Ovum pick-up (OPU) technical variability limits the efficiency and consistency of bovine in vitro embryo production (IVP). Here, we established a sequential refinement framework for OPU by systematically evaluating key technical parameters across seven cumulative experiments in Holstein cows. An 18 G aspiration needle achieved significantly higher cumulus-oocyte complex (COCs) recovery than 20 G and 22 G needles (53.05% vs. 40.03-45.79%; p < 0.05). 90° improved recovery compared with non-vertical insertion (55.64% vs. 48.86%), while insertion to one-third to one-half follicle depth and bevel-down orientation further enhanced recovery efficiency (56.52% and 57.97%, respectively; p < 0.05). Polyvinyl chloride (PVC) tubing provided superior COCs recovery (63.29%) and morphological quality, with a tubing length of 1.2-1.5 m. Vacuum pressure of 50 mmHg resulted in higher COCs recovery (70.27%) and blastocyst rate (40.65%), significantly outperforming the previously used 70 mmHg setting. In contrast, excessive aspiration pressure (80 mmHg) was associated with mitochondrial dysfunction, disrupted cortical granule distribution, and increased apoptosis (p < 0.05), indicating impaired oocyte nuclear and cytoplasmic maturation under excessive mechanical stress. Notably, recovery efficiency progressively increased from 53.05% to 70.27% throughout the sequential optimization process throughout the sequential refinement process. Collectively, this study provides a field-based refinement approach for OPU procedures and demonstrates the association between aspiration parameters and bovine oocyte developmental competence under the tested conditions.

Supplementation with Saccharomyces cerevisiae fermentation-derived postbiotics (SCFP) in calves can improve calf growth and health. This study evaluated the effects of SCFP supplementation in the calf diet on growth, health, rumen fermentation, and rumen microbiota during the first 70 d of life in 2 independent calf experiments conducted under different maternal dietary contexts. The dams of the calves consisted of 400 pregnant Holstein dry cows that received either a basal far-off and close-up TMR without SCFP (n = 200) or the same diets supplemented with SCFP (n = 200) beginning at dry-off. From these dams, 144 female calves were enrolled and allocated within maternal dietary background, blocked by birth date, and randomly assigned to receive SCFP or not in the calf diet, resulting in 2 independent experiments (n = 36 calves per group, 4 groups in total): experiment 1, CC (dam: -SCFP, calf: -SCFP) and CT (dam: -SCFP, calf: +SCFP); and experiment 2, TC (dam: +SCFP, calf: -SCFP) and TT (dam: +SCFP, calf: +SCFP). Growth performance, serum metabolites and antioxidant and anti-inflammatory indicators, diarrhea incidence, and rumen fermentation and microbial composition were measured. In experiment 1, CT calves showed greater weaning weight and preweaning ADG, reduced diarrhea incidence, and altered rumen microbial community structure at 70 d of age, with enrichment of Erysipelotrichaceae_UCG-002 and Sharpea. In experiment 2, TT calves tended to have greater preweaning ADG, whereas weaning weight was unaffected. In both experiments, calves supplemented with SCFP consistently reduced preweaning diarrhea risk. At 70 d of age, rumen microbial diversity was not affected, although calves in the TT group exhibited greater total volatile fatty acid and ammonia-nitrogen concentrations. In conclusion, direct SCFP supplementation to calves during the preweaning period improved growth performance and health and modified rumen fermentation and microbial composition. Differences in calf responses between experiments highlight the need for future studies with replicated maternal treatments to clarify the role of maternal supplementation.

