Search Results (12)

Background and Objectives: Dairy products provide vital energy, high-quality protein, and micronutrients for over six billion people worldwide, with dairy cows contributing nearly 81% of global milk production. Sustainable strategies to enhance productivity are therefore critical. Feed additives such as essential oil blends (EOB), onion peel (OPE), and prebiotics including mannan oligosaccharides (MOS) and galacto-oligosaccharides (GOS) have been proposed to improve rumen fermentation, modulate microbial ecology, and mitigate greenhouse gas emissions. This study evaluated the combined effects of EOB, OPE, MOS, and GOS on rumen metabolism using the rumen simulation technique (RUSITEC). Materials and Methods: Rumen inoculum from three cannulated Holstein Friesian cows was incubated across 16 vessels (four treatments × four replicates) for nine days. Treatments included a control (CON; TMR only), GEO (TMR + GOS + EOB + OPE), MEO (TMR + MOS + EOB + OPE), and OLEO (TMR + a 1:1 mixture of GOS and MOS + EOB + OPE). Additives were included at 3 µL/g TMR for EOB and 30 mg/g TMR (3% w/w) for OPE, GOS, MOS, or OLG. Rumen effluents were collected for untargeted metabolomic profiling by liquid chromatography–mass spectrometry, identifying 661 metabolites. Results: Partial least squares-discriminant analysis revealed clear separation between CON and additive groups, confirming distinct metabolic shifts. GEO primarily enhanced tryptophan, tyrosine, and purine metabolism; MEO stimulated phosphonate and pyrimidine pathways and bile acid biosynthesis; OLEO promoted phosphonate, nicotinamide, and taurine metabolism. Microbial analysis showed enrichment of taxa such as Lachnospira, Succinivibrionaceae, Macellibacteroides, Lysinibacillus, and Christensenellaceae, indicating complementary effects on fermentation and microbial stability. Conclusions: These results demonstrate that dietary supplementation with GEO, MEO, or OLEO modulates rumen metabolism and microbial ecology without impairing fermentation, supporting improved nutrient utilization, antioxidant defenses, and metabolic resilience in dairy cows, with potential benefits for productivity and sustainability. Full article

Background and Objectives: The potential of essential oils (EOs) and fumaric acid (FA) to modulate ruminal fermentation and mitigate greenhouse gas emissions in dairy cows has attracted significant attention. However, little is known about the specific metabolites produced as a result of their interaction. This study investigated the combined effects of essential oil blends (EOBs) and FA on rumen metabolites using a rumen simulation technique (RUSITEC) system. Materials and Methods: Three rumen-cannulated, non-lactating Holstein Friesian cows served as inoculum donors. The total mixed ration (TMR; CON) comprised corn silage (60%), alfalfa hay (20%), and concentrate (20%). Three distinct EOBs were formulated: EOB1 [Garlic, Lemongrass, Cumin, Lavender, and Nutmeg; at 4:2:2:1:1, respectively], EOB2 [Anise, Clove, Oregano, Cedarwood, and Ginger; at 4:2:2:1:1, respectively], and EOB3 [Clove, Anise, Peppermint, and Oregano; at 4:3:2:1, respectively]. Four treatments evaluated were control (CON), EFA1 (EOB1 + FA), EFA2 (EOB2 + FA), and EFA3 (EOB3 + FA). EOBs and FA were included at 10 µL/g feed and 3% of TMR, respectively. Rumen effluents were collected over 5 days for metabolome analysis using liquid chromatography-mass spectrometry (LC–MS). Results: A total of 661 metabolites were detected and identified. Volcano plot analysis revealed 13 differentially abundant metabolites for EFA1, 41 for EFA2, and 19 for EFA3 compared to CON group. PLS-DA analysis showed clear separation of treatments, indicating modifications in the rumen fluid metabolome. Conclusions: The treatments led to the enrichment of pathways involved in amino acid, nucleotide, cofactor, and energy metabolism. These additives have the potential to optimize nutrient utilization and overall animal health. Therefore, in vivo studies should be conducted to validate their efficacy. Full article

