The objective of this study was to investigate the effects of Acalypha australis L. extract (ALE) on the growth performance and intestinal health in piglets. A total of 24 weaned piglets were randomly allocated to three groups: the control group (CON), which was fed a basal diet, and the ALE0.5 and ALE1.0 groups, which were fed the basal diet supplemented with 0.5 and 1.0 g/kg of ALE, respectively. The measured variables included growth performance, digestive enzyme activity, intestinal morphology, antioxidant capacity, and intestinal microbiota and metabolites. The results showed that, compared to the CON group, supplementation of 1.0 g/kg ALE in the diets of weaned piglets significantly increased the ratio of gain to feed from 15 to 21 days (p < 0.05), decreased the diarrhea rate from Days 15 to 21 and Days 0 to 21 (p < 0.05), and increased the activities of pancreatic α-amylase, lipase, trypsin, and chymotrypsin, as well as duodenal α-amylase, lipase, and trypsin, and jejunal maltase and sucrase (p < 0.05). Additionally, supplementation of 1.0 g/kg ALE in the diet significantly improved the intestinal morphology of the duodenum and jejunum, as well as the expression of intestinal barrier-related genes in the small intestine (p < 0.05). Moreover, it significantly increased serum glutathione peroxidase activity and jejunal and ileal superoxide dismutase activities (p < 0.05), and also significantly increased the colonic propionic acid concentration of piglets (p < 0.05). The ALE supplementation increased the abundance of the colonic marker bacteria Collinsella in the piglets and influenced pathways related to amino acid metabolism, carbohydrate metabolism, and lipid metabolism. ALE can serve as a potential natural feed additive to regulate the structure of intestinal microbiota and metabolic pathways, enhance antioxidant capacity, improve intestinal health, reduce diarrhea incidence, and ultimately promote the growth performance of piglets.

As auxiliary components of the carbohydrate active enzymes (CAZymes), carbohydrate-binding modules (CBMs) influence the enzymatic hydrolysis of substrates. To investigate the role of anaerobic fungal CBMs in lignocellulose degradation, an architectural analysis of bacterial and fungal CBM-containing protein sequences was completed. Results indicated that 67.9% of the fungal CBMs were incorporated into the plant-biomass-degrading enzymes and 51.1% of the anaerobic fungal CBMs were biased to be fused with the hemicellulose-degrading enzymes. Based on the transcriptomic data of anaerobic fungus Pecoramyces ruminantium F1, three upregulated CBM-fused hemicellulose-degrading enzyme gene clusters were identified. Results suggested that the fused CBMs retained the enzymatic function of the associated CAZymes. Importantly, the CBM1 domains of acetyl xylan esterase (AxeA16138) and α-L-arabinofuranosidase (AraA02173) possessed a crucial role to promote the xylanase-mediated hydrolysis of hemicellulose. These results demonstrated that anaerobic fungal CBMs harbor substantial potential to enhance hemicellulose degradation.

Bacteriophage adhesion to intestinal mucus has been proposed as an important mechanism for preventing bacterial infection. However, whether differences in adhesion capacity among closely related phages translate into distinct protective outcomes remains unclear. In this study, we compared two T4-like bacteriophages, S143_2 and W143, to investigate how variation in mucosal adhesion influences antibacterial efficacy and host responses. Both phages exhibited comparable lytic activity against enteropathogenic Escherichia coli (EPEC143) but differed in adhesion capacity to intestinal epithelium. In vitro assays demonstrated that phages adhered to mucus-secreting IPEC-1 cells via a reversible mucus association, with S143_2 displaying significantly stronger adhesion than W143. This difference was further confirmed in vivo, where S143_2 showed higher mucosal retention and a significantly greater mucosa-to-digesta ratio of phage titers in the small intestine. Functionally, enhanced adhesion was associated with improved protective efficacy. In a prophylactic mouse model, pretreatment with S143_2 resulted in reduced body weight loss and lower intestinal pathogen load compared to W143. Furthermore, S143_2 induced a more robust systemic immune response evidenced by elevated serum cytokines and the enrichment of immune-related signaling pathways in the intestinal transcriptome. Together, these findings demonstrate that mucosal adhesion capacity is a critical determinant of phage performance in vivo. Our results highlight that, beyond host range and bactericidal capacity, adhesion properties should be considered a key criterion in the rational selection of bacteriophages for preventing and treating mucosa-associated infections.IMPORTANCEPhages are among the most promising antibiotic alternatives, yet their evaluation has largely focused on bacterial host range and lytic activity. Our findings highlight mucosal adhesion as a previously overlooked but decisive factor in phage efficacy when targeting mucosa-associated infections. Phages with stronger epithelial adhesion exhibit superior protection and immune modulation in the gut, underscoring that adhesion capacity should be integrated as a key criterion in the rational selection and engineering of therapeutic phages.

