Identification and characterization of whole blood gene expression and splicing quantitative trait loci during early to mid-lactation of dairy cattle.

BACKGROUND: Characterization of regulatory variants (e.g., gene expression quantitative trait loci, eQTL; gene splicing QTL, sQTL) is crucial for biologically interpreting molecular mechanisms underlying loci associated with complex traits. However, regulatory variants in dairy cattle, particularly in specific biological contexts (e.g., distinct lactation stages), remain largely unknown. In this study, we explored regulatory variants in whole blood samples collected during early to mid-lactation (22-150 days after calving) of 101 Holstein cows and analyzed them to decipher the regulatory mechanisms underlying complex traits in dairy cattle. RESULTS: We identified 14,303 genes and 227,705 intron clusters expressed in the white blood cells of 101 cattle. The average heritability of gene expression and intron excision ratio explained by cis-SNPs is 0.28 ± 0.13 and 0.25 ± 0.13, respectively. We identified 23,485 SNP-gene expression pairs and 18,166 SNP-intron cluster pairs in dairy cattle during early to mid-lactation. Compared with the 2,380,457 cis-eQTLs reported to be present in blood in the Cattle Genotype-Tissue Expression atlas (CattleGTEx), only 6,114 cis-eQTLs (P < 0.05) were detected in the present study. By conducting colocalization analysis between cis-e/sQTL and the results of genome-wide association studies (GWAS) from four traits, we identified a cis-e/sQTL (rs109421300) of the DGAT1 gene that might be a key marker in early to mid-lactation for milk yield, fat yield, protein yield, and somatic cell score (PP4 > 0.6). Finally, transcriptome-wide association studies (TWAS) revealed certain genes (e.g., FAM83H and TBC1D17) whose expression in white blood cells was significantly (P < 0.05) associated with complex traits. CONCLUSIONS: This study investigated the genetic regulation of gene expression and alternative splicing in dairy cows during early to mid-lactation and provided new insights into the regulatory mechanisms underlying complex traits of economic importance.

A matter of age? How age affects the adaptation of lactating dairy cows to virtual fencing.

Virtual Fencing (VF) can be a helpful technology in managing herds in pasture-based systems. In VF systems, animals wear a VF-collar using global positioning, and physical boundaries are replaced by virtual ones. The Nofence (Nofence AS, Batnfjordsøra, Norway) collars used in this study emit an acoustic warning when an animal approaches the virtual boundaries, followed by an aversive electrical pulse if the animal does not return to the defined area. The stimuli sequence is repeated up to 3 times if the animal continues to walk forward. Although it has been demonstrated that animals successfully learn to adapt to the system, it is unknown if this adaptation changes with animal age and thus has consequences for VF training and animal welfare. This study compared the ability of younger and older dairy cows to adapt to a VF system and whether age affected activity behavior, milk yield, and animal long-term stress under VF management. The study was conducted on four comparable strip-grazing paddocks. Twenty lactating Holstein-Friesian cows, divided into four groups of five animals each, were equipped with VF collars and pedometers. Groups differed in age: two groups of older cows (> 4 lactations) and two groups of younger ones (first lactation). After a 7-day training, paddock sizes were increased by successively moving the virtual fence during four consecutive grazing periods. Throughout the study, the pedometers recorded daily step count, time spent standing, and time spent lying. For the determination of long-term stress, hair samples were collected on the first and last day of the trial and the hair cortisol content was assessed. Data were analyzed by generalized mixed-effect models. Overall, age had no significant impact on animal responses to VF, but there were interaction effects of time: the number of acoustic warnings in the last period was higher in younger cows (P < 0.001), and the duration of acoustic warnings at training was shorter for older cows (P < 0.01). Moreover, younger cows walked more per day during the training (P < 0.01). Finally, no effects in milk yield or hair cortisol content were detected. In conclusion, all cows, regardless of age, adapted rapidly to the VF system without compromising their welfare according to the indicators measured.

