Progress 10/16/12 to 10/15/17
Outputs
Progress Report Objectives (from AD-416): Objective 1: Complete development and implementation of a publically available Version 2.0 of the Animal-GRIN database and initiate planning and development of a genomic component for the database: Version 2.1 of the Animal-GRIN. Objective 2: Employ current genomic and quantitative analyses to characterize genetic diversity in the germplasm collection and associated populations to guide additional germplasm sampling and diversity analyses for domestic and international germplasm stakeholders; focus on rare and economically important traits, breeds, species and populations. Objective 3: Refine cryopreservation, germplasm evaluation and reproductive technologies that facilitate efficient germplasm collection and its utilization by various livestock industries. Approach (from AD-416): Globally, 20% of the world�s livestock breeds are �at risk� of extinction -- such a contraction limits the flexibility of livestock producers to respond to future biological or economic challenges. Underlying this contraction of genetic resources are financial pressures faced by the livestock breeder coupled with greater organization of genetic selection programs. Public sector institutions are also faced with mounting pressures to reduce or eliminate research populations. This trend is of particular concern as research moves from genetic sequencing to understanding how genes function and translate into phenotypes. This project plan addresses genetic security by continuing the development of germplasm and tissues collections for all major livestock species in the U.S., so that industry and the research community can access these resources at any time. Three primary objectives address: database development, assessing in-situ and germplasm collection genetic diversity, and improvement in the efficiency and efficacy of cryopreservation protocols. All three of these objectives are highly interdependent and are required for the functionality of the repository. The expected results from this plan are a greater level of genetic security for U.S. livestock populations; information about the collection so researchers and industry can readily access the material; and improved cryopreservation protocols and germplasm evaluation methods. The successful execution of these objectives translates into greater national and global food security and greater economic vitality of the U.S. livestock sector. This is the final report for the project 3012-31000-005-00D. Research will continue on in a new project 3012-31000-006-00D, �National Animal Germplasm Program�. Significant progress in conserving animal genetic resources during the 2012 to 2017 project life cycle has been achieved with the collection growing to 929,000 samples from 48,600 animals (Objective 3a). Of the breeds or special populations in the collection, 117 have reached the minimum criteria to be considered secure and available for reconstituting a breed or population if necessary. As a result the number of animals and samples in the collection increased 105% and 14.7% respectively. The increase in animal numbers was, in part, due to the implementation of ovary and testes cryopreservation for chicken breeds. All milestones for information system development were accomplished. The second version of the publicly accessible Animal- Germplasm Resources Information Network (GRIN) database was completed and put into production (Objective 1). The genomics component for the database was also put into production, and that element of the database system contains genotypic information on more than 2,000 animals. An array of studies was completed that evaluated various aspects of genetic diversity for cattle, swine, and goats. Of particular note was the evaluation of genetic markers associated with adaptation to environmental challenges such as temperature and humidity among U.S. ecoregions for Hereford cattle. It was found that distinctive subpopulations have developed for each ecoregion, suggesting that the forces of selection are impacting various traits associated with adaptation to different production environments. Evaluating genetic diversity among swine populations we determined that feral populations of pigs on Hawaiian Islands and Guam had unique Y chromosome genetic structure not found in continental U.S. or Chinese pig breeds (Objective 2a). Across livestock species our work suggests that the U.S. has a vast array of genetic resources for industry to use, and they do use this diversity to lead the world in superior lines of cattle, chickens, and pigs. Rare breeds are often raised by producers with little access or experience with reproductive technologies, making it difficult to acquire germplasm from their livestock. A range of experiments were conducted to improve the shipment of beef and swine ovaries and the subsequent harvesting of oocytes from that tissue (Objective 3b). Implementation of a protocol for shipping beef cattle ovaries was developed. Using a similar protocol for pigs was shown to be possible and was evaluated using shipped ovaries from Guinea Hogs. The composition of the plasma membrane of trout and bull sperm was evaluated to determine if membrane composition had a measurable impact on measures of sperm motility. Differences have been found and at this point the data suggest regulation of intracellular calcium concentration could be a useful indicator for sperm motility. ARS scientist at the National Animal Germplasm Program in Fort Collins, Colorado provided expert technical input into U. S. government positions on livestock and aquatic genetic resources in conjunction with the State Department and Foreign Agricultural Service of USDA. In addition ARS represented U. S. governmental positions at the Intergovernmental Technical Working Group on Animal Genetic Resources at the Food and Agriculture Organization of the United Nations (Objective 2d). Accomplishments 01 Collection use - reconstitution of lost genetic resources. ARS is well on the way of developing a comprehensive collection of animal genetic resources, and already the collection has had tremendous positive impact on the livestock industry and the research community. To date over 6,000 animals samples in the collection have been requested by industry and the research community for different purposes, including: genomic evaluation, reconstitution of research populations, adding genetic variation to populations of rare breeds, and corrective mating. Three examples of the profound impact are: 1) Purdue University acquired pig germplasm samples to reconstitute a research line of pigs known to affect meat quality, which resulted in substantial funding and more than 10 scientific articles and prompted Virginia Tech to establish a second research population of this pig line. 2) The Angus Association obtained a sample determined to be free of a lethal mutation, which meant more than 29,000 other cattle did not have to be genotyped thereby saving the Association approximately $2 million. 3) In collaborative research, ARS scientists in Fort Collins, Colorado and researchers at Pennsylvania State University, determined there were only two different Y chromosomes in the Holstein population (our major milk producing breed) and that there were two additional Y chromosomes in the collection that were not identified in the present Holstein population. Thus, ARS and Penn State scientists worked with industry to produce bull calves with these two Y chromosomes from the collection as a first step to re-introducing the lost Y chromosomes and protecting the fertility of the Holstein population. These examples demonstrate the importance and utility of the germplasm collection to the U.S. livestock sector, not only for use in emergencies but also as a tool for industry and researchers to use in their efforts to solve a wide range of livestock industry problems. 02 Innovation fund facilitates development of animal-germplasm resources information network (GRIN) webpages. Animal-GRIN webpages were developed in 2006 and had not been updated, thereby not taking advantage of new and innovative webpage design. ARS researchers in Fort Collins, CO designed, oversaw development and implementation of new Animal-GRIN webpages. This improvement should maximize the learning opportunity of the webpage visitor, finding the desired information easier, and better demonstrate the breadth of the animal germplasm collection.
