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.

Negative impacts of endophyte-infected Lolium arundinaceum (Darbyshire) (tall fescue) are responsible for over $2 billion in losses to livestock producers annually. While the influence of endophyte-infected tall fescue has been studied for decades, mitigation methods have not been clearly elucidated. Isoflavones found in Trifolium pratense (red clover) have been the subject of recent research regarding tall fescue toxicosis mitigation. Therefore, the aim of this study was to determine the effect of ergovaline and red clover isoflavones on rumen microbial populations, fiber degradation, and volatile fatty acids (VFA) in an in vitro system. Using a dose of 1.10 mg × L-1, endophyte-infected or endophyte-free tall fescue seed was added to ANKOM fiber bags with or without 2.19 mg of isoflavones in the form of a control, powder, or pulverized tablet, resulting in a 2 × 3 factorial arrangements of treatments. Measurements of pH, VFA, bacterial taxa, as well as the disappearance of neutral detergent fiber (aNDF), acid detergent fiber (ADF), and crude protein (CP) were taken after 48 h of incubation. aNDF disappearance values were significantly altered by seed type (P = 0.003) and isoflavone treatment (P = 0.005), and ADF disappearance values were significantly different in a seed × isoflavone treatment interaction (P ≤ 0.05). A seed × isoflavone treatment interaction was also observed with respect to CP disappearance (P ≤ 0.05). Eighteen bacterial taxa were significantly altered by seed × isoflavone treatment interaction groups (P ≤ 0.05), eight bacterial taxa were increased by isoflavones (P ≤ 0.05), and ten bacterial taxa were altered by seed type (P ≤ 0.05). Due to the beneficial effect of isoflavones on tall fescue seed fiber degradation, these compounds may be viable options for mitigating fescue toxicosis. Further research should be conducted to determine physiological implications as well as microbiological changes in vivo.

Improving feed utilization in cattle is required to reduce input costs, increase production, and ultimately improve sustainability of the beef cattle indus

The prevalence of reproductive losses costing 1 billion dollars annually to the beef and dairy industries has led researchers to evaluate strategies to improve reproductive efficiency. The microbiome of the reproductive tract has been a recent focus in humans due to its effects on health and fertility; however, little research has evaluated the uterine and vaginal microbiomes of cattle. The present study evaluated the reproductive tract bacterial communities of postpartum beef cows undergoing estrus synchronization and the differences between cows who became pregnant and those who failed to conceive. Forty postpartum Angus cows were subjected to a 7 Day Co-Synch protocol with a pre-synchronization step 21 days prior (day -21) to artificial insemination (AI). Uterine and vaginal flushes were collected on day -21, -9, and -2 of the protocol for pH and bacterial DNA extraction to sequence the V1-V3 hypervariable regions of the 16S rRNA gene. Transrectal ultrasound was performed 30 days after AI for pregnancy diagnosis. Ten pregnant and non-pregnant animals were selected and used for analysis. Decreased numbers of bacterial species present in the uterus of cows were identified, independent of pregnancy status, leading up to AI with an average of 1180 ± 97 species on day -21 to 427 ± 99 on day -2 (P < 0.0001). Principal coordinate analyses depicted significant clustering of bacterial communities by day in the uterus (P = 0.001) and vagina (P = 0.001) as well as pregnancy status in the uterus at day -2 (P = 0.005). Additionally, at day -2, multiple significantly different uterine bacterial genera were present at >1% in non-pregnant cows while present at <1% in pregnant cows. Uterine pH increased in non-pregnant cows but decreased in pregnant cows. In summary, the results suggest uterine bacterial abundances prior to AI may influence the uterine environment and reproductive outcomes.

