Ruminal Bacteria Research Articles

Effects of Chitosan-Based Additive on Rumen Fermentation and Microbial Community, Nutrients Digestibility and Lactation Performance in Goats.

Recently, the potential of using chitosan (CHI) as a feed additive to enhance ruminal fermentation and improve animal performance has gained increasing attention in ruminant nutrition. This study was undertaken to investigate the effect of dietary supplementation with increasing doses of CHI on rumen fermentation attributes and microbial composition, digestibility and milk performance in Dhofari goats. Twenty-four lactating goats (27 ± 1.8 kg of initial live body weight) were fed a control diet comprising of Rhodes grass hay plus a concentrate feed mixture. Goats were assigned to one of three experimental treatments (n = 8 per treatment) as: (1) control diet with no supplement (CTRL), (2) control diet with 0.300 g/day CHI (CHI0.3) and (3) control diet supplemented with 0.600 g/day CHI (CHI0.6) for a 45-day experimental period. Dietary supplementation with increasing doses of CHI decreased (p < 0.05) linearly ruminal pH (p = 0.023), total short chain fatty acids concentrations (p = 0.011), acetate (p = 0.013) and butyrate (p = 0.042) proportions, acetate to propionate ratio (p < 0.001), estimated methane (CH4) production (p < 0.001), ammonia nitrogen concentrations (p = 0.003) and protozoa abundance (p = 0.003). However, the ruminal propionate proportion augmented (p = 0.002) linearly with increasing doses of CHI in the diet. Increasing doses of CHI linearly increased the abundance of the ruminal propionate-producing bacteria, while diminished acetate and CH4-producing bacteria (p < 0.05). Serum total protein (p = 0.037) and glucose (p = 0.042) levels linearly increased as CHI doses increased in the diet. However, serum UREA levels decreased linearly (p = 0.002) by 21% with increasing CHI amounts in the diet. The digestibility of organic matter, crude protein and neutral detergent fibre increased linearly with the increasing CHI doses (p < 0.05). Neither linear nor quadratic responses (p > 0.05) were observed in daily milk yield and feed efficiency by supplementing the diet with CHI. In conclusion, supplementing the diet with CHI at a dose of 0.600 g/day as a feed additive for dairy goats reduced estimated CH4 generation and improved fibre and protein digestion, with no influence on feed intake, milk yield or composition.

PSIV-11 The inhibitory activity of phyto-phenolic compounds on ammonia-producing ruminal bacteria

Abstract Antibiotic resistance has progressively increased over time, thus necessitating the discovery of sustainable, novel antimicrobials. Ammonia producing bacteria (APB) are predominant organisms occupying many niches in the rumens of cattle and soil. The objective of this study was to discover target phyto-phenolic compounds (PPCs) that can act as replacements for existing antimicrobials, as well as at what concentration ranges may be optimal for inhibition of APB growth. In recognition of this discovery, producers may have alternative antimicrobial compounds for use in managing APB and common silage spoilage organisms. To shed light on this matter, inhibitory concentrations (IC) and ammonia assays were implemented. ICs analyzed the inhibitory action of six structurally similar PPCs: thymol, eugenol, carvacrol, hydro-cinnamic acid (H-CA), trans-cinnamic acid (T-CA), and phloroglucinol. Five APB (SR, MD1, F, BG1, and B14) were exposed to five concentrations of phytochemicals via a 10% v/v serial dilution: 10, 1, 0.1, 0.01, 0.001 mM. Following serial dilution, the bacteria were incubated ~24h at 39°C in a water bath, and visually inspected for growth. Ammonia assays utilized the least, fully inhibitory concentration of each phytochemical identified via the inhibitory concentration experiments. Ammonia concentrations were determined using the phenolic acid/hypochlorite method. All protocols utilized anaerobic sterile technique. All experiments were repeated in triplicate and significant results (P &amp;lt; 0.05) were determined using a one-way ANOVA with Tukey’s post hoc test utilizing OriginPro statistical software. Inhibition results revealed that eugenol, thymol, and carvacrol were successful in inhibiting bacterial growth at a concentration range of 10 mM to 1 mM. However, phloroglucinol was not fully inhibitory at any concentration tested against any bacteria. Eugenol, thymol, carvacrol were fully inhibitory at either 10 mM or 1 mM for all organisms tested except BG1, where it was only inhibitory at 10 mM. Ammonia analyses determined that eugenol and carvacrol greatly reduced ammonia production for all species tested except for B14, a generalist, which had no measurable changes in ammonia for any of the treatments including control. Thymol reduced ammonia production for MD1, BG1, and SR. Additionally, H-CA and T-CA decreased ammonia production of SR compared with control but not as much as eugenol or carvacrol. Results suggested that the growth of APB was generally inhibited by the greatest concentrations of PPCs (10 mM to 1 mM) tested here. Additionally, phloroglucinol did not inhibit bacterial growth therefore it was not included in the ammonia analyses. Eugenol and carvacrol significantly reduced ammonia production in all organisms, therefore indicating possible antimicrobial candidates to be further investigated. In comparison with carvacrol and eugenol, T-CA and H-CA made a modest to negligible reduction in ammonia. Phyto-phenolic compounds that were successful in inhibiting bacterial growth and reducing ammonia production demonstrate several promising target antimicrobials.