This study evaluated the effects of diets supplemented with different doses of potassium bicarbonate (KHCO3) on lactation performance, milk fatty acid composition, nutrient digestibility, ruminal fermentation, and serum parameters in peak-lactation dairy cows. Sixty multiparous Holstein cows (69.8 ± 21.2 d in milk; 41.2 ± 5.4 kg/d milk yield) were randomly assigned to diets containing 0% (CON), 0.75%, or 1.50% KHCO3 (dry matter basis) for an 8-wk treatment period. Data were analyzed using a mixed model with dietary treatment, time, and their interaction as fixed effects and cow as a random effect. Linear and quadratic responses were assessed using preplanned contrasts. Diets supplemented with KHCO3 had no effect on dry matter intake (DMI) and milk yield. However, 3.5% fat-corrected milk (FCM) tended to increase linearly, milk fat concentration increased linearly, and FCM/DMI exhibited linear and quadratic increases with increasing KHCO3 supplementation. Apparent digestibility of neutral detergent fiber and acid detergent fiber increased linearly, accompanied by a linear increase in the molar proportion of acetate with increasing KHCO3. In contrast, total volatile fatty acid concentration exhibited a quadratic response. Ruminal pH exhibited a quadratic increase with increasing KHCO3 addition, whereas ruminal ammonia-N and the molar proportions of butyrate, valerate, and isovalerate showed linear and quadratic decreases. Blood urea nitrogen concentration decreased linearly, with no effects on serum indicators of energy metabolism, liver function, or antioxidant capacity. Milk concentrations of C10:0, C12:0, and cis-9, trans-11 CLA increased linearly, whereas the trans-10 to trans-11 C18:1 ratio decreased linearly with increasing KHCO3 supplementation. Overall, supplementation with KHCO3 during peak lactation improved fiber digestibility and favored trans-11 biohydrogenation, thereby improving milk fat content and feed efficiency. Under the conditions of this study, a dietary inclusion of 0.75% KHCO3 was sufficient to support a more favorable ruminal fermentation profile and productive performance in dairy cows.

The bovine tongue is a complex and very important muscular and gustatory organ, yet a comprehensive understanding of its gustatory apparatus across diverse genetic resources remains elusive. In this study, we conducted a multidimensional analysis of the lingual morphology and taste bud (TB) distribution in 40 specimens from 12 representative bovine breeds and species across China, encompassing Bos taurus taurus (Taurine cattle), Bos taurus indicus (Zebu cattle), Bubalus bubalis (water buffalo), and Bos grunniens (domestic yak). Morphometric measurements and histological quantifications were integrated to evaluate the influence of species, sex, age, and geographical factors. Given the relatively limited sample size per breed, these findings are presented as exploratory research. Our results revealed that yak and water buffalo showed the most distinct morphological patterns of mechanical papillae compared to the other populations. Taurine and Zebu cattle displayed more similar lingual morphology traits. Although high phenotypic correlations were observed between lingual morphometric parameters and quantitative papillae indicators, factors such as age, altitude, and feeding methods showed minimal influence on lingual phenotypic variation within this cohort (p > 0.05). Furthermore, we constructed a topological atlas of TB distribution, revealing that TB distribution patterns are decoupled from macro-anatomical dimensions, highlighting the complexity of the bovine gustatory system. These findings provide a quantitative baseline for ruminant comparative anatomy and offer structural insights into the evolutionary adaptation and nutrient regulation mechanisms of diverse bovine species in varying environments.

BACKGROUND:Metabolic disorders in peripartum ruminants affect health and productivity, with gut microbiota playing a key role in host metabolism. Therefore, our study aimed to characterize the gut microbiota of peripartum dairy cows to better understand the relationship between metabolic phenotypes and the rumen and fecal microbiomes during the peripartum period. RESULTS:In a longitudinal study of 91 peripartum cows, we analyzed rumen and fecal microbiomes via 16S rRNA and metagenomic sequencing across six time points. By using enterotype classification, ecological model, and random forest analysis, we identified distinct deterministic succession patterns in the rumen and fecal microbiomes (rumen: rapid transition-transition-stable; hindgut: stable-transition-stable). Key microbes, such as Succiniclasticum and Bifidobacterium, were found to drive microbial succession by balancing stochastic and deterministic processes. Notably, we observed that changes in gut microbiota succession patterns significantly influenced metabolic phenotypes (e.g., serum non-esterified fatty acid, glucose, and insulin levels). Mediation analysis suggested that specific gut microbes (e.g., Prevotella sp900315525 in the rumen and Alistipes sp015059845 in the hindgut) and metabolic pathways (e.g., glucose-related pathway) were associated with host metabolic phenotypes. CONCLUSIONS:Overall, utilizing a large gut microbiome dataset and enterotype- and ecological model-based microbiome analyses, we comprehensively elucidated the succession and assembly of the gut microbiota in peripartum dairy cows. We further confirmed that changes in gut microbiota succession patterns were significantly related to the metabolic phenotypes of peripartum dairy cows. These findings provide valuable insights for developing health management strategies for peripartum ruminants.