Bovine respiratory disease (BRD) is one of the most common economic and health challenges to the beef cattle industry. Prophylactic use of antimicrobial drugs can alter the microbial communities in the respiratory tract. Considering that the bovine upper respiratory tract microbiome has been associated with generalized health, understanding the microenvironment that influences this microbiome may provide insights into the pathogenesis of BRD. This study aimed to determine temporal variation in nasopharyngeal (NP) microbiome in naturally occurring BRD in newly received stocker calves. Mixed breed steers ( n = 40) were purchased from an auction market and housed in a commercial stocker farm. Clinical signs were used to identify BRD affected animals, and calves were categorized based on the number of treatments (NumTrt) received (0, 1, 2). On days 0, 7, 14, and 21, NP samples were collected, and subsequent DNA were isolated and sequenced. After sequencing, 16S rRNA V4 gene was amplified and utilized for NP bacterial determination. The difference in relative abundance based on day and NumTrt was measured using repeated measures ANOVA (PROC GLIMMIX; SAS 9.4). Firmicutes, Proteobacteria, Actinobacteriota, Bacteroidota, and Verrucomicrobiota were the top phyla and Mycoplasma , Histophilus , Geobacillus , Saccharococcus , Lactobacillus , and Pasteurella were the top genera. In healthy calves, the relative abundance of Mycoplasma differed by day ( P = 0.01), whereas on day 7, calves had five times greater abundance compared to day 0 (d 0: 0.06 ± 0.05; d 7: 0.30 ± 0.05). No differences were observed in the alpha diversity matrices based on day or NumTrt ( P > 0.05). Results of this study suggest compositional variations in NP microbial populations occur during disease conditions.

The rumen microbiome provides approximately 70% of energy required for the ruminant host through fermentation byproducts. Due to microbial impacts on host health and productivity, several studies have associated microbiome and metabolome variation with important sustainability phenotypes, such as feed efficiency and methane emissions. Recent studies have indicated the microbial communities present in rumen fluid are under low-to-moderate host genetic control. However, microorganisms present on the rumen wall have been underrepresented in research, negating their unique role in host-microbiome interactions. Current studies investigating heritability of the rumen planktonic microorganisms are plagued by small sample sizes, high management and breed heterogeneity, and low resolution for classifying microbial communities. To determine the heritability of the rumen fluid and epithelium-associated microbiome and metabolome, Angus steers (n = 350) were enrolled in a 70-d feed efficiency trial, where real-time dry matter intake was measured. Body weights were collected throughout the trial to calculate average daily gain and residual feed intake. Mid-trial, a novel surgery method was used to swab the rumen wall to capture epithelium-associated microorganisms. On d 70, 50 mL of rumen fluid was obtained from steers using orogastric tubing for collection of planktonic microbial communities. Metagenomic DNA extracted from both rumen fluid and epimural samples was used for reduced representation sequencing to predict microbial features such as diversity, composition, and putative function. Genomic DNA isolated from whole blood was used to perform low-pass whole genome sequencing for use in downstream heritability estimates and variant association analyses. Many core microbial abundances are under low-to-moderate host genetic control (h2 > 0.15; P < 0.05); however, many microorganisms had low heritability estimates (h2 < 0.10). Preliminary genome-wide associations studies are vastly underpowered to detect host genetic variants associated with microbial features, but data generated using the total number of animals will better refine these polymorphisms. Several key heritable metabolites were identified in amino acid metabolism pathways in rumen fluid (P < 0.05). The rumen microbiome appears to act as a complex polygenic trait and incorporating microbial features into prediction models may improve feed efficiency estimations. Understanding host genetic control of the rumen microbiome and metabolome may empower downstream applications to enhance genomic and phenotypic predictions for feed efficiency and other sustainability traits in beef cattle.

Microbiome studies in animal science using 16S rRNA gene sequencing have become increasingly common in recent years as sequencing costs continue to fall and bioinformatic tools become more powerful and user-friendly. The combination of molecular biology, microbiology, microbial ecology, computer science, and bioinformatics—in addition to the traditional considerations when conducting an animal science study—makes microbiome studies sometimes intimidating due to the intersection of different fields. The objective of this review is to serve as a jumping-off point for those animal scientists less familiar with 16S rRNA gene sequencing and analyses and to bring up common issues and concerns that arise when planning an animal microbiome study from design through analysis. This review includes an overview of 16S rRNA gene sequencing, its advantages, and its limitations; experimental design considerations such as study design, sample size, sample pooling, and sample locations; wet lab considerations such as field handing, microbial cell lysis, low biomass samples, library preparation, and sequencing controls; and computational considerations such as identification of contamination, accounting for uneven sequencing depth, constructing diversity metrics, assigning taxonomy, differential abundance testing, and, finally, data availability. In addition to general considerations, we highlight some special considerations by species and sample type.

