Research Project: Improving Dairy Cow Feed Efficiency and Environmental Sustainability Using Genomics and Novel Technologies to Identify Physiological Contributions and Adaptations

Location: Animal Genomics and Improvement Laboratory

2025 Annual Report

Objectives
Objective 1: Develop genomic resources and molecular tools to determine limitations to nutrient use efficiency in dairy cattle. Sub-objective 1.A: Using phenotypically well-defined lines of Holstein dairy cattle selected and monitored for feed efficiency, investigate the biological and genetic bases of nutrient use efficiency and support genomic selection studies by identifying highly affected pathways against a backdrop of known RFI phenotype and methane emissions. Sub-objective 1.B: Continue to refine and validate a short-term isolated duodenal model for assessing intestinal epithelial tissue transcriptomic changes in ruminant gastrointestinal tissues in vivo in response to changes in luminal nutrient flow. Objective 2: Develop and apply novel nutritional strategies for calf rearing and weaning to reduce feed and nutritional costs to dairy cattle production through further refining the global landscape of genomic and epigenomic regulatory elements, exploring the regulatory dynamics of chromatin states in rumen development during weaning and for complex traits. Sub-objective 2.A: Characterize molecular phenotypes of the calf rumen transcriptome through strand-specific RNA sequencing (ssRNA-seq) and single-cell RNA sequencing during development. Sub-objective 2.B: Functionally annotate the calf rumen epigenome and identify transcriptional cis-regulatory modules during development, including histone modification, chromatin accessibility, and architecture using Chromatin Immunoprecipitation-sequencing (ChIP-Seq) technologies. Objective 3: Using in vitro and in vivo gastrointestinal tissue responses (ruminal, duodenal, and colon) in lactating and non-lactating cows, investigate perturbations in luminal factors (changes in nutrient flow) at single-cell resolution. Sub-objective 3.A: Assess short-term (days) responses of metabolism- and transport-related genes and proteins of the intestinal epithelia in response to direct delivery of individual substrates or nutritional components (e.g., nutrients, metabolites, humoral factors). Sub-objective 3.B: Evaluate the impact of nutrient use efficiency, as defined by dairy efficiency as determined RFIlac, on total tissue, animal protein, and lipid carcass composition.

Approach
To improve feed efficiency and reduce methane emissions of dairy cattle through genetic selection and management understanding the basis for dairy cows which are divergent in feed efficiency is necessary. A database of the genetic and production information, including enteric methane emissions, has been compiled for more than 15 years and will continue to facilitate extensive analysis. Additionally, these highly phenotyed animals will be used to assess changes in tissues known to affect efficient use of nutrients. Methods to temporarily isolate regions of small intestine of live, adult cows will be established to study direct nutrient effects on gut function and gene expression. Moreover, epigenetic factors controlling rumen development during weaning will be investigated using state-of-the-art molecular technologies. Finally, changes in gastrointestinal cells of dairy cows related to gut growth and function during critical stages of production will be characterized by examining gene expression in gut tissues of cows under different dietary and production conditions over time.

Progress Report
Progress was made on two of the three objectives of project 8042-31310-114-000D (Improving Dairy Cow Feed Efficiency and Environmental Sustainability Using Genomics and Novel Technologies to Identify Physiological Contributions and Adaptations). This represents the third year of this project. Under Objective 1, along with gas and growth assessments, blood plasma was collected monthly from cows during the trial for analysis of indicators of inflammation, metabolism, and stress. Genomic DNA was isolated from each heifer for high- density single-nucleotide polymorphism genotyping using the Illumina BovineHD Genotyping BeadChip with over 777,000 markers. The data collected from prior years have been added to a database with information from over 400 growing heifers from the same herd to investigate the biological and genetic bases of nutrient use efficiency and to support genomic selection studies. Holstein dairy cows (n=48) from the BARC herd also were evaluated for feed efficiency during the first 100 days of lactation using RFI estimates and associated production measures. Those data were added to a database with distinct records from over 1,710 heifers and lactating cows from the same herd and are being used to investigate the genetic basis of feed efficiency in dairy cattle. Through cooperation with multiple US universities, a combined database has been developed which contains greater than 9,000 feed intake trials for the assessment of dairy cattle size on dry matter intake and used to make predictions on expected feed intake based on live bodyweight assessments. And further, in conjunction with the RFI assessments, enteric methane emissions of 350 dairy cows from 100 to 150 days of lactation have been evaluated as well as their residual feed intake to gain a better understanding of the relationship between feed efficiency during lactation as part of a $2 million Foundation for Food and Agriculture grant with Michigan State University and greenhouse gas emissions of dairy cows as part of a $2 million Greener Cattle initiative with University of Wisconsin. Finally, this data has also been shared with international partners as part of a multimillion-dollar grant led by investigators at the University of Guelph in Guelph, Canada, to improve genetic selection for feed-efficient dairy cattle. Under Objective 3, 47 Holstein dairy cows phenotypically described for RFI lac were euthanized for the collection of 57 distinct animal tissue and microbiome samples (plus 12 fetal tissues when present) in support of USDA- ARS scientists from three research units in Beltsville, Maryland and one unit in Madison, Wisconsin, as well as university collaborators (University of Delaware (1), University of Pennsylvania Vet Med (1), University of Wisconsin (2), UC Davis Vet Med.(1), University of Kentucky (1) and UMD (1)). For scientists at Beltsville this comprehensive data set is being used to provide a complete resource for the development of the FarmGTex atlas for basic functional knowledge of genome function to decipher the genotype-to-phenotype (G2P) link in farmed animals. Systematic characterization of molecular states of cells in livestock tissues is essential for understanding cellular and genetic mechanisms underlying economically and ecologically important physiological traits. As a part of a coordinated international action to accelerate genome to phenome, a comprehensive Cattle Cell Atlas (CattleCA) by generating and analyzing single-cell/nucleus RNA sequencing (sc/snRNA-seq) data has been built. Using Stereo-seq technology, we generated high-resolution spatial transcriptomic maps of the cow embryo and adult brain, as well as gastrointestinal tissues including the liver, rumen, and abomasum. Ongoing analyses aim to identify distinct cell types and states, uncover key regulators of cell differentiation, and map gene networks that shape tissue structure and function. These insights will provide a foundational framework for understanding bovine development and disease. A comprehensive cattle cell atlas is an invaluable resource for cattle genetics and genomics, developing cell-based food systems, and evolutionary cell biology.