Yaks are important livestock species on the Qinghai-Tibet Plateau (QTP), but their productivity is constrained by the harsh alpine environment and the seasonal scarcity of forage resources. Improving feed efficiency has become the key to the sustainable development of Plateau Yak breeding industry. Phytosterols have shown a good application prospect in ruminant feed efficiency improvement, but the impact on yaks is not clear. This study explored the effects of dietary phytosterols supplementation on growth performance, nutrient digestibility, rumen microbial community structure and metabolic function of yaks fed in a barn. Twenty-eight 1.5-year-old male yaks (137.10 ± 8.70 kg) were randomly divided into two groups after 28 days of pre feeding period: the control group (Con, n = 14) was fed with basic diet, and the experimental group (PS, n = 14) was added with 200 mg/kg phytosterols in concentrate. The formal test period was 60 days. Compared with Con group, PS group significantly increased average daily gain (P = 0.001), apparent digestibility of crude protein (P = 0.036) and neutral detergent fiber (P = 0.006), and reduced feed conversion rate (P = 0.002). The rumen fermentation mode of PS group changed, the proportion of propionate increased significantly (P = 0.001), while the proportion of acetate (P = 0.006), acetate to propionate ratio (P = 0.001) and lactate (P = 0.035) concentration decreased significantly. Phytosterols significantly changed the structure of rumen microbial community and significantly increased the copy number of rumen bacteria (P < 0.05). Metatranscriptomic analysis showed that PS group significantly increased the relative abundance of key bacteria, including Succiniclasticum, Faecalibacterium, Ruminococcus, Butyrivibrio, and Alistipes (P < 0.05). Microbial co-occurrence network analysis revealed that the number of edges, average degree and modular index of microbial community structure network in PS group increased. Rumen function analysis showed that glycolysis / gluconeogenesis and propionate metabolism were significantly enhanced in PS group (P < 0.05), and the expression of key enzymes such as hexokinase and fumarate hydratase were significantly enhanced (P < 0.05). Metabolomic analysis revealed that phosphoenolpyruvic acid, fumarate and methylmalonyl-CoA were significantly accumulated in the rumen of PS group (P < 0.05). Pathway analysis showed that the pathway impact value of propionate metabolism and glycolysis / gluconeogenesis was greater than 0.1. Phytosterols (200 mg/kg of concentrate) can effectively improve the feed efficiency of yaks by regulating the structure and function of rumen microorganisms and altering the fermentation patterns.

Dietary protein plays a crucial role in shaping the gut microbiome and modulating intestinal amino acid metabolism. Gut microbiome is recognized as a reservoir for carrying antimicrobial resistance genes. However, the relationship between amino acids metabolism and antibiotic resistome remains poorly understood. Here, a pig model was used to study this relationship by comparing the impact of dietary casein hydrolysate diet with those of an intact casein diet. Metabolomics analysis revealed that casein hydrolysate supplementation primarily altered amino acid metabolism, characterized by significantly reduced levels of several amino acids, including tyrosine and glutamine, accompanied by increased levels of amino acid–derived metabolites. Metagenomics analyses indicated that these metabolic shifts were closely associated with microbial changes in the gut, particularly the genera Escherichia and Bifidobacterium. Consistently, microbial genes related to amino acid transport and metabolism exhibited higher abundances. Notably, the abundances of antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs) were significantly enriched in response to casein hydrolysate supplementation. Integrated metabolome–resistome correlation analyses revealed significant associations between multiple amino acids, including tyrosine and glutamine, and distinct ARG subtypes, indicating a tight coupling between amino acid metabolism and antibiotic resistance potential. Metagenomics binning and assembly further resolved the taxonomic origins of these functional traits. Specifically, in Escherichia fergusonii and Bifidobacterium thermophilum, genes related to amino acid metabolism, ARGs, and MGEs were co-localized on the same contigs with close genomic proximity. Together, these findings highlight a strong link between microbial amino acid metabolism and the resistome, suggesting that dietary casein hydrolysate reshapes both microbial metabolic functions and antibiotic resistance potential within the intestinal ecosystem.