93 Ruminal archaea and bacteria metatranscriptomic responses to supplementation in steers fed low-quality forage

Abstract Forage comprises the majority of beef cattle diets globally, and its quality can vary. Supplementation presents a significant opportunity to improve the efficiency of production. Supplementation has been shown to increase ruminally available nitrogen, improve forage utilization, and reduce livestock methane (CH4) emissions. However, corresponding effects of supplementation on gene expression of rumen microbiota, in particular metabolic pathways involved in methanogenesis, have yet to be studied. This study aimed to evaluate the effect of supplement composition before and after feeding on rumen archaeal and bacterial gene expression in steers consuming a low-quality forage diet. Ruminal samples were collected from five ruminally cannulated Angus steers [body weight (BW) = 375 ± 45 kg] used in a 4 × 4 Latin square fed a basal diet of King Ranch Bluestem Hay (3.5% CP, 73% NDF), and one of four isonitrogenous (130 mg N/kg BW) supplements. Steers provided the two most divergent supplements (2% starch, 43% CP; 56% starch, 21% CP) were used for metatranscriptomic analysis. Rumen fluid samples were collected via rumen cannula 0 and 4 h post-feeding. Total RNA was extracted and sequenced. Differentially expressed genes (DEGs) were identified and functionally annotated. Pathway enrichment analysis was completed to determine enriched pathways (broad metabolic pathways) and modules (specific metabolic reactions). Only main effects are presented in this abstract. Archaeal hydrogenotrophic and methylotrophic methanogenesis reactions and tryptophan biosynthesis reactions were upregulated in the 2% starch supplement (P &amp;lt; 0.05). The 2% starch supplement resulted in fewer bacterial total upregulated carbohydrate-active enzymes compared with the 56% starch supplement (957 vs. 1,695 DEGs, respectively; P &amp;lt; 0.05), and an increased proportion of cellulose and hemicellulose-specific enzymes (35 vs. 29%, respectively; P &amp;lt; 0.05). The 56% starch supplement resulted in upregulation of the bacterial 3-Hydroxypropionate bi-cycle reaction (P &amp;lt; 0.05). Collection time exerted a larger effect than supplement composition on both archaeal and bacterial gene expression, with 83% and 63% of DEGs associated with time, respectively (P &amp;lt; 0.05). Archaeal hydrogenotrophic methanogenesis and carbon metabolism were upregulated 4 h post-feeding. In addition, 1,704 bacterial carbohydrate-active enzymes were upregulated 4 h post-feeding, as compared with 975 upregulated at 0 h (P &amp;lt; 0.05). However, these differences in DEGs did not correlate with significantly enriched pathways of bacterial carbohydrate metabolism. These findings demonstrate improvements to low-quality forage utilization, through CP and starch supplementation, can be correlated to relevant changes in gene expression of rumen microbiota. Furthermore, they demonstrate that supplementation influences the expression of methanogenesis reactions, suggesting an opportunity to affect CH4 production. Additional studies quantifying forage utilization, rumen microbiota gene expression and CH4 production are needed to further optimize supplementation strategies for improved efficiency of beef production whilst maintaining or reducing its carbon footprint.

Partially substituting alfalfa hay with hemp forage promotes the health and well‐being of goats via altering ruminal and plasma metabolites and metabolic pathways

AbstractHemp forage (HF) seems a suitable forage for ruminants for its high nutritional value and rich phytochemicals that exert health and growth‐promoting activities. We investigated the effects of hemp‐related phytochemicals on rumen and plasma metabolism using metabolome when partially substituting alfalfa hay with HF in goat diets. Numbers of differential metabolites linearly increased with increasing HF substituting rate, approximately 50% of which were phytochemicals. Metabolic pathway enrichment analysis showed that the inclusion of HF greatly promoted steroid hormone biosynthesis, one carbon pool by folate, and retinol metabolism pathways in both rumen and plasma, which are beneficial for promoting animal health and well‐being and enhancing the quality of animal products. Some phytochemicals showed inhibitory activities on the growth of certain ruminal bacteria; meanwhile, the detected intermediate metabolites indicated degradation of the phytochemicals by ruminal microbes. These phytochemicals work individually and synergistically to alter ruminal and plasma metabolic pathways, thus exerting benefits in promoting the health and well‐being of animals.

Differences in Lactation Performance, Rumen Microbiome, and Metabolome between Montbéliarde × Holstein and Holstein Cows under Heat Stress.

