Brief Summary of Minutes of Annual Meeting

Minutes of NC-1170/NRSP-8 Poultry Business Meeting January 10, 2010 Meeting chaired by Huanmin Zhang (NRSP-8 Poultry) and Eric Wong (NC-1170). Business meeting opened at 10:23 AM on Sunday, January 10, 2010. Members present: S. Burgess* (MS), H. Cheng* (ADOL), M. Delany* (CA), J. Dodgson* (MI), D. Foster** (MN), J. Fulton*** (Hy-Line), B.-W. Kong** (AR), S. Lamont* (IA), K. Reed* (MN), D. Rhoads* (AR), Y M. Saif (Administrative Advisor), C. Schmidt (DE)***, J. Song** (MD), E. Wong** (VI), H. Zhang* (ADOL), H. Zhou* (TX) Guests: D. Burt (Roslin/U Edinburgh), M. Hughes (USC), S. Redmond (IA), H.-C. Liu (NCSU), Peter DEustachio (NYU) * = NC-1170 and NRSP-8 Poultry ** = NC-1170 only *** = NRSP-8 only (classification per NIMMS participant list) 1. Comments from Project Administrators 1.a. Dr. M. Saif, Administrative Advisor The projects midterm review will take place this year. Progress reports and minutes must be submitted in timely fashion. The NC advisory committees look for evidence of IMPACT in their evaluation, as well as productivity and cooperation between states, evidence of leveraging of funds from other sources, and strong publication record. Dr. Saif commented that our project performance appears very good to date. 1.b. Dr. M. Qureshi, NIFA Representative Sent his regrets in advance to the membership that he was unable to attend. 2. Old business 2.a. Minutes of 2009 meeting Minutes were approved when the draft was circulated in January, 2009, and were posted in the NIMSS site as submitted. 3. New business 3.a. Next meeting venue. Motion (Delany/Schmidt) to meet on Saturday and Sunday (January 15-16, 2011) preceding the next PAG meeting in San Diego, was approved unanimously. Hans Cheng requested that comments of the program and whole PAG meeting (all aspects) be relayed to him, to pass to the PAG organizers for program and meeting improvement. 3.b. Election of officers NC-1170 Chair. Motion (Dodgson/Delany) that secretary Lamont move to the position of chair for two years was approved. NC-1170 Secretary. Motion (Dodgson/Reed) to elect Rhoads to position of secretary for two years was approved. NRSP-8 Chair. Motion (Rhoads/Delany) that secretary Zhou move to the position of chair for two years was approved. NRSP-8 Secretary. Motion (Reed/Delany) to elect Schmidt to position of secretary for two years was approved. 3.c. Status of potential new members Dr. Saif reported that several new members were added to the project recently and are included in the NIMSS database. 4. NRSP-8 Poultry Business. 4.a. NRSP-8 Poultry Genome Coordinators Report (Dodgson and Cheng). The written coordinators report was distributed in advance of this meeting, and contains the details of 2009 activities. In the past year, the NRSP8 poultry genome coordinators funds have been primarily used to provide seed money for the turkey genome sequence and SNP chips (60K) for the good of the NRSP-8 poultry research community, and about $5,000-7,000 on travel. Dodgson requested that, if not already done, each member send him an estimate of additional funds that were obtained or leveraged by NRSP-8 support, for reporting purposes. The coordinators can provide a little seed funding, but not support a whole project. Input for next years funding usage was requested. The annual budget is anticipated to be about $65,000. Within the objectives of this 5-year term is to improve populations. Discussion on research priorities continued. Do we need more poultry species sequenced? The duck sequence is already done by BGI, with manuscript to be submitted soon by Li Ning. Data are already loaded in ENSEMBL. Burt has sequenced Japanese quail. What about a new chicken genome build? Within the year, UMD will release a new build of the chicken genome (not Wash U, where priorities have changed). Although some additional regions are available now (Z and MHC), it is important to have one official assembly. Schmidt indicated willingness to post Z chromosome data (or other information) on GBrowse. It is currently in GenBank with delay pending publication. Is there any thought in Europe about how to get the missing chromosomes? No, this is not grant-fundable as a stand-alone project. The data are probably already available as a by-product of other work, but this does not cover the assembly and annotation. An activity deleted from a prior proposal was to have FHCRC flow-sort for the small chromosomes to get the sequence. Reed indicated there is money to sequence the turkey, and perhaps they should flow-sort to get good microchromosome sequence. A motion (Burgess/Schmidt) to form a subcommittee to examine the technical aspects of getting the sequence of the small chromosomes and report back to group via NIMSS email was approved. The committee members from NRSP-8 are: Delany, Reed, Cheng, Schmidt, and the following will be invited to advise (ex officio): Burt, Groenen, Warren. 5. Poultry Bioinformatics Subcommittee Report The BirdBase site has been designed and is up for a period of two months for community feedback. The NRSP-8 group will be reminded via NIMSS. Suggestions of additional links are encouraged. 6. Other new business: potential approaches to new AFRI grants program structure Schmidt suggested developing a megaproposal to maintain stocks of chicken and turkey, collect phenotypes, and characterize targeted populations, including informatics. This could address the feeding the world priority, climate change and energy usage (feed efficiency). Poultry is a winner in global feeding the world (more so than swine and cattle). For industry to use the information for genetic improvement, it needs to be out in the public and accessible for use. Saif suggests considering a CAP-like project in the area, for example, of genomics of disease resistance Motion to adjourn (Lamont/Dodgson) unanimously approved. Meeting adjourned at 11:35 a.m. Minutes respectfully submitted by Susan J. Lamont, Secretary, NC-1170, on 12 January 2010.

