Effects of biochanin A on lactational performance, nitrogen metabolism, and blood metabolites in dairy cows.

Optimizing nitrogen utilization efficiency and mitigating nitrogen losses in cows plays a pivotal role in fostering economic sustainability within contemporary agricultural systems. Biochanin A (BCA), a natural component in red clover, has the potential to improve nitrogen metabolism in dairy cows. The primary objective of this study was to probe the impact of biochanin A supplementation on lactational performance, nitrogen metabolism, and blood metabolites in dairy cows. A complete randomized block design experiment was conducted over 28 d, involving 36 multiparous Holstein cows (comparable milk yield = 37.1 ± 2.90 kg, BW = 642 ± 70.0 kg, days in milk = 92 ± 8.0 d, and parity = 2.4 ± 0.50), which were allocated to three treatment groups: the Control group (with 0 g/d BCA), the Low group (with 10 g/d per cow BCA), and the High group (with 40 g/d per cow BCA). Biochanin A supplementation improved the lactational performance of cows by increasing milk yield by 6.3% (P = 0.007) and feed efficiency by 12.7% (P = 0.009). Total intestinal apparent digestibility was unaffected by BCA supplementation (P > 0.05), but microbial nitrogen was increased by 30.0% (P = 0.002) for promoting nitrogen utilization efficiency by 20.7% (P = 0.004). Milk competent yields (protein, lactose, and non-fat milk solid) were increased with increasing BCA supplementation (P < 0.05). Urea nitrogen levels in plasma and milk were both decreased by BCA supplementation (P < 0.05). Blood routine parameters and plasma biochemical parameters both received no effect by BCA supplementation (P > 0.05). BCA did not affect body health of dairy cows. Additionally, none of the plasma endocrine hormones were affected (P > 0.05). A total of 95 significantly different metabolites were screened from the plasma metabolites of cows in the BCA-added and non-added groups. After performing an enrichment analysis of the metabolic pathways associated with the different metabolites, six specific pathways were identified: bile acid biosynthesis, aspartate metabolism, pyrimidine metabolism, arginine and proline metabolism, the urea cycle, and ammonia recycling. The inclusion of BCA is suggested to enhance milk yield and modulate nitrogen metabolism by influencing relevant metabolites within the metabolic pathways.

Succession of rumen microbiota and metabolites across different reproductive periods in different sheep breeds and their impact on the growth and development of offspring lambs.

BACKGROUND: The microbiota and metabolites in the gastrointestinal tracts of female animals at different reproductive periods are very important to the growth, development, and health of themselves and their offspring. However, the changes in the gastrointestinal microbiota and metabolites throughout reproductive period of different sheep breeds and their effects on the growth and development of offspring lambs are still unclear. Hence, this study presents an assessment of the reproductive hormone levels, immune levels, rumen microbiota, and metabolites in Hu sheep and Suffolk ewes at different reproductive periods and their effects on the growth and development of offspring lambs. RESULTS: Hu sheep and Suffolk during non-pregnancy, pregnancy, and lactation were used as the research objects to determine reproductive and immune indexes of ewes at different periods, analyze rumen microbiome and metabolome, and track the growth performance and development of offspring lambs. The results showed that the reproductive hormone and immune levels of Hu sheep and Suffolk underwent adaptive changes across different reproductive periods. Compared with non-pregnancy, the microbial energy metabolism and lipid metabolism function decreased during Hu sheep pregnancy, and energy metabolism function decreased during lactation. In Suffolk, energy metabolism, glycan biosynthesis, and metabolism function were enhanced during pregnancy, and the metabolism of cofactors and vitamins was enhanced during lactation. Prevotella increased in Suffolk during pregnancy and lactation (P < 0.05) and was positively correlated with the birth weight and body size of the lambs (P < 0.05). Moreover, the abundances of Butyrivibrio and Rikenellaceae_RC9_gut_group during pregnancy were positively correlated with the intestinal immunity of the offspring lambs (P < 0.05), thereby regulating the intestinal immunity level of the lambs. Metabolomic analysis revealed that the protein digestion, absorption, and amino acid metabolism of Hu sheep were enhanced during pregnancy, which provided amino acids for the growth and development of pregnant ewes and fetuses and was significantly correlated with the birth weight, body size, and intestinal immunity of lambs (P < 0.05). Simultaneously, there was an increase in acetate and propionate during the pregnancy and lactation period of both Hu sheep and Suffolk, providing energy for ewes during reproductive period. Moreover, the microbiota during the lactation period was significantly correlated with the milk quality and lambs daily gain (P < 0.05). CONCLUSIONS: This study revealed the characteristic succession changes in the rumen microbiota and its metabolites at different reproductive periods in sheep breeds and their regulation of reproductive hormone and immune levels and identified their potential effects on the growth and development of offspring lambs. The findings provide valuable insights into the health and feeding management of different sheep breeds during the reproductive stage. Video Abstract.