Association between serum mineral levels and reproductive performance in primiparous dairy cows during the peripartum period.

To the best of the authors' knowledge, no study has previously investigated whether the concentration of minerals is related to reproductive outcomes in primiparous cows. For this reason, two objectives were set in the present study: (i) to assess serum mineral levels, macrominerals, and trace elements during the transition period (period of high nutritional requirements) in primiparous cows, considering reproductive efficiency, and (ii) to address if the serum mineral levels of primiparous cows are related to reproductive efficiency. Blood samples were taken (i) one month before calving, (ii) one week before calving, (iii) one week postpartum, and (iv) one month postpartum. At the beginning and the end of the study, a body condition score (BCS) was assigned to each lactating cow with no clinical signs of disease. The difference between one month before and one month after calving was the body condition loss (ΔBCS). Optimal prepartum concentrations of K and Cl were associated with fewer days open and a shorter interval calving. Furthermore, macrominerals in the serum decreased immediately after calving (one week) but recovered at one month postpartum. In contrast, the highest concentration of trace elements was found at one week postpartum. Primiparous cows with higher postpartum Se, Mn, Co, and Mo concentrations exhibited better reproductive efficiency, and the concentrations of trace elements in serum were correlated with interval calving and the number of inseminations. Finally, primiparous cows with a greater ΔBCS (at least one point) in period 4 exhibited both a longer calving interval and a greater number of days open. In summary, this study showed, for the first time in primiparous cows, that the concentration of some serum minerals not only plays a crucial role during the transition period but is also related to crucial reproductive parameters, such as interval calving and days open.

Identification and Characterization of long non-coding RNAs in Mammary Gland Tissues of Chinese Holstein Cows.

LncRNAs (Long non-coding RNA) is an RNA molecule with a length more than 200bp. LncRNAs can directly act on mRNA, thus affecting the expression of downstream target genes and proteins, and widely participate in many important physiological and pathological regulation processes of the body. In this study, RNA-Seq was performed to detect lncRNAs from mammary gland tissues of 3 Chinese Holstein cows, including 3 cows at 7 days before calving and the same 3 cows at 30 days postpartum (early lactation stage). A total of 1,905 novel lncRNAs were detected, 57.3% of the predicted lncRNAs are ≥ 500bp and 612 lncRNAs are intronic lncRNAs. The exon number of lncRNAs ranged from 2 to 10. A total of 96 lncRNAs were significantly differentially expressed between two stages, which 47 were upregulated and 49 were downregulated. Pathway analysis found that target genes were mainly concentrated on the ECM-receptor interaction, Jak-STAT signaling pathway, PI3K-Akt signaling pathway and TGF-beta signaling pathway. This study revealed the expression profile and characteristics of lncRNAs in the mammary gland tissues of Holstein cows at non-lactation and early lactation period, and providing a basis for studying the functions of lncRNAs in Holstein cows during different lactation periods.

Population characteristics of pathogenic Escherichia coli in puerperal metritis of dairy cows in Ningxia region of China: a systemic taxa distribution of virulence factors and drug resistance genes.

