Publications

The rumen microbiome is critical in ruminant nutrition and contributes to nutrient utilization and feed efficiency in cattle. Therefore, the objective of this study was to interrogate microbial and biochemical factors affecting divergences in feed efficiency in Angus steers using 16S amplicon sequencing and untargeted metabolomics. Average residual feed intake (RFI) was calculated, and steers were divided into low- and high-RFI groups. Features were ranked in relation to RFI through supervised machine learning on microbial and metabolite compositions. Residual feed intake was associated with several attributes of the rumen bacterial community. Low-RFI steers were associated with decreased bacterial α- (P=0.03) and β-diversity (P<0.001). Several serum metabolites were associated with RFI. Based on fold change (high/low RFI), low-RFI steers had greater abundances of pantothenate (P=0.02). Machine learning on RFI was predictive of both serum metabolomic signature and rumen bacterial composition (AUC ≥0.7). Log-ratio proportions of the bacterial classes Flavobacteriia over Fusobacteriia were enriched in low-RFI steers (F=6.8, P=.01). Greater proportions of pantothenate-producing bacteria, such as Flavobacteriia, and/or reductions in Fusobacteriia may result in improved nutrient utilization in low-RFI steers. Pantothenate and Flavobacteriia may serve as potentially novel biomarkers to assess or predict feed efficiency in Angus steers.

Untargeted metabolomics can provide insight as to the physiological health of an animal; but is currently underutilized in animal systems, particularly with integration of multiple physiological systems. The objective of this study was to determine differences between rumen fluid and serum metabolomes using an untargeted metabolomics approach in order to provide a foundation for future research on rumen function and systemic metabolite utilization. Two fistulated Black Angus cows of approximately six years of age weighing 603.406 ± 37.5 kg were placed on a primarily corn silage-based diet and acclimated to the diet for a two week period. Feed intake was monitored using the GrowSafe system©. Rumen fluid was collected via rumen cannula and blood taken from the coccygeal vein at the beginning of the study and weekly for 12 weeks for a total of 26 samples from each animal. Untargeted serum metabolomics was conducted utilizing the Dionex UltiMate 3000 UPLC system and electrospray ionization was used to introduce the samples into an Exactive Plus Orbitrap MS. A total of 156 known metabolites were identified using MAVEN. Metabolites were median-centered, log-transformed, and range-scaled using Metaboanalyst 4.0. Orthogonal PLSDA showed distinct clustering of serum and rumen fluid metabolites with no overlap (R2Y = 0.987; Q2 = 0.952) with 1000 permutations. A total of 143 metabolites differed between serum and rumen fluid as analyzed by t-test with Benjamini-Hochberg FDR (P ≤ 0.05). The greatest fold metabolite differences (rumen fluid/serum) included 3-methylphenylacetic acid (66.1; P < 0.0001), kynurenine (44.2; P < 0.0001), kynurenic acid (86.2; P < 0.0001), indole (0.0146; P < 0.0001), and cholesterol sulfate (0.00156; P < 0.0001). These data provide evidence of metabolite disparity between blood and rumen content. Further studies may use this information and techniques to account for physiological differences in animals that may contribute to production-important phenotypes.

Beef is the most consumed red meat in the United States, and the US is the largest producer and consumer of beef cattle globally. Feed is one of the larges

Lolium arundinaceum [(Darbyshire) tall fescue] toxicosis is responsible for substantial beef production losses in the United States, due to its negative effects on reproduction, growth, and feed efficiency. These effects are consequences of toxic alkaloids within tall fescue. Interseeding legumes, such as Trifolium pratense (red clover), into pastures has been shown to mitigate a portion of these effects. Clovers contain isoflavones, which may play a role in tall fescue toxicosis mitigation. The present study utilized 36 Angus steers to determine the effects of daily supplementation with a red clover-isolated isoflavone feed additive on physiological symptoms of tall fescue toxicosis and the rumen microbial environment over a 21-d period. Angus steers were initially stratified based upon their single nucleotide polymorphism genotype at the DRD2 receptor. Treatments were then randomly assigned in a 2 × 2 factorial arrangement within a completely randomized design, where treatment factors consisted of tall fescue seed type (endophyte-infected tall fescue seed vs. endophyte-free tall fescue seed) supplemented with and without the isoflavone additive. Steers that consumed endophyte-infected tall fescue seed had lower serum prolactin concentrations (P = 0.0007), average daily gain (ADG; P = 0.003), final body weight (BW; P = 0.004), and feed efficiency (P = 0.018) when compared with steers that consumed endophyte-free tall fescue seed. Serum insulin-like growth factor-1 (IGF-1) tended to be reduced with supplementation of isoflavones (P = 0.06) but was unaffected by seed type (P ≥ 0.10) and seed by treatment interaction (P ≥ 0.10). Isoflavones reduced serum glucose levels (P = 0.023), but neither seed type, isoflavones, or their interaction affected serum urea nitrogen (SUN), nonesterified fatty acids (NEFA), or insulin (P ≥ 0.10). Volatile fatty acid concentrations, dry matter intake (DMI), ruminal pH, and overall feeding behaviors were also unaffected by seed type or isoflavone treatments (P ≥ 0.10). Twenty-eight ruminal bacteria taxa shifted as a result of seed type or isoflavone treatment (P < 0.05). In this experiment, feeding isoflavones to Angus cattle did not completely mitigate all symptoms of fescue toxicosis. However, dose-response trials may aid future research to determine if dietary supplementation with isoflavones alleviates fescue toxicosis symptoms and promotes livestock growth and performance.