Microbial-Metabolomic Exploration of Tea Polyphenols in the Regulation of Serum Indicators, Liver Metabolism, Rumen Microorganisms, and Metabolism in Hu Sheep.

This study investigated the impact of tea polyphenols on serum indices, rumen microorganisms, rumen metabolism, and liver metabolism in Hu sheep. Sixty healthy lambs, aged three months and with similar average weights, were chosen and randomly assigned to control (CON), TP400, TP800, and TP1200 groups, each consisting of fifteen lambs. The control group received a basal diet, while the experimental groups were provided with basal diet supplemented with 400 mg/kg, 800 mg/kg, and 1200 mg/kg of tea polyphenols, respectively. Compared with the CON group, the addition of tea polyphenols to the diet significantly increased serum IgA, GSH-Px, and TSOD. In addition, tea polyphenols were able to increase rumen pH but had no significant effect on the rumen NH3-N, VFA molar content, and the microbial top 10 phylum and genus levels. Moreover, Firmicutes predominated in the network map of the top 80 abundant microorganisms at the genus level, identifying 13 biomarkers at the genus level. In addition, strong correlations were observed between liver and rumen metabolites, particularly between rumen succinic acid and liver alanyl-serine and methylmalonic acid. Furthermore, tea polyphenol additions changed the enrichment of liver and rumen metabolites in the top five KEGG metabolic pathways, but 400-1200 mg/kg additions had no negative impact on the liver and rumen. In summary, TP significantly influences rumen and liver metabolites in Hu sheep, enhancing lamb immunity and antioxidant capacity, with 400 mg/kg being the most effective dosage.

Metagenomics-Metabolomics Exploration of Three-Way-Crossbreeding Effects on Rumen to Provide Basis for Crossbreeding Improvement of Sheep Microbiome and Metabolome of Sheep.

The objective of this experiment was to explore the effects of three-way hybridization on rumen microbes and metabolites in sheep using rumen metagenomics and metabolomics. Healthy Hu and CAH (Charolais × Australian White × Hu) male lambs of similar birth weight and age were selected for short-term fattening after intensive weaning to collect rumen fluid for sequencing. Rumen metagenomics diversity showed that Hu and CAH sheep were significantly segregated at the species, KEGG-enzyme, and CAZy-family levels. Moreover, the CAH significantly increased the ACE and Chao1 indices. Further, correlation analysis of the abundance of the top 80 revealed that the microorganisms were interrelated at the species, KEGG-enzyme, and CAZy-family levels. Overall, the microbiome significantly affected metabolites of the top five pathways, with the strongest correlation found with succinic acid. Meanwhile, species-level microbial markers significantly affected rumen differential metabolites. In addition, rumen microbial markers in Hu sheep were overall positively correlated with down-regulated metabolites and negatively correlated with up-regulated metabolites. In contrast, rumen microbial markers in CAH lambs were overall negatively correlated with down-regulated metabolites and positively correlated with up-regulated metabolites. These results suggest that three-way crossbreeding significantly affects rumen microbial community and metabolite composition, and that significant interactions exist between rumen microbes and metabolites.

Ssc-miR-101-3p inhibits hypoxia-induced apoptosis and inflammatory response in alveolar type-II epithelial cells of Tibetan pigs via targeting FOXO3.