Heat stress negatively affects lactation performance and rumen microbiota of dairy cows, with different breeds showing varying levels of heat tolerance. This study aimed to compare the lactation performance of Montbéliarde × Holstein (MH, n = 13) and Holstein (H, n = 13) cows under heat stress, and 16S rRNA sequencing and liquid chromatography-mass spectrometry (LC-MS) were used to determine the rumen microbiome and metabolome in experimental cows. The results indicated that during heat stress, milk yield (p = 0.101), milk fat yield, milk protein yield, milk protein, and milk lactose (p < 0.05) in Montbéliarde × Holstein cows were higher than those in Holstein cows, whereas milk yield variation and somatic cell counts (p < 0.05) were lower than those in Holstein cows. The sequencing results indicated that the rumen of Montbéliarde × Holstein cows was significantly enriched with beneficial bacteria, such as Rikenellaceae, Allobaculum, and YRC22 (p < 0.05). In addition, correlations were observed between specific ruminal bacteria and lactation performance. Ruminal metabolites related to antioxidant and anti-inflammatory properties were significantly higher (p < 0.05) in Montbéliarde × Holstein cows than in Holstein cows. Overall, Montbéliarde × Holstein cows showed higher production efficiency under heat stress, which may be related to the different rumen mechanisms of crossbred and Holstein cows in adapting to heat stress.

Feed Restriction in Angus Steers Impacts Ruminal Bacteria, Its Metabolites, and Causes Epithelial Inflammation

We identified alterations in the ruminal microbiome, metabolome, and epithelial inflammatory response due to moderate feed restriction (FR). Ruminal digesta and epithelial biopsies from seven ruminally cannulated Angus steers were initially collected during ad libitum access to feed (PRE). After a 10 day recovery, steers underwent a 3-day FR period (FRP) at 25% intake of PRE followed by a 15 day recovery (POST) phase with ad libitum access to feed. At the end of FRP and POST, ruminal digesta and epithelial biopsies were collected again for microbial DNA and tissue RNA extraction. RT-qPCR was applied for relative microbial abundance and RNA extraction. Metabolite profiling of digesta was performed via GC-MS. The abundance of Succinivibrio dextrinosolvens, Streptococcus bovis, and Bifidobacteria spp. (N124) was higher (p &lt; 0.05) during FRP than PRE and POST, while Lactobacillus spp. (C25), Escherichia coli (EC42405), Fibrobacter succinogenes, and Megaspheara elsdenii abundances were lower in FRP than PRE (p &lt; 0.05). The TNF and TLR2 mRNA abundance was greater in FRP than PRE (p &lt; 0.05). Among 15 detected amino acids, glutamine, isoleucine, lysine, phenylalanine, threonine, and valine were lower (p &lt; 0.05) in FRP than PRE. Metabolite pathway analysis revealed alterations in amino acid, fatty acid, vitamin, and energy metabolism during FRP (p &lt; 0.05). The mRNA of the proinflammatory genes TNF and TLR2 in the epithelium peaked (p &lt; 0.05) at FRP and remained higher at POST. Results indicated that a short FR influenced ruminal bacteria, reduced concentrations of most metabolites, and triggered an inflammatory response.

Rumen microbiota helps Tibetan sheep obtain energy more efficiently to survive in the extreme environment of the Qinghai-Tibet Plateau.

T-sheep and H-sheep exhibit different environmental adaptability and production performance. The rumen microbiome has co-evolved with hosts and plays a vital role in nutrient digestion and energy metabolism. In our previous study, we found that T-sheep have a higher efficiency in energy metabolism than H-sheep, but the rumen microbial community remains unclear. In this study, we determined the rumen bacterial profile and rumen fermentation parameters to reveal the bacterial profiles and predictive functions among breeds and diets with four different energy levels, as well as the correlation between bacterial profiles and rumen fermentation characteristics. The results showed that the rumen total volatile fatty acids (VFAs), acetate, butyrate, total branched-chain VFAs, iso-butyrate, and iso-valerate were higher in T-sheep than H-sheep. The alpha diversity of ruminal bacteria is not affected by dietary energy, but it shows a distinction between the sheep breeds. Specifically, T-sheep rumen bacteria exhibit higher alpha diversity than H-sheep. The beta diversity of ruminal bacteria is not influenced by dietary energy or sheep breeds, indicating similar communities of ruminal bacteria between different diets and sheep breeds. The phyla of Bacteroidetes and Firmicutes predominate in the rumen, with a higher relative abundance of Firmicutes observed in T-sheep than H-sheep. The two most abundant genera in the rumen were Prevotella 1 and Rikenellaceae RC9 gut group. Prevotella 1 is the predominant bacterial genus in the rumen of H-sheep, while the Rikenellaceae RC9 gut group dominates in the rumen of T-sheep. Microbial co-occurrence network analysis reveals that variations in rumen fermentation characteristics result from differences in module abundance, with a higher abundance of VFA-producing modules observed in the rumen of T-sheep. Microbial function prediction analysis showed that dietary energy rarely alters the functional composition of rumen bacteria. However, there were differences in the functions of rumen bacteria between sheep breeds, with T-sheep showing a greater emphasis on energy metabolism-related functions, while H-sheep showed a greater emphasis on protein metabolism-related functions. These findings provide evidence of the special rumen microbial community that helps T-sheep efficiently obtain energy from low-protein and low-energy diets, enabling them to survive in the extreme environment of the Qinghai-Tibet Plateau.