In this study, the RUSITEC system was used to study the regulation of rumen-derived Pseudomonas sp. PSC001 (PSC001) on the rumen environment contaminated by Zearalenone (ZEN). The rumen fluid of dairy cows was selected as the fermentation broth, and four experimental groups were set up: control group (CON), Pseudomonas group (PS), ZEN pollution group (ZEN), and PS and ZEN co-treatment group (PS + ZEN). The NH₃-N, microbial protein (MCP), and volatile fatty acid (VFA) in the rumen fermentation broth were measured after culturing, and the changes in microbial community structure in rumen fluid were analyzed by 16S rRNA gene sequencing. After adding PSC001, the concentration of propionic acid, valeric acid, and butyric acid increased, and the acetate to propionate ratio and concentration of isovaleric acid decreased. ZEN exposure can lead to an abnormal increase in NH₃-N, valeric acid, and isovaleric acid content and a decrease in MCP content. The content of NH₃-N, valeric acid, and isovaleric acid decreased and the content of MCP increased in the PS + ZEN combined treatment group. The addition of PSC001 and ZEN significantly or extremely significantly increased the abundance of 18 genera and significantly or extremely significantly decreased the relative abundance of 5 genera in rumen fluid, respectively. It is worth noting that with the addition of both at the same time, the abundance of four genera in the PS + ZEN group was significantly or extremely significantly increased among the five genera with decreased abundance in the ZEN group. Among the 18 genera with increased abundance in the ZEN group, 10 genera in the PS + ZEN group decreased significantly or extremely significantly. In summary, the addition of PSC001 alleviated the negative impact of ZEN on the internal environment of rumen fermentation, and it also had a positive regulatory effect on rumen fermentation. Full article

Livestock farming is a vital component of global food security, yet it remains a major contributor to greenhouse gas (GHG) emissions, particularly methane (CH₄), which has a global warming potential 28 times greater than carbon dioxide (CO₂). This review provides a comprehensive synthesis of current knowledge surrounding the sources, biological mechanisms, and mitigation strategies related to CH₄ emissions from ruminant livestock. We first explore the process of methanogenesis within the rumen, detailing the role of methanogenic archaea and the environmental factors influencing CH₄ production. A thorough assessment of both direct and indirect methods used to quantify CH₄ emissions is presented, including in vitro techniques (e.g., syringe method, batch culture, RUSITEC), in vivo techniques (e.g., respiration chambers, Greenfeed, laser CH₄ detectors), and statistical modeling approaches. The advantages and limitations of each method are critically analyzed in terms of accuracy, cost, feasibility, and applicability to different farming systems. We then examine a wide range of mitigation strategies, organized into four core pillars: (1) animal and feed management (e.g., genetic selection, pasture quality improvement), (2) diet formulation (e.g., feed additives such as oils, tannins, saponins, and seaweed), (3) rumen manipulation (e.g., probiotics, ionophores, defaunation, vaccination), and (4) manure management practices and policy-level interventions. These strategies are evaluated not only for their environmental impact but also for their economic and practical viability in diverse livestock systems. By integrating technological innovations with sustainable agricultural practices, this review highlights pathways to reduce CH₄ emissions while maintaining animal productivity. It aims to support decision-makers, researchers, and livestock producers in the global effort to transition toward climate-smart, low-emission livestock farming. Full article

In vitro methods have advanced research on rumen microbiology and fermentation. However, artificial saliva formulation may need adjustments, particularly in urea content, for modern diets, warranting further research. This study investigated the effects of different nitrogen (N) levels in artificial saliva on ruminal fermentation and digestion in diets for dairy cows using a Rusitec^® system. Eighteen fermenters tested three saliva treatments with different N levels: a standard saliva as the control and two treatments with N reduced by 15% and 30%. Data were analyzed as a completely randomized design using the MIXED procedure of SAS (v. 9.4), with linear and quadratic contrasts tested for treatment effects (significance set at p ≤ 0.05). Results showed that altering N content had no significant effect on pH, ammonia concentrations, or NH₃-N outflow, nutrient digestibility (dry matter, crude protein, fiber, and starch), gas and methane production, or volatile fatty acid concentrations. The efficiency of microbial protein synthesis and N flow exhibited quadratic responses, with the lowest values observed at the highest level of N reduction in the saliva (−30%). These findings suggest that although ruminal function and digestion remain stable with reduced N, microbial protein synthesis efficiency may decline beyond a threshold. Full article

