Effects of protein concentration and beta-adrenergic agonists on ruminal bacterial communities in finishing beef heifers.

To improve animal performance and modify growth by increasing lean tissue accretion, beef cattle production has relied on use of growth promoting technologies such as beta-adrenergic agonists. These synthetic catecholamines, combined with the variable inclusion of rumen degradable (RDP) and undegradable protein (RUP), improve feed efficiency and rate of gain in finishing beef cattle. However, research regarding the impact of beta-adrenergic agonists, protein level, and source on the ruminal microbiome is limited. The objective of this study was to determine the effect of different protein concentrations and beta-adrenergic agonist (ractopamine hydrochloride; RAC) on ruminal bacterial communities in finishing beef heifers. Heifers (n = 140) were ranked according to body weight and assigned to pens in a generalized complete block design with a 3 × 2 factorial arrangement of treatments of 6 different treatment combinations, containing 3 protein treatments (Control: 13.9% CP, 8.9% RDP, and 5.0% RUP; High RDP: 20.9% CP, 14.4% RDP, 6.5% RUP; or High RUP: 20.9% CP, 9.7% RDP, 11.2% RUP) and 2 RAC treatments (0 and 400 mg/day). Rumen samples were collected via orogastric tubing 7 days before harvest. DNA from rumen samples were sequenced to identify bacteria based on the V1-V3 hypervariable regions of the 16S rRNA gene. Reads from treatments were analyzed using the packages 'phyloseq' and 'dada2' within the R environment. Beta diversity was analyzed based on Bray-Curtis distances and was significantly different among protein and RAC treatments (P < 0.05). Alpha diversity metrics, such as Chao1 and Shannon diversity indices, were not significantly different (P > 0.05). Bacterial differences among treatments after analyses using PROC MIXED in SAS 9 were identified for the main effects of protein concentration (P < 0.05), rather than their interaction. These results suggest possible effects on microbial communities with different concentrations of protein but limited impact with RAC. However, both may potentially act synergistically to improve performance in finishing beef cattle.

Beta-Adrenergic Agonists, Dietary Protein, and Rumen Bacterial Community Interactions in Beef Cattle: A Review.

Improving beef production efficiency, sustainability, and food security is crucial for meeting the growing global demand for beef while minimizing environmental impact, conserving resources, ensuring economic viability, and promoting animal welfare. Beta-adrenergic agonists and dietary protein have been critical factors in beef cattle production. Beta-agonists enhance growth, improve feed efficiency, and influence carcass composition, while dietary protein provides the necessary nutrients for muscle development and overall health. A balanced approach to their use and incorporation into cattle diets can lead to more efficient and sustainable beef production. However, microbiome technologies play an increasingly important role in beef cattle production, particularly by optimizing rumen fermentation, enhancing nutrient utilization, supporting gut health, and enhancing feed efficiency. Therefore, optimizing rumen fermentation, diet, and growth-promoting technologies has the potential to increase energy capture and improve performance. This review addresses the interactions among beta-adrenergic agonists, protein level and source, and the ruminal microbiome. By adopting innovative technologies, sustainable practices, and responsible management strategies, the beef industry can contribute to a more secure and sustainable food future. Continued research and development in this field can lead to innovative solutions that benefit both producers and the environment.

Blood parameters associated with residual feed intake in beef heifers.

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.

The Impacts of Supplemental Protein during Development on Amino Acid Concentrations in the Uterus and Pregnancy Outcomes of Angus Heifers.

Replacement heifer development is one of the most critical components in beef production. The composition of the ideal uterine environment could maximize fertility and reproductive efficiency. Our hypothesis was that protein supplementation would affect the uterine environment of beef heifers without inhibiting development or reproduction. To test the effects of dietary supplementation on these outcomes, a randomized complete block design with repeated measures was implemented. Angus heifers (n = 60) were blocked by body weight (BW) and randomly assigned to one of three supplemental protein treatment groups (10% (CON), 20% (P20), and 40% (P40)). Mixed model ANOVAs were used to determine whether protein supplementation treatments, time, and the interaction or protein supplementation, semen exposure, and the interaction influenced uterine luminal fluid (ULF) and pregnancy outcomes. Amino acids (AAs) were impacted (p < 0.001), specifically, the essential AAs: Arg, Iso, Leu, Val, His, Lys, Met, Phe, Trp. Protein supplementation influenced multiple AAs post-insemination: Arg (p = 0.03), CC (p = 0.05), 1-MH (p = 0.001), and Orn (p = 0.03). In conclusion, protein supplementation did not affect the reproductive development via puberty attainment or the timing of conception even with alterations in growth. However, uterine AA concentrations did change throughout development and protein supplementation influenced ULF d 14 post-insemination, which may affect the conception rates.