Objective 1, Create and share data and technology to enhance the development and application of genomics and systems biology in poultry Sequencing, mapping, and functional genomics analysis of the turkey genome are being completed by the consortium (MN). Approximately 93% of the turkey genome has been sequenced with 17x coverage. Annotation identified nearly 16,000 genes and 15,093 recognized as protein coding genes and 611 RNA genes. A BAC-contig based physical and comparative map of the turkey genome has been generated (MI, TX, CA). The turkey autosomal genome is covered by 150 contigs. The BAC-contig comparative map provided the platform with which contigs and scaffolds have been assembled for the first draft of the turkey genome. About 30 rearrangements between the chicken and turkey genomes have been identified. A high density 60K chicken SNP array was developed to evaluate genome-wide marker assisted selection using the Illumina platform. The final array contained 56,702 SNPs and is available to the public (ADOL). The chicken and turkey MHC gene clusters continue to be analyzed. To further the study of the MHC-Y gene cluster, a complex of YF1*7.1 heavy chain and beta2 microglobulin was crystallized and examined at 1.32 angstrom resolution (COH). MN has completed sequencing of BACs that contain portions of the turkey MHC. A survey of sequence variation in the B-locus of commercial turkeys has been completed and found to be highly recombinant. A wider range of sequence variation and haplotype number at the MHC-B locus was observed in wild turkeys compared to commercial turkeys. Objective 2, Facilitate the creation and sharing of poultry research populations and the collection and analysis of relevant new phenotypes including those produced by gene transfer IA maintains many unique chicken research lines that serve as resources for identifying genes and QTLs of economic importance. Financial constraints resulted in the termination of 4 of the 24 lines in 2009. All lines are currently vulnerable to economic extinction. A number of the lines are being used in active collaborations with researchers from Cornell, MSU, U DE, and USDA-ARS. NC continues to characterize chicken primordial germ cells (PGCs). Epidermal growth factor stimulated the proliferation of cPGCs via activation of Ca2+ PKC through the NFKB1 signaling pathway . Retinoic acid was found to induce cPGCs of both sexes to enter meiosis. Studies using retroviral delivery of RNA interference against viral receptor and env proteins of MDV to generate viral resistance in vivo have been completed (MI). Objective 3, Elucidate genetic mechanisms that underlie economic traits and develop new methods to apply that knowledge to poultry breeding practices A number of genome wide SNP-trait association analyses have been conducted. SNPs associated with dermal hyperpigmentation, polydactyly, and other morphological traits such as silkie feathering or hookless, feathered legs, vulture hock, and duplex comb were identified in Silkie chickens (NC). Whole genome SNP analyses were also used to identify genes affecting sperm mobility and ascites susceptibility (AR, NC, ADOL). SNP association analysis for growth in two advanced intercross lines has revealed SNPs within the anti-Mullerian hormone, insulin-like growth factor 1 receptor and neurotensin receptor 1 genes (IA). MD examined an association between SNP and multiple production traits such as abdominal fat and body weight in fat and lean chicken lines. Four candidate genes (superoxide dismutase, aldo-keto reductase, glypican, and syndecan) located in chromosomes with known QTL for abdominal fat were selected. SNPs within the promoter region of these genes were found to be associated with fat yield, fat weight, and breast yield. Genomic selection (GS) studies in egg layer chickens have been conducted at IA. The GS program could obtain substantially greater responses per year than the traditional program by halving generation intervals through early selection using breeding values from dense marker data. A genome wide polymorphism association study has been conducted between broiler lines that differ in their Campylobacter jejuni colonization. A number of SNPs were detected between the resistant and susceptible lines (TX). A phenotypic database using the AR randombred population has been established (GA). These traits are grouped into 7 categories: growth, feed intake/feed efficiency, nutrient digestibility and retention, body composition, meat quality, physiological traits or hormones such as IGF1, IGF2, insulin, T3, T4, glucagon, ghrelin, and GH, and skeletal traits (GA). Association of SNPs with IGF1 and IGF2 was