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Goat milk is abundant in nutrients, particularly in milk fats, which confer health benefits to humans. Exploring the regulatory mechanism of fatty acid synthesis is highly important to understand milk composition manipulation. In this study, we used chromatin immunoprecipitation sequencing (ChIP-seq) on goat mammary glands at different lactation stages which revealed a novel lactation regulatory factor: cell death-inducing DFFA-like effector B (CIDEB). RT-qPCR results revealed that CIDEB was significantly upregulated during lactation in dairy goats. CIDEB overexpression significantly increased the expression levels of genes involved in fatty acid synthesis ( ACACA , SCD1 , p < 0.05; ELOVL6 , p < 0.01), lipid droplet formation ( XDH , p < 0.05), and triacylglycerol (TAG) synthesis ( DGAT1 , p < 0.05; GPAM , p < 0.01) in goat mammary epithelial cells (GMECs). The contents of lipid droplets, TAG, and cholesterol were increased ( p < 0.05) in CIDEB-overexpressing GMECs, and knockdown of CIDEB led to the opposite results. In addition, CIDEB knockdown significantly decreased the proportion of C16:0 and total C18:2. Luciferase reporter assays indicated that X-box binding protein 1 (XBP1) promoted CIDEB transcription via XBP1 binding sites located in the CIDEB promoter. Furthermore, CIDEB knockdown attenuated the stimulatory effect of XBP1 on lipid droplet accumulation. Collectively, these findings elucidate the critical regulatory roles of CIDEB in milk fat synthesis, thus providing new insights into improving the quality of goat milk.

Background: Goat milk has gained global attention for its unique nutritional properties and potential health benefits. Advancements in functional genomic technologies have significantly progressed genetic research on milk composition traits in dairy goats., Results: This review summarizes various research methodologies applied in this field. Genome-wide association studies (GWAS) have identified genomic regions associated with major milk components, with the diacylglycerol acyltransferase 1 ( DGAT1 ) gene and casein gene cluster consistently linked to milk composition traits. Transcriptomics has revealed gene expression patterns in mammary tissue across lactation stages, while the role of non-coding RNAs (such as miRNAs and circRNAs) in regulating milk composition has been confirmed. Proteomic and metabolomic studies have not only helped us gain a more comprehensive understanding of goat milk composition characteristics but have also provided crucial support for the functional validation of genes related to milk components. The integration of multi-omics data has emerged as an effective strategy for elucidating complex regulatory networks from a systems biology perspective., Conclusions: Despite progress, challenges remain, including refining reference genomes, collecting large-scale phenotypic data, and conducting functional validations. Future research should focus on improving reference genomes, expanding study populations, investigating functional milk components, exploring epigenetic regulation and non-coding RNAs, and studying microbiome-host genome interactions. These efforts will inform more precise genomic and marker-assisted selection strategies, advancing genetic improvements in milk composition traits in dairy goats.

Background: Milk clotting enzymes, essential for milk coagulation in cheese production, are obtained from the stomach of young ruminants, an expensive and limited source. This study was accomplished by finding a suitable alternative. Bacterial isolates recovered from honey were screened for milk clotting enzyme activity. and further, by immobilization of the microorganisms to enhance stability and facilitate their repeated use., Result: The most effective enzyme was produced by a microbe identified as Bacillus amyloliquefaciens based on 16 S rRNA sequencing. The cells were encapsulated in Ca 2+ alginate beads. These beads retained complete enzyme production after being used five times. Glucose and Soybean were selected as the most favorable carbon and nitrogen sources, respectively. The optimum temperature for activity was 35 ℃ for both free and immobilized cells but as the temperature was increased to 55 °C and above, the encapsulated form retained more activity than the free cells. The pH optimum shifted from 6.5 to 7 for the free cells to 7-7.5 for the immobilized cells. The immobilization process decreased the activation energy for enzyme production and activity, prolonged the enzyme half-life, and increased the deactivation energy. Enzyme produced by immobilized cells generated a more compact cheese., Conclusions: The finding of this study was to identify a less expensive source of milk-clotting enzymes and confirm the success of cell immobilization in improving cell rigidity and stability. Also, immobilization of this B. amyloliquefaciens strain offers an enzyme source of value for industrial production of cheese., (© 2024. The Author(s).)

The impact of nutritional modification to increase functional polyunsaturated fatty acids (PUFA), such as n-3 and n-6 fatty acids (FA) or conjugated linoleic acid (CLA), on milk proteome profile during early lactation remains largely unknown. We used an untargeted proteomics approach to investigate the impact of lactation day and PUFA supplementation on the proteome signature in skimmed milk over the course of early lactation. Sixteen Holstein dairy cows received abomasal infusion of saturated FA (CTRL) or a mixture of essential FA and CLA (EFA + CLA group) from - 63 to + 63 days relative to parturition. Using quantitative proteomics, 479 unique proteins were identified in skimmed milk at days 1, 28, and 63 postpartum. The top discriminating proteins between transition milk (day 1) and mature milk (days 28 and 63), including members of complements (i.e. C2 and C5), growth factor (TGFB2), lipoproteins (i.e. APOE and APOD), and chaperones (i.e. ST13 and CLU), are associated with calves' immune system and gut development. The EFA + CLA supplementation moderately affected a few proteins associated with regulating mammary glands' lipogenesis through the (re)assembly of lipoprotein particles, possibly under the PPAR signaling pathway. Collectively, skimmed milk proteome is dynamically regulated initially by cow's metabolic and physiological changes and to a lesser extent by nutritional PUFA modifications., (© 2024. The Author(s).)

Inflammatory diseases of the intestinal tract in piglets severely impair the economic performance of pig farms. Pig milk exosomes can encapsulate miRNAs which can then enter the piglet intestine to play an immunomodulatory role. Previously, we comparatively analyzed and identified exosomal miRNAs in the colostrum and mature milk of Bamei and Landrace pigs, and we screened for ssc-miR-22-3p, which is associated with inflammation and immune response; however, the role played by ssc-miR-22-3p in the immune response in IPEC-J2 cells is not yet clear. In this study, we first constructed a pig intestinal inflammatory response model using Lipopolysaccharide (LPS) and Polyinosinic-polycytidylic acid (Poly (I:C)), and we investigated the role of ssc-miR-22-3p targeting MAPK14 in the regulation of LPS and Poly (I:C)-induced inflammatory injury in IPEC-J2 cells by RT-qPCR, cell counting kit-8 (CCK-8), EdU staining, lactate dehydrogenase (LDH) activity assay, and dual luciferase reporter gene assay. We successfully established LPS and Poly (I:C)-induced cell damage models in IPEC-J2 cells. The immune response of IPEC-J2 cells was stimulated by induction of IPEC-J2 cells at 10 μg/mL LPS and 20 μg/mL Poly (I:C) for 24 h. Overexpression of ssc-miR-22-3p decreased cytokine expression and promoted cell viability and proliferation. The functional enrichment analysis revealed that ssc-miR-22-3p targets genes enriched in the pathways of negative regulation of inflammatory response and bacterial invasion of epithelial cells. The validity of the binding site of ssc-miR-22-3p to MAPK14 was tested by a dual luciferase reporter gene. Pig milk exosome ssc-miR-22-3p promotes cell viability and proliferation by targeting MAPK14 , and it alleviates LPS and Poly (I:C)-induced inflammatory responses in IPEC-J2 cells.