Escherichia coli (E. coli) is closely associated with the occurrence of puerperal metritis in dairy cows. E. coli carries some the virulence and multi-drug resistant genes, which pose a serious threat to the health of postpartum cows. In this study, E. coli was isolated and identified from the uterine contents of postpartum cows with puerperal metritis in the Ningxia region of China, and its phylogenetic subgroups were determined. Meanwhile, virulence and drug resistance genes carried by E. coli and drug sensitivity were detected, and the characteristics of virulence and drug resistance genes distribution in E. coli phylogroups were further analyzed. The results showed that the isolation rate of E. coli in puerperal metritis samples was 95.2%. E. coli was mainly divided into phylogroups B2 and D, followed by groups A and B1, and was more connected to O157:H7, O169:H4, and ECC-1470 type strains. The virulence genes were mainly dominated by ompF (100%), traT (100%), fimH (97%), papC (96%), csgA (95%), Ang43 (93.9%), and ompC (93%), and the resistance genes were dominated by TEM (99%), tetA (71.7%), aac(3)II (66.7%), and cmlA (53.5%). Additionally, it was observed that the virulence and resistance gene phenotypes could be divided into two subgroups, with subgroup B2 and D having the highest distributions. Drug sensitivity tests also revealed that the E. coli was most sensitive to the fluoroquinolones enrofloxacin, followed by macrolides, aminoglycosides, tetracyclines, ß-lactams, peptides and sulfonamides, and least sensitive to lincosamides. These results imply that pathogenic E. coli, which induces puerperal metritis of dairy cows in the Ningxia region of China, primarily belongs to the group B2 and D, contains multiple virulence and drug resistance genes, Moreover, E. coli has evolved resistance to several drugs including penicillin, lincomycin, cotrimoxazole, and streptomycin. It will offer specific guidelines reference for the prevention and treatment of puerperal metritis in dairy cows with E. coli infections in the Ningxia region of China.

The Effects of Lentinan on the Hematological and Immune Indices of Dairy Cows.

In this study, we investigated the effects of lentinan (LNT) on hematological parameters, immune indices, and metabolite levels in dairy cows. We randomly assigned forty Holstein cows to four treatment groups. The treatments consisted of 0, 5, 10, and 15 g/d of LNT. Compared with the control group, the addition of 10 g/d of LNT decreased the content of ALT and IL-8 but simultaneously increased the content of IL-4 in the cows' serum. Supplementation with 10 g/d of LNT decreased the levels of lymphocyte, RDW, ALT, AST, TC, IL-2, and IL-8, but, concurrently, in-creased the levels of granulocytes and IL-4 in their serum. In addition, supplementation with 15 g/d of LNT decreased the levels of RDW, TC, IL-2, and IL-8, but, at the same time, increased the levels of IL-4 and IgM in their serum. For the metabolomic analysis, cows fed with 0 and 10 g/d of LNT were selected. The results showed that 10 metabolites, including reduced nicotinamide riboside and trehalose, were upregulated in the 10 g/d group. These differential metabolites were enriched in tyrosine metabolism and trehalose degradation and altered two metabolic pathways of ubiquinone and other terpene quinone biosynthesis, as well as starch and sucrose metabolism. These findings provide evidence that LNT could be used to reduce the risk of inflammation in dairy cows.

Supplementation of Methionine Dipeptide Enhances the Milking Performance of Lactating Dairy Cows.

Methionine dipeptide (Met-Met) could improve milk protein synthesis in bovine epithelia mammary cells and lactating mice, while the effects of Met-Met on lactation performance, rumen fermentation and microbiota profile in lactating dairy cows have not been explored. For this reason, 60 Chinese lactating Holstein cows were allocated into three treatment groups: control group (CON), 6 g/d methionine dipeptide group (MM), and 6.12 g/d rumen-protected methionine dipeptide group (RPMM). The experiment lasted for 10 weeks to monitor lactation performance, plasma amino acid profile and rumen fermentation parameters and microbiota profile. Results showed that MM increased the energy-corrected milk (ECM), and RPMM increased both milk yield and ECM (p < 0.05). The milk protein concentration and yield were increased by MM and RPMM (p < 0.05). The rumen fermentation showed that RPMM increased total volatile fatty acids, acetate and valerate concentrations (p < 0.05). The relative abundance of Firmicutes, including Succiniclasticum, Selenomonas and Clostridium_XlVa, were enriched and the Prevotella was decreased by RPMM (p < 0.05). In summary, daily supplementing with 6 g of MM or RPMM in lactating dairy cows could improve milk yield and both percentage and yield of milk protein, and RPMM benefited the rumen fermentation and altered the bacterial composition. These results provided the first evidence that Met-Met supplementation can improve lactation performance of dairy cows.

Which Factors Influence the Consumption of Antibiotics in Q-Fever-Positive Dairy Farms in Northern Germany?