also conducted. The IGF-1 SNP was associated with residual feed consumption, whereas the IGF2 SNP was associated with body weight gain and feed intake. IA completed a comparison of Chi-Square, ANOVA, and MANOVA for analysis of high density SNP data from case-control pools. ANOVA was found to be an effective substitute for the Chi-square test to identify significant SNPs and MANOVA can be used to identify the joint effect of region in case-control studies across lines. Genomic selection using breeding values (GS-EBV) estimated from dense marker data is a promising approach for genetic improvement (IA). Expression analyses of economically important genes are being studied using DNA microarrays or real time PCR. NC examined the expression of genes in the duodenum of turkey embryos using a focused array. At hatch, there was an increase in enzymes to digest disaccharides and transporters to absorb monosaccharides and small peptides (i.e., PepT1). VA continues to study the regulation of expression of the peptide transporter, PepT1. The nuclear receptor peroxisome proliferator activated receptor alpha gene may play a role in the regulation of PepT1 gene expression, during fasting. NC continues to investigate the molecular basis for the sex-linked barring gene. A region of the Z chromosome was found to be significantly associated with a defect in PGGT1B, a member of the gene family involved in prenylation, a post-translational modification essential for normal melanocyte function. Genes involved in the photoneuroendocrine system were investigated in male chicks following transfer from a short to long photoperiod or treatment with sulfamethazine (SMZ), a compound that affects early sexual maturation.. Long day length in combination with SMZ significantly elevated FSH beta mRNA transcripts and increased plasma levels of FSH (protein). Long day length alone significantly increased FSH beta mRNA transcripts but not plasma FSH. Therefore SMZ was shown under the experimental conditions to release protein (FSH) into the vasculature. In contrast, prolactin mRNA transcripts and plasma prolactin concentration were stimulated solely by long day length (AR). AR investigated genes involved in the stress response. A synergistic release of the stress hormone corticosterone following peripheral administration of corticotrophin releasing hormone and arginine vasotocin appears to involve both peptides binding to their respective receptors juxtapositioned on the same pituitary corticotropes forming heterodimers. Differential expression of muscle genes in turkeys at different developmental stages has been identified (MN, OSU, MSU). MN continues to investigate the underlying genetic predisposition of turkeys to ascites and round heart disease. MN is also investigating the molecular mechanisms and genetic variation underlying aflatoxin resistance in turkeys. The role that GST genes play in this resistance is being studied. Proteomic profiling of chicken splenocytes exhibiting natural killing activity has been initiated (COH). The problem of optimizing two stage transcriptional profiling experiments involving microarrays (Stage 1) and qRT-PCR (Stage 2) in order to maximize sensitivity while controlling false discovery rate was investigated. A novel method for the analysis of relative quantification of qRT-PCR data was examined using a general linear mixed model methodology. This new method is especially useful for studies involving multiple experimental factors and complex designs. (WI) Other studies included a dynamic linear model for quantitative genetic analysis of longitudinal traits. The accuracy of prediction of phenotypes for complex traits in poultry was studied in a genome assisted context. Research continues on Mareks Disease (MD). ADOL continues to identify genes that confer resistance to MD. Utilizing QTL analysis, DNA microarrays, and 2-hybrid screens, three MD resistance genes (GH1, SCA2, and BLB) have been identified. An alternative approach using allele specific screens, has revealed many genes that respond to Mareks Disease Virus (MDV) infection, such as genes involved in T cell activation, vesicle mediated transport and cell cycling pathways. The role that the transcription factor, Meq plays in MDV continues to be investigated. Proteins that bind to Meq have been identified. The cell cycle regulatory protein CWF19L2 interacts with Meq and may be one mechanism for MDV induced transformation. The role that stem cell antigen 2 (SCA2) plays in MD resistance was further investigated. SCA2 was found to influence MDV replication and spread and is dependent on the presence of MDV US10, which expresses an enhanced EGFP fusion