Exosomes are cell-derived, membrane-surrounded particles that deliver bioactive molecules to various cells. Due to their small size, low immunogenicity, extended blood circulation, and involvement in cellular communication, they hold potential as effective drug carriers. Exosomes are present in various biological fluids, including mare's milk, a traditional drink in Central Asia. This study aims to compare exosome isolation methodologies and determine the stability of mare's milk-derived exosomes as potential therapeutic carriers. Three extraction methods-immunoprecipitation, size exclusion chromatography, and total exosome isolation-were compared in terms of exosome characteristics, purity, and content. The isolated exosomes were then loaded with quercetin, and their ability to increase its bioavailability was tested in vitro and in vivo. Total exosome isolation was identified as the most efficient method for producing high-quality exosomes. These exosomes were loaded with quercetin and compared to free quercetin and exosomes alone. Exosomes loaded with 80 µM quercetin significantly restored β-galactosidase activity and cellular viability in doxorubicin-treated cells, exhibiting similar potency to 160 µM free quercetin. In aged model animals, treatment with quercetin-loaded exosomes resulted in significantly less acute and subacute damage to the myocardium, kidneys, and liver compared to untreated control animals. This study provides a proof-of-concept that mare's milk-derived exosomes can be effectively absorbed by cells and animal tissues, supporting their potential use as drug carriers.

Genome-wide association studies (GWASs) are used to identify quantitative trait loci for phenotypic traits of interest. The use of multilocus mixed models allows to correct for population stratification and account for long-range linkage disequilibrium. In this study, GWASs were conducted to identify the genetic bases of milk production (milk yield, protein and fat composition, and yield) in two autochthonous dual-purpose cattle breeds from the Aosta Valley. Using either the breeding values or the deregressed proofs, common significative single nucleotide polymorphisms have been identified for milk yield, protein percentage, and fat percentage. Two major quantitative trait loci regions have been identified on the chromosomes 5 and 14 for the fat percentage, harbouring the MGST1, CYHR1, VPS28, and CPSF1 genes. For the protein percentage, a candidate region has been identified on BTA 6; in this region, the CSN1S1, CSN2, HSTN, CSN3, and RUFY3 genes are annotated. Most of the identified genes have already been associated with milk composition in other studies on cosmopolitan and local cattle. These results show that the genes involved in milk composition quantitative traits in the Aosta cattle are common also in other cattle breeds and they can be further investigated with the use of whole genome sequencing data., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

A meta-analysis was performed to investigate the effects of capsaicin (CAP) on lactational performance and blood chemistry and cell counts in dairy cattle. The database comprised 11 peer-reviewed studies published between 2011 and 2024. The overall effect of CAP, challenge (e.g., ketosis, glucose tolerance, LPS), and dose were considered explanatory variables in the analysis. Lactation performance response variables included DMI, milk yield (MY), ECM yield, feed efficiency (FE), milk components, and BW. Hematological response variables included red and white blood cell counts and blood BHB, total fatty acids, insulin, and glucose concentrations. Data were analyzed using random- and mixed-effect models in the "robumeta" package in RStudio. Milk yield was increased by 2.9% by CAP when compared with control. Capsaicin supplementation increased FE by 3.4% compared with control. Milk fat concentration and yield were also increased by CAP compared with control by 2.6% and 4.0%, respectively. Blood glucose concentrations were decreased 2.5% by CAP supplementation, whereas insulin levels were unaffected. Cows fed CAP during a challenge had higher MY and FE and tended to have lower blood glucose than their control counterparts. Overall, this analysis suggests that CAP supplementation may be directly affecting host physiology by altering glucose metabolism, but further research to define the mechanism is warranted., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Nitrogen (N) utilisation efficiency (NUE, milk N yield [g/d]/N intake [g/d]) is an important performance indicator in dairy farming. Determining the NUE-associated blood metabolite profile will contribute to the optimisation of nutritional strategies to further improve NUE among dairy cows. Here, 20 primiparous lactating cows with days in milk ranging from 95 to 115 days were selected from a total of 1 221 cows. Each cow's N intake and milk N yield were measured for 7 days. Subsequently, blood samples were collected before morning feeding. Based on analysis and calculations, cows were retrospectively classified into two groups based on their NUE values, namely, a low NUE group (LNUE, NUE = 24.8 ± 1.6%, n = 10, mean ± SD) and a high NUE group (HNUE, NUE = 35.2 ± 1.7%, n = 10, mean ± SD). Plasma samples were selected from six cows in each group for metabolomics and amino acid profiling. Among the 41 differential metabolites (DMs) identified in the metabolomic analysis, sucrose, MG(0:0/22:1(13Z)/0:0), 2-amino-6-hydroxyhexanoic acid, and L-glutamine exhibited significant correlations with NUE, milk yield, and BW (P < 0.05). Moreover, the five differential amino acids and amino acid metabolites (DAAs) identified in the amino acid profiling and 5 of the 6 differential amino acids and amino acid conjugates identified by plasma metabolomics were found to be less abundant in the HNUE group (P < 0.05). Specifically, there was a 39.4% decrease in L-arginine content and a 29.2% decrease in L-glutamine content (P < 0.05). Pathway analysis indicated that the DMs and DAAs were mainly involved in arginine biosynthesis, glutathione metabolism, arginine and proline metabolism, and tryptophan metabolism (pathway impact > 0.1). These results provided new insights into the new blood metabolite profile associated with NUE in dairy cows. These new insights can provide foundational information for the formulation of new strategies to further enhance NUE in dairy cows., (Copyright © 2024 The Author(s). Published by Elsevier B.V. All rights reserved.)