Influence of dietary mulberry (Morus alba L.) leaf supplementation on production performance, blood metabolites, rumen fermentation characteristics and ruminal bacteria community in lactating dairy cows

The purpose of this study was to determine the effect of mulberry (Morus alba L.) leaf (ML) supplementation on the productivity, blood metabolism, rumen fermentation characteristics and rumen microbial population in lactating dairy cows. Twelve healthy multiparous Holstein dairy cows (initial characteristics: body weight = 442.83 ± 86.87 kg; milk yield = 12.55 ± 1.91 kg/d; days in milk = 150.0 ± 47.0 d) were randomly allocated to two treatment groups of six cows each. The cows were fed two different diets: a control diet (basal diet without ML; CTRL group) and a ML-supplemented diet (basal diet + 300 g ML; ML group). The trial lasted for a total of 28 d after an adjustment period of 14 d. The total rumen volatile fatty acid (VFA) levels (p = .063) and the propionate level (p < .05) were increased in the ML group, whereas the ratio of acetate to propionate was significantly decreased (p < .05). A microbiome study demonstrated that the dietary treatments significantly influenced the composition of rumen bacteria. The ruminal populations of Paludibacter propionicigenes, Ruminococcus sp. JE7A12 and Prevotella oris were greater (p < .05) in ML group than in CTRL group, while Enterococcus faecalis numbers significantly decreased (p < .05) in the ML group. Supplementing a dairy cow diet with 300 g ML has the potential to alter the microbial community and fermentation in the rumen, leading to an increase in propionate synthesis.

Negative energy balance affects perinatal ewe performance, rumen morphology, rumen flora structure, and placental function.

This study investigated the effects of negative energy balance (NEB) on perinatal ewes, with a focus on changes in growth performance, serum biochemical parameters, rumen fermentation, ruminal bacteria composition, placental phenotype-related indicators, and expression levels of genes related to placental function. Twenty ewes at 130 days of gestation were randomly allocated to either the positive energy balance (PEB) or NEB groups. In the experiment, ewes in the PEB group were fed the same amount as their intake during the pre-feeding baseline period, while ewes in the NEB group were restricted to 70% of their individual baseline feed intake. The experiment was conducted until 42 days postpartum, and five double-lamb ewes per group were selected for slaughter. The results demonstrated that NEB led to a significant decrease in body weight, carcass weight, and the birth and weaning weights of lambs (P < 0.05). Additionally, NEB caused alterations in serum biochemical parameters, such as increased non-esterified fatty acids and β-hydroxybutyrate levels and decreased cholesterol and albumin levels (P < 0.05). Rumen fermentation and epithelial parameters were also affected, with a reduction in the concentrations of acetic acid, butyric acid, total acid and a decrease in the length of the rumen papilla (P < 0.05). Moreover, NEB induced changes in the structure and composition of ruminal bacteria, with significant differences in α-diversity indices and rumen microbial community composition (P < 0.05). Gene expression in rumen papilla and ewe placenta was also affected, impacting genes associated with glucose and amino acid transport, proliferation, apoptosis, and angiogenesis (P < 0.05). These findings screened the key microbiota in the rumen of ewes following NEB and highlighted the critical genes associated with rumen function. Furthermore, this study revealed the impact of NEB on placental function in ewes, providing a foundation for investigating how nutrition in ewes influences reproductive performance. This research demonstrates how nutrition regulates reproductive performance by considering the combined perspectives of rumen microbiota and placental function.

Saccharomyces cerevisiae and Kluyveromyces marxianus yeast co-cultures modulate the ruminal microbiome and metabolite availability to enhance rumen barrier function and growth performance in weaned lambs