Tibetan pigs are a unique swine strain adapted to the hypoxic environment of the plateau regions in China. The unique mechanisms underlying the adaption by Tibetan pigs, however, are still elusive. Only few studies have investigated hypoxia-associated molecular regulation in the lung tissues of animals living in the plateau region of China. Our previous study reported that ssc-miR-101-3p expression significantly differed in the lung tissues of Tibetan pigs at different altitudes, suggesting that ssc-miR-101-3p plays an important role in the adaptation of Tibetan pigs to high altitude. To understand the underlying molecular mechanism, in this study, the target genes of ssc-miR-101-3p and their functions were analyzed via various methods including qRT-PCR and GO and KEGG pathway enrichment analyses. The action of ssc-miR-101-3p was investigated by culturing alveolar type-II epithelial cells (ATII) of Tibetan pigs under hypoxic conditions and transfecting ATII cells with vectors overexpressing or inhibiting ssc-miR-101-3p. Overexpression of ssc-miR-101-3p significantly increased the proliferation of ATII cells and decreased the expression of inflammatory and apoptotic factors. The target genes of ssc-miR-101-3p were significantly enriched in FOXO and PI3K-AKT signaling pathways required to mitigate lung injury. Further, FOXO3 was identified as a direct target of ssc-miR-101-3p. Interestingly, ssc-miR-101-3p overexpression reversed the damaging effect of FOXO3 in the ATII cells. In conclusion, ssc-miR-101-3p targeting FOXO3 could inhibit hypoxia-induced apoptosis and inflammatory response in ATII cells of Tibetan pigs. These results provided new insights into the molecular mechanisms elucidating the response of lung tissues of Tibetan pigs to hypoxic stress.

Genetic characterization of Tibetan pigs adapted to high altitude under natural selection based on a large whole-genome dataset.

The Qinghai-Tibet Plateau is a valuable genetic resource pool, and the high-altitude adaptation of Tibetan pigs is a classic example of the adaptive evolution of domestic animals. Here, we report the presence of Darwinian positive selection signatures in Tibetan pigs (TBPs) using 348 genome-wide datasets (127 whole-genome sequence datasets (WGSs) and 221 whole-genome single-nucleotide polymorphism (SNP) chip datasets). We characterized a high-confidence list of genetic signatures related response to high-altitude adaptation in Tibetan pigs, including 4,598 candidate SNPs and 131 candidate genes. Functional annotation and enrichment analysis revealed that 131 candidate genes are related to multiple systems and organs in Tibetan pigs. Notably, eight of the top ten novel genes, RALB, NBEA, LIFR, CLEC17A, PRIM2, CDH7, GK5 and FAM83B, were highlighted and associated with improved adaptive heart functions in Tibetan pigs high-altitude adaptation. Moreover, genome-wide association analysis revealed that 29 SNPs were involved in 13 candidate genes associated with at least one adaptive trait. In particular, among the top ten candidate genes, CLEC17A is related to a reduction in hemoglobin (HGB) in Tibetan pigs. Overall, our study provides a robust SNP/gene list involving genetic adaptation for Tibetan pig high-altitude adaptation, and it will be a valuable resource for future Tibetan pig studies.

Effects of Steviol Glycosides on Growth Performance, Ruminal Fermentation and Microbial Diversity of Hu Sheep.

The experiment was conducted to investigate the potential effects of steviol glycosides on growth performance, rumen fermentation processes, and microbial diversity in Hu sheep. A single-factor design was used for the trial. Twenty healthy weaned Hu lambs, possessing comparable body weights averaging 18.31 ± 1.24 kg, were randomly allocated into two distinct groups: the control group (CON) and the experimental group (STE), with each comprising 10 lambs. The CON was fed the basal diet, and the STE was supplemented with 0.07% steviol glycosides based on the basal diet. During the experimental period, variations in body weight and feed intake were closely monitored and recorded. After feeding for 90 d, blood was collected to determine blood biochemical indices, and rumen fluid samples were gathered for an in-depth analysis of rumen fermentation parameters and microbial diversity. The outcomes revealed no statistically significant differences in growth performance or serum biochemical indices between the two groups (p > 0.05). Rumen pH in STE and CON was within the normal range. The rumen ammonia nitrogen (NH3-N) and acetic acid (AA) content of STE decreased significantly compared with CON (p < 0.05). No significant variations were observed in the levels of other volatile fatty acids (VFAs) between the two groups (p > 0.05). The rumen microbial OTUs count, as well as the Shannon, Simpson, Chao1, and Ace indices, were notably lower in the STE group compared to the CON group (p < 0.05). Additionally, at the phylum level, the relative abundance of Firmicutes, Bacteroidetes, and Proteobacteria collectively accounted for over 97% of the total phylum composition. In comparison to the CON group, the STE group exhibited an increase in the relative abundance of Proteobacteria (p < 0.05), accompanied by a significant reduction in the relative abundance of Patescibacteria and Desulfobacteria (p < 0.05). At the genus level, there was a notable increase in the relative abundance of Prevotella_7 and Succinivibrionaceae_UCG_001 in the STE group, whereas the relative abundance of Rikenellaceae_RC9_gut_group significantly decreased (p < 0.05). According to the correlation analysis between rumen microflora and VFAs, the relative abundance of Succinivibrionaceae_UCG_001 displayed a significant negative correlation with AA (p < 0.05), whereas Lactobacillus exhibited a notable positive correlation with isobutyric acid (IBA) (p < 0.05). In summary, steviol glycosides had no significant effect on the production performance and blood biochemical indexes of Hu sheep. Steviol glycosides can improve rumen fermentation parameters and rumen microflora structure of Hu sheep and have a certain effect on rumen microbial diversity and composition.