Alpha-ketoglutarate (AKG), a key intermediate in the tricarboxylic acid cycle, has been demonstrated to exert multiple physiological benefits, including promoting growth, enhancing nitrogen utilization, improving immunity, and optimizing intestinal health in monogastric livestock and aquatic animals. However, studies investigating its effects in ruminants remain limited. This study systematically evaluated the effects of AKG on rumen fermentation characteristics, production performance, nutrient digestibility, and health parameters in mid-lactation dairy cows through a combination of in vitro fermentation and in vivo feeding trials. The in vitro experiment was conducted using 4 AKG concentrations (0, 5, 15, and 45 mg/dL), with each treatment performed in quadruplicate and repeated across 3 batches. Rumen fermentation parameters and microbial crude protein (MCP) were measured to determine the optimal AKG dose. Results showed that AKG supplementation linearly increased total gas production, MCP, and total VFA concentrations, while linearly decreasing pH and NH3-N levels. All AKG supplementation levels improved in vitro rumen fermentation characteristics, with stronger responses observed at higher doses. Based on these in vitro findings, an in vivo experiment was performed using 24 multiparous Holstein cows, stratified by milk yield into 2 blocks. Within each block, cows were randomly assigned to one of 2 treatment groups (n = 12 per group): a control group fed the basal diet, and an AKG group fed the basal diet supplemented with 25 g of AKG per cow per day. The trial lasted 10 weeks, including a 2-week adaptation period and an 8-week experimental period. AKG supplementation significantly increased milk protein yield, while numerical increases were observed for DMI, milk yield, ECM, and lactose yield that did not reach statistical significance. Additionally, dietary AKG markedly enhanced ruminal MCP, butyrate, valerate, isobutyrate, isovalerate, and total branched-chain volatile fatty acid concentrations, with a tendency to increase total VFA. Compared with the control group, cows in the AKG group exhibited significantly higher apparent digestibility of DM, OM, and CP. Plasma biochemical and immune analyses revealed that AKG supplementation significantly decreased aspartate aminotransferase and γ-glutamyl transferase activities, while markedly increasing IgG levels, suggesting a more favorable hepatic metabolic profile and enhanced humoral immune response. AKG supplementation did not affect the population of bacteria, protozoa, methanogens, or fungi, but reduced the α-diversity of rumen bacteria. At the phylum level, AKG supplementation did not affect rumen bacterial abundance, but at the genus level, it tended to increase the abundance of Prevotella while decreasing that of Barnesiella, Coprobacter, and Desulfovibrio. In summary, the in vitro experiments showed that 15-45 mg/dL of AKG was identified as an appropriate supplementation level. The in vivo feeding trial further demonstrated that dietary supplementation with 25 g/d AKG enhanced rumen fermentation, improved nutrient digestibility and plasma immune-related biomarkers, and increased milk protein production in dairy cows. These changes were associated with decreased bacterial α-diversity and a trend toward increased Prevotella abundance.

The gayal (Bos frontalis), a semi-domesticated bovine species, demonstrates exceptional adaptability to lignocellulose-rich diets dominated by bamboo, suggesting the presence of a specialized gastrointestinal microbiome. However, the functional mechanisms underlying this host-microbiome interaction remain poorly understood. Here, we conducted integrated metagenomic and metatranscriptomic analyses of rumen, cecum, and colon digesta from yellow cattle and gayal raised on the same bamboo-based high-fiber diet. The results showed that gayal exhibited superior fiber-degrading capacity relative to yellow cattle, evidenced by significantly higher (P < 0.05) fiber digestibility, cellulase and xylanase activities, and increased volatile fatty acids production despite identical feed intake. Microbial community analysis revealed distinct composition in both the rumen and hindgut of gayal compared to yellow cattle, with notable enrichment of taxa specialized in lignocellulose degradation. Metatranscriptomic profiling further identified upregulation of key lignin-modification enzymes, particularly AA6, AA2, and AA3, primarily encoded by Prevotella, Cryptobacteroides, Limimorpha, and Ventricola. These enzymes are known to modify lignin structure to increase polysaccharide accessibility. These results demonstrate that gayal hosts a unique and metabolically active gastrointestinal microbiome capable of efficient lignocellulose deconstruction through a coordinated enzymatic cascade, especially effective in dismantling lignin barriers. This study provides novel insights into host-microbiome co-adaptation to fibrous feeds and highlights the potential of gayal-derived microbial consortia and enzymes for improving roughage utilization in ruminant agriculture.