It was the aim of this study to examine whether the usage of the vaccine COXEVAC® (Ceva Santé Animale) could reduce the consumption of antibiotics in Q-fever-positive dairy farms. Additionally, the effects of other herd-level factors on the consumption of antibiotics were investigated. A total of 36 farms with vaccination and 13 farms without vaccination participated in this longitudinal cohort study. In all herds, Coxiella burnetii had been directly or indirectly diagnosed. To compare the treatment frequency of antibiotics between the vaccinated group and the non-vaccinated group, the consumption of antibiotics for each farm was collected using the veterinary documents about the application and delivery of antibiotics. To gather detailed information about herd data, nutrition, milking management, housing, and animal health, the farmers were interviewed with the help of a questionnaire. The results thereof suggest that there might be an association between the vaccination against Q fever and a reduced consumption of antibiotics. Neither herd size nor milk yield level influenced the consumption of antibiotics in the study herds. Type of flooring and udder-cluster disinfection while milking were associated with a lower and higher therapy frequency, respectively. Further studies are necessary to elucidate the cause-effect relationship between vaccination and the consumption of antibiotics.

Application of Epidemiological Methods in a Large-Scale Cross-Sectional Study in 765 German Dairy Herds-Lessons Learned.

From 2016 to 2020, the "PraeRi" study, conducted by three German veterinary universities, was aimed at enhancing animal health and welfare in dairy farms. With 765 dairy farms visited and 101,307 animals examined, this study provided a basis for improving animal health and welfare. The study population comprised three different regions representing a broad variety of characteristics. To ensure representative estimates, a sample size of 250 farms was determined for each region, employing a stratified sampling plan based on farm size. According to the information provided by the farmers, the most commonly occurring disease in their herds was mastitis without general disorder (14.2% to 16.3% of the herd-depending on the region). For most disorders, prevalence data were lowest for the region South compared with the two remaining regions. Multivariable regression analyses were performed to identify risk factors for various target variables, and the results were communicated through individual reports and benchmarking flyers to participating farmers. The authors encountered challenges in management and communication due to the project's size in terms of personnel, data, and farms examined. Harmonizing data management and hypothesis testing across all involved parties added complexity.

Comparison between intrauterine application of an antibiotic and an herbal product to treat clinical endometritis in dairy cattle - A randomized multicentre field study.

Uterine diseases are main indications for antibiotic use in dairy cows. To test a non-antibiotic treatment option, we compared the effect of an intrauterine cephapirin (Metricure®; cefapirin benzathin 500 mg per dosis; CEPH) with an intrauterine applied herbal product (25 ml of EucaComp® PlantaVet containing alcoholic extracts of Calendula officinalis L., Mellissa officinalis L., Origanum majorana L. and Eucalyptus globulus Labill. (EUC)) on the clinical cure of endometritis. Examinations of 816 cows between 21 and 35 days after calving were performed and cases of clinical endometritis (n = 169) were included. Diagnosis based on a scoring system for vaginal discharge. Study animals were randomly assigned to one of two treatment groups and treated immediately. After excluding animals with incomplete datasets, 136 cows (EUC: n = 61; CEPH: n = 75) remained for the final analysis. In total, 64% (EUC: 61%, CEPH: 67%) of analysed endometritis cases were considered as clinically cured 14 ± 2 days after the first treatment, 15% stayed uncured after the application of a maximum of two consecutive treatments, leading to an overall clinical cure rate of 85% (EUC: 82%, CEPH: 88%). No statistically difference in clinical cure rates could be observed between both treatment groups nor 14 ± 2 days after the first treatment (p = 0.956) neither regarding the overall cure rate (p = 0.923). In conclusion, the clinical cure of dairy cows' endometritis after the intrauterine application of the herbal product was non-inferior to the intrauterine application of the antibiotic cephapirin. These results could contribute to reduce the antimicrobial use in the daily veterinary routine treatment of endometritis.