Rumen Biogeographical Regions and Microbiome Variation.

The rumen is a complex organ that is critical for its host to convert low-quality feedstuffs into energy. The conversion of lignocellulosic biomass to volatile fatty acids and other end products is primarily driven by the rumen microbiome and its interaction with the host. Importantly, the rumen is demarcated into five distinct rumen sacs as a result of anatomical structure, resulting in variable physiology among the sacs. However, rumen nutritional and microbiome studies have historically focused on the bulk content or fluids sampled from single regions within the rumen. Examining the rumen microbiome from only one or two biogeographical regions is likely not sufficient to provide a comprehensive analysis of the rumen microbiome and its fermentative capacity. Rumen biogeography, digesta fraction, and microbial rumen-tissue association all impact the diversity and function of the entirety of the rumen microbiome. Therefore, this review discusses the importance of the rumen biographical regions and their contribution to microbiome variation.

Preovulatory follicular fluid and serum metabolome profiles in lactating beef cows with thin, moderate, and obese body condition.

Extremes in body condition reduce fertility and overall productivity in beef cattle herds, due in part to altered systemic metabolic conditions that influence the intrafollicular and uterine environment. Follicular fluid and serum metabolome profiles are influenced by body composition in women and dairy cattle; however, such information is lacking in beef cattle. We hypothesized that body condition score (BCS)-related alterations in the metabolome of preovulatory follicular fluid and serum may influence oocyte maturation while impacting the oviductal or uterine environment. Therefore, we performed a study with the objective to determine the relationship between BCS and the metabolome of follicular fluid and serum in lactating beef cattle. We synchronized the development of a preovulatory follicle in 130 cows of varying BCS. We collected blood and performed transvaginal follicle aspirations to collect follicular fluid from the preovulatory follicle ~18 h after gonadotropin-releasing hormone administration to stimulate the preovulatory gonadotropin surge. We then selected follicular fluid and serum samples from cows with BCS 4 (Thin; n = 14), BCS 6 (Moderate; n = 18), or BCS >8 (Obese; n = 14) for ultra-high performance liquid chromatography-high resolution mass spectrometry. We identified differences in the follicular fluid or serum of thin, moderate, and obese animals based on multiple linear regression. MetaboAnalyst 5.0 was used for enrichment analysis of significant metabolites. We identified 38 metabolites in follicular fluid and 49 metabolites in serum. There were no significant differences in follicular fluid metabolite content among BCS classifications. There were 5, 22, and 1 serum metabolites differentially abundant between thin-obese, moderate-thin, and moderate-obese classifications, respectively (false discovery rate [FDR] < 0.10). These metabolites were enriched in multiple processes including "arginine biosynthesis," "arginine/proline metabolism," and "D-glutamine/D-glutamate metabolism" (FDR < 0.04). Pathways enriched with serum metabolites associated with BCS indicate potentially increased reactive oxygen species (ROS) in serum of thin cows. ROS crossing the blood follicular barrier may negatively impact the oocyte during oocyte maturation and contribute to the reduced pregnancy rates observed in thin beef cows.

Extremes in body condition affect fertility and pregnancy outcomes in beef cows. Much research has been done in women and dairy cows to evaluate body condition's effect on oocyte and embryo quality, pregnancy rates, and pregnancy outcomes. However, little work of this type has been done in beef cows and most studies do not focus on the preovulatory time period. The preovulatory time period is an essential time for the oocyte, as final stages of prematuration and the completion of oocyte maturation take place in the peri-ovulatory follicle. The follicular fluid provides the microenvironment for oocyte maturation and exchanges substances with maternal circulation at the blood follicular barrier. Alterations in maternal circulation due to extremes in body condition may pass into the follicular fluid and affect the oocyte during the preovulatory time period. Such conditions may contribute to the reduced fertility seen in beef cows with extreme body condition.

Piceatannol antagonizes lipolysis by promoting autophagy-lysosome-dependent degradation of lipolytic protein clusters in adipocytes.