Impacts
(N/A)
Publications
- Blackburn, H.D., Abdelrahman, H., Bosworth, B.G., Leeds, T.D., Lui, Z., Waldbieser, G.C., Rexroad III, C.E., Tiersch, T. 2017. Aquaculture genomics, genetics and breeding in the United States: Current status, challenges, and priorities for future research. Biomed Central (BMC) Genomics. 18:191.
- Blackburn, H.D., Krehbiel, B.C., Ericsson, S.A., Wilson, C.S., Caetano, A., Paiva, S.R. 2017. A fine structure genetic analysis evaluating ecoregional adaptability of a Bos taurus breed (Hereford). PLoS One. 12(5) : e0176474.
|
Progress 10/01/15 to 09/30/16
Outputs
Progress Report Objectives (from AD-416): Objective 1: Complete development and implementation of a publically available Version 2.0 of the Animal-GRIN database and initiate planning and development of a genomic component for the database: Version 2.1 of the Animal-GRIN. Objective 2: Employ current genomic and quantitative analyses to characterize genetic diversity in the germplasm collection and associated populations to guide additional germplasm sampling and diversity analyses for domestic and international germplasm stakeholders; focus on rare and economically important traits, breeds, species and populations. Objective 3: Refine cryopreservation, germplasm evaluation and reproductive technologies that facilitate efficient germplasm collection and its utilization by various livestock industries. Approach (from AD-416): Globally, 20% of the world�s livestock breeds are �at risk� of extinction -- such a contraction limits the flexibility of livestock producers to respond to future biological or economic challenges. Underlying this contraction of genetic resources are financial pressures faced by the livestock breeder coupled with greater organization of genetic selection programs. Public sector institutions are also faced with mounting pressures to reduce or eliminate research populations. This trend is of particular concern as research moves from genetic sequencing to understanding how genes function and translate into phenotypes. This project plan addresses genetic security by continuing the development of germplasm and tissues collections for all major livestock species in the U.S., so that industry and the research community can access these resources at any time. Three primary objectives address: database development, assessing in-situ and germplasm collection genetic diversity, and improvement in the efficiency and efficacy of cryopreservation protocols. All three of these objectives are highly interdependent and are required for the functionality of the repository. The expected results from this plan are a greater level of genetic security for U.S. livestock populations; information about the collection so researchers and industry can readily access the material; and improved cryopreservation protocols and germplasm evaluation methods. The successful execution of these objectives translates into greater national and global food security and greater economic vitality of the U.S. livestock sector. Development of a genomics component for Animal-GRIN V2 is proceeding ahead of schedule (Objective 1). The genomics component has been designed, coded, and tested using genomic data from ARS, universities, and a beef breed association. We will continue to test data entry and retrieval procedures during the next year. The network of programmers (ARS/NAGP, Canada and Brazil) was increased with the addition of computer programming support from Louisiana State University. Future efforts will focus on developing better user interfaces. Germplasm and tissue collections grew across species by approximately 1. 5% during the last year (Objective 3a). The collection now consists of more than 870,000 samples from 36,800 animals. Over 2100 animals in the collection were used to add genetic variation to a population or for genomic analyses. Important additions to the collection were made: Atlantic Salmon (Salmonidae Salris Salar), oyster tissues (Ostreidae Crassostrea virginica), yak (Bos grunniens) and honey bees (Apis melifera) . With the addition of Yak all branches of the genus Bos are now represented in the repository and two (Bos and Bison) of the three branches of taxonomic family Bovidae. In addition, there are now five branches of the family Salmonidae in the collection. Four breeds reached minimum collections goals and can be classified as secured (Polypay and St Croix sheep; and Crevecoeur and Aseel chickens). The St Croix, Crevecoeur and Aseel are classified as rare breeds. Genetic diversity evaluation of the swine collection and feral swine populations on Pacific islands were continued by exploring genotype differences (which serve to identify parental linages) on the Y chromosome (Objective 2a). Two findings stand out. First, feral pig populations on the islands of Guam, Hawaii, and Kauai were primarily from Asia and likely brought to these islands by different waves of Polynesian migration. Second, among several major and rare breeds (Duroc, Guinea Hog, Hampshire, Tamworth, and Yorkshire) sampled from the continental U.S. had only one haplotype on the y chromosome. It is hypothesized that this haplotype predates the formulation of these breeds and a common ancestral population. Previous genetic diversity studies for oysters utilized microsatellites, however the genomics community has been migrating toward the use of single nucleotide polymorphisms (SNP). Using SNPs have a number of advantages (e.g., datasets can be combined), therefore we developed with a contractor a new small SNP panel (210 SNPs) for Ostreidae Crassostrea virginica oysters (Objective 2c). With this new tool we will be able to quantify genetic differences of this species of oyster in different geographic regions along the Atlantic coast. Genotyping oysters along the various coastal regions of Louisiana will be completed this year and allow ARS to explore genetic differences and the presence of genetic by environmental interactions as they may be associated with climate change. An ARS scientist at the National Animal Germplasm Program in Fort Collins, CO provided expert technical input into U.S. government positions on livestock and aquatic genetic resources in conjunction with the State Department and Foreign Agricultural Service of USDA. In addition ARS represented U.S. governmental positions at the Intergovernmental Technical Working Group on Animal Genetic Resources at the Food and Agriculture Organization of the United Nations (Objective 2d) . ARS researchers in Fort Collins, CO were able to identify optimal culture conditions prior to freezing, freezing methods and freezing devices to produce frozen-thawed oocytes with a high enough quality that they could be used for in vitro fertilization in cattle (Objective 3b). Furthermore, because of these findings the National Animal Germplasm Program is able to collect and transport oocytes to our laboratory within 24 hours while maintaining a high sample quality. This research enables us to build a secure, quality collection of oocytes from cattle and other mammalian species for