In lambs, weaning imposes stress that can contribute to impaired rumen epithelial barrier functionality and immunological dysregulation. In this study, the effects of a yeast co-culture consisting of Saccharomyces cerevisiae and Kluyveromyces marxianus (NM) on rumen health in lambs was evaluated, with a focus on parameters including growth performance, ruminal fermentation, and epithelial barrier integrity, ruminal metabolic function, and the composition of the ruminal bacteria. In total, 24 lambs were grouped into four groups of six lambs including a control (C) group fed a basal diet, and N, M, and NM groups in which lambs were fed the basal diet respectively supplemented with Saccharomyces cerevisiae yeast cultures (30 g/d per head), Kluyveromyces marxianus yeast cultures (30 g/d per head), and co-cultures of both yeasts (30 g/d per head), the experiment lasted for 42 d. Subsequent analyses revealed that relative to the C group, the average daily gain (ADG) of lambs in the NM group was significantly greater (P<0.05) and exhibited significant increases in a range of mRNA relative expression including monocarboxylate transporter 1 (MCT1), (Na+)/hydrogen (H+) exchanger 1 (NHE1), (Na+)/hydrogen (H+) exchanger 3 (NHE3), proton-coupled amino acid transporter 1 (PAT1), vacuolar H+-ATPase (vH+ ATPase), Claudin-1, Occludin in the rumen epithelium (P<0.05). Compared with the C group, the pH of the rumen contents in the NM group was significantly decreased (P<0.05), and the concentrations of acetate, propionate, and butyrate were significantly increased (P<0.05). Analysis of the rumen bacteria showed that the NM group exhibited increases in the relative abundance of Prevotella, Treponema, Moryella, Fibrobacter, CF231 and Ruminococcus (P<0.05). Metabolomics analyses revealed an increase in the relative content of phthalic acid (P<0.001) and cinnamaldehyde (P=0.008) in the NM group as compared to the C group (P<0.05), together with the greater relative content of L-tyrosine, L-dopa, rosmarinic acid, and tyrosol generated by the tyrosine metabolic pathway (P<0.05). Spearman’s correlation analyses revealed relative abundance levels of Fibrobacter and Ruminococcus were positively correlated with the mRNA relative expression levels of PAT1, NHE3, and zonula occluden-1 (ZO-1), as well as with tyrosol, phthalic acid, and cinnamaldehyde levels (P<0.05). Ultimately, these results suggest that dietary supplementation with NM has a wide range of beneficial effects on weaned lambs and is superior to single bacterial fermentation. These effects include improvements in daily gain and rumen epithelial barrier integrity, as well as improvements in the composition of the rumen microbiome, and alterations in tyrosine metabolic pathways.

Screening and evaluation of the ruminal cellulolytic bacteria and their potential application as probiotics.

Rumen microbiologists are looking for new probiotics to improve the digestibility of livestock diets. This study intended to screen and evaluate the ruminal cellulolytic bacteria (CBs) and their potential application as probiotics. Microbial culture and molecular techniques performed to isolate CBs from the rumen of camels, deer and rams. Their antibacterial and antibiogram tests were done using disc diffusion method. Their potential to degrade cellulose, starch, tannin and protein were investigated using clear zone halo, and spectrophotometric techniques. Bilious, saline, and acidic broth media were used to study the resistance of isolates in intestinal conditions. The phylogenetic analysis revealed that the strains belonged to Firmicutes and Proteobacteria phyla, Citrobacter murliniae, Ornithinibacillus bavariensis, C. braakii, and Bacillus subtilis. The highest cellulase (CAS) activity was recorded by C. murliniae Dez wildlife13A (2.98 UmL-1), whereas C. braakii Loot desert 111A (1.14 Uml-1) was produced the lowest enzyme. The isolates were highly resistant to synthetic conditions of intestine (pH 2.5-3.5, bile 0.3-2%), as well as tolerated higher concentrations of NaCl (up to 10%). They effectively inhibited standard pathogen strains, and showed sensitivity to the used antibiotics. This study reports the cellulolytic O. bavariensis Tabbas desert 32A for the first time from the rumen, which will have potential biotechnological applications.

Exploration of mobile genetic elements in the ruminal microbiome of Nellore cattle

Metagenomics has made it feasible to elucidate the intricacies of the ruminal microbiome and its role in the differentiation of animal production phenotypes of significance. The search for mobile genetic elements (MGEs) has taken on great importance, as they play a critical role in the transfer of genetic material between organisms. Furthermore, these elements serve a dual purpose by controlling populations through lytic bacteriophages, thereby maintaining ecological equilibrium and driving the evolutionary progress of host microorganisms. In this study, we aimed to identify the association between ruminal bacteria and their MGEs in Nellore cattle using physical chromosomal links through the Hi-C method. Shotgun metagenomic sequencing and the proximity ligation method ProxiMeta were used to analyze DNA, getting 1,713,111,307 bp, which gave rise to 107 metagenome-assembled genomes from rumen samples of four Nellore cows maintained on pasture. Taxonomic analysis revealed that most of the bacterial genomes belonged to the families Lachnospiraceae, Bacteroidaceae, Ruminococcaceae, Saccharofermentanaceae, and Treponemataceae and mostly encoded pathways for central carbon and other carbohydrate metabolisms. A total of 31 associations between host bacteria and MGE were identified, including 17 links to viruses and 14 links to plasmids. Additionally, we found 12 antibiotic resistance genes. To our knowledge, this is the first study in Brazilian cattle that connect MGEs with their microbial hosts. It identifies MGEs present in the rumen of pasture-raised Nellore cattle, offering insights that could advance biotechnology for food digestion and improve ruminant performance in production systems.