Overly elevated circulating non-esterified fatty acids (NEFAs) is an emerging health concern of obesity-associated energy disorders. However, methods to reduce circulating NEFAs remain elusive. The present study determined the effect of piceatannol, a naturally occurring stilbene, on adipocyte lipolysis and its underlying mechanism. Differentiated 3T3-L1 adipocytes, brown adipocytes and isolated white adipose tissue were treated with various concentrations of piceatannol for 1.5-h both in the basal and stimulated lipolysis conditions. Piceatannol significantly inhibited NEFAs and glycerol release with a concomitant reduction of ATGL, CGI-58 and PLIN1 expression in adipocytes. Using a series of inhibitor assays, piceatannol-induced degradation of these proteins was found to be mediated by upregulation of the autophagy-lysosome pathway. Moreover, we demonstrated that piceatannol is capable of stimulating autophagy in vitro. Importantly, piceatannol administration tended to lower fasting-induced serum glycerol levels in healthy mice. Furthermore, piceatannol administration lowered lipolysis, central adiposity and hyperinsulinemia in diet-induced obese mice. Our study provides profound evidence of a novel inhibitory role of piceatannol in lipolysis through autophagy-lysosome-dependent degradation of the key lipolytic proteins in adipocytes. This study offers a mechanistic foundation for investigating the potential of piceatannol-containing foods in reducing lipolysis and its associated metabolic disorders.

Considerations and best practices in animal science 16S ribosomal RNA gene sequencing microbiome studies.

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.

Data of bacterial community dynamics resulting from total rumen content exchange in beef cattle.

OBJECTIVES: Extensive efforts have been made to characterize the rumen microbiome under various conditions. However, few studies have addressed the long-term impacts of ruminal microbiome dysbiosis and the extent of host control over microbiome stability. These data can also inform host-microbial symbioses. The objective was to develop preliminary data to measure the changes that occur in the rumen bacterial communities following a rumen content exchange to understand the effects major perturbations may impart upon the rumen microbiome, which may be host-driven. DATA DESCRIPTION: We report here an initial rumen content exchange between two SimAngus (Simmental/Angus) non-pregnant, non-lactating cows of ~ 6 years of age weighing 603.4 ± 37.5 kg. To measure bacterial community succession and acclimation following the exchange, rumen content was collected via rumen cannula at the beginning of the study immediately prior to and following the rumen content exchange, and weekly for 12 weeks. The V4 hypervariable region of the 16S rRNA gene was targeted for DNA sequencing and bacterial analysis. Over 12 weeks, numerous genera and diversity varied, before partial return to pre-exchange metrics. These preliminary data help support potential host control for the rumen microbiome, aiding in efforts to define bovine host-microbe relationships.

Ruminal Protozoal Populations of Angus Steers Differing in Feed Efficiency.

Feed accounts for as much as 70% of beef production costs, and improvement of the efficiency with which animals convert feed to product has the potential to have substantial financial impact on the beef industry. The rumen microbiome plays a key role in determining feed efficiency; however, previous studies of rumen microbiota have not focused on protozoal communities despite the estimation that these organisms represent approximately 50% of rumen content biomass. Protozoal communities participate in the regulation of bacterial populations and nitrogen cycling-key aspects of microbiome dynamics. The present study focused on identifying potential associations of protozoal community profiles with feed efficiency. Weaned steers (n = 50) 7 months of age weighing approximately 260 kg were adapted to a growing ration and GrowSafe for 2 weeks prior to a 70-day feed efficiency trial. The GrowSafe system is a feeding system that monitors feed intake in real time. Body weights were collected on the first day and then every 7 days of the feed efficiency trial, and on the final day, approximately 50 mL of rumen content were collected via orogastric tubing and frozen at -80 °C. Body weight and feed intake were used to calculate residual feed intake (RFI) as a measure of feed efficiency, and steers were categorized as high (n = 14) or low (n = 10) RFI based on ±0.5 standard deviations about the mean RFI. Microbial DNA was extracted, and the eukaryotic component profiled by amplification and sequencing of 18S genes using degenerate primers that can amplify this locus across a range of protists. The taxonomy of protozoal sequences was assigned using QIIME 1.9 and analyzed using QIIME and SAS 9.4 with significance determined at α ≤ 0.05. Greater abundances of unassigned taxa were associated with high-RFI steers (p = 0.03), indicating a need for further study to identify component protozoal species. Differences were observed between low- and high-RFI steers in protozoal community phylogenetic diversity, including weighted beta-diversity (p = 0.04), Faith's phylogenetic diversity (p = 0.03), and observed Operational taxonomic unit (OTU) (p = 0.03). The unassigned taxa and differences in phylogenetic diversity of protozoal communities may contribute to divergences observed in feed efficiency phenotypes in beef steers.