Improving the efficiency of estrus synchronization in cows.

Objective: This study aimed to determine the effects of biologically active substances and electrical stimulation of the uterus in cows on the effectiveness of estrus synchronization. Materials and Methods: Ninety (n = 90) Kazakh white-headed cows were synchronized with two injections of gonadotropin-releasing hormone on days 0 and 9 and prostaglandin F2α on day 7. The cows were divided into six groups and, during the protocol, treated with biologically active substances (Tetramag, Selevetum, antiseptic-stimulator Dorogov 2 fraction, groups 2, 3, and 4). Cows in groups 5 and 6 were treated with the same substances but additionally had electrical stimulation of the uterus, while cows in group 1 were left untreated and served as a control. Results: The results have shown that on Day 0, no differences were observed in E2 concentrations between the groups. However, on the 10th day, a significant disparity was noted in the E2 level among cows in group 6 compared to groups 2, 3, 4, and the control group. Conversely, no significant differences were observed between groups 5 and 6. Likewise, the fertility rate in cows from group 6 was significantly higher compared to groups 2, 3, 4, and the control group, with no significant differences between groups 5 and 6. Conclusion: It can be concluded that the utilization of electrical stimulation of the uterus and the inclusion of certain biological substances during the estrus synchronization protocol demonstrate a positive effect on the reproductive performance of beef cattle in Kazakhstan.

Effect of dietary supplementation of plantain herb, lemongrass and their combination on milk yield, immunity, liver enzymes, serum, and milk mineral status in dairy cows.

Objective: This research aimed to assess the effects of dried plantain herb, lemongrass, and their combination on milk yield, immunological, liver enzymatic, serum, and milk mineral status in dairy cows. Materials and Methods: Twenty cows were arbitrarily assigned to 4 diets. Cows were given a basal ration considered as control diet (CL-D) having 14.93% crude protein (CP)and 10.96 MJ ME per kg dry matter (DM). Each cow was given 100 gm plantain, 100 gm lemongrass, and 50 gm plantain + 50 gm lemongrass with CL-D and taken as plantain diet (PT-D), lemongrass diet (LG-D), and plantain-lemongrass diet (PL-D), daily for 63 days, respectively. Blood and milk samples were taken four times at an interval of 14 days. Data were analyzed using a two-way repeated measures analysis of covariance. Results: Better DM consumption and milk yield were observed in the PT-D and LG-D compared to the CL-D (p ≤ 0.05). LG-D improved the milk's total solids, protein, and fat compared to CL-D (p < 0.05). Substantially, herbal groups improved serum albumin and reduced globulin concentrations compared to CL-D. LG-D had the highest serum immunoglobulin G, while herbal groups effectively reduced the liver enzymes compared to CL-D. Herbal groups did not affect serum and milk's calcium and phosphorus concentrations, while LG-D and PL-D substantially improved serum and milk zinc concentrations. Conclusions: Both plantain and lemongrass improved dairy cows' DM consumption and milk yield. Plantain and/or lemongrass enhanced the immune system and liver health, but not serum and milk calcium and phosphorus level. Lemongrass and a combination of plantain and lemongrass increased the serum and milk zinc concentrations.

Health-Related Outcomes and Molecular Methods for the Characterization of A1 and A2 Cow's Milk: Review and Update.

A new trend in cow's milk has emerged in the market called type A1 and A2 milk. These products have piqued the interest of both consumers and researchers. Recent studies suggest that A2 milk may have potential health benefits beyond that of A1 milk, which is why researchers are investigating this product further. It is interesting to note that the A1 and A2 milk types have area-specific characteristics compared to breed-specific characteristics. Extensive research has focused on milk derivatives obtained from cow's milk, primarily through in vitro and animal studies. However, few clinical studies have been conducted in humans, and the results have been unsatisfactory. New molecular techniques for identifying A1 and A2 milk may help researchers develop new studies that can clarify certain controversies surrounding A1 milk. It is essential to exercise extreme caution when interpreting the updated literature. It has the potential to spread panic worldwide and have negative economic implications. Therefore, this study aims to investigate the differences between A1 and A2 milk in various research areas and clarify some aspects regarding these two types of milk.