Dairy cows experiencing heat stress (HS) during the precalving portion of the transition period give birth to smaller calves and produce less milk and milk protein. Supplementation of rumen-protected methionine (RPM) has been shown to modulate protein, energy, and placenta metabolism, making it a potential candidate to ameliorate HS effects. We investigated the effects of supplementing RPM to transition cows under HS induced by electric heat blanket (EHB) on cow-calf performance. Six weeks before expected calving, 53 Holstein cows were housed in a tiestall barn and fed a control diet (CON, 2.2% Met of MP) or a CON diet supplemented with SmartamineM (MET, 2.6% Met of MP, Adisseo Inc., France). Four weeks precalving, all MET and half CON cows were fitted with an EHB. The other half of the CON cows were considered thermoneutral (TN), resulting in 3 treatments: CONTN (n = 19), CONHS (n = 17), and METHS (n = 17). Respiratory rate (RR), skin temperature (ST), and rectal temperature (RT) were measured thrice weekly and core body temperatures recorded biweekly. Postcalving BW and BCS were recorded weekly, and DMI was calculated and averaged weekly. Milk yield was recorded daily and milk components were analyzed every third DIM. Biweekly AA and weekly nonesterified fatty acids (NEFA), BHB, insulin, and glucose were measured from plasma. Calf birth weight and 24 h growth, thermoregulation, and hematology profile were measured and apparent efficiency of absorption (AEA) of immunoglobulins was calculated. Data were analyzed using the MIXED procedure of SAS with 2 preplanned orthogonal contrasts: CONTN versus the average of CONHS and METHS (C1) and CONHS versus METHS (C2). Relative to TN, EHB cows had increased RT during the postcalving weeks and increased RR and ST during the entire transition period. Body weight, BCS, DMI, and milk yield were not affected by the EHB or RPM. However, protein percentage and SNF were lower in CONHS, relative to METHS cows. At calving, METHS dams had higher glucose concentrations, relative to CONHS, and during the postcalving weeks, the EHB cows had lower NEFA concentrations than TN cows. Calf birthweight and AEA were reduced by HS, whereas RR was increased by HS. Calf withers height tended to be shorter and RT were lower in CONHS, compared with METHS heifers. Overall, RPM supplementation to transition cows reverts the negative effect of HS on blood glucose concentration at calving and milk protein percentage in the dams and increases wither height while decreasing RT in the calf., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Early lactation is a critical period for dairy cows, as energy requirements rapidly increase with the onset of lactation; however, early-lactation DMI in pasture-based systems are under measured. The objectives of this study were (1) to measure and profile total DMI (TDMI) and animal performance of dairy cows during early lactation in a pasture-based system, (2) to investigate early-lactation energy balance in pasture-based systems, and (3) to examine production efficiencies, including TDMI and milk solids production per 100 kg of BW. Eighty spring-calving dairy cows were allocated to a grazing group as they calved over a 2-yr period (2021 and 2022). Cows were offered a daily herbage allowance to achieve a postgrazing sward height of 4 cm, with silage supplementation when necessary due to inclement weather. Total DMI was measured using the n-alkane technique over a 12-wk period from February 1, 2021, to April 23, 2022. Total DMI and daily milk yield were significantly affected by parity with both variables being greatest for third-parity animals (17.7 kg of DM and 26.3 kg/cow per day, respectively), lowest for first parity (13.2 kg of DM and 19.6 kg/cow per day, respectively) and intermediate for second-parity animals (16.8 kg of DM and 24.1 kg/cow per day, respectively). Peak TDMI was reached on wk 10 for first-parity animals (14.6 kg of DM), wk 11 for second parity animals (19.3 kg of DM) and wk 12 for third-parity animals (19.9 kg of DM). Parity also had a significant effect on unité fouragère lait (UFL; feed units for milk) feed balance as first-parity animals experienced a greater degree of negative energy balance (-3.2 UFL) compared with second- and third-parity animals (-2.3 UFL). Breed and parity had an effect on production efficiencies during the first 12 wk of lactation as Jersey × Holstein Friesian cows had greater TDMI/100 kg of BW and milk solids/100 kg of BW compared with Holstein Friesian cows., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Determination of energy requirements for growth depends on measuring the composition of BW gain. Previous studies have shown that the composition of gain can be altered in young dairy calves by the composition of the milk replacer diet. Here, our objective was to determine body composition and the composition of empty body gain in young calves fed increasing amounts of a milk replacer containing adequate CP. Male Holstein calves underwent an adjustment period of 14 d after birth in which they were fed whole-waste milk at 10% of BW. Calves were then stratified by BW and randomly assigned to either an initial slaughter group (n = 11) or to groups fed 1 of 3 milk replacer amounts and slaughtered after 35 d of growth. Calves in all treatments consumed the same milk replacer containing 24.8% CP (DM basis; from all milk proteins) and 18.9% fat, reconstituted to 12.5% solids. Treatments were milk replacer fed at 1.25% of BW (DM basis; n = 6), 1.75% of BW (n = 6), or 2.25% of BW (n = 8), adjusted weekly as calves grew. Calves fed at 1.25% or 1.75% of BW were fed twice daily and those fed 2.25% of BW were fed 3 times daily. No starter was offered. Postslaughter, the bodies of calves were separated into 4 fractions: carcass; total viscera minus digesta; head, hide, feet, and tail;, and blood. The sum of those 4 fractions was empty BW (EBW), which increased linearly as the amount of milk replacer increased. Final heart girth and body length, but not withers height, increased linearly as intake increased. Gain:feed increased linearly with increasing milk replacer. Feeding more milk replacer increased the amounts of lean tissue and fat in the body. The percentages of water and protein in the final body decreased linearly, whereas fat percentage and energy content increased linearly as intake increased. As gain increased, the percentage of protein in gain decreased and the percentage of fat increased, resulting in an increase of energy content of EBW gain. Efficiency of energy use (retained energy:gross energy intake) increased linearly but retained energy:ME available for growth was not different among treatments. Efficiency of protein use increased quadratically as feeding rate increased; there was no further increase at 2.25% of BW. Plasma insulin-like growth factor 1, insulin, and glucose increased linearly, whereas urea-N decreased linearly, as milk replacer intake increased. Our data document changes in body composition that affect estimates of retained energy in the bodies of calves slaughtered at a common age. These data are important for calculations of energy requirements for young calves., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

The study investigated the effects of dietary protein level and the inclusion of hydroponic barley sprouts (HB) on lactation performance, blood biochemistry and N use efficiency in mid-lactation dairy cows. Treatments were arranged in a 2 × 2 factorial design with 2 CP levels (16.8% and 15.5% of DM), with HB (4.8% of DM, replacing 4.3% of alfalfa hay and 0.5% of distillers dried grains with solubles [DDGS]) or without HB. Forty-eight multiparous Holstein dairy cows (146 ± 15 DIM, 40 ± 5 kg/d of milk) were randomly allocated to 1 of 4 diets: high-protein diet (16.8% CP, HP), HP diet with HB (HP+HB), low-protein diet (15.5% CP, LP), or LP diet with HB (LP+HB). An interaction between CP × HB on DMI was detected, with DMI being unaffected by HB inclusion in cows fed the high-protein diets, but was lower in cows fed HB when the low-protein diet was fed. A CP × HB interaction was also observed on milk and milk protein yield, which was higher in cows fed HB with HP, but not LP. Inclusion of HB also tended to reduce milk fat content, and feeding HP resulted in a higher milk protein and MUN content, but lower milk lactose content. Feed efficiency was increased by feeding HP or HB diets, whereas N use efficiency was higher for cows fed LP or HB diets. There was an interaction on the apparent total-tract digestibility of DM and CP, which was higher when HB was fed along with HP, but reduced when fed with LP, whereas the digestibility of ADF was increased by feeding low-protein diets. In conclusion, feeding a low-protein diet had no adverse effect on cow performance, while feeding HB improved milk and milk component yield, and N efficiency when fed with a high-CP diet, but compromised cow performance with a low-CP diet., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