The effects of feeding ferric citrate on ruminal bacteria, methanogenic archaea and methane production in growing beef steers.

Methane produced by cattle is one of the contributors of anthropogenic greenhouse gas. Methods to lessen methane emissions from cattle have been met with varying success; thus establishing consistent methods for decreasing methane production are imperative. Ferric iron may possibly act to decrease methane by acting as an alternative electron acceptor. The objective of this study was to assess the effect of ferric citrate on the rumen bacterial and archaeal communities and its impact on methane production. In this study, eight steers were used in a repeated Latin square design with 0, 250, 500 or 750 mg Fe/kg DM of ferric iron (as ferric citrate) in four different periods. Each period consisted of a 16 day adaptation period and 5 day sampling period. During each sampling period, methane production was measured, and rumen content was collected for bacterial and archaeal community analyses. Normally distributed data were analysed using a mixed model ANOVA using the GLIMMIX procedure of SAS, and non-normally distributed data were analysed in the same manner following ranking. Ferric citrate did not have any effect on bacterial community composition, methanogenic archaea nor methane production (P>0.05). Ferric citrate may not be a viable option to observe a ruminal response for decreases in enteric methane production.

The effects of protein level on cytokines and chemokines in the uterine environment of beef heifers during development.

The development of replacement heifers is crucial for breeding success and herd efficiency. Nutritional management can affect not only reproductive development but also the inflammatory status of the uterine environment, which may impact reproductive functions such as pregnancy establishment and development. The study herein evaluated the concentration of cytokines and chemokines in the uterus of heifers supplemented with different levels of protein. Angus heifers (n = 60) were blocked by body weight (BW) and randomly assigned to 1 of 3 treatments based on protein supplementation level: control of 10% crude protein (CON), 20% crude protein (P20), or 40% crude protein (P40). BW, body condition score, and blood samples were taken every 2 wk for 140 d to monitor development. Uterine flushes were performed monthly and concentrations of cytokines (IL-1α, IL-1ß, TNF-α, IFN-γ, IL-10, VEGF-α, IL-17A, and IL-36RA) and chemokines (IL-8, MCP-1, MIP-1α, and MIP-1ß) were quantified via ELISA multiplex. To test if there were mean differences in cytokines between the treatment groups or over time, PROC GLIMMIX (SAS v 9.4) was utilized. Concentrations of all cytokines and chemokines, except IL-1α, changed throughout heifer development (P < 0.05). Heifers in the P40 treatment group displayed reduced concentrations of MCP-1 (P = 0.007) and tended to have decreased concentrations of IFN-γ (P = 0.06). Cytokine IL-36RA tended (P = 0.06) to be affected by protein level, with the lowest concentrations observed in CON heifers. Most cytokines and chemokines increased following the initial month of supplementation (P < 0.05). The increase in concentrations after 1 mo may indicate an adaptive response in the uterus to diet change. Cytokines and chemokines fluctuated due to physiological changes occurring during development. Further research is needed to determine the influence of nutrition on uterine inflammation and long-term impacts on reproductive function.

Rumen and Serum Metabolomes in Response to Endophyte-Infected Tall Fescue Seed and Isoflavone Supplementation in Beef Steers.

Fescue toxicosis impacts beef cattle production via reductions in weight gain and muscle development. Isoflavone supplementation has displayed potential for mitigating these effects. The objective of the current study was to evaluate isoflavone supplementation with fescue seed consumption on rumen and serum metabolomes. Angus steers (n = 36) were allocated randomly in a 2 × 2 factorial arrangement of treatments including endophyte-infected (E+) or endophyte-free (E-) tall fescue seed, with (P+) or without (P-) isoflavones. Steers were provided a basal diet with fescue seed for 21 days, while isoflavones were orally administered daily. Following the trial, blood and rumen fluid were collected for metabolite analysis. Metabolites were extracted and then analyzed by UPLC-MS. The MAVEN program was implemented to identify metabolites for MetaboAnalyst 4.0 and SAS 9.4 statistical analysis. Seven differentially abundant metabolites were identified in serum by isoflavone treatment, and eleven metabolites in the rumen due to seed type (p < 0.05). Pathways affected by treatments were related to amino acid and nucleic acid metabolism in both rumen fluid and serum (p < 0.05). Therefore, metabolism was altered by fescue seed in the rumen; however, isoflavones altered metabolism systemically to potentially mitigate detrimental effects of seed and improve animal performance.