Mycobiome of the external ear canal of healthy cows.

Malassezia yeasts belong to the normal skin microbiota of a wide range of warm-blooded animals. However, their significance in cattle is still poorly understood. In the present study, the mycobiota of the external ear canal of 20 healthy dairy Holstein cows was assessed by cytology, culture, PCR, and next-generation sequencing. The presence of Malassezia was detected in 15 cows by cytology and PCR. The metagenomic analysis revealed that Ascomycota was the predominant phylum but M. pachydermatis the main species. The Malassezia phylotype 131 was detected in low abundance. Nor M. nana nor M. equina were detected in the samples.

The mycobiota of the external ear canal of healthy cows was assessed by cytology, culture, PCR, and NGS. The presence of Malassezia was detected by cytology and PCR. Ascomycota was the main phylum and M. pachydermatis the main species. The Malassezia phylotype 131 was also detected in the samples.

Harnessing Vaginal Probiotics for Enhanced Management of Uterine Disease and Reproductive Performance in Dairy Cows: A Conceptual Review.

Uterine disease in cattle impairs reproductive performance and profitability and increases antibiotic use and antimicrobial resistance. Thus, probiotics offer a promising alternative therapy. This review presents conceptual findings on the efficacy of probiotics in managing uterine diseases and fertility in cows. Probiotics containing Lactobacillus spp. and Bifidobacterium spp. individually or as composite formulations are known to improve fertility. Strategic intravaginal administration of these formulations would likely enhance uterine immunity, particularly during the postpartum period. While current findings on the benefits to uterine health are encouraging, there is still significant knowledge missing, including a lack of empirical information from large-scale field trials. This review underscores the need for evidence-based guidelines for probiotics, such as genomic selection of formulations, targeted delivery, or potential synergy with other interventions. Future research should address these gaps to maximize the potential of probiotics in managing uterine diseases and enhancing the reproductive health of dairy cattle.

The cooperative regulatory effect of the miR-130 family on milk fat metabolism in dairy cows.

Objective: There is a strong relationship between the content of beneficial fatty acids in milk and milk fat metabolic activity in the mammary gland. To improve milk quality, it is therefore necessary to study fatty acid metabolism in bovine mammary gland tissue. In adipose tissue, peroxisome proliferator-activated receptor gamma (PPARG), the core transcription factor, regulates the fatty acid metabolism gene network and determines fatty acid deposition. However, its regulatory effects on mammary gland fatty acid metabolism during lactation have rarely been reported. Methods: Transcriptome sequencing was performed during the prelactation period and the peak lactation period to examine mRNA expression. The significant upregulation of PPARG drew our attention and led us to conduct further research. Results: According to bioinformatics prediction, dual-luciferase reporter system detection, qRT‒PCR and Western blotting, miR-130a and miR-130b could directly target PPARG and inhibit its expression. Furthermore, triglyceride and Oil Red O staining proved that miR-130a and miR-130b inhibited milk fat metabolism in BMECs, while PPARG promoted this metabolism. In addition, we also found that the coexpression of miR-130a and miR-130b significantly enhanced their ability to regulate milk fat metabolism. Conclusion: In conclusion, our findings indicated that miR-130a and miR-130b could target and repress PPARG and that they also have a functional superposition effect. miR-130a and miR-130b seem to synergistically regulate lipid catabolism via the control of PPARG in BMECs. In the long-term, these findings might be helpful in developing practical means to improve high-quality milk.

Clinicopathological studies on ulcerative lymphangitis in cattle: Alterations in serum inflammatory cytokines, anti-microbial, organs functions, and oxidative stress-related biomarkers.