Accomplishments
1. Defining the gastrointestinal response to lactation in high producing dairy cattle. Holstein cattle are a cornerstone of global dairy production, providing a substantial portion of the milk and dairy products consumed worldwide. To better understand how these animals meet the intense physiological demands of lactation, USDA-ARS scientists in Beltsville, Maryland, conducted a comprehensive study of to identify all biologically active RNA gene products (transcriptome) present in key gastrointestinal tissues, including rumen, duodenum, and colon, across the full lactation cycle. Using state-of-the-art technology (RNA-seq), epithelial cells from these tissues at eight critical time points were analyzed, spanning early lactation through mid and late stages to the dry period, in five multiparous cows. The study revealed changes occurring over time as well as physiological status underlying nutrient digestion, absorption, and immune regulation that support milk production. This resource offers one of the most detailed views to date of the gastrointestinal transcriptomic landscape in dairy cattle, providing essential insights for researchers future genetic and functional studies aimed at improving lactation efficiency and animal performance.

Review Publications
Gao, Y., Liu, G., Ma, L., Li, C., Baldwin, R.L. 2025. A resource of longitudinal RNA-seq data of Holstein cow rumen, duodenum, and colon epithelial cells during the lactation cycle. BMC Genomic Data. 26. Article e9. https://doi.org/10.1186/s12863-025-01295-5.
Wang, X., Ding, Y., Li, C., Ma, Q., Shi, Y., Liu, G., Li, C., Kang, X. 2024. Effects of rumen metabolite butyric acid on bovine skeletal muscle satellite cells proliferation, apoptosis, and transcriptional states during myogenic differentiation. Domestic Animal Endocrinology. https://doi.org/10.1016/j.domaniend.2024.106892.
Su, Z., Li, C., Yang, C., Ding, Y., Zhou, X., Xu, J., Qu, C., Shi, Y., Li, C., Kang, X. 2025. Identification of single nucleotide polymorphisms (SNPs) potentially associated with residual feed intake in Qinchuan beef cattle by hypothalamus and duodenum RNA-Seq data. PeerJ. 13. Article e19270. https://doi.org/10.7717/peerj.19270.
Cavani, L., Parker Gaddis, K.L., Baldwin, R.L., Santos, J.E., Koltes, J.E., Tempelman, R.J., VandeHaar, M.J., White, H.M., Penagaricano, F., Weigel, K.A. 2024. Genetic characterisation of feeding patterns in lactating Holstein cows and their association with feed efficiency traits. Journal of Animal Breeding and Genetics. https://doi.org/10.1111/jbg.12911.
Kramer, A.E., Ellwood, K., Guarina, N., Li, C., Dutta, A. 2024. Transcriptomic data reveals MYC as an upstream regulator in laying hen follicular recruitment. Poultry Science. https://doi.org/10.1016/j.psj.2024.104547.
Wang, X., Zhou, X., Li, C., Qu, C., Shi, Y., Li, C., Kang, X. 2024. Integrative analysis of whole genome bisulfite and transcriptome sequencing reveals the effect of sodium butyrate on DNA methylation in the differentiation of bovine skeletal muscle satellite cells. Genomics. https://doi.org/10.1016/j.ygeno.2024.110959.
