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United States Department of Agriculture

Agricultural Research Service

Research Project: NATIONAL ANIMAL GERMPLASM PROGRAM (NAGP)
2010 Annual Report


1a.Objectives (from AD-416)
1. Develop germplasm and DNA/tissue collection coupled with measurement of diversity. o Continue to develop germplasm collections across species and associated information. o Evaluate, refine and implement pedigree clustering approach for germplasm collection. o Pursue approaches to compare collection to in-situ populations using quantitative and/or molecular approaches. o Develop collections of DNA and/or tissues containing DNA and associated information. 2. Further develop the animal section of the GRIN network. o Develop database information system that documents the germplasm/tissue collection (Version.
2)and has multi-location capacity. o Expand descriptors for all species as defined by species committees, and substantially increase data collection efforts. 3. Develop methods for population regeneration. o Computationally determine approaches for population regeneration and their management. 4. Improve cryopreservation methods for tissues. o Development of predictors/evaluation of post-thaw semen viability. o Procedures for collection and freezing of small ruminant and/or beef oocytes. o Determination of optimal semen cryopreservation diluents and freezing methodologies.


1b.Approach (from AD-416)
The over-arching goal of the National Animal Germplasm Program (NAGP) is to increase the security of U.S. livestock genetic resources by the development of a repository containing diverse livestock genetic resources. The proposed objectives of this plan are important because they will: strengthen the genetic diversity contained in the collection; improve the ability and efficiency of reconstituting populations through improved cryopreservation procedures and reconstitution strategies; and provide potential collection users with a more comprehensive understanding of what is contained in the collection through the GRIN database. Executing these objectives will require the utilization of quantitative and molecular genetics, reproductive biology, cryopreservation, and information systems science. The beneficiaries of this effort cover a wide spectrum including: livestock breeders; researchers reconstituting populations and performing various types of molecular studies; and the American public at large which benefits from the increased food security the program provides.


3.Progress Report
Genetic security of livestock populations has been improved as the total collection increased beyond 575,000 samples, an increase of approximately 6% from the previous year. Importantly, the collection has not only grown but during FY2010 914 samples from 507 animals left the repository for animal generation, germplasm evaluation, or DNA studies. Significant collection efforts were placed on acquiring embryos and semen from Angora goats. This breed is of economic significance particularly in Texas and due to weather related mortalities and market forces a significant contraction in numbers and genetic diversity for this breed has occurred. Additional collections are planned in FY2011.

Work performed under Objective 3 has shown that reconstitution of breeds or special populations can be accomplished with fewer units of germplasm. Therefore, the majority of Species Committees have agreed to reduce Core Collection requirements (the quantity of semen needed to reconstitute a breed or population by 150%). The new values will, in general, reduce target quantities by more than 50%.

Using the results of Objective 1b, pedigree clustering, the International Brangus Breeders Association was shown the status of the germplasm collection for that breed of cattle. As a result new acquisitions were made and an analysis of the breed’s inbreeding levels were calculated. This analysis showed that the rate of inbreeding is increasing, at an increasing rate, and that effective population size (a measure of a population’s genetic health which FAO has recommended a minimum target of 50 head) has dropped below 30 head (the breed registered over 17,000 animals in 2009).

Database development has been advanced with programmers from Brazil and Canada working at their home institutions or working at our laboratory. A substantial review of the progress in database development was held with programmers and geneticists from all three countries meeting at our lab. The database users felt that the programming team is making substantial progress and the level of sophistication in database functionality is exceeding expectations.

Harvesting and cryopreserving chicken ovaries was initiated. This approach has been shown to have relatively high success rates and allows for all of the chicken genome to be secured.


4.Accomplishments
1. Genetic security increased. Globally, genetic diversity is contracting across livestock species. Development of comprehensive ex-situ-cryopreserved germplasm collections in genebanks offers a level of genetic security for the livestock industry and research community. By building and expanding the germplasm collection the livestock industry and research community are able to access the collection’s genetic resources as needed.

2. Combining and analyzing sheep DNA from Brazil and the US. Understanding the genetic distinctness of US sheep breeds in relation to those in other countries can impact conservation strategies and breeding programs. Using genotypes generated in our respective countries, ARS scientists at the National Animal Germplasm Program in Fort Collins, Colorado and Brazilian scientists explored methods to combine and analyze genetic differences of sheep breeds found in the US and Brazil. The approach used to merge the data was successful and we were able to determine genetic distances and show how breeds were grouped together based upon physiological function (meat vs wool/hair production). Importantly this study showed that the Brazilian and US hair breeds are substantially different from one another suggesting different countries of origin. As a result, US breeders interested in Brazilian breeds (and vice versa) could expect substantial hybrid vigor by importing and crossing these breeds; and that countries can leverage information generated independently and use that information in their conservation programs.