inclusion in the germplasm collection. Quantifying semen sample quality for the ability to fertilize gametes impacts the efficiency of collection development and assisting users in obtaining viable offspring from the collected germplasm. Using flow cytometry and spectrofluorometry, we investigated the effects of sperm activation on plasma membrane phospholipid organization, intracellular calcium concentrations, and protein organization and determined how these relate to trout fertility. The analyses demonstrated that when samples from multiple trout are compared, individuals with higher and lower chances of fertilizing eggs in vitro, based on their plasma membrane phospholipid organization and intracellular calcium concentrations, can be identified. Application of these findings will be used to evaluate samples in the repository so that users can achieve highr levels of fertility from the genetic resources obtained from the collection. Accomplishments 01 Gene banking livestock pays dividends. Development of a genetically diverse collection of animal genetic resources secures our most important populations of livestock and affords industry and the research community with opportunities to use the collection for a wide range of purposes. For example, a lethal mutation was found in a prominent U.S. cattle breed. Animals carrying the mutation can be determined using a genetic test, and if found to be carriers the animal can be identified as such and not used for breeding. The breed association was able to obtain the only known tissue sample of a prominent bull from the ARS collection. The sample was genotyped and it was determined that the targeted bull did not carry the gene for the lethal disease. As a result the producers did not have to genotype 29, 000 other cattle related to the bull and this saved the association and breeders approximately $640,000. This is an example of how genetically diverse collections held in a USDA gene bank can be used to address current problems confronting the livestock industry. 02 Genomics component of Animal-GRIN Version 2 information system introduced. Prior to the development of this software the livestock research community did not have any mechanism to permanently store genotypes derived from publically funded livestock genomics work. As a result the long-term security of expensive data was at risk and it was difficult for other researchers to access and leverage this information in other experiments. An ARS led team developed a genomics component which is part of Animal-GRIN Version 2. With this component it is possible for public sector researchers to enter their genomic data into the database and the community at large to have access to those genotypes. The developed component also makes possible, for the first time, the linkage and access to genomic, phenotypic, management, and environmental information with a physical tissue sample from the individual animal. This approach serves to protect the public investment in genomic research and affords a broad range of users with the opportunity to leverage previous investments made in genotyping.
Impacts
(N/A)
Publications
- Carvalho, C.M., Paiva, S., Araujo, A., Mariante, A., Blackburn, H.D. 2015. Genetic structure of goat breedsfrom Brazil and the United States: Implications for conservation and breeding programs. Journal of Animal Science. 93:4629-4636.
- Purdy, P.H., Wilson, C.S., Spiller, S.F., Blackburn, H.D. 2015. Biobanking genetic resources: Challenges and implementation at the USDA National Animal Germplasm Program. Reproduction, Fertility and Development. 28:1072- 1078.
- Purdy, P.H., Barbosa, E.A., Praamsma, C.J., Schisler, G.J. 2016. Modification of trout sperm membranes associated with activation and cryopreservation. Implications for fertilizing potential. Cryobiology. 73:73-79.
- Inskeep, K., Keisler, D.H., Purdy, P.H. 2016. Reproduction. Pp. 1000-1044. In: American Sheep Industry Association Inc (eds.). Sheep Production Handbook. Volume 8. Book Chapter. ADS Publishing, Fort Collins, Colorado.
|
Progress 10/01/14 to 09/30/15
Outputs
Progress Report Objectives (from AD-416): Objective 1: Complete development and implementation of a publically available Version 2.0 of the Animal-GRIN database and initiate planning and development of a genomic component for the database: Version 2.1 of the Animal-GRIN. Objective 2: Employ current genomic and quantitative analyses to characterize genetic diversity in the germplasm collection and associated populations to guide additional germplasm sampling and diversity analyses for domestic and international germplasm stakeholders; focus on rare and economically important traits, breeds, species and populations. Objective 3: Refine cryopreservation, germplasm evaluation and reproductive technologies that facilitate efficient germplasm collection and its utilization by various livestock industries. Approach (from AD-416): Globally, 20% of the world�s livestock breeds are �at risk� of extinction -- such a contraction limits the flexibility of livestock producers to respond to future biological or economic challenges. Underlying this contraction of genetic resources are financial pressures faced by the livestock breeder coupled with greater organization of genetic selection programs. Public sector institutions are also faced with mounting pressures to reduce or eliminate research populations. This trend is of particular concern as research moves from genetic sequencing to understanding how genes function and translate into phenotypes. This project plan addresses genetic security by continuing the development of germplasm and tissues collections for all major livestock species in the U.S., so that industry and the research community can access these resources at any time. Three primary objectives address: database development, assessing in-situ and germplasm collection genetic diversity, and improvement in the efficiency and efficacy of cryopreservation protocols. All three of these objectives are highly interdependent and are required for the functionality of the repository. The expected results from this plan are a greater level of genetic security for U.S. livestock populations; information about the collection so researchers and industry can readily access the material; and improved cryopreservation protocols and germplasm evaluation methods. The successful execution of these objectives translates into greater national and global food security and greater economic vitality of the U.S. livestock sector. Animal-GRIN Version 2 was released last fiscal year and has reached full implementation in the U.S. and Brazil during FY15. Under Objective 1 a user�s manual has been prepared for publication. System use and the User�s Manual have been tested on various users not familiar with the system�s development. The development of the genomics database component Animal GRIN Version 2 under the specific cooperative agreement with Colorado State University has progressed ahead of schedule. Development has reached a point where genomic data in different formats