Rumen microbiota succession throughout the perinatal period and its association with postpartum production traits in dairy cows: A review.

The transition period for dairy cows usually refers to the 3 weeks pre-calving to the 3 weeks post-calving. During this period, dairy cows undergo metabolic and physiological adaptations because of their susceptibility to metabolic and infectious diseases. Poor feeding management under these circumstances may adversely affect the health and subsequent production performance of the cows. Owing to long-term adaptation and evolution, the rumen has become a unique ecosystem inhabited by a complex microbial community closely associated with its natural host. Dietary components are metabolized by the rumen microbiota, and volatile fatty acids and microbial protein products can be used as precursor substances for synthesizing meat and milk components. The successful transition of perinatal dairy cows includes changes in diet, physiology, and the rumen microbiota. Rumen microbial profiles have been confirmed to be heritable and repairable; however, adverse circumstances affect rumen microbial composition, host digestion and metabolism, as well as postpartum production traits of dairy cows for a certain period. Preliminary evidence indicates a close relationship between the rumen microbiota and animal performance. Therefore, changes in rumen microbes during the transition period and the intrinsic links between the microbiota and host postpartum phenotypic traits need to be better understood to optimize production performance in ruminants.

Unveiling the Influence of Copy Number Variations on Genetic Diversity and Adaptive Evolution in China's Native Pig Breeds via Whole-Genome Resequencing.

Copy number variations (CNVs) critically influence individual genetic diversity and phenotypic traits. In this study, we employed whole-genome resequencing technology to conduct an in-depth analysis of 50 pigs from five local swine populations [Rongchang pig (RC), Wuzhishan pig (WZS), Tibetan pig (T), Yorkshire (YL) and Landrace (LR)], aiming to assess their genetic potential and explore their prospects in the field of animal model applications. We identified a total of 96,466 CNVs, which were subsequently integrated into 7112 non-redundant CNVRs, encompassing 1.3% of the swine genome. Functional enrichment analysis of the genes within these CNVRs revealed significant associations with sensory perception, energy metabolism, and neural-related pathways. Further selective scan analyses of the local pig breeds RC, T, WZS, along with YL and LR, uncovered that for the RC variety, the genes PLA2G10 and ABCA8 were found to be closely related to fat metabolism and cardiovascular health. In the T breed, the genes NCF2 and CSGALNACT1 were associated with immune response and connective tissue characteristics. As for the WZS breed, the genes PLIN4 and CPB2 were primarily linked to fat storage and anti-inflammatory responses. In summary, this research underscores the pivotal role of CNVs in fostering the diversity and adaptive evolution of pig breeds while also offering valuable insights for further exploration of the advantageous genetic traits inherent to China's local pig breeds. This facilitates the creation of experimental animal models tailored to the specific characteristics of these breeds, contributing to the advancement of livestock and biomedical research.

Relationship between Rumen Microbial Differences and Phenotype Traits among Hu Sheep and Crossbred Offspring Sheep.