Insights on kraft lignin degradation in an anaerobic environment

Lignin is an aromatic macromolecule and one of the main constituents of lignocellulosic materials. Kraft lignin is generated as a residual by-product of the lignocellulosic biomass industrial process, and it might be used as a feedstock to generate low molecular weight aromatic compounds. In this study, we seek to understand and explore the potential of ruminal bacteria in the degradation of kraft lignin. We established two consortia, KLY and KL, which demonstrated significant lignin-degrading capabilities. Both consortia reached maximum growth after two days, with KLY showing a higher growth and decolorization rate. Additionally, SEM analysis revealed morphological changes in the residual lignin from both consortia, indicating significant degradation. This was further supported by FTIR spectra, which showed new bands corresponding to the C-H vibrations of guaiacyl and syringyl units, suggesting structural transformations of the lignin. Taxonomic analysis showed enrichment of the microbial community with members of the Dickeya genus. Seven metabolic pathways related to lignin metabolism were predicted for the established consortia. Both consortia were capable of consuming aromatic compounds such as 4-hydroxybenzoic acid, syringaldehyde, acetovanillone, and syringic acid, highlighting their capacity to convert aromatic compounds into commercially valuable molecules presenting antifungal activity and used as food preservatives as 4-hydroxyphenylacetic, 3-phenylacetic, and phenylacetic acids. Therefore, the microbial consortia shown in the present work are models for understanding the process of lignin degradation and consumption in bacterial anaerobic communities and developing biological processes to add value to industrial processes based on lignocellulosic biomass as feedstock.

227 Comparative analysis of rumen bacterial communities from bison heifers fed two forage-based diets of different quality

Abstract Bison have a demonstrated ability to thrive on low-quality forage, an attribute that distinguishes them from their domestic ruminant counterparts. This could be attributed to the capability of their rumen symbiotic microorganisms in extracting more nutrients out of plant biomass. To gain further insight into the role of the bison microbiome in this beneficial adaptation, the primary objective of this study was to determine the diversity and composition of ruminal bacteria in bison fed poor versus good quality forage-based diets. Ruminal content was collected by stomach tubing from 25 bison heifers fed good quality forage (GQG) during the summer (last week of June 2023) at the McGinley Ranch (Gordon, Nebraska, USA). The same individuals were then transitioned to low quality hay (PQG) in pen confinement for approximately 4 to 5 wk before resampling. Data generated from Illumina MiSeq (2×300) sequencing of PCR amplicons targeting the V1-V3 region of the 16S rRNA gene were analyzed using a combination of custom Perl scripts, and publicly available software (Mothur v.1.40, RDP classifier and NCBI Blast). Principal Coordinate Analysis (PCoA) showed a clear distinction in bacterial composition between the two types of forages (Figure 1). Further analysis of species-level operational taxonomic units (OTUs) was performed, and a total of 78,808 OTUs were identified across all samples. From the 20 most highly represented OTUs, the respective abundance of twelve OTUs showed significant differences between the two forage types (Kruskal-Wallis’s sum-rank test; P &amp;lt; 0.05). For example, OTUs SD_Bb-00014 (µGQG = 1.06% vs µPQG = 0.19%), and SD_Bb-00049 (µGQG = 0.98% vs µPQG = 0.08%) showed significant decreases in abundance when animals switched to PQG. Conversely, OTUs SD_Bb-00071 (µGQG = 0.2% vs µPQG = 2.71%), SD_Bb-00049 (µGQG = 0.68% vs µPQG = 1.67%) and OTUs SD_Bb-00148 (µGQG = 0.19% vs µPQG = 1.40%) exhibited a significant increase in abundance in response to the forage transition. Notably, only one of the assessed OTUs, SD_Bb-00071, showed high sequence identity to its closest valid relative, Fibrobacter succinogenes (98.41%). In contrast, most of the other highly represented OTUs were found to be very dissimilar to known species, suggesting that they might represent novel or currently uncharacterized bacterial species (Table 1). Together, these results demonstrate the significant impact of variations in nutritional value and forage quality on the composition of rumen bacterial communities in the North American bison. Poor forage quality appeared to favor bacterial species such as Fibrobacter succinogenes, known for its cellulolytic capabilities, presumably because of its greater efficiency in breaking down more resistant plant cell walls. Its ability and that of other abundant OTUs to efficiently convert plant fibers into assimilable nutrients is likely critical in allowing bison to perform better on poor-quality forage compared with domesticated ruminants.

Enhancing Rumen Fermentation and Bacteria Community in Sika Deer (Cervus nippon) through Varying Levels of Dragon Fruit Peel Polyphenolic Extract: An In Vitro Study.

The aim of this study is to investigate the effect of dragon fruit peel polyphenolic extract (DFPE) on gas production, rumen fermentation, and bacterial communities in sika deer using an in vitro technique. Three treatments with different DFPE levels (DFPE0, base diet; DFPE5, base diet + 5 mg/g DFPE; DFPE10, base diet + 10 mg/g DFPE, respectively; n = 6) were implemented. The phenolic composition of DFPE, gas production (GP), ammonia nitrogen (NH3-N), volatile fatty acid (VFA), and bacteria communities was evaluated after 24 h of incubation. The results showed that GP and NH3-N were reduced by DFPE supplementation. Total VFA, isovaleric acid, and valeric acid were increased (p < 0.05) by the addition of DFPE. No changes (p > 0.05) were observed in pH, acetic acid, propionic acid, isobutyric acid, butyric acid, and the ratio of acetic acid to propionic acid. Additionally, the alpha indexes, including Sobs, Shannon, and Ace, were increased by DFPE supplementation. Moreover, at the phylum level, DFPE supplementation increased (p = 0.01) Bacteroidota but reduced (p < 0.01) Firmicutes. At the genus level, compared to DFPE0, the DFPE10 had increased relative abundances of Rikenellaceae_RC9_gut_group (p < 0.01), norank_f_Muribaculaceae (p = 0.01), Lachnospiraceae_NK3A20_group (p < 0.01), Christensenellaceae_R-7_group (p < 0.01), and NK4A214_group (p < 0.01), decreased relative abundances of Streptococcus (p < 0.01), Oribacterium (p = 0.01), and Enterococcus (p < 0.01). Compared to DFPE0, DFPE5 had no change (p > 0.05) in all bacteria at the genus level except for decreased relative abundance of Enterococcus (p < 0.01). These results indicated that DFPE may be able to be used as a feed additive to enhance fermentation parameters and improve ruminal bacteria communities in Sika deer.

