Location: Agricultural Genetic Resources Preservation Research
2025 Annual Report
Objectives
Objective 1: Build, secure, manage, and facilitate the use of the animal genetic resource collection.
Subobjective 1A. Operate species committees to advise NAGP on collection development and use.
Subobjective 1B. Build, secure and manage genetic resource collections using quantitative and molecular analyses to further enhance the germplasm collection.
Sub-objective 1C. Engagement and representation in the United Nation’s Food and Agriculture Organization’s (FAO) Intergovernmental Technical Working Group on Animal Genetic Resources (ITWG-AnGR).
Objective 2: Development, maintenance, and implementation of the publicly accessible Animal-GRIN database Version 2.
Subobjective 2A. Redesign and/or necessary software upgrades to database and public webpages.
Objective 3: Characterize genetic diversity to guide collection development and increase its utility.
Subobjective 3A. Quantify genetic differences within and among breeds across different environmental zones.
Subobjective 3B. Quantify genetic diversity of minor, rare, or heritage chicken breeds.
Objective 4: Rejuvenate poultry research lines from cryopreserved semen using both surgical and non-surgical approaches.
Objective 5: Develop and refine assisted reproductive technologies that enable the efficient and effective collection, evaluation, and utilization of germplasm.
Subobjective 5A. Visualization of computer aided sperm analysis (CASA) results.
Subobjective 5B: Develop improved preservation and utilization methodologies for sperm.
Sub-objective 5C. Adapt and improve chicken primordial germ cell collection, preservation, utilization, and quality evaluation methods for use by gene banks and industry.
Approach
The United States is the epicenter of improved livestock genetics. American poultry breeding companies distribute genetic stocks to more than 110 countries, and the Holstein is the premier genetic package used globally for milk production and to a level that more dairy bull semen is exported than used domestically. The breeding complex, using genetic diversity of our breeds, fuels increased animal productivity and economic activity on a global scale and underpins the global food supply of animal protein. However, there has been a contraction of genetic variability and the number of farm enterprises engaged in livestock breeding. This draws into question the stability of the breeding complex and access to the full complement of genetic resources and thereby hindering the U.S. competitive position, and our ability to reintroduce lost genetic variability and address challenges such as climate change. Therefore, this project’s primary goal is to secure genetic resources and make the collected material and information accessible in national emergencies, for research purposes, or industry’s desire to utilize the USDA National Animal Germplasm Program (NAGP) collection for genetic improvement programs. Developing robust collections requires quantification of genetic variability, publicly accessible information, and cryopreservation of tissues used to reconstitute animals of interest. The project and its framework is established and operational. We will use this framework to acquire germplasm/tissue from targeted animals within a breed or commercial line, evaluate various attributes of genetic diversity, and document the collection in the Animal-Genetic Resources Information Network (Animal-GRIN) information system. As demonstrated, there have been short-term research activities (by the project or stakeholders accessing the collection) that transformed the livestock sector, and long-term the project can facilitate corrective mating or reintroduction of genetics absent from populations. The proposed project plan will provide tools to stakeholders and will continue developing the collection, thus providing the U.S. with greater security for its animal genetic resources.
Progress Report
Global demands on animal genetic diversity are increasing, and the U.S. plays a major role in providing the world with improved genetic resources. 60 to 70% of the world’s broiler and shrimp genetics are the product of U.S. companies. This is in addition to supplying over 700,000 livestock producers with improved genetics by which they earn a living. Therefore, proactively taking steps to protect U.S. animal genetics, is core to the USDA, ARS National Animal Germplasm Program’s (NAGP) mission and program objectives.
To better secure our animal genetic resources, the NAGP has in the past year captured a wide range of genetic diversity across species (Subobjective 1B), performed genomic analyses to quantify the genetic variability within and among our breeds (Subobjective 1B & Subobjective 3A), taken emergency steps to protect chicken genetic resources with new techniques (Subobjective 5C), explored new methods to combine sperm evaluation methods for more accurate post-thaw assessments (Subobjective 5A), and refined the information system Animal-Genetic Resources Information Network (Animal-GRIN) (Objective 2). Due to the global importance of U.S. genetic resources, NAGP was also engaged with the United Nation’s Food and Agriculture Organization to ensure that U.S. positions on the development of policies and application of relevant technology concurs with U.S. perspectives (Subobjective 1C).