This experiment was conducted to investigate the effect of three-way hybrid sheep and Hu sheep on serum indicators, rumen fermentation, rumen enzyme activity, and microorganisms in sheep. Healthy and similar birth weights from three groups (Hu, n = 11; Charolais × Australian White × Hu, CAH, n = 11; Charolais × Dorper × Hu, CDH, n = 11) were selected to be fed by the ewes until 45 days of age. Subsequently, they were weaned intensively and underwent short-term fattening for 3 months along with selected male lambs fed intensively. During this period, they were fed and watered ad libitum. Blood and rumen fluid were collected and analyzed for serum indicators and rumen fluid microorganisms, enzyme activity, and VFA, respectively, at the end of the fattening period. Compared with Hu lamb, the offspring of the three-way hybrid lamb showed significant improvements in body weight, serum lactate dehydrogenase, and creatinine content. However, there was no significant effect on serum immunity and antioxidant indices. In addition, the rumen fluid volatile fatty acid (VFA) molar concentration and microcrystalline cellulose and lipase content were significantly lower in the three-way hybrid lamb compared to Hu lamb, but ß-glucosidase, amylase, pepsin, and VFA molar ratio were not significantly affected. Subsequently, 16S rRNA sequencing diversity analysis revealed that three-way hybrid lamb significantly increased rumen microbial ACE and Chao1 indices compared to Hu lamb. Meanwhile, the abundance of Verrucomicrobiota and Synergistota significantly increased at the phylum level. Correlation analysis showed that Prevotella had the highest proportion, while Rikenellaceae_RC9_gut_group correlated most closely with others genus. The microbial communities isovaleric acid molar concentration and proportion were strongly correlated. In addition, there were significant differences in correlations between microbial communities and isobutyric acid, butyric acid and valeric acid content, and their molar proportion, but they were not significantly correlated with digestive enzymes. From the functional enrichment analysis, it was found that hybrid progeny were mainly enriched in the pyruvate metabolism, microbial metabolism in diverse environments, carbon metabolism, and quorum sensing pathways. In contrast, the Hu sheep were primarily enriched in the cysteine and methionine, amino sugar and nucleotide sugar, and biosynthesis of secondary metabolite pathways. These results suggest that hybridization can play a role in regulating organismal metabolism and improve animal production performance by influencing the structure and characteristics of microbial communities.

Effect of commercial fibrolytic enzymes application to normal- and slightly lower energy diets on lactational performance, digestibility and plasma nutrients in high-producing dairy cows.

The inclusion of fibrolytic enzymes in the diet is believed to have positive effects on animal production. Hence, the objective of this study was to investigate the impact of supplementing diets with a commercial fibrolytic enzyme preparation (Vistamax; mixture of xylanase and cellulase) derived from Trichoderma reesei on lactational performance, digestibility, and plasma nutrient levels in high-producing dairy cows. Two dietary energy levels were considered: a normal energy diet (metabolizable energy = 2.68 Mcal/kg) and a slightly lower energy diet (metabolizable energy = 2.55 Mcal/kg). A total of 120 lactating Holstein cows (parity = 2; Days in Milk = 113 ± 23) were randomly assigned to four treatment groups using a 2 * 2 factorial arrangement. The dietary treatments consisted of: (1) normal energy diet without enzyme supplementation (NL); (2) normal energy diet with enzyme supplementation (NLE); (3) slightly lower energy diet without enzyme supplementation (SL); and (4) slightly lower energy diet with enzyme supplementation (SLE). The amount of enzyme added to the diets was determined based on previous in vitro studies and supplier recommendations. The enzyme and premix were mixed prior to the preparation of the total mixed ration, and the trial lasted for a duration of 42 days. The results indicated that the application of the fibrolytic enzyme did not have a significant effect on dry matter intake (DMI), but it did enhance the digestibility of dry matter (DM), neutral detergent fiber (NDF), potentially digestible NDF (pdNDF), organic matter (OM), milk production, milk urea nitrogen (MUN), and blood urea nitrogen (BUN). On the other hand, the slightly lower energy diet resulted in a decrease in DMI, milk production, milk protein yield, plasma free amino acids (FAA), and an increase in plasma B-hydroxybutyrate (BHBA). In conclusion, the inclusion of the fibrolytic enzyme in the diets of dairy cows led to improvements in the digestibility of DM, NDF, pdNDF, OM, milk production, and feed efficiency. Furthermore, the application of the enzyme to the slightly lower energy diet resulted in milk production levels comparable to those observed in cows fed the untreated normal energy diet.

Epiberberine: a potential rumen microbial urease inhibitor to reduce ammonia release screened by targeting UreG.