protein. Genetic resistance to MD consists of both MHC and non-MHC genes. Using unique genetic lines at ADOL, statistical analyses showed that both vaccination and genetic line were important main effects that affected MD incidence and survival post infection. Epigenetic alterations induced by MDV exposure is being examined by MD. Methylation patterns and array based comparative genomic hybridization data have been analyzed. A 225 bp insert in the 3 untranslated region of the MHC-B haplotypes has a large effect on the occurrence of MD. The incidence of MD was 47% in congenic birds bearing the BG1*R4 allele and only 19% in congenic birds bearing the BG1*R2 allele (COH). Analysis of telomeres-telomerase regulation and function in MDV is ongoing (CA). The timing of MDV integration into the host chromosome has been investigated. At 7 days post infection, the MDV integrates into the genome. The integration profile is not uniform within or among tissues. Studies of the molecular mechanism of infection or resistance/susceptibility for other avian diseases caused by avian metapneumovirus (AMPV), avian influenza virus (AIV), infectious laryngotracheitis virus (ILTV), Salmonella and Eimeria have been conducted. An infectious AMPV that contains biomarkers was developed using reverse genetics to use as a vaccine virus. Individual AMPV genes were cloned and cotransfected into cells in order to produce infectious virions that contain the biomarkers (GFP and 6xHis tag). This allows the biomarker AMPV vaccines to be readily detected from wild type AMPV. TX has examined associations between polymorphisms in the chicken Mx gene with antiviral responses in avian influenza infected embryos and broilers. A non synonymous substitution (S631N) in the chicken Mx gene has been reported to be associated with resistance to AIV. There was a tendency for NN genotypes to have lower virus titer than SS gentoypes. An integrated analysis of micro RNA expression and mRNA transcription in AIV infected chicken lungs was examined. There were 91 miRNAs and 146 genes that were differentially expressed between infected and non infected chicken lungs (TX). AR has identified and characterized the expression of seven ILTV encoding microRNAs. Immunogens that protect against cellulitis in turkeys are being developed at MN. Recombinant fusion proteins of the alpha subunit of Clostridium septicum will be used as a vaccine in poults to reduce C. septicum exotoxin and thus reduce cellulitis. A comparison of differentially expressed genes in bursa between chickens infected with wild type Campylobacter jejuni and non-infected chickens was conducted (TX). There were 2591 and 2936 genes significantly changed between 1 and 4 hours post innoculation in infected and non-infected chickens. Genome wide transcriptome profiles of chicken macrophages have been completed using DNA microarrays following exposure to Salmonella derived endotoxin (IA). 10% of the differentially expressed genes were involved in the inflammatory response. NFKBIA, IL-1B, IL8, and CCL4 genes were consistently induced following endotoxin treatment (IA). Differential expression of intestinal genes following an Eimeria maxima challenge has been investigated (VA). Downregulation of the liver expressed antimicrobial peptide-2 (LEAP-2) gene, which is part of the innate immune system, was found to be correlated with high lesion scores in chicks. IN continues to investigate physiological mechanisms associated with aggression and stress in poultry using a functional genomics approach. Two lines of chickens, KGB and Dekalb, have been subjected to environmental stressors and the transcriptome response in the hypothalamus has been measured by microarray. Involvement of dopamine and serotonin pathways was revealed. Genomic selection has been used to select for animal well being concerns, such as livability in layers and leg angle in broilers. IN is also mapping QTL for growth using Paul Siegels high and low weight selected lines. Four interacting QTLs have been identified and are predicted to cause nearly half of the 9-fold difference in body weight. These 4 QTL alleles from the low line are being introgressed into the high line using marker assisted back-crossing to the high line. Telomere length variation of different genotypes and cell lines is an ongoing project (CA). Significant variation exists within and among chicken genotypes for chromosome-specific telomeric array organization and total genome telomeric sequence content. The molecular basis for the wingless-2 mutation continues to be investigated with attention focused on the TSEN-2 gene, which plays a role in the removal of introns from specific pre tRNAs.