Pasteurized colostrum has significantly contributed to improving the health and growth of newborn calves by reducing total bacterial count. However, previous research on animal responses to pasteurized colostrum has primarily focused on physiological functioning and production performance, especially during the preweaning period, with limited attention to any postweaning effects from the feeding of pasteurized colostrum at birth. We conducted a comprehensive investigation into the growth, health, blood immunity, and microbiota responses of dairy calves in these two groups from birth to 180 d of age. In this study, a total of 32 healthy female Holstein calves [mean birth weight = 39.8 ± 1.22 kg (mean ± standard deviation)] were selected and divided into two groups (n = 16; fed either pasteurized or unpasteurized colostrum at birth). The results demonstrated that calves fed pasteurized colostrum exhibited enhanced growth performance as indicated by higher body weight (BW) and average daily gain (ADG) compared to those fed unpasteurized colostrum (p < 0.05). Calves fed pasteurized colostrum displayed higher lymphocyte ratio (W-SCR) and platelet distribution width (PDW), along with lower neutrophil ratio (W-LCR) and neutrophil count (W-LCC) (p < 0.05). Additionally, substantial differences were identified in microbial richness and diversity between the pasteurized and unpasteurized colostrum-fed groups (p < 0.05). Distinct microbial communities were observed in the ruminal and fecal regions (p < 0.05), and we detected shared beneficial microbiota (Alloprevotella, Parabacteroides, and unidentified_Prevotellaceae) and metabolic functions (metabolism of energy, amino acids, and glycan) in both gut regions of the pasteurized group. Furthermore, our study revealed intricate and robust interactions among microbiota, volatile fatty acid (VFA) and blood indicators (|r| > 0.5 and p < 0.05). In conclusion, the findings in the present experiment suggest that the positive effects from d 0 pasteurized colostrum feeding may be seen up to d 180, including improved growth performance, health, and blood immunity, and these may be attributed to modifications in microbiota development induced by pasteurized colostrum.

IntroductionAlpha-linolenic acid (ALA), an essential polyunsaturated fatty acid, modulates gastrointestinal microbiota and host immunity, yet its regulatory mechanisms in dairy cows remain unclear.MethodsThis study investigated how dietary ALA influenced gut microbiota, metabolome, and immune function in lactating Holstein cows. Ten cows were randomly assigned to two groups (n = 5) receiving either a low-ALA (LALA, 5.02 ± 0.09% ALA of total fatty acids) or high-ALA diet (HALA, 32.04 ± 1.55% ALA of total fatty acids). Rumen fluid, feces, and blood samples were analyzed post-intervention.ResultsThe HALA group exhibited increased ruminal abundance of Eubacterium coprostanoligenes group and Ruminococcus (p < 0.05), alongside reduced proinflammatory metabolites including dodecanoic acid, myristic acid, and prostaglandin I2 in the rumen. Plasma leukotriene C4 levels were also decreased (p < 0.05). Metabolomic enrichment analysis revealed significant downregulation of arachidonic acid metabolism. Correlation analyses demonstrated that Eubacterium coprostanoligenes group negatively associated with suppressed prostaglandin I2 (rumen metabolite) and leukotriene C4 (plasma metabolite), but positively correlated with enriched fecal Clostridia UCG-014 and Ruminococcus.DiscussionThese findings indicate that high dietary ALA reshapes gastrointestinal microbiota and attenuates inflammatory responses by inhibiting microbial-metabolite-driven arachidonic acid metabolism, thereby enhancing immune regulation in dairy cows.