Red clover extract (RCE) contains isoflavone phytoestrogens as its primary bioactive components, which have been shown to modulate rumen microbial metabolism and improve animal performance. This study investigated the effects of dietary RCE supplementation on growth performance, nutrient digestibility, antioxidant status, rumen fermentation, and rumen microbiota in fattening Hu sheep. Twenty-four Hu sheep, comprising 12 males (22.28 +/- 1.32 kg) and 12 females (20.80 +/- 1.74 kg), were randomly allocated to two dietary treatments: a control group (CON) fed a basal diet and an RCE group receiving the basal diet supplemented with 0.4% RCE (dry matter basis). The study consisted of a 2-week adaptation period and a 7-week trial period. Dietary RCE supplementation did not affect dry matter intake, average daily gain, or feed conversion efficiency (G:F) (P > 0.05). However, RCE supplementation significantly increased ruminal acetate concentration (P < 0.05) and tended to elevate total volatile fatty acid concentration (P = 0.091). The apparent digestibility of dry matter, organic matter, neutral detergent fiber, and acid detergent fiber was significantly enhanced in the RCE group compared to the CON group (P < 0.05). Dietary RCE supplementation significantly decreased plasma urea nitrogen and creatinine concentrations (P < 0.05), while significantly increasing superoxide dismutase and catalase activities (P < 0.05). Rumen microbiota analysis showed that RCE supplementation significantly decreased the relative abundances of Ruminococcus and Faecalibacterium while increasing those of Treponema, Selenomonas, Carboxylicivirga, and Bacteroides (P < 0.05). Additionally, significant treatment & times; sex interactions were detected for plasma glucose, total cholesterol, and glutathione peroxidase activity (P < 0.01), as well as for the relative abundances of Acetitomaculum, Marvinbryantia, Sedimentibacter, Erysipelothrix, Fretibacterium, and Anaerovorax (P < 0.05), indicating sex-specific responses to RCE in these variables. In conclusion, dietary supplementation with 0.4% RCE improved fiber digestibility, antioxidant capacity, and modulated rumen microbiota composition without affecting growth performance in fattening Hu sheep, while sex-related differences were limited to selected metabolic and microbial parameters.

Background and Aim: Enteric methane emission from dairy cows contributes substantially to greenhouse gas production and represents an inefficient loss of dietary energy. Phytosterols are plant-derived bioactive compounds with lipid-modulating and rumen fermentation-regulating properties; however, their effects on methane emission intensity and rumen microbial ecology in lactating dairy cows remain insufficiently explored. This study evaluated the effects of dietary phytosterols supplementation on lactation performance, nutrient digestibility, serum biochemical parameters, rumen fermentation characteristics, methane emission intensity, and rumen microbial composition in mid-lactation Holstein dairy cows. Materials and Methods: Thirty-four multiparous Holstein dairy cows with similar days in milk and milk yield were randomly assigned to either a control (CON) group or a phytosterols (PHY) group receiving 15 g/d of a commercial phytosterols product containing 5% active phytosterols. The experimental period lasted 50 days, including 7 days of adaptation and 43 days of data collection. Feed intake and milk yield were recorded daily. Milk composition, apparent nutrient digestibility, serum biochemical indices, rumen fermentation parameters, methane emission intensity, quantitative polymerase chain reaction, and 16S rRNA gene sequencing were analyzed. Methane and carbon dioxide emissions were measured using an automated head-chamber system. Results: Dietary phytosterols supplementation significantly improved milk yield, milk fat percentage, milk protein percentage, energy-corrected milk, and 3.5% fat-corrected milk compared with the CON group (p < 0.05). Apparent digestibility of organic matter, crude protein, neutral detergent fiber, and ether extract was also significantly enhanced. Serum glucose and blood urea nitrogen concentrations increased, whereas total cholesterol and low-density lipoprotein cholesterol concentrations decreased in the PHY group. Phytosterols supplementation significantly reduced methane emission intensity per kilogram of energy-corrected milk. Ruminal acetate proportion and acetate-to-propionate ratio decreased, whereas microbial crude protein and branched-chain volatile fatty acids increased. In addition, phytosterols altered rumen microbial composition by increasing the abundance of beneficial bacterial genera, including Succinivibrionaceae UCG-001 and Prevotella, while reducing methanogenic archaea, particularly Methanobacteriota and Methanimicrococcus. Conclusion: High-dose phytosterols supplementation improved lactation performance, enhanced nutrient utilization, modulated rumen microbial communities, and reduced methane emission intensity in mid-lactation dairy cows. These findings indicate that phytosterols may serve as a promising natural feed additive for improving dairy production efficiency while supporting methane mitigation strategies in sustainable dairy farming.