The aim of this study was to evaluate the effects of different anti-mycotoxin feed additives on the concentration of mycotoxins in milk, urine, and blood plasma of dairy cows fed diets artificially contaminated with mycotoxins. Secondarily, performance, total-tract apparent digestibility of nutrients, and blood parameters were evaluated. Twelve multiparous cows (165 ± 45 DIM, 557 ± 49 kg BW, and 32.1 ± 4.57 kg/d milk yield at the start of the experiment) were blocked according to parity, milk yield, and DIM and used in a 4 × 4 Latin square design experiment with 21-d periods, where the last 7 d were used for sampling and data analysis. Treatments were (1) mycotoxin group (MTX), basal diet (BD) without anti-mycotoxin feed additives; (2) hydrated sodium calcium aluminosilicate (HSCA), HSCA added to the BD at 25 g/cow per day; (3) mycotoxin deactivator (MD; Mycofix Plus, dsm-firmenich) added to the BD at 15 g/cow per day (MD15); and (4) MD added to the BD at 30 g/cow per day (MD30). Cows from all treatments were challenged with a blend of mycotoxins containing 404 μg of aflatoxin B 1 , 5,025 μg of deoxynivalenol (DON), 8,046 μg of fumonisins (FUM), 195 μg of T2 toxin (T2), and 2,034 μg of zearalenone (ZEN) added daily to the BD during the last 7 d of each period. Neither performance (milk yield and composition) nor nutrient digestibility was affected by treatments. All additives reduced aflatoxin M 1 (AFM1) concentration in milk, whereas MD15 and MD30 group had lower excretion of AFM1 in milk than HSCA. Deoxynivalenol, FUM, T2, or ZEN were not detected in milk of MD15 and MD30. Concentrations in milk of DON, FUM, T2, and ZEN were similar between MTX and HSCA. Except for AFM1, none of the analyzed mycotoxins were detected in urine of MD30 group. Comparing HSCA to MD treatments, the concentration of AFM1 was greater for HSCA, whereas MD30 was more efficient at reducing AFM1 in urine than MD15. Aflatoxin M1, DON, FUM, and ZEN were not detected in the plasma of cows fed MD30, and DON was also not detected in MD15 group. Plasma concentration of FUM was lower for MD15, similar plasma FUM concentration was reported for HSCA and MTX. Plasma concentration of ZEN was lower for MD15 than MTX and HSCA. Serum concentrations of haptoglobin and hepatic enzymes were not affected by treatments. Blood concentration of sodium was lower in HSCA compared with MD15 and MD30 groups. In conclusion, the mycotoxin deactivator proved to be effective in reducing the secretion of mycotoxins in milk, urine, and blood plasma, regardless of the dosage. This reduction was achieved without adverse effects on milk production or total-tract digestibility in cows fed multi-mycotoxin-contaminated diets over a short-term period. Greater reductions in mycotoxin secretion were observed with full dose of MD., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Brown Swiss (BS) cows have greater urea concentrations in milk and blood compared with Holstein (HO) cows. We tested the hypothesis that BS and HO cows differ in kidney function and nitrogen excretion. Blood, saliva, urine, and feces were sampled in 31 multiparous BS and 46 HO cows kept under identical feeding and management conditions. Samples were collected at different lactational stages after the monthly DHIA control test-day. To test the glomerular filtration rate (GFR) and urea excretion, concentrations of creatinine and urea were measured in serum, urine, and saliva. As an additional marker to estimate GFR, we determined symmetric dimethylarginine (SDMA) in serum. Feces were analyzed for DM content and nitrogen concentration. Data on milk urea and protein concentrations and daily milk yield were obtained from the monthly DHIA test-day records. The effects of breed, time, and parity number on blood, saliva, urine, feces, and milk parameters were evaluated with the GLM procedure of SAS, with breed, time, and parity number as fixed effects. Differences between BS and HO were assessed by the Tukey-corrected t-test. Concentrations of urea, creatinine, and SDMA in serum, were greater in BS than in HO cows: 5.46 ± 0.19 versus 4.72 ± 0.13 mmol/L (urea), 105.96 ± 2.23 versus 93.07 ± 1.50 mmol/L (creatinine), and 16.78 ± 0.69 versus 13.39 ± 0.44 µg/dL (SDMA). We observed a greater urea concentration in BS cows (25.8 ± 0.7 vs. 21.8 ± 0.7 mg/dL) and protein content in milk (3.70% ± 0.08% vs. 3.45% ± 0.07%) than in HO cows. Urea and creatinine concentrations in urine and saliva did not differ between breeds. No differences between BS and HO were observed for milk yield, fecal DM, and fecal nitrogen content. Dry matter intake and BW were similar in BS and HO cows. Despite greater urea, creatinine, and SDMA concentrations in blood, as well as a higher milk urea content in BS compared with HO, respective concentrations in urine did not differ between breeds. In conclusion, our results demonstrate a lower renal GFR in BS compared with HO cows, thereby contributing to the greater plasma urea concentration in BS cows. However, estimation of nitrogen excretion via milk, urine, and feces does not entirely reflect nitrogen turnover within the animal., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Dietary carbohydrate manipulation can be used to reduce enteric CH 4 emission, but few studies have focused on the interaction of the different types of carbohydrates that can affect feed intake and ruminal fermentation. Understanding this interaction is necessary to make the most out of CH 4 mitigation feeding strategies using different dietary carbohydrates. The aim of this study was to test the effect on enteric CH 4 emission, feed intake, and milk production response when cows were fed either grass-clover silage (GCS) or corn silage (CS) as the sole forage source (55% of dry matter, DM), in combination with either barley (BAR) or dried beet pulp (DBP) as a concentrate (21.5% of DM). A total of 24 (half first-parity and half second-parity) cows were used in a crossover design with 2 periods of 21 d each, receiving 2 of 4 diets obtained from a 2 × 2 factorial arrangement of the experimental diet. Feed intake, CH 4 emission metrics, and milk production were recorded at the end of the experimental periods. The diets had NDF concentrations between 258 and 340 g/kg of DM and starch concentrations between 340 and 7.45 g/kg of DM (CS-BAR and GCS-DBP, respectively). The effects of silage and concentrate on dry matter intake (DMI) were additive, with the highest feed intake in cows fed CS-BAR, followed by cows fed CS-DBP, GCS-BAR, and GCS-DBP (21.2, 19.9, 19.1, and 18.3 kg/d, respectively). Energy corrected milk (ECM) yield was not affected by silage source in first parity cows, but it was higher for cows fed CS than cows fed GCS in second parity. The effects of silage and concentrate on CH 4 production (g/d), yield (g/kg of DMI), and intensity (g/kg of ECM) were not additive, as cows fed GCS had similar responses regardless of the concentrate used, but cows fed CS had lower CH 4 production, yield, and intensity when fed BAR instead of DBP. The lower CH 4 production, yield, and intensity in cows fed CS-BAR compared with other diets could be partially explained by the nonlinear relationship between ruminal VFA and carbohydrates (NDF and starch) concentration reported in the literature; however, we observed a linear relationship between the acetate/propionate ratio and CH 4 yield, suggesting possible other effects. The effects of silage and concentrate on the ruminal VFA were additive in first parity cows, but not in second parity cows. The interaction between dietary carbohydrate type and parity might indicate an effect of feed intake or the energy balance of the cow. Feeding cows silage and concentrate both rich in starch can result in the lowest enteric CH 4 emission., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