Cloning, expression and application of a novel laccase derived from water buffalo ruminal lignin-degrading bacteria

Water buffalo is the only mammal found to degrade lignin so far, and laccase plays an indispensable role in the degradation of lignin. In this study, multiple laccase genes were amplified based on the water buffalo rumen derived lignin-degrading bacteria Bacillus cereus and Ochrobactrum pseudintermedium. Subsequently, the corresponding recombinant plasmids were transformed into E. coli expression system BL21 (DE3) for induced expression by Isopropyl-β-D-thiogalactopyranoside (IPTG). After preliminary screening, protein purification and enzyme activity assays, Lac3833 with soluble expression and high enzyme activity was selected to test its characteristics, especially the ability of lignin degradation. The results showed that the optimum reaction temperature of Lac3833 was 40 °C for different substrates. The relative activity of Lac3833 reached the highest at pH 4.5 and pH 5.5 when the substrates were ABTS or 2,6-DMP and guaiacol, respectively. Additionally, Lac3833 could maintain high enzyme activity in different temperatures, pH and solutions containing Na+, K+, Mg2+, Ca2+ and Mn2+. Importantly, compared to negative treatment, recombinant laccase Lac3833 treatment showed that it had a significant function in degrading lignin. In conclusion, this is a pioneering study to produce recombinant laccase with lignin-degrading ability by bacteria from water buffalo rumen, which will provide new insights for the exploitation of more lignin-degrading enzymes.

Targeted and untargeted metabolomics reveals meat quality in grazing yak during different phenology periods on the Qinghai-Tibetan Plateau

Yak meat is more popular among consumers because of its high nutritional value, but little attention has been paid to its meat quality, which is affected by different phenology periods grass. We hypothesized that seasonal variations in grass composition influenced the ruminal bacteria community, and eventually affected the meat quality of yaks. This study aims to investigate the relationship of meat quality in grazing yak as well as the key rumen bacteria using targeted and untargeted metabolomics and 16S rRNA during different phenology periods. The main three altered metabolic pathways in grazing yak, including amino acids biosynthesis, glutathione metabolism, and fatty acids biosynthesis, were found in the grass period (GP) group compared to the regreen period (RP) and hay period (HP) groups. The GP group had higher concentrations of flavor amino acids (FAA), polyunsaturated fatty acids (PUFA), and a lower ratio of n-6/n-3 compared with the RP group. Correlation analysis results showed that Rikenellaceae_RC9_gut_group was positively correlated with fatty acids and lipid metabolites, which might be involved in lipid metabolism. Pediococcus had a positive correlation with biological peptides, which could be involved in the metabolism of bioactive compounds. In conclusion, grass in different phenology periods was associated with modified amino acids and fatty acids composition of yak meat as well as altered regulation of biological pathways, which was correlated with changes in rumen bacterial communities.

Feeding a Saccharomyces cerevisiae fermentation product during a gut barrier challenge in lactating Holstein cows impacts the ruminal microbiota and metabolome