BACKGROUND:The purpose of this study was to investigate the effects of changes in rumen microbial adaptability on rumen fermentation and nutrient digestion in Horqin beef cattle during different seasons of grazing and supplementary feeding. Four healthy female Horqin beef cattle were selected as experimental animals for grazing and supplementary feeding. The feed, feces, and rumen fluid were collected in spring (March), summer (June), autumn (September), and winter (December) to determine nutrient digestibility, rumen fermentation parameters and microflora. The microflora was sequenced with 16S rRNA. RESULTS:The results revealed that the rumen volatile fatty acid, ammonia nitrogen and microbial protein concentrations in summer were significantly higher than those in the other seasons (P < 0.05). The digestibility of dry matter, organic matter, crude protein and ether extracts decreased significantly in autumn and winter (P < 0.05), especially in winter. Compared with autumn, the rumen bacterial diversity of beef cattle increased significantly in spring (P < 0.05). The results of the rumen microflora revealed that Prevotella was the most dominant bacteria in the rumen of beef cattle in summer, and Rikenellaceae_RC9_gut_group was the most dominant bacteria in the other seasons. The low quality of forage in spring, autumn and winter resulted in an increase in the relative abundance of Rikenellaceae_RC9_gut_group, NK4A214_group, and Christensenellaceae_R-7_group, etc., which are associated with fibre decomposition. In addition, nucleotide metabolism and glycan biosynthesis and metabolism increased significantly in bacteria in the summer (P < 0.05). CONCLUSIONS:The results revealed the dynamic adaptability of the rumen microbiota to seasonal and dietary changes and provide references for the grazing and supplementary feeding of Horqin beef cattle in different seasons.

This study aimed to investigate the effects of daily weight gain and feed intake of calves on first-lactation milk yield and composition using a meta-analysis. A total of 57 treatments from 18 studies were included in the study. Univariate and multivariate mixed models were constructed for calf ADG, liquid DMI (LDMI), starter DMI (SDMI), 305-d milk, milk fat, and protein yield data to gain insight into the effects of preweaning calf daily gain and feed intake on first-lactation performance. Univariate mixed models revealed ADG was significantly positively correlated with 305-d milk, milk fat, and protein yields during the first-lactation period. This indicates that ADG is a significant determinant of enhanced production performance during the first-lactation period. Furthermore, a significant quadratic correlation was observed between LDMI and 305-d milk, milk fat, and protein yields during the first-lactation period. The optimal performance during the first lactation was achieved when LDMI was maintained at 0.79 to 0.80 kg/d. In contrast, no significant association was observed between SDMI and production performance during the first-lactation period. Further multivariate mixed model analyses demonstrated that, when the effects of the 3 independent variables were considered collectively, only ADG exhibited a significant positive effect on 305-d milk yield and fat production during the first-lactation period. However, the modeling of milk protein yield revealed that ADG and LDMI exerted a significant influence, whereas the effect of SDMI remained insignificant. This study emphasized the significant effect of ADG and LDMI in optimizing the future lactation performance of calves, providing a crucial foundation for the development of scientific feeding management strategies.

Approximately, one-third of dairy cows suffer from postpartum diseases. Ketosis is considered an important inducer of other postpartum diseases by disrupting energy metabolism. Although the rumen microbiome may be involved in the etiology of ketosis by supplying volatile fatty acids, the rumen environmental dynamics of ketosis cows are unclear. Using multi-omics, this study aimed to elucidate changes in the rumen microbiome during parturition of ketosis cows and the association between the rumen microbiome and host energy metabolism. The study included 810 rumen content samples and 789 serum samples from day − 21 and 21 relative to calving day from 61 ketosis cows and 84 healthy cows. In ketosis cows, the rumen bacterial composition after parturition changed dramatically and needed a longer time to restore. The molar proportions of propionate were lower in ketosis cows than those in healthy cows on days 3 and 7 and negatively correlated with the serum β-hydroxybutyrate (BHBA) levels. The fermentation sub-pathway of propionate metabolism and partial glucogenic amino acid pathways were downregulated on day 3. Prevotella, UBA1066, and microbiota diversity indices regulate serum BHBA and glucose (GLU) levels via arginine, alanine, glycine, or propionate. Propionate administration to ketosis cows potentially decreased the serum BHBA concentration. Collectively, we found rumen disruption happened after calving among ketosis cows, and insufficient glycogenic substrates, such as propionate, may be related to ketosis development. The study findings have implications for the relationship between rumen microbiome dynamics and host energy metabolism, which lays the foundation for the future rumen microbiome investigation for improving postpartum management in cows.