Cavani, L., Novo, L.C., Reyes, F.S., Nascimento, B.M., VandeHaar, M.J., Tempelman, R.J., Parker Gaddis, K.L., Baldwin, R.L., Santos, J.E., Koltes, J.E., White, H.M., Weigel, K.A., Penagaricano, F. 2025. Associations between body temperature and feed efficiency traits in lactating Holstein cows. Journal of Dairy Science Communications. 6(2):256-260. https://doi.org/10.3168/jdsc.2024-0701.
Fang, L., Teng, J., Lin, Q., Bai, Z., Liu, S., Guan, D., Li, B., Gao, Y., Hou, Y., Gong, M., Pan, Z., Yu, Y., Clark, E., Smith, J., Rawlik, K., Xiang, R., Chamberlain, A.J., Goddard, M.E., Littlejohn, M., Larson, G., Machugh, D.E., O’Grady, J.F., Sorensen, P., Sahana, G., Lund, M., Jiang, Z., Pan, X., Gong, W., Zhang, H., He, X., Zhang, Y., Gao, N., He, J., Yi, G., Liu, Y., Zhao, P., Zhou, Y., Wang, X., Young, R.S., Xia, C., Cheng, H., Ma, L., Cole, J.B., Baldwin, R.L., Li, C., Van Tassell, C.P., Lunney, J.K., Liu, W., Guan, L., Zhao, X., Ibeagha-Awemu, E., Luo, O., Lin, L., Canela-Xandri, O., Derks, M., Crooijmans, R., Godia, M., Madsen, O., Groenen, M., Koltes, J.E., Tuggle, C.K., Mccarthy, F.M., Rocha, D., Amills, M., Clop, A., Ballester, M., Tosser-Klopp, G., Li, J., Fang, C., Fang, M., Wang, Q., Hou, Z., Wang, Q., Zhao, F., Jiang, L., Zhao, G., Zhou, Z., Zhou, R., Liu, H., Li, M., Mo, D., Liu, X., Chen, Y., Yuan, X., Li, J., Zhao, S., Ding, X., Sun, D., Sun, H., Li, C., Jiang, Y., Wu, D., Wang, W., Fan, X., Zhang, Q., Li, K., Yang, N., Hu, X., Liu, G. 2025. The farm animal genotype-tissue expression (FarmGTEx) project. Nature Genetics. https://doi.org/10.1038/s41588-025-02121-5.
Gao, Y., Liu, G., Ma, L., Fang, L., Li, C., Baldwin, R.L. 2024. Transcriptomic profiling of gastrointestinal tracts in dairy cattle during lactation reveals molecular adaptations for milk synthesis. Journal of Advanced Research. https://doi.org/10.1016/j.jare.2024.06.020.
Gao, Y., Yang, L., Kuhn, K.L., Li, W., Zanton, G.I., Bowman, M.E., Zhao, P., Zhou, Y., Fang, L., Cole, J.B., Rosen, B.D., Ma, L., Li, C., Baldwin, R.L., Van Tassell, C.P., Zhang, Z., Smith, T.P., Liu, G. 2025. Long read and preliminary pangenome analyses reveal breed-specific structural variations and novel sequences in Holstein and Jersey cattle. Journal of Advanced Research. 79:137-150. https://doi.org/10.1016/j.jare.2025.04.014.
Hu, Z., Boschiero, C., Neupane, M., Bhowmik, N., Yang, L., Kilian, L., Dejarnette, M., Sargolzaei, M., Harstine, B., Li, C., Tuo, W., Baldwin, R.L., Van Tassell, C.P., Sattler, C.G., Liu, G.E. 2025. Exploring tissue- and sex-specific DNA methylation in cattle using a pan-mammalian infinium array. International Journal of Molecular Sciences. 26(9).Article e4284. https://doi.org/10.3390/ijms26094284.
Boschiero, C., Neupane, M., Yang, L., Schroeder, S.G., Tuo, W., Ma, L., Baldwin, R.L., Van Tassell, C.P., Liu, G. 2024. A pilot detection and associate study of gene presence-absence variation in Holstein cattle. Animals. https://doi.org/10.3390/ani14131921.
Wu, X., Caputo, M.J., Wiggans, G.R., Norman, H.D., Miles, A.M., Van Tassell, C.P., Baldwin, R.L., Schutz, M.M., Burchard, J., Durr, J.W. 2025. Transforming the tradition of discrete milk yield correction factors: A continuous 1-step DeLorenzo-Wiggans method. Journal of Dairy Science Communications. 6(1):84-89. https://doi.org/10.3168/jdsc.2024-0583.