Objective 1: Develop resources, tools, and selectable markers to improve nutrient use efficiency in dairy cattle. Tools and resources will be developed including: 1) genetically and phenotypically characterized lines of cattle divergently selected for feed efficiency to support genomic selection for greater efficiency and lower CH4 emissions, and identify possible negative consequences of selection on production performance; 2) whole tissue models of cattle intestine (mini-guts, or enteroids) to support tissue-specific investigations ex vivo; and 3) an ‘isolated’ small intestine model to study effects of specific nutrients on intestinal development, function, and gene expression of mature dairy cows in vivo. Sub-objective 1.A: Develop and phenotypically characterize lines of Holstein dairy cattle divergently selected for RFI during growth (RFIgrowth) to investigate the biological and genetic bases of nutrient use efficiency, and to support genomic selection studies. Sub-objective 1.B: Characterize and exploit relationships between RFI (RFIgrowth and RFIlac) and enteric CH4 emissions of dairy cattle. Sub-objective 1.C: Develop ruminant organoids to study gut health and nutrient use efficiency of dairy cattle ex vivo. Sub-objective 1.D: Develop and validate a short-term isolated duodenal model for the assessment of intestinal epithelial tissue transcriptomic response to alterations in nutrient delivery in vivo. Objective 2: Evaluate and develop novel dietary strategies to reduce feed and nutritional costs to dairy cattle production. Studies of newborn dairy calves will be conducted to characterize molecular changes controlling gene expression during rumen development and differentiation, and evaluate novel feed additives as alternatives to antibiotics to improve calf health and production performance. Sub-objective 2.A: Evaluate the effects of non-nutritive feed additives (e.g., phytochemicals) on gut health and nutrient use efficiency of dairy cattle. Sub-objective 2.B: Characterize molecular phenotypes of the calf rumen transcriptome through strand-specific RNA sequencing (ssRNA-seq) during development. Sub-objective 2.C: 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: Evaluate in vivo gastrointestinal tissue responses (ruminal and duodenal) of lactating and dry dairy cows to perturbations in luminal factors (changes in nutrient flow) and physiological stressors (transition cow and early lactation). Molecular mechanisms regulating cell proliferation and development of rumen and intestinal epithelia during critical changes in nutrient delivery and the dairy cow production cycle (transition into lactation, ration changes, stage of lactation) will be identified and characterized through transcriptomic studies of serial biopsies from live animal models. Subobjective 3A, 3B- See project plan
To improve feed efficiency and reduce methane emissions of dairy cattle through genetic selection and management, dairy cows divergent in feed efficiency will be developed, and a database of their genetic and production information, including enteric methane emissions, will be compiled for extensive analysis. Whole tissue models of intestine (mini-guts) will be developed from calves to study gut function and nutrient use, and methods to temporarily isolate regions of small intestine of live, adult cows will be established to study nutrient effects on gut function and gene expression. In addition, novel plant-derived compounds will be evaluated as alternatives to antibiotics to improve gut function, disease resistance, and feed efficiency of dairy calves. Epigenetic factors controlling calf 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 was made on all three objectives of project 8042-31310-078-00D (Improving Feed Efficiency and Environmental Sustainability of Dairy Cattle through Genomics and Novel Technologies). Under Objective 1 (develop resources, tools, and selectable markers to improve nutrient use efficiency in dairy cattle), an additional 69 Holstein dairy heifers from the Beltsville Agricultural Research Center (BARC) herd were evaluated for feed efficiency during a 91-day growth trial using an estimate called residual feed intake (RFI). Daily enteric methane and carbon dioxide production also were measured for each heifer using an automated monitoring system called GreenFeed to determine relationships between feed efficiency of dairy cattle and their contribution to greenhouse gas emissions. Blood plasma was collected monthly from heifers during the growth trial to analyze inflammation, metabolism, and stress indicators. Genomic DNA was isolated from each heifer for high-density genotyping using an array with over 777,000 markers. The collected data were added to a database with information from over 300 growing heifers from the same herd to investigate nutrient use efficiency's biological and genetic bases and support genomic selection studies. All heifers are being categorized as either high- or low-RFI and bred to specific Holstein sires with high or low genetic merit for RFI to develop lines of Holstein cattle divergently selected for feed efficiency. Holstein dairy cows 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 information from over 1,400 lactating cows from the same herd and are being used to investigate the genetic basis of feed efficiency in dairy cattle. All data were 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. Enteric methane emissions of dairy cows from 100 to 150 days of lactation were evaluated as well as their residual feed intake to gain a better understanding of the relationship between feed efficiency during lactation and greenhouse gas emissions of dairy cows as part of a $2 million Foundation for Food and Agriculture grant with Michigan State University. Under Objective 3 (evaluate in vivo gastrointestinal tissue responses of lactating and dry dairy cows to perturbations in luminal factors and physiological stressors), tissue biopsies from 118 samples from lactating cows at the longitudinal lactation stage have been collected and sequenced for transcriptomic analysis. Collected samples were from three tissue sources (rumen, duodenum, and colon) representative of the lactation cycle to assess changes as lactational needs and rations change. The RNA sequencing data currently are being processed. Single-cell RNA sequencing was performed for Holstein ruminal epithelial cells during weaning and is the first reported single-cell transcriptomic analysis in cattle. Rumen single-cell transcriptomes were successfully generated and revealed major and some novel cell types. Cell cycles, components, relative timings, and regulatory networks of rumen epithelial cells were characterized along with co-expression and gene function patterns. With the proposed cell lineage development model, six-cell types identified across their temporal and spatial distributions were revealed and appear to be correlated with the underlying layers, structures, and functions of the rumen epithelium.
1. First dairy cattle ruminal cell atlas using single-cell RNA sequencing technology. Rumen epithelial cell differentiation and development during weaning in newborn ruminants is necessary for the efficient digestion of solid feed and optimal growth performance. Because of the ruminal wall's complex and variable physical composition, investigating specific effects of changing ruminal environments on all aspects of rumen functions has been challenging. Recent breakthroughs in single-cell RNA-sequencing technologies enabled ARS researchers in Beltsville, Maryland, to establish an atlas of ruminal cells for dairy cattle at single-cell resolution. Scientists were able to clarify the complexity of rumen epithelial diversity and explain cell identity, fate, and function. This first effort in implementing single-cell RNA sequencing technologies in cattle opens the door for discoveries about the roles of tissue and cell types in complex traits at single-cell resolution.
Gao, Y., Fang, L., Baldwin, R.L., Connor, E.E., Cole, J.B., Van Tassell, C.P., Ma, L., Li, C., Liu, G. 2021. Single-cell transcriptomic analyses of cattle ruminal epithelial cells before and after weaning. Genomics. 113(4):2045-2055. https://doi.org/10.1016/j.ygeno.2021.04.039.
Zhou, Y., Liu, S., Hu, Y., Fang, L., Gao, Y., Xia, H., Schroeder, S.G., Rosen, B.D., Connor, E.E., Li, C., Baldwin, R.L., Cole, J.B., Van Tassell, C.P., Yang, L., Ma, L., Liu, G. 2020. Comparative whole genome DNA methylation profiling of cattle tissues reveals global and tissue-specific methylation patterns. BMC Biology. 18(1):85. https://doi.org/10.1186/s12915-020-00793-5.