Rumen microbial urease inhibitors have been proposed for regulating nitrogen emission and improving nitrogen utilization efficiency in ruminant livestock industry. However, studies on plant-derived natural inhibitors of rumen microbial urease are limited. Urease accessory protein UreG, plays a crucial role in facilitating urease maturation, is a new target for design of urease inhibitor. The objective of this study was to select the potential effective inhibitor of rumen microbial urease from major protoberberine alkaloids in Rhizoma Coptidis by targeting UreG. Our results showed that berberine chloride and epiberberine exerted superior inhibition potential than other alkaloids based on GTPase activity study of UreG. Berberine chloride inhibition of UreG was mixed type, while inhibition kinetics type of epiberberine was uncompetitive. Furthermore, epiberberine was found to be more effective than berberine chloride in inhibiting the combination of nickel towards UreG and inducing changes in the second structure of UreG. Molecular modeling provided the rational structural basis for the higher inhibition potential of epiberberine, amino acid residues in G1 motif and G3 motif of UreG formed interactions with D ring of berberine chloride, while interacted with A ring and D ring of epiberberine. We further demonstrated the efficacy of epiberberine in the ruminal microbial fermentation with low ammonia release and urea degradation. In conclusion, our study clearly indicates that epiberberine is a promising candidate as a safe and effective inhibitor of rumen microbial urease and provides an optimal strategy and suitable feed additive for regulating nitrogen excretion in ruminants in the future. KEY POINTS: • Epiberberine is the most effective inhibitor of rumen urease from Rhizoma Coptidis. • Urease accessory protein UreG is an effective target for design of urease inhibitor. • Epiberberine may be used as natural feed additive to reducing NH3 release in ruminants.

Relationships among bacterial cell size, diversity, and taxonomy in rumen.

Introduction: The rumen microbial community plays a crucial role in the digestion and metabolic processes of ruminants. Although sequencing-based studies have helped reveal the diversity and functions of bacteria in the rumen, their physiological and biochemical characteristics, as well as their dynamic regulation along the digestion process in the rumen, remain poorly understood. Addressing these gaps requires pure culture studies to demystify the intricate mechanisms at play. Bacteria exhibit morphological differentiation associated with different species. Based on the difference in size or shape of microorganisms, size fractionation by filters with various pore sizes can be used to separate them. Methods: In this study, we used polyvinylidene difluoride filters with pore sizes of 300, 120, 80, 40, 20, 8, 6, 2.1, and 0.6 µm. Bacterial suspensions were successively passed through these filters for the analysis of microbial population distribution using 16S rRNA gene sequences. Results: We found that bacteria from the different pore sizes were clustered into four branches (> 120 µm, 40-120 µm, 6-20 µm, 20-40 µm, and < 0.6 µm), indicating that size fractionation had effects on enriching specific groups but could not effectively separate dominant groups by cell size alone. The species of unclassified Flavobacterium, unclassified Chryseobacterium, unclassified Delftia, Methylotenera mobilis, unclassified Caulobacteraceae, unclassified Oligella, unclassified Sphingomonas, unclassified Stenotrophomonas, unclassified Shuttleworthia, unclassified Sutterella, unclassified Alphaproteobacteria, and unclassified SR1 can be efficiently enriched or separated by size fractionation. Discussion: In this study, we investigated the diversity of sorted bacteria populations in the rumen for preliminary investigations of the relationship between the size and classification of rumen bacteria that have the potential to improve our ability to isolate and culture bacteria from the rumen in the future.

Effects of Isochlorogenic Acid on Ewes Rumen Fermentation, Microbial Diversity and Ewes Immunity of Different Physiological Stages.

The effects of isochlorogenic acid (ICGA) on ewes rumen environment, microbial diversity, and immunity at different physiological stages (estrus, pregnancy and lactation) were studied in this experiment. Twenty healthy female Hu lambs of 1.5 months with similar body weight (17.82 ± 0.98 kg) and body condition were selected and randomly divided into two groups: the control group (CON) and the ICGA group (ICGA). The lambs of CON were fed a basal diet, while the lambs of ICGA were supplemented with 0.1% ICGA based on the basal diet. Lambs rumen fermentation characteristics, microbial diversity and immunity at estrus, pregnancy, and lactation stages were determined and analyzed, respectively. The results showed that the rumen pH in CON increased first and then decreased as lambs grew (p < 0.05). However, it showed the opposite change in ICGA. The content of ammonia nitrogen (NH3-N) showed the highest at estrus stage in both groups, but it was significantly higher in ICGA than that in CON (p < 0.05). The Acetic acid/propionic acid (A/P) ratio at estrus stage and the volatile fatty acids (VFAs) at pregnancy stage in ICGA were significantly higher than those of the CON (p < 0.05). The 16S rDNA sequencing analysis showed that the Shannon, Chao 1 and ACE indexes of the ICGA were significantly higher than those of the CON both at estrus and lactation stages (p < 0.05), while they showed higher at the pregnancy stage in CON (p > 0.05). Principal component analysis (PCA) showed that there were significant differences in rumen microorganism structure between CON and ICGA at all physiological stages (p < 0.01). At the phylum level, compared with the CON, Firmicutes relative abundance of three physiological stages decreased (p > 0.05) while Bacteroidota increased (p > 0.05). The relative abundance of Synergistota at estrus stage and Patescibacteria at the lactation stage increased significantly too (p < 0.05). At the genus level, compared with the CON, the relative abundance of Prevotella at three stages showed the highest (p > 0.05), while the relative abundance of Succiniclasticum, unclassified_Selenomonadaceae and Rikenellaceae_RC9_gut_group showed different abundances at different physiological stages in ICGA. Compared with the CON, the lambs of the ICGA showed higher blood IgG, IgM, and TNF- α contents at three physiological stages and higher IL-6 contents at pregnancy stage (p < 0.05). Conclusion: Adding ICGA could regulate ewes rumen fermentation mode at different physiological stages by increasing rumen NH3-N at estrus, VFAs at pregnancy, and the ratio of A/P at lactation. It optimizes rumen microbial flora of different physiological stages by increasing Bacteroidota relative abundance while reducing Firmicutes relative abundance, maintaining rumen microbial homeostasis at pregnant stage, increasing the number of beneficial bacteria in later lactating and ewes blood immunoglobulins content at three physiological stages.