can be up and down loaded and validated for accuracy. For the first time a large and significant proportion of a gene bank�s swine germplasm collection, representing 18 breeds, has been genotyped using the 60K SNP chip for evaluation of genetic diversity and comparing the collection to in-situ populations (Sub-objective 2a). Included in the breeds genotyped were three Chinese pig breeds and samples from feral pigs on Guam and Hawaiian islands. Results to date suggest major breeds (Hampshire, Yorkshire and Duroc) are genetically distinct among themselves and other breeds evaluated. The rare and minor breeds Hereford, Tammworth, Large Black, Guinea Hog, and Ossabaw Island were found not to be as distinctive from each other as originally assumed. The feral populations were found to be distinct, as well as, showing some association with the Minzhu breed from China. These finding suggest there are unique genetic resources on the U.S. territorial island in the pacific warranting additional collection activities. Two analyses determined the impact of genetic drift and/or selection on farm animal breed evolution, population differentiation and distinctness (subobjective 2b). These were: an evaluation of U. S. and Brazilian Braford, a relatively new composite breed of cattle, using a customized SNP panel; and determination of allelic frequency differences for SNPs associated with adaptation to climate among Hereford cattle raised in five U.S. ecoregions (in coming agreement 49479). Major findings for composite breed formation show that the proportional composition of parental breeds has remained stable and as the breed grew older homozygosity increased. Using SNPs it was possible to determine how popular sires can change breed structure, which also confirms our pedigree based evaluations. Genetic differences, based upon allele frequencies were evaluated by ecoregions subpopulations of Hereford cattle were identified. Ecoregions with the largest differences in allele frequencies were the warm-arid and warm-humid zones and suggests cattle from these regions may have adapted to the climates where they were produced. The transition-zone (e.g, Kentucky, Tennessee) was interesting as cattle in that region tended to be intermediate in allele frequencies for the SNPs associated with environmental stress. These findings suggest substantial genetic variability of environmental stress exists in the breed and this variability can be used to adapt cattle to climate change. Gene banks need to better quantify genetic differences among the germplasm collection given the substantial numbers of animals collected (sub-objective 2c). Therefore, small SNP panels which can identify diverse populations and provide insight about the genetic diversity of production characteristics are important. Initial analysis to develop a small SNP panel started with the identification of SNPs associated with litter size in sheep. A panel of 43 SNPs was selected based upon known polymorphisms in major genes affecting prolificacy (GDF9, BMP15 and BMPR1B). The identified SNPs were tested on samples from 188 animals from 15 breeds in the gene bank. Additional polymorphisms mined from the International Sheep Genome Consortium were used. Ten SNPs were found to be polymorphic. One SNP with a known mutation associated with prolificacy was observed in the Polypay breed. The remaining nine polymorphic SNPs were located in exons and five of them with amino acid change but no association with prolificacy identified as of yet. The derived panel represents the first time a multi-genic panel for prolificacy was used and researchers may find it useful in studying genetic/allelic interaction governing prolificacy and as a mechanism to improve the characterization and management of gene bank sheep collections at low cost test. During FY15 the US country report on US animal genetic resources was developed and submitted to FAO for inclusion into the 2nd State of the World�s Report on Farm Animal Genetic Resources (Sub-objective 2d). Additionally, the chapter entitled �Conservation� was co-authored as part of the report. An interagency meeting was held to discuss USG response to the FAO led effort to assess the status of aquatic genetic diversity. Technical advice was provided to DOS concerning the impact of various reporting mechanisms on biodiversity which were being negotiated upon in various fora. National Animal Germplasm Program Species Committees met at various venues and provided input into collection development and program activities. A key meeting among committee chairs was held in Baton Rouge, LA, in addition to the regular program of work, the honey bee community were invited to the meeting to discuss how to coordinate making the collection of bee germplasm operational. (Sub-objective 2e). There has been a substantial increase in the collection (Subobjective 3a) of rare chicken breeds due to the ability to harvest and cryopreserve ovaries and testes. Filling gaps in the pig collection continued across a wide range of breeds. An important acquisition of semen from the United Kingdom was made for the Large Black and Gloucestershire Old Spots breeds. The addition of tissue samples from feral pigs in Hawaii and Guam to the collection facilitated evaluating diversity of US pig breeds (see Subobjective 2a). The first Atlantic salmon from Maine also entered the repository in this fiscal year. Developing a germplasm collection of mammalian female gametes or embryos increases the robustness of the germplasm collection. While embryos can be routinely harvested and frozen they are costly and difficult to obtain. Oocytes may provide a less expensive option and offer more flexibility in creating an embryo but it has been unclear whether slow freezing or vitrification should be used and what role the maturity of an oocyte plays in the cryopreservation process (Subobjective 3b). Experiments were designed to explore these issues with cattle oocytes. The cryoprotectants ethylene glycol, dimethylsulfoxide and a combination of the two were used in slow freezing and vitrification techniques. Each combination of cryoprotectant(s) and freezing methods was tested with 24 hour matured and immature oocytes. Viability of treatment options are based upon fluorescence microscopy as well as in-vitro fertilization rates and embryonic development. The results of these experiments will provide insight about the cryopreservation protocols and the preservation of oocytes at different levels of maturity. Understanding the physiologic interactions of cells with the medium used to freeze them is an essential part of understanding how to properly cryopreserve and utilize a sperm sample for artificial insemination (Subobjective 3c). Using methods to evaluate cellular quality which were developed in our laboratory (plasma membrane phospholipid and protein organization, calcium activity, protein tyrosine phosphorylation), which were established in previous years using ram sperm, we were able to identify affects attributed to the type of medium used for boar sperm