3. Securing Holstein germplasm - Collections in France, The Netherlands, and US. Globally, Holstein is the predominant dairy cattle breed and has been shown to have a small effective population size (< 36 animals). Therefore the three countries wanted to know how well Holstein is secured and the diversity represented in the three countries’ gene banks. Working with scientists from France and the Netherlands an ARS scientist provided information on the US Holstein collection, suggested analytical approaches, interpreted the results, and co-authored a proceedings paper. This project indicated that among the three gene banks the US collection was the largest in terms of number of bulls represented, and had the lowest genetic relationships when compared to the French and Dutch collections. Using the effective number of founders to compare gene bank collections to each country’s set of active bulls it was determined that the US collection with 784 founders, exceeding our current population of active bulls with 115 founders. Given these results, we can state that the US collection is the largest and most diverse collection among the three countries. In addition, the US collection is more diverse than the current breeding population of US Holsteins and therefore the gene bank has secured the Holstein breed in the event of a disaster or need to reintroduce genetic variation.

4. Efficient Use of Cryopreserved Boar Semen. Germplasm collected and stored in a gene bank will become a limited resource as it is used over time. Therefore, optimizing the number of motile cells to inseminate in a sow is important. To address this issue, an experiment evaluating the optimal insemination dose (1.0, 0.75, 0.5, and 0.25 billion sperm) for boar semen was performed. Scientists with National Animal Germplasm Program, in Fort Collins, co-designed, cryopreserved all boars’ samples, performed pre-freeze and post-thaw analysis of the semen samples, provided input into statistical analysis and prepared an abstract with university collaborators. The experiment found no statistical differences in pregnancy rate. However, the 0.25 treatment had numerically fewer pregnant sows. To ascertain the dose most appropriate to use if semen is limited, the total number of fetuses per sow inseminated was multiplied by the number of potential doses available if only one billion motile sperm cells existed. The results suggest that the optimal dose of semen to utilize per insemination was between 0.50 and 0.75 billion motile sperm cells. Such information significantly aids the National Animal Germplasm Program in planning germplasm collections and assures that effective use of stored germplasm can be made when needed. From an industry perspective the results suggest that inseminating sows with one or two billion cells may exceed what is needed to create a pregnancy with a sufficient litter size.


5.Significant Activities that Support Special Target Populations
Many rare breeds are owned by small scale farmers. To capture this diversity for the repository we perform field collections on small farms. Such action provides us with an opportunity to provide small scale producers with information about the breeding soundness of their animals and management of genetic diversity. Significant collaborations exist with a number of historically black colleges and universities in the collection of germplasm. Substantial collaboration and dialog exists between the primary non-governmental organization working in the area of conservation of animal genetic resources. These linkages support germplasm collection, interaction on research problems, and management of in-situ populations for genetic diversity.


Review Publications
Blackburn, H.D., Silversides, F., Purdy, P.H. 2009. Inseminating fresh or cryopreserved semen for maximum efficiency: implications for gene banks and industry. Poultry Science. 88:2192-2198.

Purdy, P.H., Song, Y., Silversides, F.G., Blackburn, H.D. 2009. Evaluation of Glycerol Removal Techniques, Cryoprotectants, and Insemination Methods for Cryopreserving Rooster Sperm with Implications for Breed and/or Line Regeneration. Poultry Science. 88:2184-2191

Purdy, P.H., Moce, E., Stobart, R., Murdoch, W.J., Moss, G.E., Larson, B., Ramsey, S., Graham, J.K., Blackburn, H.D. 2009. The fertility of ram sperm held for 24 hours at 5 ºC prior to cryopreservation. Animal Reproduction Sciences. 118:231-235.

Moce, E., Purdy, P.H., Graham, J.K. 2009. Treating ram sperm with cholesterol-loaded cyclodextrins improves cryosurvival. Animal Reproduction Sciences. 118:236-247.

Welsh, C.S., Stewart, T.S., Schwab, C., Blackburn, H.D. 2010. Pedigree Analysis Of Five U.S. Swine Breeds And The Implications For Genetic Conservation. Journal of Animal Science. 88:1610–1618.

Spencer, K.W., Purdy, P.H., Blackburn, H.D., Spiller, S.F., Stewart, T.S., Knox, R.V. 2010. Effect of number of motile frozen-thawed boar sperm and number of fixed-time inseminations on fertility in estrous-synchronized gilts. Animal Reproduction Sciences. 121:259-266.

Last Modified: 10/20/2014
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