We determined the effects of increasing the dietary inclusion of whole cottonseed (WCS) on nutrient digestibility and the milk production responses of high-producing dairy cows. Twenty-four multiparous Holstein cows (mean ± SD; 52.7 ± 2.63 kg/d milk; 104 ± 23 DIM) were randomly assigned to treatment sequences in a replicated 4 × 4 Latin square design with 21-d periods. Treatments were increasing doses of WCS at 0%, 8%, 16%, and 24% DM, with WCS replacing soybean meal and hulls to maintain similar diet nutrient composition (% DM) of NDF (32%), forage NDF (21%), starch (27%), and CP (17%). Total fatty acid (FA) content of each treatment was 1.70%, 2.96%, 4.20%, and 5.40% DM, respectively. Three preplanned contrasts were used to test the linear, quadratic, and cubic effects of increasing dietary WCS. Increasing dietary WCS from 0% to 24% DM quadratically influenced the intake of DM and NDF, with the highest value being the inclusion of 8% WCS, and intakes of 16- and 18-carbon, and total FA, with maximum values obtained up to 24% WCS. Increasing dietary WCS affected digestibility of DM (cubic) and NDF (quadratic), with the lowest values being the inclusion of 8% WCS. Increasing WCS increased 16-carbon digestibility (quadratic) but decreased digestibility of 18-carbon and total FA (both quadratic), with highest and lowest values for the inclusion of 24% WCS, respectively. Increasing dietary WCS quadratically increased absorbed 16- and 18-carbon, and total FA, with maximum values obtained for 24% WCS. Increasing dietary WCS quadratically increased yields of milk, milk fat, milk protein, milk lactose, 3.5% FCM, and ECM, and linearly increased BW gain. The source of milk FA was affected by dietary WCS, with a quadratic decrease in the yield of de novo and mixed milk FA and a quadratic increase in preformed milk FA. Increasing dietary WCS linearly increased trans-10 C18:1 milk FA content. As dietary WCS increased, plasma insulin linearly decreased, and plasma gossypol levels linearly increased. Despite the decrease in total FA digestibility, increasing dietary WCS from 0% to 24% DM increased FA absorption. Increasing the dietary inclusion of WCS up to 16% DM increased milk production responses and DM intake. Under the current dietary conditions, high-producing dairy cows benefited best from a diet containing an inclusion of 8% to 16% WCS in DM., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

The objectives of this study were to determine the farm-level hyperketolactia (HKL) prevalence, as diagnosed from the milk BHB concentration, on dairy farms milking with an automated milking system (AMS) and to describe the farm-level housing, management, and nutritional risk factors associated with increased farm-average milk BHB and the within-herd HKL prevalence in the first 45 DIM. Canadian AMS farms (n = 162; eastern Canada, n = 8; Quebec, n = 24; Ontario, n = 75; western Canada n = 55) were visited once between April and September 2019 to record housing and herd management practices. The first test milk data for each cow under 45 DIM were collected, along with the final test of the previous lactations for all multiparous cows, from April 1, 2019, to September 30, 2020. The first test milk BHB was then used to classify each individual cow as having or not having HKL (milk BHB ≥0.15 mmol/L) at the time of testing. Milk fat and protein content, milk BHB, and HKL prevalence were summarized by farm and lactation group (all, primiparous, and multiparous). During this same time period, formulated diets for dry and lactating cows, including ingredients and nutrient composition, and AMS milking data were collected. Data from the AMS were used to determine milking behaviors and milk production of each herd during the first 45 DIM. Multivariable regression models were used to associate herd-level housing, feeding management practices, and formulated nutrient composition with first test milk BHB concentrations and within-herd HKL levels separately for primiparous and multiparous cows. The within-herd HKL prevalence for all cows was 21.8%, with primiparous cows having a lower mean prevalence (12.2 ± 9.2%) than multiparous cows (26.6 ± 11.3%). Milk BHB concentration (0.095 ± 0.018 mmol/L) and HKL prevalence for primiparous cows were positively associated with formulated prepartum DMI and forage content of the dry cow diet; however, they were negatively associated with formulated postpartum DMI, the major ingredient in the concentrate supplemented through the AMS, and the partially mixed ration to AMS concentrate ratio. Multiparous cows' milk BHB concentration (0.12 ± 0.023 mmol/L) and HKL prevalence were positively associated with the length of the previous lactation, milk BHB at dry-off, prepartum diet nonfiber carbohydrate content, and the major forage fed on farm, while tending to be negatively associated with feed bunk space during lactation. This study is the first to determine the farm-level risk factors associated with herd-level prevalence of HKL in AMS dairy herds. The results may help to optimize management and guide diet formulation and thus promote the reduction of HKL prevalence., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

The objective of this study was to evaluate the effect of energy and protein in the diet on the recovery of milk ethanol stability (MES) induced by feed restriction. Twelve Holstein and Holstein × Jersey crossbred cows with an average of 146 ± 50 DIM, 575.4 ± 70 kg of BW, and 18.93 ± 5.46 kg/d of milk yield were distributed in a 3 × 3 Latin square design with 3 treatments and 3 experimental periods. Each experimental period lasted 24 d, comprising 3 phases: a 13-d adaptation phase (100E+100P), a 4-d induction phase for milk ethanol instability (50E+50P), and a 7-d recovery phase for MES (3 treatments). The 3 treatments during the recovery phase consisted of 3 diets aiming to meet the requirements of energy and protein (100E+100P), only energy (100E+50P), or only protein (50E+100P). The diet during the adaptation and induction phases was common for all cows. The energy and protein levels to meet each cow's requirements were based on the group average. Restriction of energy and protein reduced DM, CP, and total digestive nutrient intake for cows fed 100E+50P and 50E+100P. The lowest BW was observed for cows fed 50E+100P, with no difference for BCS. During the induction phase, MES was reduced by 9 percentage units. Cows fed 100E+100P recovered MES in the first days of the recovery phase, whereas 100E+50P slightly improved MES, and 50E+100P had a constant decrease in MES. Cows fed 100E+50P and 50E+100P produced, respectively, 3.6 and 5.9 kg less milk than those fed 100E+100P. The 50E+100P treatment exhibited the highest milk fat content and SCS, along with the lowest milk lactose content. Protein content was higher in the 100E+100P treatment. Cows fed 50E+100P showed higher serum albumin levels compared with those on the 100E+100P treatment, not differing from the 100E+50P treatment. We concluded that the complete recovery of MES in cows with feed restrictions is possible only by supplying both the energy and protein requirements in the cows' diet. However, restricting energy intake poses a greater limitation on MES recovery compared with restricting protein., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Mare milk has a unique protein composition that makes it a preferred option for adult and infant nutrition. Several functional properties have been attributed to this milk, but with little current evidence. In fact, knowledge on mare milk composition is still limited. In particular, studies addressing the performance of mare milk proteins during human gastrointestinal digestion are scarce, which limits the understanding of mare milk nutritional quality and functionality. For this reason, the present study describes the digestibility of mare milk proteins and the release of peptides as affected by management and lactation stage, factors which are known to affect milk composition. Mare milk samples from 3 different farms collected during 6 mo of lactation (n = 54) were subjected to a static in vitro gastrointestinal model to measure peptide release and protein digestibility. In the present study, a detailed description of protein and individual amino acid behavior during the digestion process was given. For the first time, digestion of the 2 equine β-lactoglobulin isoforms (I and II) was described individually. In addition, it was found that lactation stage and management system can significantly affect protein digestibility and peptide release during gastrointestinal digestion of mare milk. Presumably, differences in the composition of mare milk influence the protein structure and enzyme accessibility, which might have an impact on digestion behavior. Although no specific bioactive peptides were identified, several precursors of previously described bioactive peptides were found. These findings could support the idea of mare milk as a food with added value., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