This study investigated how sorghum-based diets utilizing white (WSD) and red sorghum (RSD) with distinct polyphenol profiles influence nutrient digestibility, gut microbiota, and metabolic responses in growing pigs compared with corn-based diet (CON). An in vitro trial was first conducted using a simulated digestion model, followed by an in vivo trial involving 24 growing pigs (initial body weight: 20.84 +/- 0.68 kg), with 8 pigs per dietary treatment. In the in vitro trial, RSD showed the lowest nutrient digestibility during pepsin and pepsin-pancreatin hydrolysis (P < 0.05), produced less cumulative gas after 18 h fermentation relative to CON and WSD (P < 0.05). In the in vivo trial, RSD reduced apparent ileal and total-tract digestibility of nutrients compared with CON and WSD (P < 0.05). Additionally, RSD reduced villus height and villus-tocrypt ratio (VCR) in the jejunum (P <0.05) and tended to decrease ileal VCR (P = 0.09). Notably, RSD altered colonic microbiota composition, enriching genera including, Ligilactobacillus and Akkermansia, and functional taxa (LDA > 2, P < 0.05). Untargeted metabolomics revealed more differential metabolites in RSD, with KEGG enrichment highlighting phenylpropanoid biosynthesis and multiple amino acid-associated pathways. The RSD group exhibited elevated levels of polyphenol-derived metabolites (e.g., epicatechin) and reduced concentrations of several proteinderived metabolites (P < 0.05). Overall, red sorghum reduces nutrient digestibility but remodels colonic microbial composition and metabolic output; these shifts suggest potential prebiotic-like effects that merit further study to determine implications for intestinal health and production.

Antimicrobial resistance (AMR) poses a growing threat to global health, and increasing evidence reveals a substantial overlap in resistance genes between the gut microbiota of humans and food-producing animals, suggesting potential for cross-species transmission. Understanding the early-life development of the gut resistome is essential for designing effective AMR prevention strategies. This review synthesizes current knowledge on the age-dependent assembly of the gut resistome in both humans and food-producing animals, highlighting a consistent pattern of high antimicrobial resistance genes (ARGs) loads at birth followed by a gradual decline with age. We emphasize the critical role of diet in shaping resistome dynamics, formula feeding and high-fat, high-protein diets are associated with increased ARGs burden, whereas breastfeeding and diverse, fiber-rich diets are linked to reduced ARG prevalence. Furthermore, we discuss the potential of probiotics and prebiotics to mitigate gut AMR, while underscoring the importance of assessing resistance gene transfer risk in functional food development. Finally, we outline key knowledge gaps and propose future research directions within the framework of “One Health”. This review provides a comprehensive foundation for policy and intervention strategies to control gut-derived AMR and protect public health.