The use of single-wall carbon nanotubes (SWNTs) in manufacturing and biomedical applications is increasing at a rapid rate; however data on the effects of a potential environmental release of the materials remain sparse. In this study, soils with either low or high organic matter contents as well as pure cultures of E. coli are challenged with either raw as-produced SWNTs (AP-SWNTs) or SWNTs functionalized with either polyethyleneglycol (PEG-SWNTs) or m-polyaminobenzene sulfonic acid (PABS-SWNTs). To mimic chronic exposure, the soil systems were challenged weekly for six weeks; microbial activities and community structures for both the prokaryote and eukaryote community were evaluated. Results show that repeated applications of AP-SWNTs can affect microbial community structures and induce minor changes in soil metabolic activity in the low organic matter systems. Toxicity of the three types of SWNTs was also assessed in liquid cultures using a bioluminescent E. coli-O157:H7 strain. Although decreases in light were detected in all treated samples, low light recovery following glucose addition in AP-SWNTs treatment and light absorption property of SWNTs particles suggest that AP-SWNTs suppressed metabolic activity of the E. coli, whereas the two functionalized SWNTs are less toxic. The metals released from the raw forms of SWNTs would not play a role in the effects seen in soil or the pure culture. We suggest that sorption to soil organic matter plays a controlling role in the soil microbiological responses to these nanomaterials.

Blood chemistry may provide indicators to greater feed efficient cattle. As a side objective to previous research, 17 Angus heifers approximately two years old underwent a feed efficiency trial to determine residual feed intake (RFI) and identify variation in blood chemistry in beef cattle divergent in feed efficiency. Heifers were categorized as high- or low-RFI based ± 0.25 standard deviations around mean RFI. Blood samples were analyzed using an i-STAT handheld blood analyzer to measure sodium, potassium, glucose, blood urea nitrogen (BUN), creatinine, hematocrit, and hemoglobin. BUN was greater in high-RFI heifers (µ = 8.7 mg/dL) contrasted to low-RFI heifers (µ = 6.5 mg/dL; P = 0.01), whereas glucose was greater in low-RFI heifers (µ = 78.1 mg/dL) contrasted to high-RFI heifers (µ = 82.0 mg/dL; P = 0.05). No other blood chemistry parameters differed by RFI. The greater abundance of BUN in high-RFI heifers may indicate inefficient utilization of protein or mobilization of tissue protein for non-protein use. Greater blood glucose concentrations in low-RFI heifers may indicate greater utilization of energy precursors, such as volatile fatty acids, or metabolites. These data suggest there are readily measurable indicators of physiological variation in nutrient utilization; however, this warrants additional studies to explore.

As the global population is expected to exceed 9 billion people by 2050, finding novel methods of improving food production is imperative. 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 Black Angus steers. Fifty Black Angus steers of 7 months of age, weighing 264 ± 2.7 kg were acclimated to the GrowSafe© feeding system for 10d prior to intake measurement, and fed a step-up receiving diet 14d before receiving a growing ration (11.57% CP and 76.93% TDN DM) with 28 mg monensin/kg DM. Steers were maintained on the diet for 70d. Weekly BW was measured, serum collected, and rumen content was obtained via gastric tubing. Based on performance and FI measured from 0 to 70d, the average RFI was calculated and steers were divided into low- (n=14) and high-RFI (n=15) groups based on 0.5 SD below and above the mean RFI, respectively. 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. Genomic DNA was extracted from rumen content and the amplified V1-V3 hypervariable region of the bacterial 16S rRNA gene was sequenced for analyses. Missing values were approximated through matrix completion and data was normalized using a centered log-ratio transformation. Random Forests supervised machine learning and feature selection was performed on the bacterial compositions. Residual feed intake was associated with several attributes of the rumen bacteriome. Low-RFI steers were associated with decreased bacterial α- (P = 0.03) and β- diversity (R2 = 1, 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 (0.375; P = 0.04) and reduced abundances of glucose-6-phosphate (2.13; P = 0.02) and glucose-1-phosphate (2.13; P = 0.03). Machine learning on RFI was highly predictive of both serum metabolomic signature and rumen bacterial composition (accuracy ≥0.7). Fold change Flavobacteriia abundances were greater with increased pantothenate contrasted to reduced pantothenate (5.06; P = 0.04). Greater abundances of pantothenate-producing bacteria, such as Flavobacteriia, may result in improved nutrient utilization in low-RFI steers. Pantothenate and/or Flavobacteriia may serve as potentially novel biomarkers to assess or predict feed efficiency in Black Angus steers on a backgrounding diet.