Subacute ruminal acidosis (SARA) has emerged as a prevalent digestive disorder that significantly affects the overall health of ruminants, with notable links to various inflammatory diseases. Throughout the progression of SARA, elevated lipopolysaccharide (LPS) levels in the rumen play a crucial role in initiating the innate immune response. In this review, we evaluate the recent insights into the pathways associated with SARA-induced inflammatory responses, with a specific focus on LPS. It is important to recognize the variation in the immune response activation potential of LPS derived from different bacterial sources. This variability aligns with the widespread detection of LPS in the rumens of ruminants with SARA. Nonetheless, trained immunity is expected to become a novel strategy for the prevention and control of SARA. This mechanism offers a rapid response to secondary stimuli, including LPS, effectively preventing inflammation. Ultimately, this review establishes a comprehensive system integrating SARA, LPS, and trained immunity. Through this integrated approach, we aim to provide innovative solutions to the challenges associated with SARA.

Milk is a valuable source of essential nutrients for humans. In dairy cows, the rumen—the largest digestive organ—plays a critical role in supporting high productivity. The postpartum period marks a crucial transition to peak lactation, characterized by a rapid rise in milk and milk solids yield. However, the microbial mechanisms underlying this transition remain poorly understood. Previous studies of periparturient cows' rumen microbiota have primarily relied on relative microbiome profiles (RMP). Due to high false-positive rates of relative abundance (RA), RMP are unsuitable for capturing longitudinal microbial dynamics. In contrast, quantitative microbiome profiles (QMP), which focuses on absolute abundance (AA), offers a more accurate assessment of microbial community changes over time. Using QMP, we identified two distinct enterotypes (Clusters 1 and 2) across all samples, with Cluster 2 increasing progressively during lactation. Subsequently, through differential analysis, correlation analysis, and contribution assessment, we identified three genera—Succinivibrionaceae_UCG-001, Lachnospira, and Ruminococcus—were strongly associated with the rapid increase in milk yield. Among them, Succinivibrionaceae_UCG-001 showed the most significant contribution to changes in milk yield and energy corrected milk (ECM), with its abundance markedly rising as lactation progressed. Additionally, we observed that the effect of Ruminococcus on ECM may be mediated by changes in ruminal propionate e level. These findings highlight Succinivibrionaceae_UCG-001 as a key taxon driving the early lactation surge in milk production and provide novel insights into the role of the rumen microbiome in optimizing dairy performance.

Antibiotic resistance genes (ARGs) and virulence factor genes (VFGs) have become a global public health threat. As one of the main types of livestock, dairy cows under intensive farming practices pose a greater risk as the gastrointestinal tract of subclinically diseased cows serves as a "silent" gene reservoir. However, the distribution, transmission, and impacts of ARGs and VFGs in the gastrointestinal tract of subclinically diseased cows on human health and their own metabolism remain unclear. In this study, a nested case-control study was conducted within a prospective cohort of 211 periparturient dairy cows, collecting a total of 58 samples of rumen fluid, feces, and blood from subclinical ketosis and healthy cows. The results demonstrated that compared to healthy cows, the gastrointestinal microbiome of diseased cows contained a greater number of ARGs and VFGs exhibiting significantly increased abundance (ARGs in rumen: 30 vs 10, ARGs in feces: 66 vs 42; VFGs in rumen: 6 vs 1, VFGs in feces: 30 vs 10). Meanwhile, analysis of mobile genetic elements (MGEs) and MetaCompare2 database further indicated that the gastrointestinal microbiome of diseased cows poses a greater potential risk to human health. Additionally, specific ARGs/VFGs (e.g., FosM1) were found to be involved in the pathological process of ketosis of cow and were significantly associated with endogenous pathways such as bile acid metabolism. This highlights the important role of the ARGs-VFGs-CAZymes (Carbohydrate-active enzymes) co-occurrence network in host metabolic diseases. Our study highlights the human health risks of subclinically diseased cows as a reservoir of resistance genes, providing a theoretical basis and practical direction for improving the "One Health" prevention and control strategy.