cryopreservation that result in differences in cellular function, quality and fertilizing potential. Knowledge gained from these analyses is now being applied to additional species to increase our understanding of their sperm physiology and to increase their fertilizing potential when frozen-thawed sperm is used for artificial insemination. Our use of flow cytometry to evaluate cellular quality is effective but because this requires the analysis of multiple characteristics unique to sperm at once it would be convenient to minimize the number of analyses per male by combining analyses into one sample with multiple fluorescent stains (Subobjective 3d). Multiplexing, which is the combination of fluorescent stains to identify multiple cellular characteristics in one analysis, proved to be achievable but because of a limited availability of stains across the fluorescent spectra and because of the limitations of our flow cytometer we deemed this to be inefficient and not cost effective. Still, the investigation of these stains enabled us to identify and optimize the use of three additional stains that evaluate plasma membrane phospholipid, or protein organization and protein tyrosine phosphorylation and have proven valuable in identifying key aspects of sperm physiology. The use of these fluorescent stains is under further investigation and our hope is that they will continue to prove simple to use and valuable for assessing sperm quality and physiology across species. Accomplishments 01 Implementation of Animal-GRIN Version 2 information system. A multi- national internet based information system for animal genetic resources was launched by ARS scientists in Fort Collins, CO in collaboration with scientists from Empresa Brasileria Pesuisa Agropecuaria, and Agriculture and Agri-Foods Canada. The information system allows gene bank managers, government and university scientists, and various stakeholder groups to view germplasm and tissue collections for potential use. Gene bank managers can use the system to assess the status of the collection and identify collection gaps and thereby plan future collection activities. This new version has the ability to compare production parameters of animals with samples in the collection and living populations. Furthermore, this version of the system will allow the gene bank managers in the three countries to compare various aspects of the collection. 02 ARS scientist, various university and industry customers developed the U.S. country report for FAO�s 2nd State of the World�s Animal Genetic Resources. The forth coming publication from FAO will provide a global assessment the conservation and utilization of farm animal genetic resources. In addition, an ARS scientist in Fort Collins, CO and collaborator from Wageningen University co-authored the Conservation chapter for the 2nd report. This chapter will provide a current assessment of the science and application of in-situ and gene banking practices for the global community to utilize. 03 Genetic diversity assessment of pigs. ARS scientists in Fort Collins, CO collaborated with scientists from Empresa Brasileria Pesuisa Agropecuaria, via a Brasilian-ARS exchange program, to compare the genetic diversity of U. S. major, minor or rare breeds plus feral pigs from Hawaii and Guam with three Chinese breeds. Results showed a unique genetic structure among the major commercially important breeds. In addition, the data suggest feral pigs in Hawaii and Guam originated from two places in Asia. These findings suggest the feral populations on Pacific islands may possess unique alleles or gene combinations which could be useful to mainland production. 04 Subpopulations of Hereford cattle exist in varying ecoregions. Using single nucleotide polymorphisms previously identified as having a role in an animal�s ability to cope with environmental stress, an assessment was performed that identified subpopulations of Hereford cattle in specific ecoregions of the United States. The findings suggest that as the full impact of climate change occurs there are groups of cattle which may have a genetic structure that enables them to better cope with such changes.
Impacts
(N/A)
Publications
- Purdy, P.H., Graham, J.K. 2014. Membrane modification strategies for cryopreservation. In: Wolkers, W.F., Oldenhof, H., editors. Cryopreservation and Freeze-Drying Protocls. Third Edition. The Lab Protocol Series Methods in Molecular Biology. Book Chapter. New York: Springer. p. 337-342.
- Daigneault, B.W., Mcnamara, K.A., Purdy, P.H., Krisher, R.L., Knox, R.V., Rodriguez-Zas, S.L., Miller, D.J. 2015. Enhanced fertility prediction of cryopreserved boar spermatozoa using novel sperm function assessment. Journal of Andrology. 3:558-568.
|
Progress 10/01/13 to 09/30/14
Outputs
Progress Report Objectives (from AD-416): Objective 1: Complete development and implementation of a publically available Version 2.0 of the Animal-GRIN database and initiate planning and development of a genomic component for the database: Version 2.1 of the Animal-GRIN. Objective 2: Employ current genomic and quantitative analyses to characterize genetic diversity in the germplasm collection and associated populations to guide additional germplasm sampling and diversity analyses for domestic and international germplasm stakeholders; focus on rare and economically important traits, breeds, species and populations. Objective 3: Refine cryopreservation, germplasm evaluation and reproductive technologies that facilitate efficient germplasm collection and its utilization by various livestock industries. Approach (from AD-416): Globally, 20% of the world�s livestock breeds are �at risk� of extinction -- such a contraction limits the flexibility of livestock producers to respond to future biological or economic challenges. Underlying this contraction of genetic resources are financial pressures faced by the livestock breeder coupled with greater organization of genetic selection programs. Public sector institutions are also faced with mounting pressures to reduce or eliminate research populations. This trend is of particular concern as research moves from genetic sequencing to understanding how genes function and translate into phenotypes. This project plan addresses genetic security by continuing the development of germplasm and tissues collections for all major livestock species in the U.S., so that industry and the research community can access these resources at any time. Three primary objectives address: database development, assessing in-situ and germplasm collection genetic diversity, and improvement in the efficiency and efficacy of cryopreservation protocols. All three of these objectives are highly interdependent and are required for the functionality of the repository. The expected results from this plan are a greater level of genetic security for U.S. livestock populations; information about the collection so researchers and industry can readily access the material; and improved cryopreservation protocols and germplasm evaluation methods. The successful execution