Through its influence on the gut microbiota, feeding of Saccharomyces cerevisiae fermentation products (SCFP) has been a successful strategy to enhance the health of dairy cows during periods of physiological stresses. Although production and metabolic outcomes from feeding SCFP are well-known, combined impacts on the ruminal microbiota and metabolome during gut barrier challenges remain unclear. To address this gap in knowledge, multiparous Holstein cows (97.1 ± 7.6 DIM; n = 8/group) fed a control diet (CON) or CON plus 19 g/d SCFP for 9 wk were subjected to a feed restriction (FR) challenge for 5 d, during which they were fed 40% of their ad-libitum intake from the 7 d before FR. DNA extracted from ruminal fluid was subjected to PacBio Full-Length 16S rRNA gene sequencing, RT-PCR of 12 major ruminal bacteria, and metabolomics analysis of up to 189 metabolites via GC-MS. High-quality amplicon sequence analyses were performed with Targeted Amplicon Diversity Analysis (TADA), MicrobiomeAnalyst, PICRUSt2, and STAMP software, while metabolomics data were analyzed via MetaboAnalyst 5.0. Ruminal fluid metabolites from the SCFP group exhibited a greater α diversity Chao 1 (P = 0.03) and Shannon indices (P = 0.05), and the PLS-DA analysis clearly discriminated metabolite profiles between dietary groups. The abundance of CPla_4_termite_group, Candidatus_Saccharimonas, Oribacterium, and Pirellula genus in cows fed SCFP was greater. In the SCFP group, concentrations of ethanolamine, 2-amino-4,6-dihydroxypyrimidine, glyoxylic acid, serine, threonine, cytosine, stearic acid, and pyrrole-2-carboxylic acid were greater in ruminal fluid. Both Fretibacterium and Succinivibrio abundance were positively correlated with metabolites across various biological processes: gamma-aminobutyric acid, galactose, butane-2,3-diol, fructose, 5-amino pentanoic acid, β-aminoisobutyric acid, ornithine, malonic acid, 3-hydroxy-3-methylbutyric acid, hexanoic acid, heptanoic acid, cadaverine, glycolic acid, β-alanine, 2-hydroxybutyric acid, methyl alanine, and alanine. In the SCFP group, compared with CON, the mean proportion of 14 predicted pathways based on metabolomics data was greater, while 10 predicted pathways were lower. Integrating metabolites and upregulated predicted enzymes (NADP+-dependent glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, serine: glyoxylate aminotransferase, and D-glycerate 3-kinase) indicated that the pentose phosphate pathway and photorespiration pathway were most upregulated by SCFP. Overall, SCFP during FR led to alterations in ruminal microbiota composition and key metabolic pathways. Among those, there was a shift from the tricarboxylic acid (TCA) cycle to the glyoxylate cycle and nitrogenous base production was enhanced.

Impact of High-Moisture Ear Corn on Antioxidant Capacity, Immunity, Rumen Fermentation, and Microbial Diversity in Pluriparous Dairy Cows

Due to the increasing costs of livestock farming, it is important to find cost-effective alternatives of feed stuffs. This study investigated the effects of high-moisture ear corn (HMEC) feeding on the production performance, serum antioxidant capacity, immunity, and ruminal fermentation and microbiome of dairy cows. Thirty pluriparous Chinese Holstein cows were randomly allocated to two groups: steam-flaked corn (SFC) and HMEC (replacement of 2 kg equal dry matter SFC) and fed for a 60 day trial. The results showed replacing SFC with HMEC significantly increased dry matter intake, milk yield, and 4% fat-corrected milk yield (p &lt; 0.05). Serum levels of superoxide dismutase, glutathione peroxidase, and immunoglobulins G, M, and A were significantly higher, and those of creatinine and cholesterol were significantly lower, in the HMEC group than in the SFC group (p &lt; 0.05). HMEC also significantly increased total volatile fatty acid and acetate (p &lt; 0.05) concentrations. In both groups, the dominant phyla of ruminal bacteria were Bacteroidetes, Firmicutes, and Actinobacteria, and the dominant genera were Prevotella, NK4A214-group, and Succiniclasticum. Mogibacterium, Eubacterium nodatum group, norank-f-Lachnospiraceae, and Eubacterium brachy group were significantly enriched in the ruminal fluid of HMEC-group cows (p &lt; 0.05). In conclusion, replacing SFC with HMEC improved production performance, antioxidant capacity, and immunity, while regulating both ruminal fermentation and the composition of the ruminal microbiome in dairy cows.

Feeding Pellets Containing Agro-Industrial Waste Enhances Feed Utilization and Rumen Functions in Thai Beef Cattle.

The objective of this research was to investigate the effects of citric waste fermented with yeast waste pellet (CWYWP) supplementation on feed intake, rumen characteristics, and blood metabolites in native Thai beef cattle that are fed a rice-straw-based diet. Four native male Thai beef cattle (1.0-1.5 years old) with an initial body weight (BW) of 116 ± 16 kg were held in a 4 × 4 Latin square design within 21-day periods. The animals were assigned to receive CWYWP supplementation at 0%, 2%, 4%, and 6% of the total dry matter (DM) intake per day. The results indicate that feeding beef cattle with CWYWP leads to a linear increase in the total intake as well was the intake of crude protein (CP) and the digestibility of CP, with the maximum levels observed at 6% CWYWP supplementation (p < 0.05). Rumen characteristics, including pH, blood urea-nitrogen concentration, and protozoal population, showed no significant alterations in response to the varying CWYWP dosages (p > 0.05). In addition, the CWYWP supplementation resulted in no significant changes in the concentration of ammonia-nitrogen, remaining within an average normal range of 10.19-10.38 mg/dL (p > 0.05). The inclusion of 6% CWYWP resulted in the highest population of ruminal bacteria (p < 0.05). Additionally, the CWYWP supplementation led to a statistically significant increase in the mean propionic acid concentration as compared to the group that did not receive the CWYWP supplementation (p < 0.05). In conclusion, this experiment demonstrates that supplementing Thai native beef cattle with CWYWP at either 4% or 6% DM per day can enhance their total CP intake as well as the CP digestibility and rumen bacterial population, and can increase propionate concentration.