Peripartum dairy cows commonly experience energy metabolism disorders, which lead to passive culling of postpartum cows and a decrease in milk quality. By using ketosis peripartum dairy cows as a model, this study aims to elucidate the metabolic mechanism of peripartum cows and provide a novel way for managing energy metabolic disorders. From a cohort of 211 cows, we integrated multi-omics data (metagenomics, metabolomics, and transcriptomics) to identify key microbes and then utilized an in vitro rumen fermentation simulation system and ketogenic hepatic cells to validate the potential mechanisms and the effects of postbiotics derived from key microbes. Postpartum cows with metabolic disorders compensate for glucose deficiency through mobilizing muscle proteins, which leads to marked decreases in milk protein content. Concurrently, these cows experience rumen microbiota disturbance, with marked decreases in the concentrations of volatile fatty acids and microbial protein, and the deficiency of alanine (Ala) in microbial protein is correlated with the metabolic disorder phenotype. Metagenomic binning and in vitro fermentation assays reveal that Ruminococcus_E bovis (MAG 189) is enriched in amino acid biosynthesis functions and responsible for Ala synthesis. Furthermore, transcriptomic and metabolomic analyses of the liver in metabolic disorder cows also show impaired amino acid metabolism. Supplementation with Ala can alleviate ketogenesis in liver cell models by activating the gluconeogenesis pathway. This study reveals that Ruminococcus_E bovis is associated with host energy metabolism homeostasis by supplying glucogenic precursors to the liver and suggests the use of Ala as a method for the treatment of energy metabolism disorders in peripartum cows.

Postpartum energy metabolism disorders pose a significant challenge to the health and productivity of dairy ruminants, yet their underlying pathogenesis remains poorly understood. The critical role of the gut microbiota in regulating host metabolic processes via the “gut-liver axis” has garnered increasing attention, but its specific mechanisms in dairy ruminant energy metabolism disorders are still unclear. This study uses dairy cows as a model and employs a large-scale case–control analysis to systematically investigate the pathophysiological basis of postpartum energy metabolism disorders through the lens of the “gut-liver axis.” Postpartum energy metabolism disorders in dairy cows are characterized by elevated blood β-hydroxybutyrate (BHB) and aspartate aminotransferase (AST) levels, and hepatic steatosis. A random forest model based on gut microbiota effectively predicts disease occurrence (AUC = 0.74). Multi-omics (metagenomics, metabolomics, and transcriptomics) analysis further identified key microbes, including Faecousia species (sp017465625 and sp017380435), Methanosphaera species (sp016282985), and Bifidobacterium globosum. These microbes regulate acetate concentration in the gut, which is significantly correlated with key genes in the hepatic PPAR and PI3K-AKT pathways, as well as with blood BHB levels. Primary hepatocyte culture experiments further confirmed that sodium acetate effectively inhibits hepatic fat deposition induced by mixed fatty acids through the hepatic AMPK-PPARA axis and reduces the production of BHB in the culture medium. This study demonstrates that key gut microbes and their metabolic product (acetate) inhibit the occurrence of metabolic disorders through the hepatic AMPK-PPARA axis. These findings provide new insights and potential therapeutic targets for understanding and mitigating postpartum metabolic disorders in dairy ruminants.