of these objectives translates into greater national and global food security and greater economic vitality of the U.S. livestock sector. Animal-Genetic Resources Information Network provides a transparent portal to the national collection of animal germplasm and tissue for gene bank managers and stakeholders. Animal-GRIN V2.1 database has been launched and has replaced Animal-GRIN V1 in the US (objective 1). This transition was seamless and allowed ARS and industry groups to use the system without interruption. New features such as comparing collection phenotypes to those from the in-situ population of a breed serve to inform collection managers and stakeholders about the utility of the collection. For example, with this information we have shown that bulls from major beef cattle breeds collected 30 years ago have estimated breeding values greater than the current breed average for production traits like: weaning weight and yearling weight. The collaboration between Agriculture and Agri-Foods Canada and Empresa Brasileira de Pesquisa Agropecuaria continues as those countries bring Animal-GRIN V2 into routine use. Work was initiated on developing a genomics component for the database, which was originally scheduled to start in project plan year 5, which exceeded project plan expectations. This element was designed and programming initiated. The primary purpose of this element is to store genotypic information on animals in the collection and as a repository for genomic information developed by industry and USDA funded projects. A specific cooperative agreement with Colorado State University is facilitating genomic database development. The addition of this element to the database will provide stakeholders more information on the animals in the collection and increase the collection�s utility. Under objective 2a pedigree analysis to determine inbreeding levels and rates and effective population size have been performed for Alpine, Lamancha, Nigerian Dwarf, Nubian, Oberhasli, Saanen, Sable, and Toggenburg goats. Most of the breeds have inbreeding levels ranging from 10 to 15% for the current generation but the Oberhasli had a large and substantial inbreeding level (33%). The computed effective population size of 21 animals for the breed is below the recommended minimum of 50 set by the Food and Agriculture Organization of the United Nations (FAO- UN). This suggests breeders need to lower inbreeding rates by carefully designing mating strategies but this will be difficult as our cluster analysis indicates the average genetic relationship between clusters is greater than 25%. Also for objective 2a microsatellite analysis of genetic variation of meat and fiber goats was performed and combined with Brazilian meat goats. Among US breeds the Spanish and Angora had the greatest levels of heterozygosity and average number of alleles per locus. While population size for both breeds has contracted during the past two decades the results suggest these breeds have substantial genetic variability. Lamancha, Myotonic and Boer goats had smaller levels of heterozygosity and average number of alleles per loci, but have ample genetic variability for producers to utilize. Compared to Brazilian breeds, US breeds had higher measures of genetic diversity but the US Spanish goat was genetically similar to some Brazilian breeds. US goat production is conducted by minority and underserved farmers so these results will provide underserved producers with information which they can use to better manage genetic diversity. The results will also be used by Fort Collins scientists to develop the goat germplasm collection. Understanding breed uniqueness at the genetic level has yet to be fully understood and has ramifications in international agreements. Two biological factors, genetic drift and selection, impact the unique structure of a breed and its change over time. These factors act independently or in conjunction with one another to differentiate breeds from each other and subpopulations within a breed (objective 2b). Microsatellite analysis of Meishan pigs illustrated large and substantial genetic drift occurred when experimental populations were mated randomly and without selection. Analysis of the population differentiation over 20 years demonstrated that the fixation index (Fst) had increased to 0.05. This finding underscores the impact genetic drift acting in the absence of selection can have on imported populations. The NAGP working with the Bovine Genomics Laboratory explored how US Jersey has become differentiated from Jersey on Jersey Isle using the high density bovine SNP chip. This breed was developed on Jersey Isle but the Isle has not allowed importation of Jersey cattle back onto the island. The comparison is important as this allows the US to evaluate how imported populations have become �Americanized� during the past 100 or more years. Quantifying such differences can be used in international forums on issues like access and benefit sharing of genetic resources. The study found a Fst of 0.07 between US and Jersey Isle populations suggesting a modest but important genetic separation. While the two Jersey populations have become separated due to selection and genetic drift they still are uniquely Jersey when compared to other dairy cattle breeds based upon principal component analysis. Runs of homozygosity showed how the two populations changed over time. The Jersey Isle cattle had shorter runs of homozygosity (indicating older or less intense selection) while the US population had larger runs of homozygosity indicative of more recent selection pressure. NAGP developed and submitted the US Report on Animal Genetic Resources to the FAO-UN as a contribution to the forthcoming State of the World�s Animal Genetic Resources Report (objective 2d). The report updates the status of our animal genetic resources situation by providing appraisals of the current breed census data, measures of genetic diversity, changes in policies, and organizations involved in managing and utilizing animal genetic resources. The US was invited to submit text boxes for the full State of the World Report on the role of genetic drift on imported populations (objective 2b) and to draft a chapter on conservation efforts through gene banking. The species committees of the NAGP play a critical role in advising the program on a range of important issues (objective 2e). These committees review the germplasm collection status and assist in linking the program to various elements of the breeding industry for specific species. These committees met and their input has been solicited for collection and protocol development and use, and genetic assessments. The germplasm collection affords an opportunity to explore how genetic differences impact physiological processes. Industry and researchers often suggested that inbreeding depression lowers the fertilizing capacity of various cattle breeds. We explored the post-thaw performance of sperm from various inbred cattle (objective 2f) and observed that inbreeding levels had no impact