Project description:The structure and function of the microbiome inhabiting the rumen are, amongst other factors, mainly shaped by the animal’s feed intake. Describing the influence of different diets on the inherent community arrangement and associated metabolic activities of the most active ruminal fractions (bacteria and archaea) is of great interest for animal nutrition, biotechnology and climatology. Samples were obtained from three fistulated Jersey cows rotationally fed with corn silage, grass silage or hay, each supplemented with a concentrate mixture. Samples were fractionated into ruminal fluid, squeezed solid and solid matter. DNA, proteins and metabolites were analyzed subsequently. DNA extracts were used for Illumina sequencing of the 16S rRNA gene and the metabolomes of rumen fluids were determined by 500MHz-NMR spectroscopy. Tryptic peptides derived from protein extracts were measured by LC-ESI-MS/MS and spectra were processed by a two-step database search for quantitative metaproteome characterization. Protein- and DNA-based datasets revealed significant differences between sample fractions and diets and affirmed similar trends concerning shifts in phylogenetic composition. Ribosomal genes and proteins belonging to the phylum of Proteobacteria, particularly Succinivibrionaceae, exhibited a higher abundance in corn silage-based samples while fiber-degraders of the Lachnospiraceae family emerged in great quantities throughout the solid phase fractions. The analysis of 8163 quantified bacterial proteins revealed the presence of 166 carbohydrate active enzymes in varying abundance. Cellulosome affiliated proteins were less expressed in the grass silage, glycoside hydrolases appeared in slightest numbers in the corn silage. Most expressed glycoside hydrolases belonged to families 57 and 2. Enzymes analogous to ABC transporters for amino acids and monosaccharides were more abundant in the corn silage whereas oligosaccharide transporters showed a higher abundance in the fiber-rich diets. Proteins involved in carbon metabolism were detected in high numbers and identification of metabolites like short-chain fatty acids, methylamines and phenylpropionate by NMR enabled linkage between producers and products. This study forms a solid basis to retrieve deeper insight into the complex network of gut microbial adaptation.

Project description:Beef represents a major diet component and source of protein in many countries. With an increment demand for beef, the industry is currently undergoing changes towards natural produced beef. Consumers not only concern about product quality, but also for the well-being of animals. Therefore, the consumption of grass-fed meat is continuously growing. However, the nutritional true differences between feeding systems are still unclear. The aim of this study was to examine latissimus dorsi muscle quality and animal welfare by transcriptome and metabolome profiles, and to identify biological pathways related to the differences between grass- and grain-fed Angus steers. By RNA-Seq analysis of latissimus dorsi muscle, we have recognized 241 differentially expressed genes (FDR < 0.1). The metabolome examination of muscle and blood revealed 163 and 179 altered compounds in each tissue (P-value < 0.05), respectively. Accordingly, alterations in glucose metabolism, divergences in free fatty acids and carnitine conjugated lipid levels, and altered Î²-oxidation, have been observed. In summary, this study demonstrates a unique transcriptomic and metabolic signature in the muscle of grain and grass finished cattle. Results support the accumulation of anti-inflammatory n3 polyunsaturated fatty acids in grass finished cattle, while higher levels of n6 PUFAs in grain finished animals may promote inflammation and oxidative stress. Furthermore, grass-fed animals produce tender beef with lower total fat and higher omega3/omega6 ratio than grain fed animals, which could potentially benefit consumer health. Finally, blood cortisol levels strongly indicate that grass fed animals experience less stress than the grass fed individuals The steers came from a closed Wye Angus herd with very similar genetics. The grass-fed group was comprised of steers that received alfalfa and orchard grass hay, clover and orchard grass pasture, or orchard grass and alfalfa pasture. The grass-fed individuals consumed grazed alfalfa upon availability and bales during winter and were not exposed to any corn, any form of grain or feed by-products. The alfalfa and grass hay were harvested from land that has had minimal fertilizer and no application of pesticides or inorganic chemicals. The control group was fed a conventional diet consisting of corn silage, soybean, shelled corn and minerals. The pastures were managed as organic landsâwithout fertilizers, pesticides or any chemical additives. At the slaughter plant, 10 ml whole blood sample from the jugular vein was collected in EDTA tubes and directly storage at -80Â°C. Then, a small piece of longissimus dorsi muscle was obtained from each hot carcass at the level of the 12th intercostal space and immediately frozen in dry ice for posterior analysis.