Methane is an important component of greenhouse gases, and ruminant production is a significant source of methane emissions. At present, flavonoid feed additives have certain applications in methane inhibition in ruminants. However, the effects of different doses of quercetin on rumen fermentation parameters, rumen bacteria and archaea are still unclear. Therefore, this study investigated the effects of quercetin on in vitro rumen fermentation parameters, methane production, and microflora in beef cattle. A completely randomized design was adopted. Quercetin was added to the fermentation substrates at 0% (group C), 0.5% (group Q1), 1% (group Q2) and 1.5% (group Q3). Anaerobic fermentation was carried out at 39°C for 48 h, gas production (GP) was recorded at different times, gas composition was determined, and methane (CH4) production was calculated. Fermentation parameters and dry matter digestibility (DMD) were determined after 48 h. Moreover, rumen fluid was collected for rumen bacterial and archaeal flora determination. The results were as follows: (1) After 32 h of fermentation, the GP decreased in response to the addition of quercetin. With increasing quercetin concentration, the theoretical maximum gas production decreased quadratically before 20 h (Pquadratic = 0.032). There was a quadratic increase in gas production (Pquadratic = 0.024). With increasing quercetin supplementation, the NH3-N content increased quadratically (Pquadratic = 0.027). MCP increased linearly and quadratically with quercetin (Plinear = 0.002, Pquadratic = 0.005), whereas DMD decreased linearly and quadratically with quercetin (Plinear = 0.013, Pquadratic = 0.032). Both 0.5 and 1% quercetin significantly reduced the butyrate content (Pquadratic = 0.002). With the addition of quercetin, the levels of butyrate, isobutyrate, isovalerate, and total volatile fatty acid (TVFA) first decreased but then increased (Pquadratic < 0.05). (2) With increasing quercetin concentration, methane production (Pquadratic = 0.009) and the methane proportion (Pquadratic < 0.001) decreased quadratically. (3) The ACE index and Chao1 index increased quadratically with quercetin supplementation (Pquadratic < 0.05). The relative abundance of Succiniclasticum in groups Q1 and Q3 increased, whereas the relative abundances of norank_f__norank_o__Rickettsiales and Curtobacterium decreased in all quercetin groups at the genus level (P < 0.05). (4) Quercetin supplementation did not affect the diversity of the archaeal community, but the relative abundance of Methanobrevibacter in group Q2 decreased. Overall, quercetin influenced in vitro rumen fermentation and the bacterial flora to decrease methane production and promote rumen nitrogen utilization and MCP synthesis.

Fatty liver syndrome is a prevalent metabolic disorder in transition dairy cows, characterized by excessive hepatic lipid accumulation that impairs liver function and leads to systemic metabolic disturbances. Docosahexaenoic acid (DHA), a prominent n-3 polyunsaturated fatty acid (PUFA), not only exhibits anti-inflammatory and anti-oxidative properties, but also holds potential in ameliorating lipid metabolism. This study integrated in vitro bovine primary hepatocyte models and in vivo dairy cow trials to investigate the regulatory effects of DHA on hepatic lipid deposition. In vitro, 40 μmol/L DHA significantly reduced triglyceride (TAG) accumulation in steatotic hepatocytes by downregulating genes involved in fatty acid transport (FABP-1, CD36) and lipogenesis (DGAT2, FAS, SREBP-1C), while upregulating markers of lipolysis (CGI-58, ATGL) and fatty acid oxidation (ACADL, CPT1A, CPT2). Transmission electron microscopy (TEM) confirmed DHA-mediated restoration of mitochondrial ultrastructure and enhanced lipid droplet (LD)-mitochondria interactions. In vivo, dietary rumen-protected DHA (180 g/d) supplementation reduced hepatic lipid deposition, improved liver function (evidenced by decreased total bilirubin and alanine aminotransferase), reduced oxidative stress and inflammation (suppressed malondialdehyde, glutathione peroxidase, and lipopolysaccharide), coincided with relieving insulin resistance (reduced insulin and glucose, as well increased adiponectin) in dairy cows with fatty liver. These improvements may be attributed to increased expression of TOMM20 and MtCo-1, promoting mitochondrial biogenesis and β-oxidation, along with an elevated plasma n-3/n-6 ratio. Collectively, these findings suggest that DHA supplementation represents a promising nutritional strategy for preventing spontaneous fatty liver in transition dairy cows by enhancing hepatic lipid clearance and restoring metabolic homeostasis.