ARG allele predictions. A listing of all predicted candidate antibiotic resistance gene (ARG) alleles in the short- and long-read assemblies. (XLSX 17 kb)

This study determined potential microbial and metabolic biomarkers of feed efficiency in Angus heifers. Seventeen ruminal cannulated Angus heifers underwent a 70-day feed efficiency trial. Residual feed intake was used to determine high and low feed efficient heifers. On day 70 of the trial, rumen content and blood were collected and used for microbial and metabolomic analyses, respectively. Bacterial populations were examined by targeting the V4 region of the 16S rRNA gene and analyzed using QIIME and SAS. Rumen fluid and serum metabolites were analyzed using MetaboAnalyst. No microbial taxa differed after false discovery rate correction, but seven did differ (p ≤ 0.05) prior to correction, including Lachnospiraceae (Other), Desulfobulbaceae, Neisseriaceae, Shuttleworthia, Corynebacterium, p-75-a5, and L7A-E11. No differences were observed in alpha diversity metrics. Beta diversity utilizing unweighted UniFrac distances analyzed via PERMANOVA was significant (p = 0.03). Several metabolites in rumen fluid metabolites were correlated with bacteria that differed by feed efficiency phenotype. The metabolites correlated with bacteria were primarily involved in nutrient signaling and microbial crude protein availability. These data suggest variation in the availability of nutrients, primarily amino acids, as well as a relationship among microbiota, metabolome, and host feed efficiency phenotypes in heifers.

Hypergeometric test of contig alignment depth. These are the results of an enrichment test designed to identify differences in community abundance/composition between several public rumen datasets (see Additional file 1: Supplementary methods). The short-read assembly and long-read assembly results are listed on separate tabs. Enrichment was determined by the Hypergeometric mean test using a Benjamini-Hochberg-corrected alpha. (XLSX 224 kb)

Abstract Tall fescue [Schedonorus arundinaceus (Shreb.) Dumort.; (TF)] is the primary forage species used by cow-calf producers in grazing systems in Tennessee. It is an excellent cool-season perennial grass due to its great forage mass (FM), nutritive value, and extensive growing season. Orchardgrass [Dactylis glomerata L.; (OG)] is also a widely used cool-season grass in the U.S. This study compared TF and OG cultivars under continuous stocking in terms of FM, nutritive value, and animal performance over two grazing seasons (2022 to 2023) in Spring Hill, Tennessee, U.S. Treatments were: 1) OG cv. Persist I (OG-P1), 2) OG cv. Persist II (OG-P2), 3) TF cv. Kentucky 31’ (TF-K31), and 4) a novel endophyte TF cv. Max Q (TF-NE). Forage mass was not affected by treatments (P = 0.0519), with an average of 2979 kg DM ha−1. Crude protein (CP) and neutral detergent fiber (NDF) concentrations were also unaffected by treatments (P = 0.6728, P = 0.1300), averaging 106 and 687 g kg–1, respectively. However, TF-NE had the lowest acid detergent fiber (ADF) (P = 0.0258; 380 g kg–1) and the greatest in vitro dry matter digestibility in 48 hours (INVDMD48) (P &amp;lt; 0.0001; 680 g kg–1), while OG-P2 had the greatest ADF (400 g kg–1) and the lowest INVDMD48 (661 g kg–1). Total gain and average daily gain (ADG) were affected by treatment × year interaction (P = 0.0314 and P = 0.0161, respectively). In 2022, TF-NE, TF-K31, and OG-P1 outperformed OG-P2, but no differences were observed in 2023 (total gain: 78.6 kg animal–1; ADG: 0.78 kg animal–1 day–1). Ergovaline concentrations in TF-K31 were low, which likely minimized its negative effects. These findings indicate that both TF and OG can sustain beef cattle production under continuous stocking, with TF offering potential advantages in botanical composition and forage nutritive value over time.

Once thought to be sterile, the reproductive tract microbiome has been characterized due to the transition from culture-dependent identification of bacteria to culture-independent sequencing methods. The urogenital microbiome was first identified in women through the Human Microbiome Project, which led to research in other species such as the bovine. Previous research focused on uterine bacteria associated with postpartum disease, but next generation sequencing methods identified a normal, healthy bacterial community of the reproductive tract of cows and heifers. Bacterial communities are now understood to differ between the uterus and vagina, and throughout the estrous cycle with changes in hormone dominance. In a healthy state, the bacterial communities largely interact with the uterine environment by assisting in maintaining the proper pH, providing and utilizing nutrients and metabolites, and influencing the immunological responses of the reproductive tract. If the bacterial communities become unbalanced due to an increase in potentially pathogenic bacteria, the health and fertility of the host may be affected. Although the presence of a reproductive tract microbiome has become widely accepted, the existence of a placental microbiome and in utero colonization of the fetus is still a popular debate due to conflicting study results. Currently, researchers are evaluating methods to manipulate the reproductive bacterial communities, such as diet changes and utilizing probiotics, to improve reproductive outcomes. The following review discusses the current understanding of the reproductive tract microbiome, how it differs between humans and cattle, and its relationship with the uterine environment.