Schizochytrium sp. powder (SP) is widely used to enrich docosahexaenoic acid in bovine milk; however, its effects on milk quality, volatile flavor profiles, and underlying regulatory mechanisms remain poorly understood. Using an integrated multi-omics approach, this study investigated how uncoated SP (USP) and coated SP (CSP) supplementation differentially influence milk antioxidant capacity, lipid composition, and volatile flavor compounds. CSP improved milk antioxidant capacity and omega-3 polyunsaturated fatty acids levels, reduced saltiness, umami, and richness perception, and induced extensive lipidome remodeling mainly involving triacylglycerols, glycosphingolipids, and betaine lipids, whereas USP caused relatively minor lipidomic changes. Regarding volatile profiles, both treatments elevated 2,3-butanedione and 2,3-pentanedione; CSP further enriched nonanal, 2-undecanone, and 2-octanone (fatty/fruity notes), while USP increased aromatic volatiles alongside the fishy-odor marker 1-octen-3-ol. Correlation analysis identified C4:0, PI(16:0_18:3;O), and TG(18:0_18:0_22:3) as potential lipid precursors of volatile flavor formation.

Enhancing milk nutritional quality through increased ω-3 polyunsaturated fatty acid (PUFA) content and a reduced ω-6/ω-3 PUFA ratio represents a significant opportunity for improving dairy products. While ruminal biohydrogenation substantially influences milk fatty acid (FA) composition, the specific microbial mechanisms regulating the milk fat ω-6/ω-3 PUFA ratio remain poorly characterized. This study aimed to identify key microbial taxa and metabolic pathways controlling this nutritionally relevant parameter, thereby establishing a foundation for targeted microbiome interventions to optimize milk FA profiles. Analysis of 95 Holstein cows revealed that rumen bacterial community composition explained 41.0

Ruminant epithelia preferentially catabolize butyrate to fuel ketogenesis, yet the mechanism by which the rumen epithelium establishes this preference remains unclear. Here, we identify ACSF2 as a mitochondrial acyl-CoA synthetase (ACS) that catalyzes the activation of butyrate to butyryl-CoA, thereby enabling ketogenesis and accounting for this bias. We found that ACSF2 is markedly enriched in the forestomachs across ovine organs, with expression far exceeding other ACSs in the rumen epithelium, and it is rising during postnatal establishment of fermentative function. Single-cell transcriptomics and immunostaining localize ACSF2 to the mitochondria-rich layers, where it is co-expressed in mitochondria with ketogenesis genes, notably the rate-limiting enzyme HMGCS2. Further gain- and loss-of-function experiments show that ACSF2 activates butyrate to butyryl-CoA, enhances butyrate-supported growth, and is required for efficient butyrate consumption and cell fitness under butyrate-dependent conditions. These findings define ACSF2 as a key mitochondrial gatekeeper for butyrate utilization and ketogenesis in the rumen epithelium, providing a molecular mechanism for butyrate-biased energy metabolism during rumen maturation.

Rosmarinic acid (RA) is a phenolic compound with antioxidant and intestinal barrier-protective effects, yet the underlying molecular mechanisms remain incompletely understood. This study aimed to investigate the antioxidant mechanisms of RA and dihydrorosmarinic acid (DHRA) in chickens. In vivo experiments revealed that administration of 100 mg/kg RA for 21 days activated the AHR and Nrf2 pathways, and upregulated antioxidant enzymes and tight junction proteins in the jejunum of chickens (P < 0.05). For in vitro experiments, chicken intestinal epithelial cells (cIECs) were cultured with 100 μM RA or DHRA, with or without Nrf2 and AHR inhibitors. RA and DHRA significantly activated AHR and Nrf2 pathways (P < 0.05), while the Nrf2 inhibitor blocked Nrf2 activation (P < 0.05), and the AHR inhibitor abrogated the activation of both AHR and Nrf2 pathways (P < 0.05). RA and DHRA significantly upregulated the expression of intestinal tight junction proteins (P < 0.05), an effect that was reversed by both AHR and Nrf2 inhibitors (P < 0.05). The activation of AHR and Nrf2 by RA and DHRA was associated with their oxidative products, such as RA-quinone, DHRA-quinone, and H₂O₂ in the culture medium. Both GSH and catalase decreased the concentration of H₂O₂ and quinones in cell culture medium and attenuated the activation of Nrf2 and AHR pathways in cIECs. In summary, the oxidation products of RA and DHRA act as dual activators of the AHR and Nrf2 pathways, enhancing antioxidant defenses and barrier function in cIECs.