Artificial gastrointestinal tract models have become essential tools in veterinary medicine, providing alternatives to in vivo studies, which are labor-intensive, costly, and under certain circumstances even ethically challenging. These in vitro models facilitate the study of digestion, enable disease and host–pathogen interaction modeling, and allow for the investigation of nutrient absorption, microbiota, and pharmacokinetics. Considering the One Health concept, the application of gastrointestinal tract systems in investigations for animals can clearly reflect human health, and thus, it is pointing to the relevance of the adaptation of already existing models and the development of new models to meet the needs of veterinary and animal farming practices. This review explores and compares the various types of gastrointestinal tract models, including static and dynamic systems, and their applications across different animal species. Specific technical and methodological considerations are discussed for core animal-developed and -tested artificial systems and their integration with common ‘omics’ techniques. Dynamic models, such as RUSITEC and PolyFermS, more accurately simulate in vivo processes, including peristalsis, enzymatic activity, and microbial fermentation. The studies employing tools for ‘omics’ approaches have been conducted with more understanding analysis and comprehensive discussion and results. Full article

This study evaluated the synergistic effects of prebiotics containing galacto-oligosaccharides (GOS) and/or mannan oligosaccharides (MOS), essential oil blend (EOB), and onion peel (OPE) on fermentation characteristics using the rumen simulation technique (RUSITEC) system. Three rumen-cannulated, non-lactating Holstein Friesian cows were the inoculum donors. The substrate used for the study was a total mixed ration (TMR), which consisted of corn silage, alfalfa hay, and concentrate at 6:2:2, respectively. Sixteen fermentation vessels were randomly allotted to four treatments with four replicates each over a 9-day period in a completely randomized design. The treatments assessed include: control [TMR only], GEO [TMR + GOS + EOB + OPE], MEO [TMR + MOS + EOB + OPE], and OLEO [TMR + OLG + EOB + OPE]. OLG comprises GOS and MOS in equal proportion. EOB was included at 3 µL/g, while OPE, GOS, MOS, and OLG were added at 30 mg/g TMR. Results showed that pH, gas volume, effluent volume, and ammonia-N were not affected (p > 0.05) by the different additives. Similarly, greenhouse gas (GHG) emissions and nutrient digestibility were not affected by the treatments. Compared to the control, total volatile fatty acids (VFA) were decreased (p < 0.05) by 14.8, 10.8, and 8.5% with GEO, MEO, and OLEO inclusion, respectively, while the molar proportion of acetate was increased (p = 0.011) by 3.3, 1.1, and 3.8% with GEO, MEO, and OLEO inclusion, respectively. MEO increased isobutyrate (p = 0.001) and branched chain VFA (p = 0.013) contents; however, GEO and OLEO inclusion reduced them. Overall, the interaction of EOB, OPE, GOS, and/or MOS did not affect nutrient digestibility or GHG emissions but reduced VFA production. Further research is recommended to assess the dose effect of the additives on GHG emissions and VFA production; and to determine the long-term effects of these interventions on the rumen microbiome and animal performance. Full article