on motility or progressive motility. Because each bull has several hundred sperm evaluated a variance can be computed and used for assessment. Post-thaw results indicate as inbreeding increased the variance in cell size decreased. This result suggests loci controlling cell size may be more homozygous as a result of inbreeding. Germplasm acquisition of six rare chicken breeds and the conservation of one research population at the Avian Disease and Oncology Laboratory were completed. Increased collection efforts were also initiated for swine and we continue to acquire unique cattle breeds and subpopulations. Over 6, 000 animals from the collection were accessed to date for research or reconstitution projects. In order to efficiently include DNA from female agricultural species in the repository it is necessary to preserve eggs, but the methods associated with preservation can be difficult and expensive to use in a field setting (objective 3b). However, adoption of protocols from recently published research and development of an inexpensive device to hold and move eggs during the freezing process enabled collection and preservation of the first oocytes by Fort Collins scientists in a field setting for inclusion in the repository. Further improvement of these methods for freezing eggs enables the NAGP to create a more complete and secure collection of germplasm. Preserving semen for artificial insemination requires dilution of the semen in a medium to enable it to survive the freezing process (objective 3c). In order to achieve the highest fertility with frozen-thawed semen it is necessary to identify the medium that provides the most protection to the sperm during freezing and still allows them to function in a normal physiologic manner. Fort Collins research with ram sperm demonstrated that the medium can have a large impact on the function of the individual sperm and on actual fertility. Similar research is now being conducted on boar sperm in order to understand the impact of the semen cryopreservation medium on that species as well. Completion of these analyses will enable ARS and others to utilize a semen cryopreservation medium that results in higher levels of fertility when frozen-thawed semen is used for artificial insemination. It is important to understand how a sperm functions in order to assess sample quality and predict fertilizing potential (objective 3d). To achieve this we utilized a number of fluorescent stains that can be used to indicate relative values of function and consequently a high or low level of quality. Our research focused on establishing methods of analysis using multiple fluorescent stains and most importantly identifying those stains which provide the most meaningful information about a sample. By optimizing these methods we will be able to understand the impact of a freezing medium on a sperm sample, determine the quality of samples, and potentially predict the fertilizing potential of sperm prior to artificial insemination. Significant Activities that Support Special Target Populations: Efforts to find a low cost approach to artificial insemination sheep are ongoing. We are in the process of refining and adapting this technology to the needs of NAGP and private producers. Much of the germplasm collection on rare breeds is done in conjunction with underserved minority farmers. As they have supported the collection efforts we initiate various types of technology transfer that they can use in their operations. During FY2014 57 tours totaling 900 people viewed the national collections maintained by the Plant and Animal Genetic Resources Preservation Unit. Included were a group of Native American extension agents, three separate women�s groups interested in agriculture, and approximately 450 college, high school and elementary students. Accomplishments 01 Conserving chicken genetic diversity. The genetic resources in the national collection held in Ft. Collins underpin the livestock sector and provide researchers with a tool for their research. Collection size has increased; specifically six rare breeds of chickens have been secured in the collection, as well as, a valuable research line of chickens. This was made possible by cryopreserving ovaries and testes from day old chicks from the respective breeds. Underserved chicken producers, who typically raise these breeds, now have secured populations. 02 Genetic diversity of dairy, meat and mohair goats. Twelve goat breeds were evaluated by ARS scientists in Fort Collins, CO for genetic diversity. Eight of the 12 were dairy goats and for these breeds pedigree analysis was performed. Meat and fiber goats were evaluated using microsatellite markers and compared to Brazilian meat goats. Microsatellite analysis indicated that US meat goats have sufficient genetic diversity and even though there has been a contraction in population size this contraction has not negatively impacted genetic diversity. Seven of eight dairy breeds exhibited no contraction of genetic diversity based upon computed inbreeding levels and effective population size. However, the Oberhasli breed had a large inbreeding coefficient (33%) and low effective population size that requires development of breeding plans to reverse slow or stop the loss of genetic variability. Goat producers are from an underserved group of minority farmers, therefore these types of analysis are not performed by the industry. This information will be presented to the various organizations representing goat breeders. 03 Genetic change of imported animal populations. ARS scientists in Fort Collins, CO have evaluated how genetic diversity has changed when animal populations are imported into a new country by evaluating molecular marker data from Meishan pigs and Jersey dairy cattle. After importation breeds remain true to breed type but there is a substantial amount of change in allele frequencies due to genetic drift and selection. These results suggest that when discussing the international exchange of genetic resources it will not be possible to attribute genetic gains and long term compensation schemes to the original genetic composition which was imported.
Impacts
(N/A)
Publications
- Blackburn, H.D., Plante, Y., Rohrer, G.A., Welch, E.W., Paiva, S. 2014. Impact of genetic drift on access and benefit sharing under the Nagoya protocol: The case of the Meishan pig. Journal of Animal Science. 92(4) :1405-1411.
- Blackburn, H.D., Plante, Y. 2014. North American animal breeding and production: meeting the needs of a changing landscape. Journal of Animal Breeding and Genetics. 131:247-248.
- Long, J.A., Purdy, P.H., Zuidberg, K., Sipke-Joost, H., Velleman, S.G., Woelders, H. 2014. Cryopreservation of turkey semen: effect of breeding line and freezing method on post-thaw sperm quality, fertilization, and hatching. Cryobiology. 68:371-378.
- Daigneault, B.W., Mcnamara, K.A., Purdy, P.H., Krisher, R.L., Knox, R.V., Miller, D.J. 2014. Novel and traditional traits of frozen-thawed porcine sperm related to in vitro fertilization success. Theriogenology. 82:266- 273.