The purpose of this experiment was to determine if nicotinic acid (NA) effects on dairy cows and rumen microbial characteristics are forage-type dependent (corn silage, CS; grass silage, GS). Four late-lactation (DIM = 225 ± 12 d) Holstein cows were used in a 4 × 4 Latin square design with a 2 × 2 factorial arrangement of treatments. The main effects were a CS (66.10% CS) based diet or a GS (79.59%) based diet with or without 12 g/d NA. Each experimental period lasted for 28 d. Milk production and milk components, blood metabolites, apparent total-tract nutrient digestibilities, minutes rumen pH were below 5.8 as an indicator of ruminal acidosis, and body temperature changes were analyzed as indicators of heat stress. Nicotinic acid supplementation did not improve apparent total-tract nutrient digestibility. Feeding a GS-based diet improved NDF and hemicellulose digestibility. Feeding a CS-based diet increased the apparent total-tract digestibility of fat, and minutes rumen pH below 5.8 for a greater proportion of the time. The CS-based diet also improved milk yield, milk fat and protein yields, and ECM yield; however, somatic cell count and BHB were also increased. Supplementing NA tended to decrease nonesterified fatty acids, especially when combined with GS where DMI was low. There was a trend for the total protozoa population to increase when GS and NA were fed but decreased when CS and NA were fed. Nicotinic acid tended to decrease rumen protozoal populations of Dasytricha, but increased populations of Ophryoscolex and Diplodiniinae with GS diets and decreased with CS diets. Entodiniinae were increased with CS but NA had no effect. Body temperature was increased when a CS-based diet was fed when compared with a GS-based diet. More research is needed to determine how NA can affect rumen microbial protein synthesis and what kind of diets will provide the optimum effect., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