Ruminant epithelia preferentially catabolize butyrate to fuel metabolism, yet the mechanism by which the rumen epithelium establishes this preference remains unclear. Here, we identify ACSF2 as a mitochondrial acyl‑CoA synthetase that catalyzes the activation of butyrate to butyryl‑CoA, thereby enabling rumen butyrate preference. We found that ACSF2 is markedly enriched in the forestomachs across ovine organs, with expression far exceeding other acyl‑CoA synthetases in the rumen epithelium, and it is rising during postnatal establishment of fermentative function. Single‑cell transcriptomics and immunostaining localize ACSF2 to the mitochondria‑rich layers, where it is co‑expressed in mitochondria with ketogenesis genes, notably the rate‑limiting enzyme HMGCS2. Further gain‑ and loss‑of‑function experiments show that ACSF2 activates butyrate to butyryl‑CoA, enhances butyrate‑supported growth, and is required for efficient butyrate consumption, cell fitness, and ketogenesis under butyrate‑dependent conditions. These findings define ACSF2 as a key mitochondrial gatekeeper for butyrate utilization in the rumen epithelium, providing a molecular mechanism for butyrate‑biased energy metabolism during rumen maturation.

Background : Methane emissions from livestock are one of the most significant contributing factors to global warming. Although it is now understood that methanogenic archaea in the rumen of ruminants utilize hydrogen, formate, and methyl compounds produced by bacteria, fungi, and ciliate protozoa to generate methane, their relative contributions remain unclear. Results : This study aims to identify priority microbial targets for methane mitigation strategies in ruminant systems, thereby supporting more sustainable and low-carbon animal production. After retrieval and screening in the Web of Science database and Google Scholar database, 210 studies containing the required data were obtained. The obtained data were constrained using Z-score and then reanalyzed. The results showed that there were significant positive correlations between methane production and bacteria and fungi ( P < 0.05). According to the correlation analysis between methane production and rumen microbiota, four regression models were constructed. Equation iv demonstrated a favorable fit (Adj. R 2 = 0.74, P < 0.01). Significant differences in the copy numbers of bacterial 16S rRNA, fungal 18S rRNA, and protozoal 18S rRNA were observed between high and low methane production conditions. When the dietary NDF content increased, the relative contribution of bacteria decreased, while those of fungi and protozoa increased relatively. However, overall, the contribution to methane production followed the order: bacteria > fungi > protozoa, and the driving factor for methane production contribution was primarily determined by microbial abundance. Conclusion : These findings facilitate the prioritization of rumen bacteria as microbial targets for methane mitigation, thereby enhancing feed energy utilization efficiency and reducing agricultural greenhouse gas emissions.

The potential role of human milk oligosaccharides (HMOs) in regulating intestinal epithelial cell proliferation and differentiation in mammals has attracted considerable attention. Fucosylated HMO can provide fucosylated glycans to the intestine of newborns, an outcome similar to genetic overexpression of fucosyltransferase. However, how HMOs, such as 2'-fucosyllactose (2'-FL), may impact small intestinal fucosylation to regulate intestinal stem cell (ISC)-mediated intestinal epithelial development remains unknown. In the present study, we employed a model of small intestinal fucosylation inhibition in pups by orally administering a fucosylation inhibitor, 2 F-peracetyl-fucose (2 F-Fuc) from postnatal day 4 (PN4) to postnatal day 21 (PN21). Inhibition of jejunal fucosylation of pups during the breastfeeding stage significantly promoted ISC differentiation toward the secretory lineage by inhibiting Notch signaling pathways. Furthermore, fucosylation inhibition caused ISC differentiation deregulation in the jejunum that persisted until PN day 42, even though 2 F-Fuc was no longer administered after weaning at PN21. Supplementation of 1,2-fucosylated oligosaccharide by 2'-FL after weaning ameliorated the long-term effects of fucosylation inhibition in rats on ISC differentiation and changes in mucus-degrading microbiota. These results demonstrate that intestinal fucosylation in pups plays an important role in maintaining the balance between ISC proliferation and differentiation. Our study offers new insights into the interactions between intestinal fucosylation and ISC function. It also establishes a research foundation for using 2'-FL as a donor for α1,2-fucosylated glycans, enabling targeted modifications in intestinal fucosylation to enhance gut health, particularly small intestine health.NEW & NOTEWORTHY Inhibition of jejunal fucosylation of pups during the breastfeeding stage significantly promoted ISC differentiation toward the secretory lineage, and this effect persisted until PN day 42 via inhibiting Notch signaling pathways. In addition, supplementation with 2'-FL after weaning ameliorated the long-term effects of fucosylation inhibition in rats on ISC differentiation and the dysregulation of mucus-degrading microbiota.