Background: Affection with Corynebacterium pseudotuberculosis (C. pseudotuberculosis) and development of cellulitis and/or abscess formation with cutaneous lymphangitis in cattle is rare to some extent, so literature about the biochemical changes that would accompany this infection is rare. Aim: In this context, the present study was designed to screen the effect of the infection with C. pseudotuberculosis cutaneous lymphangitis on the release of some immune molecules, organ functions, and redox state in Baladi cows. Methods: Fourteen Baladi cows from a small dairy farm in El-Behira, Egypt, were selected to complete this study. After bacteriological culture confirmation, seven of them were found suffering from cutaneous lesions due to infection with C. pseudotuberculosis (Diseased group), while the others were healthy (Healthy group). Serum samples were obtained to evaluate the presumptive changes in some clinicopathological parameters. Results: Serum analysis revealed a significant decrease in the levels of interferon-gamma and interleukin-17 as well as a significant decrement in the concentration of beta-defensin (ß-defensin) and lipocalin-2. While serum level of interleukin-10 recorded a significant increase in these animals when compared to healthy control animals. Concurrently, the affected animals recorded a significant elevation in serum levels of hepato-cardiac enzymes, urea, and creatinine in addition to disturbance in the serum redox state. Conclusion: In conclusion, infection with C. pseudotuberculosis cattle may disturb the defensive immune state, body organ function, and redox state of the animals.

Feeding rumen-protected methionine during the peripartum period improved milk fat content and reduced the culling rate of Holstein cows in a commercial herd.

Researchers have reported the benefits of feeding rumen-protected methionine (RPM) during the peripartum on the health parameters of dairy cows. Rumen-protected methionine has reportedly improved milk yield, milk components and liver health, but the literature is scarce on its effects in commercial herds. Therefore, we aimed to determine the effects of feeding RPMet (Smartamine M®, Adisseo Inc., Antony, France) prepartum (8 g per cow per day) and postpartum (15 g per cow per day) on performance, metabolic profile, and culling rate of Holstein cows in a commercial herd. One-hundred and 66 (n = 166) Holstein cows, 58 nulliparous and 108 parous, were randomly assigned to 1 of 2 dietary treatments, consisting of TMR top-dressed with RPMet (2.35 and 2.24% Met of MP for close-up and fresh cows, respectively) or without (control, CON, (2.03 and 1.89% Met of MP for close-up and fresh cows, respectively), fed from 21 ± 6 d prepartum until 16 ± 5 d postpartum. From 17 d in milk (DIM) until dry-off, all cows received RPMet. Daily milk yield was recorded, and milk samples were collected in the first and second weeks after calving to determine their composition. Blood samples were collected before the morning feeding on -14, -7, +1, +7, and +14 d relative to calving. Mortality and morbidity were recorded during the first 60 DIM. Cows supplemented with RPMet had greater milk yield during the first 16 DIM (31.76 vs. 30.37 kg/d; SEM = 1.04, respectively), and had greater milk fat content (4.45 vs. 4.10%; SEM = 0.11, respectively), but not milk total protein (3.47 vs. 3.39%; SEM = 0.04, respectively) and casein contents (2.74 vs. 2.66%; SEM = 0.04, respectively) than CON cows. Cows in RPMet had increased plasma Met concentrations than cows in CON (24.9 vs. 21.0 µmol/L; SEM = 1.2, respectively). Although morbidity was similar between treatments, the culling rate from calving until 60 DIM was lower for RPMet cows than for CON cows (2.4 vs. 12.1%; SEM = 0.02). In conclusion, cows receiving RPMet have greater milk yield, improved milk fat content, and a lower culling rate at 60 DIM than CON cows.

Combined Metabolomics and Biochemical Analyses of Serum and Milk Revealed Parity-Related Metabolic Differences in Sanhe Dairy Cattle.