Enrichment of milk antioxidant activity by dietary supplementation of red clover isoflavone in cows and its improvement on mice intestinal health.

Red clover (Trifolium pratense) isoflavone was supplemented to dairy cows, and antioxidant capacity of milk was assessed. Treated cows increased the activities of antioxidant enzymes, reduced production of oxidation products, and enhanced the concentrations of vitamin E and vitamin C. Moreover, milk fatty acid profile was positive influenced by 8 g/kg red clover isoflavone, with changes in the lower saturated and higher unsaturated fatty acids. We further demonstrated the efficacy of antioxidant capacity of milk in mice, found that milk from cows feeding red clover isoflavone increased the expressions of antioxidant enzymes, and alleviated lipopolysaccharide (LPS)-stimulated tissue damage of duodenum and jejunum, which was related to upregulated metabolism pathways of carbohydrate, lipid, and amino acid, as well as downregulated inflammatory related pathways. Together, dietary supplementation of red clover isoflavone is an effective way to improve milk antioxidant capacity, providing a natural strategy for developing functional foods.

Ratio-fluorescence detection of tert-butylhydroquinone based on non-conjugated polymer dots and gold nanoclusters.

A novel ratiometric fluorescent probe based on non-conjugated polymer dots (NCPDs) and gold nanocluster (AuNCs) was constructed to determine tert-butylhydroquinone (TBHQ). The probe exhibited dual emission peaks at 480 nm and 630 nm under 370 nm excitation. The fluorescence of AuNCs was quenched by TBHQ due to strong electrostatic interactions, whereas the emission of NCPDs increased. The ratio of fluorescence intensity at 480 nm to 630 nm (F480 / F630) was monitored as analytical signal response. The probe have been utilized for the detection of TBHQ with good linear relationship in the range of 0.2 to 60 µg/mL. The limit of detection (LOD) and the limit of quantitation (LOQ) were 0.048 µg/mL and 0.159 µg/L, respectively. Three levels of spiked-in TBHQ concentrations were obtained with recovery rates from 80 % to 102 %. The present study provided an effective ratiometric fluorescence method for selective screening of TBHQ in food samples.

Multi-omics revealed rumen microbiota metabolism and host immune regulation in Tibetan sheep of different ages.

The rumen microbiota and metabolites play an important role in energy metabolism and immune regulation of the host. However, the regulatory mechanism of rumen microbiota and metabolite interactions with host on Tibetan sheep's plateau adaptability is still unclear. We analyzed the ruminal microbiome and metabolome, host transcriptome and serum metabolome characteristics of Tibetan sheep at different ages. Biomarkers Butyrivibrio, Lachnospiraceae_XPB1014_group, Prevotella, and Rikenellaceae_RC9_gut_group were found in 4 months, 1.5 years, 3.5 years, and 6 years Tibetan sheep, respectively. The rumen microbial metabolites were mainly enriched in galactose metabolism, unsaturated fatty acid biosynthesis and fatty acid degradation pathways, and had significant correlation with microbiota. These metabolites further interact with mRNA, and are co-enriched in arginine and proline metabolism, metabolism of xenobiotics by cytochrome P450, propanoate metabolism, starch and sucrose metabolism, gap junction pathway. Meanwhile, serum metabolites also have a similar function, such as chemical carcinogenesis - reactive oxygen species, limonene and pinene degradation, and cutin, suberine and wax biosynthesis, thus participating in the regulation of the body's immune and energy-related metabolic processes. This study systematically revealed that rumen microbiota, metabolites, mRNA and serum metabolites of Tibetan sheep were involved in the regulation of fermentation metabolic function and immune level of Tibetan sheep at different ages, which provided a new perspective for plateau adaptability research of Tibetan sheep at different ages.