Duroc, a uniquely American pig breed, plays a major role in global pig production systems. An industry wide genomic study of Duroc genetic diversity was performed with six U.S. subpopulations, three Canadian subpopulations, and three Australian subpopulations from 10 international breeding companies and a subpopulation sampled from smaller individual U.S. breeders. Analyses suggest that corporate subpopulations are separating from each other due to differing selection criteria and genetic drift. When ADMIXTURE, which infers population structure and individual ancestry, was used it suggested three global groups of Durocs; the three Australian subpopulations, a U.S. group with three subpopulations, and a Canadian-U.S. group comprised of seven subpopulations. Effective population size (Ne) for the subpopulations ranged from 17 to 47 animals. The corporation with the smallest Ne was notified so they might consider mitigating approaches to genetic diversity loss. Upon pooling subpopulations, Duroc had an effective population size of 172 animals, well above the critical threshold of 50 animals, suggesting ample genetic diversity for corporations to continue selection programs as needed to meet customer demands. The separation of Duroc subpopulations suggests a de-facto industry-based approach for conserving genetic diversity. While individual populations are subject to genetic drift, there is an opportunity to increase genetic variation by mating different subpopulations. Currently the germplasm collection is well positioned to protect Duroc genetic diversity by holding samples from five of the seven U.S. subpopulations. (Subobjectives 1B & 3A)
U.S. shrimp breeding companies play a major role in driving genetic progress in global shrimp genetics and productivity with the genetic diversity they hold. Previously no assessment within and among companies had been conducted. Quantifying genetic diversity aids in future selection options and guides gene banks in germplasm collection strategies. For this study, we obtained 878 animal samples from three corporations, denoted as C1 (n = 278), C2 (n = 300), and C3 (n = 300). DNA extraction and genotyping were performed using 192 Single Nucleotide Polymorphisms (SNPs) evenly spaced across the shrimp genome. First and second principal components accounting for 19.5% of the variation, suggest that corporate populations were genetically distinct. This finding was supported by other analytical approaches, which estimated population structure using the fixation index (Fst) and the STRUCTURE program results. Low Ne for C1 and C2 suggest future management of genetic diversity is needed to preserve selection options. Genetic relationships (ranging from 12.5% (first cousins) to 25% (half-sibs)) found among individual shrimp within corporation were high, ranging from 41 to 87% of the individuals tested. Our findings suggest corporate lines are genetically diverse from each other. Within corporations, genetic diversity estimates suggest vigilance should be employed to minimize inbreeding and genetic relationship. When comparing the corporate populations to commercially sampled shrimp from Asia and Ecuador, the corporate lines were distinct, suggesting selection pressure for various traits is creating unique lines for the marketplace. (Subobjectives 1B & 3A)
Reintroduction of lost-Y chromosomes in Holstein cattle continues by evaluating the performance and genomic structure of the second generation (G2) progeny. These animals were the product of artificial insemination matings of first-generation bulls (sons of collection bulls) to commercial cows in herds located in Pennsylvania and New Mexico. Within-herd phenotypic evaluations suggest G2 305-day mature equivalent for milk is nearly equal to cows representing modern genetic combinations. At the molecular level, collection bull progeny (G1) and grand progeny (G2) had lower likelihood of possessing two identical alleles of a particular gene(s) than the modern contemporaries (8.75%, 11.95%, & 18.75%, respectively) which could explain higher than expected G2 performance for milk production. Surprisingly, some of the G2 cows had 305-day mature equivalents that were in the upper 25% of the cows evaluated. The G2 cows will continue to be monitored for life-time productivity and ability to remain in the test herds. Semen samples on G2 bulls have been cryopreserved and placed in the germplasm collection. (Subobjectives 1B)
Artificial insemination with frozen-thawed rooster sperm is problematic because of the low quality of sperm resulting from exposure to cryoprotectants during the freezing process. Consequently, overcoming these damaging effects was attempted by using non-traditional chemicals that do not require removal prior to insemination. Following freezing and thawing, the motility of rooster sperm across treatments was not different from the samples frozen in glycerol, the treatment that has proven to consistently provide some level of fertility, but the fertility in all treatments was very low. Additional analyses will be performed to understand the effects of the freezing treatments on rooster sperm quality and to determine if an improved freezing methodology can be identified. (Subobjective 5B)
