in gene expression between resistant and susceptible pigs was determined using microarray gene expression analyses. Identification and mapping of genes responding to Salmonella infection. Iowa State University reported their first study on Affymetrix-based transcriptional profiling in mesenteric lymph node for response to Salmonella infection, as well as a second paper on comparing the expression of genes responding to S. enterica serovar Cholerasuis (SC) versus S. enterica serovar Typhimurium (ST). The group has completed an Affymetrix-based transcriptional profiling study comparing the host response to infection by ST, and planned to submit a manuscript for publication in January 2008. They have identified genes that specifically respond to only one of these two infections, and they have evidence that many genes that respond to SC do not respond to ST infection. The Station has confirmed the microarray data for >20 genes tested by Q-PCR for both SC and ST infections, and has identified many regulatory pathways affected by Salmonella that have been shown to be involved in pathogen response in other species, as well as novel genes that indicates our understanding of pathogen response pathways needs to be expanded significantly. Characterization of the porcine circovirus associated disease complex caused by porcine circovirus type 2. The condition exists at the University of Nebraskas research farm. During the last three generations, the Station has scored all pigs of different lines from weaning through approximately 180 d of age for symptoms of PCVAD. Samples of pigs showing symptoms were submitted for necropsy to confirm typical symptoms in lymph and other tissues. PCR was conducted to confirm presence of circovirus. Pigs of Lines 1 and 2 from Generations 25-27 and from Lines 6 and 45, also from Generations 25-27, were scored for PCVAD at weaning (18 d), and at 60, 90, and 130 d of age. At each age, pigs were weighed and blood samples were collected. Serum collected at each age from all pigs scored positive for PCVAD and from a littermate or penmate without symptoms of PCVAD was submitted to the Iowa State University Veterinary Diagnostic Laboratory where PCV2 ELISA (antibody) and PCR (viremia) diagnostics were conducted. Analyses are complete for 3-4 samples from approximately 1000 pigs. Association of diagnostic data with PCVAD scores and pig weights will be used to determine degree of genetic influence in response to virus.

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