Tall fescue toxicosis negatively impacts blood flow, elevates body temperature, and reduces beef cattle’s average daily gain (ADG). In previous studies, isoflavones have diminished the symptoms of tall fescue toxicosis in ruminants. Therefore, this dataset determined the impact of low concentrations of isoflavone doses on animal vasculature, body temperature, ADG, and rumen microbial communities in beef cattle. A 21-day experiment with Angus cattle consisted of four isoflavone doses: 0 g, 2 g, 4 g, and 6 g, along with a control group. Isoflavones were mixed with 0.5 kg of dried distiller’s grains (DDGs). Daily individual rectal temperatures were recorded. Weekly blood serum was collected via coccygeal venipuncture, blood vasculature data were measured via color Doppler ultrasound, and body weight (BW) was recorded. Approximately 100 mL of rumen content was collected at the end of the trial. The pulsatility index (PI) decreased in the control group compared to the 2 g and 4 g groups (p = 0.01). Animals in the isoflavone treatment groups recorded a higher rectal temperature (p &lt; 0.05). ADG was reduced in animals undergoing isoflavone treatments (p &lt; 0.001). Finally, there was no impact on the rumen microbial communities (p &gt; 0.05). Isoflavone supplementation may mitigate tall fescue toxicosis and improve animal performance at greater doses.

Beef cattle is the most consumed red meat in the United States, and the United States is the largest producer and consumer of beef cattle globally. Feed is one of the largest input costs for the beef cattle industry, accounting for 40–60% of the total input costs. Identifying methods for improving feed efficiency in beef cattle herds could result in decreased cost to both producers and consumers, as well as increased animal protein available for global consumption. Identification of feed efficiency biomarkers could aid in the selection of more feed-efficient animals. To identify potential biomarkers of feed efficiency, rumen fluid was collected from low- (n = 14) and high-RFI (n = 15) Angus steers based on RFI ± 0.5 SD around the mean following a 70-d feed efficiency trial to determine rumen metabolites associated with variations in feed efficiency. Steers were 7 mo of age, weighing 264±2.7 kg. Rumen fluid was collected via gastric tubing, filtered through a 0.22 µM syringe filter, extracted for metabolomic analysis using 0.1% formic acid in acetonitrile:water:methanol (2:2:1), and injected into the Dionex UltiMate 3000 UHPLC system with an Exactive Plus Orbitrap MS. Peaks for known metabolites were identified using MAVEN and analyzed using MetaboAnalyst 4.0 and SAS. Significance was determined using an α ≤ 0.05. Based on fold change (high/low RFI), eight metabolites were greater in low-RFI steers compared to high-RFI steers, including 3,4-dihydroxyphenylacetate, 4-pyridoxate, citraconate, hypoxanthine, succinate/methylmalonate, thymine, uracil, and xylose (P ≤ 0.05), while 12 metabolites differed by ≥ 2-fold. These metabolites were predominantly involved in amino acid and lipid metabolism. Rumen fluid metabolomes differed in steers of varying feed efficiencies. These metabolites may be used as biomarkers of feed efficiency and may provide insight as to factors contributing to differences in feed efficiency that may be exploited to improve feed efficiency in beef cattle herds.

The milk microbiome in swine has been recently demonstrated to develop and alter the growing gut microbiome in offspring. Milk is the primary nutritional source for piglets and the established microbial communities from initial diet remain stable over time, thus being critical for long-term effects on the animal. Since little is known about the ontogeny of the milk microbiome, the objective of this study was to determine differences in the sow milk microbiome among multiple parities and days in lactation. A total of 32 sows were grouped by parity, 1–4, with 8 sows per parity. Sows were on the same diet and had ad libitum access to water. At sampling, teats were cleaned and stripped. Milk samples were manually collected on one side from a fore, mid, and hind teat, then combined for one whole milk sample per animal. Milk was sampled on days 2 and 16 to determine differences between days in lactation. Samples were sequenced using an Illumina MiSeq for V1-V3 regions of the bacterial 16S rRNA gene and downstream analyses were conducted in the R environment using ‘phyloseq’ and ‘DADA2’ packages. Statistical analyzes utilized a completely randomized design and a mixed model analysis of variance, with fixed effects of parity and days in lactation and examining a parity by day interaction. The genera Corynebacterium were greater (P &amp;lt; 0.05) abundance in parity 4. There was an increase (P &amp;lt; 0.05) in Staphylococcus at day 2 compared to day 16. Core microbial communities, Lactobacillus and Romboutsia increased (P &amp;lt; 0.05) by day 16. Identifying microbial communities present in the milk microbiome in sows can allow producers to understand how these microbes may influence the gut microbiome in growing piglets to augment gain. USDA is an equal opportunity provider and employer.