Effect of red clover isoflavones on hormone, immune, inflammatory, and plasma biochemistry in lactating dairy cows.

This study was to conducted to investigate the effect of red clover isoflavones on the health indicated by immune status and blood biochemistry in dairy cows. Sixty-eight healthy Holstein lactating cows were randomly divided into four treatments (n = 17 per treatment) from 5 blocks according to milk yield using a randomized complete block design. No initial differences in parity (2.13 ± 1.21), days in milk (165 ± 21 d), and milk yield (33.93 ± 3.81 kg/d) between groups. Cows were fed the basal diet supplemented with 0, 2, 4, or 8 g/kg red clover extract (RCE) in diet (dry matter based). Feeding, refusal feed weights, and milk yield were recorded three consecutive days in weeks 0, 4, 8, and 12. Blood was collected from the tail vein of the cows on the last day of weeks 4, 8 and 12, 1 h after the morning feeding, and analyzed for hormones, immunoglobulins, inflammatory markers, and markers of liver and kidney activities. The dry matter intake was significantly decreased by 3.7% in the 8 g/kg group (P < 0.05). The fat-corrected milk yield was significantly higher in both of the 2 and 4 g/kg groups (P < 0.01). Plasma estradiol and prolactin showed a quadratic effect with increasing RCE levels, with the highest in the 4 g/kg group (P < 0.05). Plasma tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1ß levels decreased linearly with increasing dietary RCE levels. Plasma IL-18 levels showed a quadratic effect with increasing dietary RCE levels, with significantly lower levels in both of the 2 and 4 g/kg groups (P < 0.05). Plasma immunoglobulin A and D-lactic acid levels showed a quadratic effect with increasing dietary RCE levels, with significantly higher level in the 4 g/kg group (P < 0.05). The liver function and kidney activity makers were similar (P > 0.05). These results recommend the supplementation of RCE at a level from 2 to 4 g/kg DM.

Identification of the genetic characteristics of copy number variations in experimental specific pathogen-free ducks using whole-genome resequencing.

BACKGROUND: Specific pathogen-free ducks are a valuable laboratory resource for waterfowl disease research and poultry vaccine development. High throughput sequencing allows the systematic identification of structural variants in genomes. Copy number variation (CNV) can explain the variation of important duck genetic traits. Herein, the genome-wide CNVs of the three experimental duck species in China (Jinding ducks (JD), Shaoxing ducks (SX), and Fujian Shanma ducks (SM)) were characterized using resequencing to determine their genetic characteristics and selection signatures. RESULTS: We obtained 4,810 CNV regions (CNVRs) by merging 73,012 CNVs, covering 4.2% of the duck genome. Functional analysis revealed that the shared CNVR-harbored genes were significantly enriched for 31 gene ontology terms and 16 Kyoto Encyclopedia of Genes and Genomes pathways (e.g., olfactory transduction and immune system). Based on the genome-wide fixation index for each CNVR, growth (SPAG17 and PTH1R), disease resistance (CATHL3 and DMBT1), and thermoregulation (TRPC4 and SLIT3) candidate genes were identified in strongly selected signatures specific to JD, SM, and SX, respectively. CONCLUSIONS: In conclusion, we investigated the genome-wide distribution of experimental duck CNVs, providing a reference to establish the genetic basis of different phenotypic traits, thus contributing to the management of experimental animal genetic resources.

Characterizing the dynamics of the rumen microbiota, its metabolites, and blood metabolites across reproductive stages in Small-tailed Han sheep.

IMPORTANCE: Our study illustrates the succession of the rumen microbiota and its metabolites in Small-tailed Han sheep at different reproductive stages. Among them, Firmicutes and Prevotella, which are related to energy metabolism, increased in abundance during pregnancy, while Fibrobacter, a fiber-degrading bacterium, increased in abundance during lactation. At the same time, the microbial metabolic profile and serum metabolic profile characteristics of different reproductive stages were revealed, and some functional pathways and metabolites related to energy and immunity were found. This study provides a reference for the health management of ruminants during non-pregnancy, pregnancy, and lactation.