Rejuvenation of chicken lines using primordial germ cells (PGCs) requires use of a host that has been sterilized to either completely or partially remove their native PGCs so that donor PGCs can be injected and populate the host’s gonads. Ideally, a completely sterilized host should be used, but hurdles have arisen making use of that model not feasible at this stage of experimentation. Consequently, an alternative approach using busulfan to sterilize chicken embryos was explored. While this method has been previously demonstrated as operational and provided some successful rejuvenation of chicken lines, our experimentation focused on improving the means of sterilizing the embryo and increasing the resulting fertility of the donor PGCs. Although the experimentation is still in progress, the initial results demonstrate that the busulfan treatment we developed provides efficacy for sterilizing chicken embryos that is comparable to the original reports. However, as we can now inject 50 times more PGCs into hosts than what the original reports used, we are optimistic that our fertility should be greater than what was originally achieved. (Subobjective 5C)
Substantial efforts were made to upgrade the Animal Genetic Resources Information Network (Animal-GRIN) software to the most current version of the coding languages used in the program. In addition, repeated measures were taken to comply with updates required by USDA cybersecurity. A meeting was held with an Animal-GRIN user group to evaluate steps needed to make the public facing pages easier to navigate. (Subobjective 2A)
This fiscal year the USDA ARS National Animal Germplasm Program (NAGP) increased the nation’s store of animal genetic resources by more than 21,500 samples from 952 animals (Subobjective 1B & Subobjective 3A). In addition, 11 new breeds were added to the collection, bringing the total number of breeds in the collection to 211. Major emphasis was placed upon acquiring primordial germ cell samples from university research populations and private industry. This was viewed as critical due to the ongoing threat of highly pathogenic avian influenza. As requested by the Aquaculture breeding programs, increased the number of lines and samples cryopreserved, sent, and stored at NAGP. Similarly, honeybee submissions by the Product Quality and New Uses program, and industry were made. As demonstrated with Jersey cattle (2024 annual report), it is essential for NAGP to refresh its collection of germplasm, particularly among high selection intensity breeds. Therefore, samples from new animals from breeds such as Hereford, Angus, Guernsey, Ayrshire, Charolais, Duroc, Berkshire, and Holstein were acquired. In some instances, industry imported new animals which were added to the collection from countries such as Canada and Australia; therefore, we expect that such additions will add new elements of genetic variation to the collection. Using Ward’s statistical clustering approach with pedigree data continues to provide a cost-effective method to compare in-situ populations to the collection, as it helps to identify gaps in the collection and the dynamic nature of subpopulations within a breed and has been of utility to breed associations, as our analysis of Hereford cattle pedigrees demonstrates.
Accomplishments
1. Effective use of cryopreserved dairy and beef bulls born before 2000. Breeders change selection goals, create genetic bottlenecks, and experience detrimental genetic mutations creating a need for different genetic combinations that may be found in the USDA National Animal Germplasm Program collection of genetic resources. Since the collection’s inception in 1999, it has been commonly presumed that progeny from older animals stored in the collection would have inferior performance compared to the current industry populations. ARS scientists in Fort Collins, Colorado, explored this question using highly selected cattle breeds - Holstein and Angus. Our results showed phenotypic performance (e.g., milk or beef production) of individual animals from the first and second generation of progeny sired by collection bulls or their sons could meet or exceed the population average, which was counter to the common presumption. For both breeds the key to making competitive progeny from older genetics requires three elements: 1) creation of 15-25 progeny from a collection bull to increase the probability that desirable genes will come together through recombination; 2) mate collection bulls to cows with performance levels in the upper 60-70th percentile for the traits of interest; and 3) select progeny for future matings on phenotypic performance and level of heterozygosity. These findings point to the value of the USDA National Animal Germplasm Program collection to assist both U. S. beef and dairy cattle breeders develop breed improvement that increase productivity and profitability.