Nitrogen metabolism in pigs is closely associated with gut microbiota, but the differential effects of dietary fiber sources on nitrogen metabolism and gut microbiota remain unclear. This study investigated the effects of pectin, β-glucan, arabinoxylan, and cellulose on nitrogen metabolism, fecal microbiota, and metabolic characteristics in growing-finishing pigs. Sixty healthy castrated male pigs (initial body weight 44.32 ± 0.29 kg) were randomly assigned to five groups (n = 12 per group) for a 28-day trial, receiving a basal diet supplemented with 7.0% wheat bran (CON group), 3.5% pectin (PC group), 3.5% β-glucan (BG group), 3.5% arabinoxylan (AX group), or 3.5% cellulose (CL group). Compared with the CL group, the PC and AX groups had significantly higher blood ammonia (BLA) and urea nitrogen (BUN) levels (P < 0.05). Compared with the CON and CL groups, the PC, BG, and AX groups had significantly higher fecal nitrogen (N) content (P < 0.05). Different fiber sources significantly altered the composition and functional potential of the fecal microbiota, leading to changes in key genera including Limosilactobacillus, Lactobacillus, and Escherichia-Shigella (P < 0.05). Compared with the CL group, the PC group exhibited higher microbial nitrogen cycling-related functions, fecal microbial protein (MCP), and ammonia nitrogen concentrations (P < 0.05). In conclusion, different dietary fiber sources exert differential effects on nitrogen metabolism and fecal microbiota in growing-finishing pigs. Pectin and cellulose induce distinct changes in nitrogen excretion through modulation of the gut microbiota, offering insights into using plant fiber byproducts to reduce nitrogen pollution in swine production.

This study aimed to investigate the effects of dietary supplementation with a triple-strain Bacillus-based probiotic (BP) on growth performance, intestinal barrier function, and gut microbial composition in weaned pigs. A total of 160 piglets (initial body weight, 8.0 ± 0.25 kg; 28 d of age) were randomly assigned to four treatments: a basal diet (CON) or the basal diet supplemented with 100 mg/kg (BP100, 4 × 107 colony-forming unit [CFU]/kg feed), 200 mg/kg (BP200, 8 × 107 CFU/kg feed), and 400 mg/kg (BP400, 1.6 × 108 CFU/kg feed) a triple-strain Bacillus probiotic containing Bacillus subtilis PB6, Bacillus subtilis FXA, and Bacillus licheniformis G3. Each treatment included 8 replicate pens with 5 pigs per pen. Over the 35-d feeding period (Phase 1: d 0-14; Phase 2: d 15-35), increasing dietary BP supplementation linearly improved average daily gain and feed efficiency (P < 0.05) and reduced the diarrhea index by up to 49.3% relative to the CON (P < 0.05), whereas average daily feed intake was unaffected. Apparent nutrient digestibility increased, whereas organic matter, gross energy, and nitrogen excretion decreased linearly with increasing BP supplementation (P < 0.10). Dietary BP supplementation linearly increased villus height, villus height-to-crypt depth ratio, goblet cell numbers, and tight junction protein expression, while reducing crypt depth in the small intestine (P < 0.05). In addition, increasing dietary BP supplementation linearly increased colonic Lactobacillus abundance and the concentrations of acetic acid, propionic acid, butyric acid, and total short-chain fatty acids, with the greatest values observed in the BP400 (P < 0.05). In conclusion, dietary BP supplementation improved growth performance in weaned pigs by enhancing intestinal morphological development and barrier integrity, increasing nutrient digestibility, and modulating the composition and metabolic activity of the gut microbiota. These findings highlight the potential of BP as an effective functional feed additive to mitigate weaning stress through improvements in intestinal barrier function and modulation of gut microbiota.