With the anticipated population growth by 2050, the demand for high-quality protein for human consumption is set to rise. To enhance the sustainability of U.S. cattle production, producers and researchers have traditionally concentrated on improving the feed efficiency of steers through advancements in genetics, nutrition, and microbiome tools, resulting in a more marketable beef product. However, without successful pregnancies, there would be no marketable animals to feed. Despite extensive research on hormonal impacts on cattle physiology, including nutrition and reproduction, there is limited knowledge about how the rumen microbial environment is impacted by pregnancy and feed efficiency in female beef cattle. Understanding the rumen microbiome’s role in feed efficiency and its response to hormonal changes during pregnancy is crucial for advancing sustainable beef cattle production. Therefore, this review highlights the importance of understanding the microbial dynamics in the rumen of pregnant beef females. Although progress has been made, gaps remain in understanding how varying nutritional requirements throughout pregnancy affect the rumen microbiome, highlighting the need for continued research. Addressing these areas will lead to more efficient and sustainable cattle production practices, benefiting beef production and contributing to global food security.

Reproductive inefficiency in cattle has major impacts on overall productivity of cattle operations, increasing cost of production, and impacting the sustainability of the cattle enterprise.Decreased reproductive success and associated disease states have been correlated with the presence of specific microbes and microbial community profiles, yet details of the relationship between microbial communities and host physiology are not well known.The present study profiles and compares the microbial communities in the bovine uterus and vagina using 16S rRNA sequencing of the V1-V3 hypervariable region at the time of artificial insemination.Significant differences (p < 0.05) between the vaginal and uterine communities were observed at the level of α-diversity metrics, including Chao1, Shannon's Diversity Index, and observed OTU.Greater clustering of vaginal OTU was apparent in principal coordinate analysis compared to uterine OTU, despite greater diversity in the vaginal community in both weighted and unweighted UniFrac distance matrices (p < 0.05).There was a significantly greater relative abundance of unassigned taxa in the uterus (p = 0.008), otherwise there were few differences between the overall community profiles.Both vaginal and uterine communities were dominated by Firmicutes, although the relative abundance of rRNA sequences corresponding to species in this phylum was significantly (p = 0.007) lower in the uterine community.Additional differences were observed at the genus level, specifically in abundances within Clostridium (p = 0.009), Anaerofustis (p = 0.018), Atopobium (p = 0.035), Oscillospira (p = 0.035), 5-7N15 (p = 0.035), Mycoplasma (p = 0.035), Odoribacter (p = 0.042), and within the families Clostridiaceae (p = 0.006), Alcaligenaceae (p = 0.021), and Ruminococcaceae (p = 0.021).Overall, the comparison revealed differences and commonalities among bovine reproductive organs, which may be influenced by host physiology.The increased abundance of unassigned taxa found in the uterus may play a significant biological role in the reproductive status of the animal.The study represents an initial dataset for comparing bacterial communities prior to establishment of pregnancy.

Short-read assembly HQ bins. A tab-delimited text file listing the number of contigs, total sizes, and summary statistics for short-read assembly HQ bins. (TAB 4 kb)