Classification of 16S rRNA reads is improved using a niche-specific database constructed by near-full length sequencing.

Surveys of microbial populations in environmental niches of interest often utilize sequence variation in the gene encoding the ribosomal small subunit (the 16S rRNA gene). Generally, these surveys target the 16S genes using semi-degenerate primers to amplify portions of a subset of bacterial species, sequence the amplicons in bulk, and assign to putative taxonomic categories by comparison to databases purporting to connect specific sequences in the main variable regions of the gene to specific organisms. Due to sequence length constraints of the most popular bulk sequencing platforms, the primers selected amplify one to three of the nine variable regions, and taxonomic assignment is based on relatively short stretches of sequence (150-500 bases). We demonstrate that taxonomic assignment is improved through reduced unassigned reads by including a survey of near-full-length sequences specific to the target environment, using a niche of interest represented by the upper respiratory tract (URT) of cattle. We created a custom Bovine URT database from these longer sequences for assignment of shorter, less expensive reads in comparisons of the upper respiratory tract among individual animals. This process improves the ability to detect changes in the microbial populations of a given environment, and the accuracy of defining the content of that environment at increasingly higher taxonomic resolution.

Effect of stocking density and effective fiber on the ruminal bacterial communities in lactating Holstein cows.

Overstocking can be a major issue in the dairy cattle industry, leading to negative changes in feeding and resting behavior. Additional stress imposed and alterations in feeding behavior may significantly impact the rumen microbiome. The rumen microbiome is responsible for the successful conversion of feed to usable energy for its host. Thus, understanding the effects of stocking density on the rumen microbiome is imperative for further elucidation of potentially negative consequences of overstocking in dairy cattle. This study implemented a Latin Square design accounting for four pens of cattle and four treatment periods so that all treatment combinations were assigned to every pen during one period of the study. Two treatment factors, including two levels of physically effective neutral detergent fiber, achieved with addition of chopped straw, and stocking density (100% vs. 142%) of freestalls and headlocks, were combined and tested within a factorial treatment design. Within each pen, three or four cannulated cows (n = 15 total) were sampled for rumen content on the final day of each treatment period. Each treatment was randomly assigned to a single pen for a 14-day period. The V1-V3 hypervariable regions of the 16S rRNA gene were targeted for bacterial analyses. Variables with approximately normally-distributed residuals and a Shapiro-Wilk statistic of ≥0.85 were analyzed using a mixed model analysis of variance with the GLIMMIX procedure with fixed effects of feed (straw vs. no straw), stocking density (100% vs. 142%), and the interaction of feed × stocking density, and random effects of pen, period, feed × stocking × pen × period. Pen was included as the experimental unit in a given period and the sampling unit as cow. Variables included Shannon's Diversity Index, Faith's phylogenetic diversity index, chao1, observed OTU, and Simpson's evenness E as well as most individual taxa. Data were analyzed in SAS 9.4 utilizing the GLIMMIX procedure to perform mixed model analysis of variance. If data were not normally distributed, a ranked analysis was performed. No differences were observed in α-diversity metrics by fiber or stocking density (P > 0.05). Beta diversity was assessed using weighted and unweighted Unifrac distances in QIIME 1.9.1 and analyzed using ANOSIM. No differences were observed in weighted (P = 0.6660; R = -0.0121) nor unweighted (P = 0.9190; R = -0.0261) metrics and R values suggested similar bacterial communities among treatments. At the phylum level, Tenericutes differed among treatments with an interaction of stocking density by feed (P = 0.0066). At the genus level, several differences were observed by treatment, including Atopobium (P = 0.0129), unidentified members of order RF39 (P = 0.0139), and unidentified members of family Succinivibrionaceae (P = 0.0480). Although no diversity differences were observed, taxa differences may indicate that specific taxa are affected by the treatments, which may, in turn, affect animal production.

Rumen fluid metabolomics of beef steers differing in feed efficiency.

INTRODUCTION: 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 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. METHODS: In this study, rumen fluid was collected from low- (n = 14) and high-RFI (n = 15) steers. Rumen fluid was filtered through a 0.22 µM syringe filter, extracted 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 were identified using MAVEN and analyzed using MetaboAnalyst 4.0 and SAS. Significance was determined using an α ≤ 0.05. RESULTS: 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). These metabolites were predominantly involved in amino acid and lipid metabolism. CONCLUSIONS: Rumen fluid metabolomes differ 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.