The study aimed to investigate the effect of dietary chitosan oligosaccharides (COS) meal levels on the nutrient disappearance rate, rumen fermentation, and microflora of beef cattle in vitro. A total of 24 fermentation tanks were randomly divided into four treatments containing 0% COS (CON), 0.02% COS, 0.04% COS, and 0.08% COS for an 8-day experiment period, with each treatment comprising six replicates. The disappear rates of DM, CP, EE, and total gas production were quadratically increased with increasing COS levels. The disappear rates of DM, CP, EE, and ADF were greatest, whereas the total gas production was lowest in the 0.08% COS group. The pH, NH₃-N, MCP, the content of propionate, isobutyrate, butyrate, valerate, and the A/P were quadratically increased with increasing COS levels, while the A/P were linearly decreased. The pH, MCP, and the content of propionate, and butyrate were highest, whereas the NH₃-N and the content of acetate, isobutyrate, valerate, and the A/P were lowest in the 0.08% COS group. Microbiomics analysis showed that the rumen microbial diversity was not altered between the CON and the 0.08% COS group. However, the relative abundance of Methanosphaera, Ruminococcus, Endomicrobium, and Eubacterium groups was increased, and the relative abundance of pathogenic bacteria Dorea and Escherichia-Shigella showed a decrease in the 0.08% COS group. Overall, the 0.08% COS was the most effective among the three addition levels, resulting in an increase in the disappearance rate of in vitro fermented nutrients and improvements in rumen fermentation indexes and microbial communities. This, in turn, led to the maintenance of rumen health. Full article

This study investigated the combined impact of essential oil blends (EOBs) and fumaric acid (FA) on ruminal fermentation in dairy cows using the rumen simulation technique (RUSITEC) system. Three rumen-cannulated, non-lactating Holstein Friesian cows served as inoculum donors. The substrate, a total mixed ration (TMR), comprised corn silage, alfalfa hay, and a concentrate mix in a 3:1:1 ratio. The four treatments evaluated were Control (TMR without additives), EFA1 (TMR + EOB1 + FA), EFA2 (TMR + EOB2 + FA), and EFA3 (TMR + EOB3 + FA). Sixteen fermentation chambers were randomly assigned to the treatments, each with four replicates, following a completely randomized design during a 9-day experimental period. EOBs and FA were added at 10 µL/g feed and 3% of TMR, respectively. After a 4-day adaptation, samples were collected for 5 days. Results revealed that EFA1 significantly reduced (p = 0.0351) CH₄ emissions by 60.2% without negatively impacting dry matter disappearance, fiber fraction digestibility, pH, or gas volume. All EFAs increased (p < 0.001) the propionate molar proportion and decreased (p < 0.001) the acetate-to-propionate ratio. EFA2 decreased (p < 0.05) the acetate proportion by 3.3% compared to the control. In conclusion, EFA1 is recommended as an effective nutritional intervention to mitigate CH₄ emissions and optimize ruminal fermentation in dairy cows. Full article

The aim of this study was to distinguish effects due to diet composition from those triggered by ruminal pH on fermentation patterns and microbial profiles in a continuous culture system (RUSITEC). The study followed a 2 × 2 factorial design, with two diets varying in the proportions of forage and concentrate and two pH levels in the culture medium. RUSITEC fermenters were used to simulate rumen fermentation and feed digestibility, fermentation end-products, microbial protein synthesis, microbial community, and long-chain fatty acid profiles in the digesta were determined. Multivariate analyses were applied to summarize the overall results. High concentrate (34% cereal grain, 32% hay) diets were more digestible (p < 0.05) than high forage (10% cereal grain, 78% hay) diets, resulting in a greater (p < 0.05) formation of most fermentation end-products and microbial protein in the rumen. However, there were no significant (p > 0.05) differences between diets in methane production. Ciliate protozoa, anaerobic fungi, some fibrolytic bacteria, hydrogenation of oleic acid, and relative proportion of conjugated linoleic acid were increased (p < 0.05) with high forage diets. A decline in rumen pH from 6.8 to 6.4 decreased (p < 0.05) feed digestibility, protein degradability, and the daily outputs of some fermentation end-products (gas, VFA, acetate, ammonia) but had no effect (p > 0.05) on the synthesis of microbial protein, and on the output of methane, propionate, butyrate or lactate. Minor changes in microbial community profile or the fatty acid relative proportions were observed within this pH range. The overall multivariate analysis revealed a clear discrimination between high-concentrate and high-forage diets, with subtler and less-defined pH effects on ruminal fermentation and microbial communities. Full article

