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.
Relative to Objective 1, 56 Holstein dairy heifers from the BARC herd were evaluated for feed efficiency using a measurement called residual feed intake measured during a 91-day growth trial (RFIgrowth) as well as their methane emissions from eructations. Information was combined with data collected previously from over 190 growing heifers from the same herd to investigate the biological and genetic bases of nutrient use efficiency, and to support genomic selection studies. Plasma was collected monthly from heifers during the growth trial for analysis of indicators of inflammation, metabolism, and stress. Using RFIgrowth information, 69 females (including 22 extreme “high” and 24 extreme “low” RFIgrowth) were successfully bred (as of early June 2018) to specific sires with “high” and “low” genetic merit for RFI to develop lines of Holstein cattle divergently selected for feed efficiency. The first selected calves will be born in late June 2018. Heifers resulting from these matings will begin to be evaluated in the coming year for growth performance, feed efficiency, and plasma indicators of inflammation, metabolism, and stress. Also relative to Objective 1, relationships between RFIgrowth and methane emissions as well as subsequent feed efficiency measured during the first 100 days of lactation (RFIlactation) are being analyzed on all available data from the BARC herd. Current results from 130 animals show that heifers exhibit 14% divergence in RFIgrowth (i.e., difference of 1.2 kg/d dry matter intake (DMI) between the least and most efficient heifers) and this divergence in RFI is maintained during early lactation at about 5% (i.e., difference of 1.0 kg/d in RFIlactation) with no impact on milk production or cow body weight. Thus, selection based on RFIgrowth in heifers should result in cows with greater feed efficiency during lactation with no changes in milk yield or body weight, meaning substantial reductions in feed costs to the producer without impacting milk profits. Lastly relative to Objective 1, initial isolation of stem cells from small intestine crypts and primary development of enteroids from a single cow intestinal biopsy sample was successful. Creation and maintenance of stable enteroids from multiple individual cows will be attempted in the coming year for additional study and characterization. Relative to Objective 2, a primary cell culture of cattle rumen epithelium was used to initiate chromatin-immunoprecipitation-sequencing (ChIP-seq) and transcriptomic profiling by RNA-seq and ACAT-seq. These data will be used to functionally annotate the calf rumen epigenome and identify transcriptional cis-regulatory modules during development, including histone modification, and chromatin accessibility and architecture. A significant amount of data has been obtained to date, and data analysis is in progress. Our goal is to build a resource that encompasses the effective diversity of the epigenome of cattle rumen development. Relative to Objective 3, duodenal and ruminal cannulation surgeries of cows are ongoing and initial sampling for ruminal and duodenal tissues are underway. Biopsies from duodenally cannulated cows were successfully obtained and subjected to a pilot RNA-Seq experiment to assess the validity of the approach and to support Objective 1 for enteroid development. Results to date demonstrated that developmental functions of the tissue are greatly affected by alterations in intestinal lumen nutrient concentrations. Moreover, putative major transcription factors (PTH, JUN, and WNT) were identified by biological system pathway analyses. Finally, biopsies from cow duodenum were used in short-term incubations and were histologically sectioned to begin characterization of their cellular composition and function.
1. Evaluated changes in the transcriptomes of dairy cow rumen epithelia during milking and non-milking (“dry”) periods. Milk synthesis greatly increases the nutritional demands on a cow. Cow metabolism must adapt to meet this demand and requires major functional adjustments to the digestive tract, including the rumen. The rumen is the largest compartment of the digestive tract where microbial fermentation of the feed occurs, providing energy to the cow to support milk production. ARS scientists at Beltsville, Maryland, using next-generation sequencing technology, assembled and profiled the transcriptome of rumen epithelial tissue collected from milking and nonmilking (“dry”) cows. Transcriptomics profiling and comparison revealed extensive changes in gene expression related to metabolism to support lactation. The work provides crucial knowledge to promote our understanding of functional adaptations of critical tissues that support milk production in dairy cows.
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