Microbiome and resistome transmission from mother to child, as well as from animal to environment, has been widely discussed in recent years. Dairy cows mainly provide milk and meat. However, in the dairy production system, the characteristics and transmission trends of resistome assembly and the microbiome in the gastrointestinal tract (GIT) remain unclear. In this study, we sequenced the GIT (rumen fluid and feces) microbiome of dairy cow populations from two provinces in China (136 cows and 36 calves), determined the characteristics of their resistome profiles and the distribution of antibiotics resistance genes (ARGs) across bacteria and further tracked the temporal dynamics of the resistome in offspring during early life using multi-omics technologies (16S ribosomal RNA [rRNA] sequencing, metagenome, and metatranscriptome). We characterized the GIT resistome in cows, distinguished by gut sites and regions. The abundance of ARGs in calves peaked within the first 3 days after birth, with Enterobacteriaceae as the dominant microbial host. As calves aged, resistome composition stabilized, and overall ARG abundance gradually decreased. Both diet and age influenced carbohydrate-active enzymes and ARG profiles. Resistance profiles in ecological niches (meconium, colostrum, soil, and wastewater) were unique, resembling maternal sources. Mobile genetic elements (MGEs), mainly found in soil and wastewater, played an important role in mediating these interactions. Multidrug resistance consistently emerged as the most significant form of resistance at the both the metagenome and metatranscriptome levels. Several antibiotic classes showed higher proportions at the RNA level than at the DNA level, indicating that even low-abundance gene groups can have a considerable influence through high expression. This study broadens our understanding of ARG dissemination in livestock production systems, providing a foundation for developing future preventive and control strategies.

Diurnal oscillations have been reported on ruminal prokaryotes, but the daily rhythmicity of eukaryotes remains unknown. This study investigated diurnal oscillations of ruminal prokaryotes and eukaryotes under three different feeding managements and rumen fluid transplantation conditions, aiming to elucidate the regulatory mechanisms influencing the dynamic shifts of rumen microbiome through the daily feeding cycle. Quantification and profiling of the microbiota of 288 rumen samples collected from lactating dairy cows (n = 12) every 6-h over 48-h feeding cycles under ad libitum, restricted feeding at daytime and nighttime, respectively, revealed the rhythmicity in the population and abundance of ruminal bacteria, archaea, and protozoa. Under restricted-feeding regimes, 61.99 The classified feeding-time responsive, multi-factor responsive, consistent, and inconsistent circadian rhythm of microbial taxa underscore the driven factors behind the daily dynamics of rumen microbes, which also filled the gaps for targeting specific microbial taxa for better animal production.

It has been reported that rumen microbiota affects the cattle's milk-yield productivity, but the gut microbiota's contribution to the individualized performance and its associated mechanism have not been well defined. In this study, microbiota of 222 rumen and hindgut respective samples collected from 74 cows throughout the prepartum, postpartum, and peak-lactation periods were assessed using 16S rRNA gene amplicon analysis and were evaluated whether they affected inter-individual microbial interactions, assembly, functions, and contributed to host milk production and serum parameters. Prevotella-dominated (R-Prevot, n = 27) and Butyrivibrio-dominated (R-Butyri, n = 47) enterotypes were identified for rumen microbiota, and Prevotellaceae_UCG-003-dominated (H-Prevot, n = 33) and Paeniclostridium-dominated (H-Paenic, n = 41) enterotypes were identified for fecal microbiota. Positive cohesion (cooperative behaviour) was higher, while negative cohesion (competitive behavior) was lower in R-Prevot compared to R-Butyri enterotype throughout the three lactation periods. For H-Prevot enterotype, positive cohesion was higher at prepartum and peak-lactation, but lower at postpartum; and negative cohesion was lower at prepartum and postpartum with no difference detected at peak-lactation. Both deterministic and stochastic processes contributed to the rumen and hindgut microbiota assembly process with the proportion of dispersal limitation process being higher in R-Butyri than in R-Prevot, as well as in H-Prevot than in H-Paenic enterotype at peak-lactation. Additionally, the cows with R-Prevot/H-Prevot enterotypes (n = 15) had higher milk yield and lower serum non-esterified fatty acid concentration than the cows with R-Butyri/H-Paenic enterotypes (n = 29) during lactation. These findings provide evidence that enterotype could affect microbial interactions and assembly processes, as well as the cows' productivity.