Rumen Bacteria and Serum Metabolites Predictive of Feed Efficiency Phenotypes in Beef Cattle.

The rumen microbiome is critical to nutrient utilization and feed efficiency in cattle. Consequently, the objective of this study was to identify microbial and biochemical factors in Angus steers affecting divergences in feed efficiency using 16S amplicon sequencing and untargeted metabolomics. Based on calculated average residual feed intake (RFI), steers were divided into high- and low-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 features of the bacterial community in the rumen. Decreased bacterial α- (P = 0.03) and ß- diversity (P < 0.001) was associated with Low-RFI steers. RFI was associated with several serum metabolites. Low-RFI steers had greater abundances of pantothenate (P = 0.02) based on fold change (high/low RFI). Machine learning on RFI was predictive of both rumen bacterial composition and serum metabolomic signature (AUC ≥ 0.7). Log-ratio proportions of the bacterial classes Flavobacteriia over Fusobacteriia were enriched in low-RFI steers (F = 6.8, P = 0.01). Reductions in Fusobacteriia and/or greater proportions of pantothenate-producing bacteria, such as Flavobacteriia, may result in improved nutrient utilization in low-RFI steers. Flavobacteriia and Pantothenate may potentially serve as novel biomarkers to predict or evaluate feed efficiency in Angus steers.

Assignment of virus and antimicrobial resistance genes to microbial hosts in a complex microbial community by combined long-read assembly and proximity ligation.

We describe a method that adds long-read sequencing to a mix of technologies used to assemble a highly complex cattle rumen microbial community, and provide a comparison to short read-based methods. Long-read alignments and Hi-C linkage between contigs support the identification of 188 novel virus-host associations and the determination of phage life cycle states in the rumen microbial community. The long-read assembly also identifies 94 antimicrobial resistance genes, compared to only seven alleles in the short-read assembly. We demonstrate novel techniques that work synergistically to improve characterization of biological features in a highly complex rumen microbial community.

Temporal Stability of the Ruminal Bacterial Communities in Beef Steers.

Nutritional studies involving ruminants have traditionally relied on relatively short transition or wash-out periods between dietary treatments, typically two to four weeks. However, little is known about adequate adaptation periods required to reach stabilization of the rumen microbiome that could provide more accurate results from nutritional studies in ruminants. This study determined the rumen bacterial communities and rumen environment parameters over ten weeks following transition from a forage-based to concentrate-based diet. Several α-diversity metrics, including observed OTUs and Simpson's Evenness fluctuated throughout the trial, but were typically either greatest (observed OTUs) or lowest (Simpson's) at week 5 of the trial contrasted from weeks 1 and 10 (P < 0.05). At week 4, several orders associated with the shift to the final bacterial community composition, including Pasteurellales, Aeromonadales, and Bacteroidales. At week 5, rumen pH was correlated with α-diversity (P = 0.005) and predictive of the rumen microbiome signature at week 10 (R2 = 0.48; P = 0.04). Rumen microbiome stability did not occur until approximately 9 weeks following adaptation to the diet and was associated with changes in specific bacterial populations and rumen environment. The results of this study suggest that adaptation and wash-out periods must be re-evaluated in order to accommodate necessary rumen microbiome acclimation.

Uterine and vaginal bacterial community diversity prior to artificial insemination between pregnant and nonpregnant postpartum cows1.

The present study evaluated the bovine vaginal and uterine bacterial community diversity and its relationship to fertility. Postpartum beef cows (n = 68) were synchronized beginning on day -21 and ending with timed artificial insemination (TAI) on day 0. Pregnancy was diagnosed 30 d after TAI. Uterine and vaginal flushes were collected on day -21, -9, and -2 for bacterial DNA extraction to sequence the V1 to V3 hypervariable regions of the 16S rRNA gene. Results indicated a decrease in the number of bacterial species over time in the uterus of resulting pregnant and nonpregnant beef cows (P < 0.0001). Principal coordinate analyses (PCoA) depicted clustering of samples, indicating closely related bacterial communities, by day in the uterus and vagina (P < 0.0001). At day -2, uterine samples from nonpregnant and pregnant animals clustered separately (P < 0.0001), with nonpregnant animal samples clustering tightly together. Overall, the current study suggests the shift in the reproductive bacterial communities' diversity and phylogenetic relationship leading up to the time of breeding may contribute to successful pregnancy establishment.