The objectives of this observational cohort study were to evaluate the associations of rumination time (RT) in the last week of pregnancy with transition cow metabolism, inflammation, health, and subsequent milk production, reproduction, and culling. Pregnant nulliparous (n = 199) and parous (n = 337) cows were enrolled 21 d before expected calving. Rumination time and physical activity were monitored automatically by sensors from d -21 to 15 relative to calving. Blood samples were collected on d -14, -5, 4, 8, and 12 ± 1 relative to calving. Diagnoses of clinical health problems were performed by researchers from calving to 15 DIM. In classification 1, cows were ranked based on average daily RT in the last week of pregnancy and classified into terciles as short RT (SRT), moderate RT (MRT), or long RT (LRT) for association analyses. In classification 2, RT deviation from the parity average was used in a receiver operating characteristic curve to identify the best threshold to predict postpartum clinical disease. Cows were then classified as above the threshold (AT) or below the threshold (BT). Compared with cows with LRT, cows with SRT had greater serum concentrations of nonesterified fatty acids (0.47 vs. 0.40 ± 0.01 mmol/L), BHB (0.58 vs. 0.52 ± 0.01 mmol/L), and haptoglobin (0.22 vs. 0.18 ± 0.008 g/L) throughout the transition period, and reduced concentrations of glucose, cholesterol, albumin, and magnesium in a time-dependent manner. Parous cows with SRT had higher odds of postpartum clinical disease (adjusted odds ratio [AOR] = 3.7; 95% CI: 2.1-6.4), lower odds of pregnancy by 210 DIM (AOR = 0.34; 95% CI = 0.15-0.75), and lower milk production (46.9 vs. 48.6 ± 0.5 kg/d) than parous cows with LRT. Deviation in prepartum RT had good predictive value for clinical disease in parous cows (area under the curve = 0.65; 95% CI = 0.60-0.71) but not in nulliparous (area under the curve = 0.51; 95% CI = 0.42-0.59). Separation of parous cows according to the identified threshold (≤-53 min from the parity average) resulted in differences in serum concentrations of nonesterified fatty acids (AT = 0.31 ± 0.006, BT = 0.38 ± 0.014 mmol/L), BHB (AT = 0.49 ± 0.008, BT = 0.53 ± 0.015 mmol/L), and globulin (AT = 32.3 ± 0.3, BT = 34.8 ± 0.5 g/L) throughout the transition period, as well as in serum cholesterol, urea, magnesium, albumin, and haptoglobin in a time-dependent manner. Below threshold parous cows had higher odds of clinical disease (AOR = 3.7; 95% CI = 2.1-6.4), reduced hazard of pregnancy (adjusted hazard ratio = 0.64, 95% CI: 0.47-0.89), greater hazard of culling (adjusted hazard ratio = 2.1, 95% CI: 1.2-3.6), and lower milk production (46.3 ± 0.7 vs. 48.5 ± 0.3 kg/d). External validation using data from 153 parous cows from a different herd and the established threshold in RT deviation (≤-53 min) resulted in similar predictive value, with the odds of postpartum disease 2.4 times greater in BT than AT (37.5% vs. 20.1%). In conclusion, RT in the week preceding calving was a reasonable predictor of postpartum health and future milk production, reproduction, and culling in parous cows but not in nulliparous cows., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Most trace minerals (TM) are fed above dairy cow requirements in commercial herds but their fate and effects on dairy cows have not been well documented. In this study, we evaluated the effects of feeding short-term sulfate TM above recommendations on apparent total-tract digestibility of nutrients, rumen fermentation characteristics, serum concentrations, and milk yield and composition, as well as milk, fecal, and urinary TM excretion in midlactation dairy cows. Eight multiparous Holstein cows with an average body weight (± SD) of 684 ± 29 kg at 82 ± 10 DIM in a quadruple 2 × 2 crossover design were fed a basal diet, differing in sulfate TM supplement concentrations, to provide either 0.11, 17, and 63 (control; CON) or 0.95, 114, and 123 (high trace minerals; HTM) mg of dietary Co, Mn, and Zn per kilogram of DM, respectively. Each experimental period had a 21-d adaptation to the diet, followed by a 10-d sample collection period. Feed ingredients and total feces and urine were collected during 4 consecutive d and rumen fluid was collected 0, 1, 2, 4, and 6 h relative to feeding. Milk yield was recorded daily, and milk samples were collected on 4 consecutive milkings. Ingestion of Co, Mn, and Zn was higher for the HTM group compared with the CON group by 216%, 233%, and 93%, respectively. Dry matter intake averaged 25.0 (SE = 0.6) kg/d, and apparent total-tract digestibility of major nutrients was similar between treatments. High trace minerals had no measurable effect on ruminal pH, major volatile fatty acids, and protozoa counts. Isovalerate molar proportion was 9.4% greater for the HTM group compared with the CON group. Neither milk yield (43.5 kg/d; SE = 0.8) nor milk fat and protein concentrations differed between treatments. Milk urea nitrogen concentration was significantly higher for HTM (11.7 mg/dL) compared with CON (9.7 mg/dL; SE = 0.7). Fecal excretion of Co, Mn, and Zn increased by 223%, 198%, and 75%, respectively, for the HTM group compared with the CON group. Urinary excretions of TM were marginal compared with feces, and only urinary Co and Mn were significantly higher for HTM cows than CON cows, as was similarly obtained for serum Co and Mn concentrations. Milk TM yields were not modified by treatments. In summary, short-term dietary sulfate TM supply over the recommendation did not improve cow performance but significantly increased fecal TM excretion, which could affect TM accumulation in soils where manure is applied and could potentially result in leaching into nearby watersheds. Further studies are needed to evaluate the impact of high fecal TM excretion on the environment using the One Health approach. Moreover, the effects of TM oversupply on milk production and cow health should be evaluated by long-term experiments., (Copyright © 2024 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Omitting or shortening the dry period may result in a fairly constant ration throughout the transition period of dairy cows, reducing the need for adaptation of cow metabolism and rumen function to a new lactation. The objective of this study was to determine the effect of dry period length (DP) on rumen adaptation and cow metabolic state during the transition period. Twelve pregnant, rumen-cannulated Holstein Friesian dairy cows at the end of their first lactation were assigned to 1 of 3 treatments: a conventional (60 d), short (30 d), or no dry period (0 d). At dry-off, cows received a dry-cow ration until calving. Lactating cows received a lactation ration. Cows were monitored from 8 wk before calving until 8 wk after calving for milk yield and DMI. Rumen biopsies were taken from 3 locations in the rumen at 60, 40, and 10 d before calving and 3, 7, 14, 28, and 56 d after calving to assess papillae dimensions. Blood was sampled weekly from 3 wk before until 8 wk after calving, and liver biopsies were taken at wk -2, 2, and 4 relative to calving. Prepartum, DMI, and milk yield were greater for cows with a short or no dry period, compared with cows with a conventional dry period. Postpartum, DMI was greater for cows with a short dry period compared with cows with a conventional dry period. Plasma glucose concentration was greater for cows without a dry period, compared with the other DP postpartum. Plasma concentrations of nonesterified fatty acids and BHB, and liver triglyceride content, did not differ among dry period. Rumen papillae differed in size based on biopsy location, but there was no interaction between biopsy location and the effect of DP. Rumen papillae surface area for cows managed for a 30- or 60-d dry period decreased toward calving. At 40 d prepartum, papillae surface area was greater for short and no dry period treatment compared with a conventional dry period. At 10 d prepartum, papillae surface area was greater for the no dry period treatment compared with both other treatments, and this difference was still present 3 d postpartum. Cows managed for a short dry period showed faster increase in papillae dimensions after calving compared with cows managed for a conventional dry period. From d 28 onwards, no differences in papillae surface area were observed. The faster rumen adaptation postpartum may be related to the increased DMI during the first weeks postpartum for cows managed for a short dry period. However, this did not result in improved metabolic status or milk yield. The results from the present study demonstrate that the dietary changes related to a conventional DP affected rumen papillae development, not only prepartum but also early postpartum. Further optimization of DP as well as dietary composition throughout the transition period may support cows in their adaptation to a new lactation., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Animals vary in the way in which they use energy due to diet, genetics, and management. Energy consumed by the animal supports milk production, but considerable variation among animals in energy utilization is thought to exist. The study objective was to estimate the among-animal variance in energy utilization in data collected from Jersey cows using indirect calorimetry. Individual animal-period data from 15 studies (n = 560) were used. The dataset included 115 animals from 44 to 410 DIM producing 11.5 to 39.1 kg/d of milk. On average, the 63 treatments in the dataset ranged 14.8% to 19.5% CP, 21.4% to 43.0% NDF, 16.2% to 33.3% starch, and 2.21% to 6.44% crude fat. Data were analyzed with the GLIMMIX procedure of SAS (9.4) with random effects of cow, treatment nested within period, square, and experiment. The percentage of among-animal, dietary treatment, and experimental variance was calculated as the variance associated with each fraction divided by the sum of variance from animal, dietary treatment, experiment, and residual, which was considered the total variance. The percentage of among-animal variance was characterized as high or low when the value was greater than or less than the mean value of 29.2%. Among-animal variance explained approximately 29.3% to 42.5% of the total variance in DMI, gross energy (GE), digestible energy (DE), ME, and NE L in megacalories per day. When energetic components of feces, urine, heat, and lactation in megacalories per day were expressed per unit of DMI, the among-animal variance decreased by 20.4, 4.82, 9.55, and 3.80 percentage units, respectively. However, among-animal variance explained 4.80 and 5.02 percentage units more of the total variation for methane energy and tissue energy in megacalories per day when expressed per unit of DMI. Variance in energetic efficiencies of DE/GE, ME/GE, and ME/DE were explained to a lesser extent by among-animal variance (averaging 17.8 ± 1.95%). The among-animal contribution to total variance in milk energy was 28.8%. Milk energy was a large proportion of the energy efficiency calculation, which included milk energy plus corrected tissue energy over net energy intake, which likely contributed to the 22.2% of total among-animal variance in energy efficiency. Results indicate that among-animal variance explains a large proportion of the total variation in DMI. This contributes to the variance observed for energy fractions as well as energy components when expressed in megacalories per day. Variation in energetic loss associated with methane was primarily explained by differences among animals and was increased when expressed per unit of DMI, highlighting the role of inherent animal differences in these losses., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Improving the low productivity levels of native cattle breeds in smallholder farming systems is a pressing concern in Pakistan. Crossbreeding high milk-yielding holstein friesian (HF) breed with the adaptability and heat tolerance of Sahiwal cattle has resulted in offspring that are well-suited to local conditions and exhibit improved milk yield. The exploration of how desirable traits in crossbred dairy cattle are selected has not yet been investigated. This study aims to provide the first overview of the selective pressures on the genome of crossbred dairy cattle in Pakistan. A total of eighty-one crossbred, thirty-two HF and twenty-four Sahiwal cattle were genotyped, and additional SNP genotype data for HF and Sahiwal were collected from a public database to equate the sample size in each group. Within-breed selection signatures in crossbreds were investigated using the integrated haplotype score. Crossbreds were also compared to each of their parental breeds to discover between-population signatures of selection using two approaches: cross-population extended haplotype homozygosity and fixation index. We identified several overlapping genes associated with production, immunity, and adaptation traits, including U6, TMEM41B, B4GALT7, 5S&#95;rRNA, RBM27, POU4F3, NSD1, PRELID1, RGS14, SLC34A1, TMED9, B4GALT7, OR2AK3, OR2T16, OR2T60, OR2L3, and CTNNA1. Our results suggest that regions responsible for milk traits have generally experienced stronger selective pressure than others., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Recent research has highlighted the importance of dietary fatty acid profile of fatty acid supplements on production responses of high-producing dairy cows. Conventional soybeans contain ∼15% oleic acid and ∼50% linoleic acid whereas high oleic acid soybeans (HOSB) contain ∼70% oleic acid and ∼7% linoleic acid. We determined the effect of increasing dietary inclusion of roasted and ground HOSB on production responses of high-producing dairy cows. Twenty-four multiparous Holstein cows (mean ± SD; 50.7 ± 4.45 kg/d of milk; 122 ± 57 DIM) were randomly assigned to treatment sequences in a replicated 4 × 4 Latin square design with 21-d periods. Treatments were increasing doses of HOSB at 0%, 8%, 16%, and 24% of DM. The HOSB replaced conventional soybean meal and hulls to maintain similar diet nutrient composition (% of DM) of 27.4% to 29.4% NDF, 20.6% forage NDF, 27.5% starch, and 15.9 to 16.5% CP. Total fatty acid content of treatments was 1.65%, 3.11%, 4.52%, and 5.97% of DM, respectively. Preplanned polynomial orthogonal contrasts included the linear, quadratic, and cubic effects of increasing HOSB. Increasing dietary inclusion of HOSB linearly decreased DMI and MUN and increased yields of milk, 3.5% FCM, ECM, and milk fat, and quadratically increased milk protein. The increased response to milk fat was due to an increase in preformed milk fatty acids. Due to the increase in milk component yields and decrease in DMI, there was an increase in feed efficiency. Increasing HOSB inclusion linearly decreased plasma BUN concentration and tended to decrease plasma insulin. Increasing HOSB had no effect on BW change or BCS change. In summary, increasing dietary inclusion of HOSB up to 24% of DM increased production responses of high-producing dairy cows and did not affect body reserves., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Feed efficiency is important for economic profitability of dairy farms; however, recording daily DMI is expensive. Our objective was to investigate the potential use of milk mid-infrared (MIR) spectral data to predict proxy phenotypes for DMI based on different cross-validation schemes. We were specifically interested in comparisons between a model that included only MIR data (model M1); a model that incorporated different energy sink predictors, such as body weight, body weight change, and milk energy (model M2); and an extended model that incorporated both energy sinks and MIR data (model M3). Models M2 and M3 also included various cow-level variables (stage of lactation, age at calving, parity) such that any improvement in model performance from M2 to M3, whether through a smaller root mean squared error (RMSE) or a greater squared predictive correlation (R 2 ), could indicate a potential benefit of MIR to predict residual feed intake. The data used in our study originated from a multi-institutional project on the genetics of feed efficiency in US Holsteins. Analyses were conducted on 2 different trait definitions based on different period lengths: averaged across weeks versus averaged across 28 d. Specifically, there were 19,942 weekly records on 1,812 cows across 46 experiments or cohorts and 3,724 28-d records on 1,700 cows across 43 different cohorts. The cross-validation analyses involved 3 different k-fold schemes. First, a 10-fold cow-independent cross-validation was conducted whereby all records from any one cow were kept together in either training or test sets. Similarly, a 10-fold experiment-independent cross-validation kept entire experiments together, whereas a 4-fold herd-independent cross-validation kept entire herds together in either training or test sets. Based on cow-independent cross-validation for both weekly and 28-d DMI, adding MIR predictors to energy sinks (model M3 vs. M2) significantly (P < 10 -10 ) reduced average RMSE to 1.59 kg and increased average R 2 to 0.89. However, adding MIR to energy sinks (M3) to predict DMI either within an experiment-independent or herd-independent cross-validation scheme seemed to demonstrate no merit (P > 0.05) compared with an energy sink model (M2) for either R 2 or RMSE (respectively, 0.68 and 2.55 kg for M2 in herd-independent scheme). We further noted that with broader cross-validation schemes (i.e., from cow-independent to experiment-independent to herd-independent schemes), the mean and slope bias increased. Given that proxy DMI phenotypes for cows would need to be almost entirely generated in herds having no DMI or training data of their own, herd-independent cross-validation assessments of predictive performance should be emphasized. Hence, more research on predictive algorithms suitable for broader cross-validation schemes and a more earnest effort on calibration of spectrophotometers against each other should be considered., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