The production performance of dairy cattle is closely related to their metabolic state. This study aims to provide a comprehensive understanding of the production performance and metabolic features of Sanhe dairy cattle across different parities, with a specific focus on evaluating variations in milk traits and metabolites in both milk and serum. Sanhe dairy cattle from parities 1 to 4 (S1, n = 10; S2, n = 9; S3, n = 10; and S4, n = 10) at mid-lactation were maintained under the same feeding and management conditions. The milk traits, hydrolyzed milk amino acid levels, serum biochemical parameters, and serum free amino acid levels of the Sanhe dairy cattle were determined. Multiparous Sanhe dairy cattle (S2, S3, and S4) had a greater milk protein content, lower milk lactose content, and lower solids-not-fat content than primiparous Sanhe dairy cattle (S1). Moreover, S1 had a higher ratio of essential to total amino acids (EAAs/TAAs) in both the serum and milk. The serum biochemical results showed the lower glucose and total protein levels in S1 cattle were associated with milk quality. Furthermore, ultra-high-resolution high-performance liquid chromatography with tandem MS analysis (UPLC-MS/MS) identified 86 and 105 differential metabolites in the serum and milk, respectively, and these were mainly involved in amino acid, carbohydrate, and lipid metabolism. S1 and S2/S3/S4 had significantly different metabolic patterns in the serum and milk, and more vitamin B-related metabolites were significantly higher identified in S1 than in multiparous cattle. Among 36 shared differential metabolites in the serum and milk, 10 and 7 metabolites were significantly and strongly correlated with differential physiological indices, respectively. The differential metabolites identified were enriched in key metabolic pathways, illustrating the metabolic characteristics of the serum and milk from Sanhe dairy cattle of different parities. L-phenylalanine, dehydroepiandrosterone, and linoleic acid in the milk and N-acetylornithine in the serum could be used as potential marker metabolites to distinguish between Sanhe dairy cattle with parities of 1-4. In addition, a metabolic map of the serum and milk from the three aspects of carbohydrates, amino acids, and lipids was created for the further analysis and exploration of their relationships. These results reveal significant variations in milk traits and metabolites across different parities of Sanhe dairy cattle, highlighting the influence of parity on the metabolic profiles and production performance. Tailored nutritional strategies based on parity-specific metabolic profiles are recommended to optimize milk production and quality in Sanhe cattle.

Lipopolysaccharide-binding protein in follicular fluid is associated with the follicular inflammatory status and granulosa cell steroidogenesis in dairy cows.

Metabolic stress and subsequent hepatic dysfunction in high-producing dairy cows are associated with inflammatory diseases and declining fertility. Lipopolysaccharide (LPS)-binding protein (LBP) is produced by hepatocytes and controls the immune response, suggesting that it is involved in the pathophysiology of inflammation-related attenuation of reproductive functions during metabolic stress. This study investigated the effect of LBP on the inflammatory status, oocyte quality, and steroidogenesis in the follicular microenvironment of dairy cows. Using bovine ovaries obtained from a slaughterhouse, follicular fluid and granulosa cells were collected from large follicles to evaluate the follicular status of metabolism, inflammation, and steroidogenesis. Cumulus-oocyte complexes were aspirated from small follicles and subjected to in vitro embryo production. The results showed that follicular fluid LBP concentrations were significantly higher in cows with fatty livers and hepatitis than in those with healthy livers. Follicular fluid LBP and LPS concentrations were negatively correlated, whereas LPS concentration showed a positive correlation with the concentrations of non-esterified fatty acids (NEFA) and ß-hydroxybutyric acid in follicular fluid. The blastulation rate of oocytes after in vitro fertilization was impaired in cows in which coexisting large follicles had high NEFA levels. Follicular fluid NEFA concentration was negatively correlated with granulosa cell expression of the estradiol (E2) synthesis-related gene (CYP19A1). Follicular fluid LBP concentration was positively correlated with follicular fluid E2 concentration and granulosa cell CYP19A1 expression. In conclusion, follicular fluid LBP may be associated with favorable conditions in the follicular microenvironment, including low LPS levels and high E2 production by granulosa cells.