2. Protecting research investments in chicken populations. Highly pathogenic avian influenza and increased financial pressures have raised concerns by universities and USDA about their ability to maintain viable research populations of poultry developed and used for more than 50 years. To protect the valuable genetic diversity in these populations and the research investment, ARS scientists in Fort Collins, Colorado, implemented novel procedures to collect, preserve, and increase post-thaw populations of primordial germ cells from poultry. This important advancement has increased USDA National Animal Germplasm Program capacities to safeguard such populations with increased speed and lower cost. As a result, entire poultry collections and multiple poultry populations have been collected and cryopreserved (e.g., ARS National Poultry Research Center, Iowa State University, University of Arkansas, University of California – Davis, Virginia Tech University), which ensures the U.S. poultry industry will have at its disposal the genetic diversity needed to withstand disease, pest, and production threats.
3. U.S. Mashona cattle diversity is equal to the progenitor population in Zimbabwe. U.S. Mashona cattle breeders are working toward establishing a breed association and pedigree recording system. Mashona populations in the U.S. and Zimbabwe are small but those in Zimbabwe are at greater risk of being lost. This prompted them to approach the USDA National Animal Germplasm Program for assistance in assessing this breed’s purity and genetic diversity as a baseline for determining breed registration criteria. To address this need, ARS scientists in Fort Collins, Colorado, used genotypic data and a new computational method, which allows estimation of effective population size as far back as 200 generations. This method showed that the current U.S. population is as genetically diverse as the ancestral population in Zimbabwe. This important discovery shows that the U.S. Mashona population can serve as a critical backup and source of selected genetics (and new markets) for the Zimbabwe population. Therefore, the USDA National Animal Germplasm Program collection provides important support to U.S. Mashona breeders in addressing genetic challenges and in establishing a breed association.
4. Bacon and shrimp – industry’s role in conserving genetic resources. Shrimp and pig breeding companies are structured to develop proprietary genetic lines selected for traits of interest. ARS scientists in Fort Collins, Colorado, evaluated genetic diversity among corporate breeding populations of shrimp (three companies) and Duroc swine (10 companies) showing a similar trend of corporate lines becoming separate and distinct over time for both species, which is caused by varying selection criteria within lines and genetic drift. These monumental findings show that corporate lines can serve as a de-facto in-situ conservation strategy for these species. When coupled with comprehensive collections of germplasm in the USDA National Animal Germplasm Program, a robust conservation strategy is in place for U.S. shrimp and swine industries.
Review Publications
Srikanth, K., Jaafar, M.A., Neupane, M., Zaabza, H.B., McKay, S.D., Wolfe, C.W., Metzger, J.S., Huson, H.J., Van Tassell, C.P., Blackburn, H.D. 2024. Assessment of genetic diversity, inbreeding and collection completeness of Jersey bulls in the US National Animal Germplasm Program. Journal of Dairy Science. 107(12):11283-11300. https://doi.org/10.3168/jds.2024-25032.
Azevedo, H.C., Blackburn, H.D., Lozada-Soto, E.A., Spiller, S.F., Purdy, P.H. 2024. Enhancing evaluation of bull fertility through multivariate analysis of sperm. Journal of Dairy Science. 107(12):11774-11778. https://doi.org/10.3168/jds.2024-25163.
Hu, T., Purdy, P.H., Blank, M.H., Muhonja, C.K., Pereira, R.J., Tiambo, C.K., McGrew, M.J. 2024. Direct in vitro propagation of avian germ cells from an embryonic gonad biorepository. Poultry Science. Article e104260. https://doi.org/10.1016/j.psj.2024.104260.
Bolarin, A., Berndtson, J., Tejerina, F., Cobos, C., Pomarino, C., D'Alessio, F., Blackburn, H.D., Kaeoket, K. 2024. Boar semen cryopreservation: State of the art, and international trade vision. Animal Reproduction Science. 269. Article e107496. https://doi.org/10.1016/j.anireprosci.2024.107496.