Project description:Beef represents a major diet component and one of the major sources of protein in human. The beef industry in the United States is currently undergoing changes and is facing increased demands especially for natural grass-fed beef. The grass-fed beef obtained their nutrients directly from pastures, which contained limited assimilable energy but abundant amount of fiber. On the contrary, the grain-fed steers received a grain-based regime that served as an efficient source of high-digestible energy. Lately, ruminant animals have been accused to be a substantial contributor for the green house effect. Therefore, the concerns from environmentalism, animal welfare and public health have driven consumers to choose grass-fed beef. Rumen is one of the key workshops to digest forage constituting a critical step to supply enough nutrients for animals’ growth and production. We hypothesize that rumen may function differently in grass- and grain-fed regimes. The objective of this study was to find the differentially expressed genes in the ruminal wall of grass-fed and grain-fed steers, and then explore the potential biopathways. In this study, the RNA Sequencing (RNA-Seq) method was used to measure the gene expression level in the ruminal wall. The total number of reads per sample ranged from 24,697,373 to 36,714,704. The analysis detected 342 differentially expressed genes between ruminal wall samples of animals raised under different regimens. The Fisher’s exact test performed in the Ingenuity Pathway Analysis (IPA) software found 16 significant molecular networks. Additionally, 13 significantly enriched pathways were identified, most of which were related to cell development and biosynthesis. Our analysis demonstrated that most of the pathways enriched with the differentially expressed genes were related to cell development and biosynthesis. Our results provided valuable insights into the molecular mechanisms resulting in the phenotype difference between grass-fed and grain-fed cattle. Ruminal wall samples from two randomly chosen animals per group were obtained, totaling four samples. The animals were born, raised and maintained at the Wye Angus farm. This herd, which has been closed for almost 75 years and yielded genetically similar progenies, constitutes an excellent resource to perform transcriptomic analysis. The genetic resemblance among individuals permits us to better control the cause of variation between experimental clusters and individuals. The randomly chosen pairs of animals were part of larger sets of steers that received a particular treatment. All animals received the same diet until weaning. The grain group received conventional diet consisting of corn silage, shelled corn, soy bean and trace minerals. The grass fed steers consumed normally grazed alfalfa; during wintertime, bailage was utilized. The alfalfa has been harvested from land without any fertilizers, pesticides or other chemicals. The steers ate no animal, agricultural or industrial byproducts and never receive any type of grain. Then, the calves were randomly assigned to one diet and exclusively received that regimen until termination. Grain–fed animals reached the market weight around the age of 14 month-old, however, grass-fed steers required approximately 200 additional days to achieve the same weight. Immediately after termination at the Old Line Custom Meat Company (Baltimore, MD) a small piece of ruminal wall was excised, cleaned and preserved at -80°C for posterior processing.

Project description:The grass-fed cattle obtain nutrients directly from pastures containing limited assimilable energy but abundant amount of fiber; by contrast, grain-fed steers receive a diet that is comprised mainly of grains and serves as an efficient source of high-digestible energy. Besides energy, these two types of diet differ in a large number of nutritional components. Additionally, animals maintained on rich-energy regimen are more likely to develop metabolic disorders and infectious diseases than pasture raised individuals. Thus, we hypothesize that spleenâ??the main immune organâ??may function differently under disparate regimes. The objective of this study was to find the differentially expressed genes in the spleen of grass-fed and grain-fed steers, and furtherly explore the potential involved biopathways. Through RNA sequencing (RNA-Seq), we detected 123 differentially expressed genes. Based on these genes, we performed an Ingenuity Pathway Analysis (IPA) and identified 9 significant molecular networks and 13 enriched biological pathways. Two of the pathways, Nur77 signaling in T lymphocytes and calcium-induced T lymphocyte apoptosis which are immune related, contain a pair of genes HLA-DRA and NR4A1 with dramatically altered expression level. Collectively, our results provided valuable insights into understanding the molecular mechanism of spleen under varied feeding regimens. We collected spleen samples from two randomly chosen animals per group, totaling four samples. The animals were born and raised at the Wye Angus farm, which has produced genetically similar progenies. The genetic resemblance among individuals permitted us to better control the variation between experimental individuals, constituting an excellent resource to perform scientific research. All animals included in this study received the same diet until weaning. Next, we assigned the animals to one certain diet at random, and exclusively raised them under that regimen until termination. The diet of grain-fed group consisted of soybean, shelled corn, corn silage and trace minerals. The grass-fed steers normally received alfalfa harvested from land without any fertilizers, pesticides or other chemicals; during wintertime, bailage was supplied. Grass-fed individuals ate no animal, agricultural or industrial byproducts and never consumed any type of grain. Grain-fed animals reached the market weight around 14 month-old; however, grass-fed steers needed approximately 200 additional days to achieve the same weight. Immediately after termination at the Old Line Custom Meat Company (Baltimore, MD), a small piece of spleen was incised, washed and frozen at -80Â°C for posterior processing.

Project description:Two-stage two-phase biogas reactor systems consisting each of one batch downflow hydrolysis reactor (HR, vol. 10 L), one process fluid storage tank (vol. 10 L), and one downstream upflow anaerobic filter reactor (AF, vol. 10 L), were operated at mesophilic (M, 37 °C) and thermophilic (T, 55 °C) temperatures and over a period of > 750 d (Figure 1, Additional file 1). For each reactor system and for each process temperature, two replicates were conducted in parallel, denominated further as biological replicates. Further process details were as previously published. Start-up of all fermenters were performed using liquid fermenter material from a biogas plant converting cattle manure in co-digestion with grass and maize silage and other biomass at varying concentrations and at mesophilic temperatures. Silage of perennial ryegrass (Lolium perenne L.) was digested as sole substrate in batches of varying amounts with retention times of 28 d (storage of bale silage at -20 °C, cutting length 3 cm, volatile substances (VS) 32 % of fresh mass (FM), total Kjeldahl nitrogen 7.6 g kgFM-1, NH4+-N 0.7 g kgFM-1, acetic acid 2.6 g kgFM-1, propionic acid < 0.04 g kgFM-1, lactic acid 2.6 g kgFM-1, ethanol 2.2 g kgFM-1, C/N ratio 19.3, chemical oxygen demand (COD) 357.7 g kgFM-1, analysis of chemical properties according to [6]. No spoilage was observed in the silage. Biogas yields were calculated as liters normalized to 0 °C and 1013 hPa (LN) per kilogram volatile substances (kgVS). For chemical analysis, samples were taken from the effluents of HR and AF. For sequencing of 16S rRNA gene amplicon libraries, microbial metagenomes, and microbial metatranscriptomes, samples were taken from the silage digestate in the HR digested for 2 d. At this time point, high AD rates were detected as indicated by the fast increase of volatile fatty acids (VFA), e.g., acetic acid. Sampling was performed at two different organic loading rates (OLR), i.e., batch-fermentation of 500 g (denominated as “low OLR”, samples MOLR500 and TOLR500) and 1,500 g silage (denominated as “increased OLR”, samples MOLR1500 and TOLR1500).

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