This study aimed at characterizing changes in rumen bacteria abundance and fermentation profiles by artificial saliva (AS) pH, and at evaluating the potential modulatory role of Aspergillus oryzae culture (AOC) in a rumen simulation technique (RUSITEC) system. The treatment included high AS pH (pH 6.8) or low AS pH (pH 5.5) according to the McDougall’s method, and low AS pH was sustained by changing the composition of the AS (NaHCO₃ from 9.8 to 1.96 g/L, Na₂HPO₄ from 9.3 to 1.86 g/L). In low AS pH condition, the diets contained either 0% AOC, 1.25% AOC, or 2.5% AOC. Therefore, there are four treatments: (1) high AS pH, 0% AOC (HASP); (2) low AS pH, 0% AOC (AOC0); (3) low AS pH, 1.25% AOC (AOC1); (4) low AS pH, 2.5% AOC (AOC2), respectively. The experimental diets were supplemented with 16 g basic diets with the forage to concentrate ratio of 40:60. The experiments were conducted two independent 13 days, with 9 days adaption periods and 4 days sample collection. The results showed that low AS pH decreased the degradabilites of dry matter (DM), organic matter (OM), crude protein (CP), neutral detergent fiber (NDF), and acid detergent fiber (ADF) (p < 0.05), which occurred due to a decreased abundance of fibrolytic Ruminococcus albus (p < 0.001). The total concentration of volatile fatty acid (VFA) and proportion of propionate were decreased in the low AS pH (p = 0.026) and tended to increase the molar proportion of butyrate (p = 0.086) and the ratio of acetate to propionate (p = 0.088). The abundances of phylum Firmicutes (p = 0.065) and Proteobacteria (p = 0.063) tended to be greater in low AS pH group than high AS pH group. Low AS pH increased the abundance of phylum Actinobacteria (p = 0.002) compared to the high AS pH and decreased the abundances of phylum Spirochaetes (p = 0.032). Compared with the high AS pH, low AS pH increased the abundances of Prevotella (p = 0.003), Pseudoscardovia (p = 0.001), Mitsuokella (p = 0.005), and Dialister (p = 0.047), and decreased the abundances of Olivibacter (p = 0.026), Ruminobacter (p = 0.025), Treponema (p = 0.037), and Sphaerochaeta (p = 0.027) at genus level. Under a severe SARA in RUSITEC, supplementation of 2.5% AOC increased OM degradability, the copy numbers of Selenomonas ruminantium and Fibrobacter succinogenes. These findings indicate that the reduction AS pH at 5.5 caused a strong shift in bacterial composition in rumen. In addition, the addition of AOC in diets increased the growth rate of certain rumen bacteria that digest fiber or utilize lactate under SARA condition in RUSITEC system. Full article

The Australian red meat industry has set a goal to be carbon neutral by 2030. Reaching this goal will be a challenge and will involve targeting ways to increase carbon in the landscape, improve efficiency of production and reduce methane emissions from ruminants. There are a number of different options the industry can pursue to try and achieve its goal, including changing grazing management practices and land-use to changing the animal, what it eats and the microbial ecology in their rumen. No single one of these options will enable the red meat industry to become carbon neutral by 2030, it will take a combination of all of them to help meet the challenge. We have been using an in vitro batch fermentation system and a Rusitec system as a quick, relatively inexpensive, way to screen; plants that already exist in our grazing systems, novel plants, plant extracts and organic waste products from the horticultural industry, for their potential to improve the efficiency of fermentation and reduce methane production in the rumen. We have also used these systems to provide an initial clue about the mechanism of action at the level of the ruminal microorganisms. We have identified variation in these traits amongst the plants, plant extracts and horticultural waste products we have tested that could help develop systems that reduce the environmental footprint of ruminants in tropical production systems in Australia and in other parts of the world. Full article