|
Progress 10/01/12 to 09/30/13
Outputs
Progress Report Objectives (from AD-416): Objective 1: Complete development and implementation of a publically available Version 2.0 of the Animal-GRIN database and initiate planning and development of a genomic component for the database: Version 2.1 of the Animal-GRIN. Objective 2: Quantify genetic diversity in the germplasm collection, and in-situ populations, and compare their similarities via genomic or quantitative approaches and use the results to guide germplasm collection development and provide genetic diversity assessments to breed associations. Objective 3: Refine cryopreservation, germplasm evaluation and reproductive technologies that facilitate efficient germplasm collection and its utilization by various livestock industries. Approach (from AD-416): Globally, 20% of the world�s livestock breeds are �at risk� of extinction -- such a contraction limits the flexibility of livestock producers to respond to future biological or economic challenges. Underlying this contraction of genetic resources are financial pressures faced by the livestock breeder coupled with greater organization of genetic selection programs. Public sector institutions are also faced with mounting pressures to reduce or eliminate research populations. This trend is of particular concern as research moves from genetic sequencing to understanding how genes function and translate into phenotypes. This project plan addresses genetic security by continuing the development of germplasm and tissues collections for all major livestock species in the U.S., so that industry and the research community can access these resources at any time. Three primary objectives address: database development, assessing in-situ and germplasm collection genetic diversity, and improvement in the efficiency and efficacy of cryopreservation protocols. All three of these objectives are highly interdependent and are required for the functionality of the repository. The expected results from this plan are a greater level of genetic security for U.S. livestock populations; information about the collection so researchers and industry can readily access the material; and improved cryopreservation protocols and germplasm evaluation methods. The successful execution of these objectives translates into greater national and global food security and greater economic vitality of the U.S. livestock sector. Database development is the glue for managing large genetic resource collections. This tool enables gene bank managers and the public to understand the content of the animal collections and stimulates interest in collection use. To this end progress in developing Animal-Genetic Resources Information Network Version 2 by the joint Brazilian, Canadian, and US programming team achieved the milestone for Objective 1 and real time testing of the information system has commenced. Evaluation of genetic diversity and change can provide policy makers with insights for decision making (subobjective 2b and 2d). Using germplasm samples from the repository, we documented large and substantial levels of genetic drift which had occurred among current Meishan pig populations vs samples from the original Chinese importation in 1989. This information has important ramifications about access and benefit sharing arrangements between countries as they consider the adaptation of the Nagoya Protocol. In an analysis comparing plains and wood bison we determined that the genetic distance between these two groups is small and therefore breeding programs by the National Park Service do not need to treat the two groups as separate populations or classify these two groups as subspecies. Under subobjective 2a we compared performance measures (e.g., weaning weight, fertility traits, carcass characteristics) of animals contributing samples to the repository with breed averages from 1950 to 2010 for Hereford, Angus, Charolais, and Brangus breeds. It was determined that the collection mirrors the breed average for the measured traits over time. It was also determined that the variability of the collection for these performance characteristics is large and substantial which better insures the collections use over time. Ovaries and testes from three industry lines of chickens were collected and cryopreserved. Based upon the successes with chicken ovary cryopreservation, experimentation on various cryopreservation techniques on other species was initiated under subobjective 3b. Results to date indicate that extraction and utilization of oocytes from vitrified mammalian tissue is feasible. The efficiencies of the techniques, coupled with the labor and reagent expenses, make the techniques completely impractical for cryo storage. However, emerging cryopreservation techniques for roosters by Japanese researchers may offer increases in the efficiency of using cryopreserved sperm (subobjective 3c) and are planned for FY2014. Significant Activities that Support Special Target Populations: Most cattle breed association members fall into the category of small farms. Therefore these groups need assistance in planning how they will become engaged in the new genomic era. We assisted two such associations the American Hereford Association and the American Salers Association in determining which animals or groups of animals they might sample for genotyping in a cost effective manner. Plans were developed based upon an assessment of their pedigree structure and samples that were already present in the repository. This information was presented to association executives for their use in communicating to members the proposed plan of action for acquiring and genotyping their populations; the results of which will be the development of selection tools that incorporate animal genotypes. This project continues to work closely with HBCU and Hispanic Serving institutions in managing small ruminant and cattle genetics and jointly developing research projects that are of utility to small farms and historically underserved producers. Efforts continue with Virginia State University to increase the effectiveness of germplasm collection, cryopreservation, and utilization of AI in small ruminants. With Sul Ross University we have developed a joint project evaluating different era (1970�s vs present day) Hereford cattle performance in arid environments. Accomplishments 01 U. S. genetic resources secured. Nationally genetic diversity of livestock populations is contracting. The development and maintenance of a broad based collection of germplasm at the ARS location, Fort Collins, CO, from diverse livestock populations contributes to U.S. and global food security by making genetic variability readily available for the livestock industry. The germplasm collection gives the U. S. livestock sector an unprecedented level of security in terms of genetic diversity collected and quantity of germplasm acquired. The addition of three private sector lines of chickens validates this assertion. The germplasm collection has been shown to be robust and used by producers, and public and private researchers. In addition, it serves as a ready resource in the event of a critical national need. 02 Random genetic change and policy development. Many countries are considering Nagoya Protocol ratification for the access and benefit sharing of genetic resources. But, little information about genetic change of imported populations has been available for developing a policy with impact on international germplasm exchange. The Fort Collins gene bank contains original samples from Meishan pigs imported from China in 1989 and samples from two present day research populations of Meishan. Using molecular genetic approaches, ARS researchers at Fort Collins found the impact of genetic drift (random change) was large and substantial, thereby drawing into question the utility of long term agreements and payments for genetic resources with governments tracking payments and genetic resource use. Due to an inability to effectively monitor gene flow of imported populations, the current method of exchange, private treaty contracts, are the most effective mechanism for exchanging genetic resources and promoting access and benefit sharing.
Impacts
(N/A)
Publications
- Silversides, F., Purdy, P.H., Blackburn, H.D. 2013. Comparative costs of programmes to conserve chicken genetic variation based on maintaining living populations or storing cryopreserved material. British Poultry Science. 53:599-607.
- Liu, J., Cheng, K.M., Purdy, P.H., Silversides, F.G. 2012. A simple vitrification method for cryobanking avian testicular tissue. Poultry Science. 91:3209-3213.
- Freking, B.A., Purdy, P.H., Spiller, S.F., Welsh, C.S., Blackburn, H.D. 2012. Boar sperm quality in lines of pigs selected for either ovulation rate or uterine capacity. Journal of Animal Science. 90(8):2515-2523.
- Cronin, M., Macneil, M., Ninh, V., Leesburg, V.L., Blackburn, H.D., Derr, J. 2013. Genetic variation and differentiation of bison (Bison bison) subspecies and cattle (Bos taurus) breeds and subspecies. Journal of Heredity. 104:500-509.
|
|