At the onset of lactation in dairy cows, inflammation and oxidative stress may occur and result in a risk of pathologies and lower milk yield. To propose an innovative management strategy for cows during this period, it is essential to better understand these physiological variations. Our objective was to evaluate the metabolic, redox, and immune status of 7 primiparous and 8 multiparous Holstein cows during late gestation and the first months of lactation. Blood samples were collected between 3 wk before calving until 12 wk postpartum. Milk samples were also collected, but only at the time points after calving. The metabolic (nonesterified fatty acids [NEFA], BHB, glucose, urea, calcium) and redox (reactive oxygen metabolites [ROM], oxidative stress index [OSI], glutathione peroxidase activity, vitamin E) statuses were analyzed in plasma or erythrocytes. The expression of genes related to antioxidant functions was determined in leukocytes collected from milk. For immune status, plasma cytokine levels and the production of reactive oxygen species (ROS) in classical and regulatory neutrophils were measured in 2 whole blood ex vivo challenges. The data were analyzed using a mixed model that included the fixed effects of parity and week and their interaction. Milk yield, plasma NEFA, and BHB in wk 2 and 4 after calving were higher in multiparous cows than in primiparous cows, whereas glucose and calcium tended to be lower. Plasma ROM and OSI levels in wk 8 were higher in multiparous than in primiparous cows. Multiparous cows also displayed higher glutathione peroxidase activity in erythrocytes, and antioxidant transcription factor and superoxide dismutase-1 expression levels in milk leukocytes. Moreover, multiparous cows had higher plasma concentrations of vitamin E but lower plasma levels of cytokines CXCL10, CCL2, IL1Rα, and IFNγ. Following ex vivo whole blood stimulation with Escherichia coli, lower IL1α and TNFα levels were measured in multiparous than in primiparous cows. Intracellular ROS production by neutrophils was lower in multiparous than in primiparous cows. These results thus indicated marked physiological changes in wk 8 compared with wk 2 and 4 of lactation. These differences in the physiological status of primiparous and multiparous cows offer interesting perspectives for potential dietary strategies to prevent pathologies which take account of parity and week relative to calving., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Nutrition and physiological state affect hepatic metabolism. Our objective was to determine whether feeding flaxseed oil (∼50% C18:3n-3 cis), high oleic soybean oil (∼70% C18:1 cis-9), or milk fat (∼50% C16:0) alters hepatic expression of PC, PCK1, and PCK2 and the flow of carbons from propionate and pyruvate into the tricarboxylic acid (TCA) cycle in preruminating calves. Male Holstein calves (n = 40) were assigned to a diet of skim milk with either: 3% milk fat (MF; n = 8), 3% flaxseed oil (Flax; n = 8), 3% high oleic soybean oil (HOSO; n = 8), 1.5% MF + 1.5% high oleic soybean oil (MF-HOSO; n = 8), or 1.5% MF + 1.5% flaxseed oil (MF-Flax; n = 8) from d 14 to d 21 postnatal. At d 21 postnatal, a liver biopsy was taken for gene expression and metabolic flux analysis. Liver explants were incubated in [U- 13 C] propionate and [U- 13 C] pyruvate to trace carbon flux through TCA cycle intermediates or with [U- 14 C] lactate, [1- 14 C] palmitic acid, or [2- 14 C] propionate to quantify substrate oxidation to CO 2 and acid-soluble products. Compared with other treatments, plasma C18:3n-3 cis was 10 times higher and C18:1 cis-9 was 3 times lower in both Flax (Flax and MF-Flax) treatments. PC, PCK1, and PCK2 expression and flux of [U- 13 C] pyruvate as well as [U- 13 C] propionate were not different among treatments. PC expression was negatively correlated with the enrichment of citrate M+5 and malate M+3, and PCK2 was negatively correlated with citrate M+5, suggesting that when expression of these enzymes is increased, carbon from pyruvate enters the TCA cycle via PC-mediated carboxylation, and then oxaloacetate is converted to phosphoenolpyruvate via PCK2. Acid-soluble product formation and PC expression were reduced in HOSO (MF-HOSO and HOSO) treatments compared with Flax (MF-Flax and Flax), indicating that fatty acids regulate PC expression and carbon flux, but that fatty acid flux control points are not connected to PC, PCK1, or PCK2. In conclusion, fatty acids regulate hepatic expression of PC, PCK1, and PCK2, and carbon flux, but the point of control is distinct., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Milk production and overall dairy farm economics depend on rearing dairy heifers. This study investigated the presence of a genotype by environment interaction in Holstein (HOL), Nordic Red dairy cattle (RDC), and their F 1 crossbreeds (HOL × RDC) when provided different feed rations. The aim of our study was to assess how different energy concentrations in feed rations affect growth, BCS, feed intake, and feed efficiency in the 3 groups during the prepubertal period. The 3 breed groups were randomly allocated to receive either a standard or a low-energy feed ration. Holstein heifers exhibited reduced growth and a lower BCS when they were fed the low-energy feed ration. In contrast, the RDC heifers demonstrated similar growth rates with the different feed rations and maintained similar BCS irrespective of feed energy concentration. The HOL × RDC crossbred heifers performed as an intermediate between the HOL and RDC groups. Significant differences were observed in DMI and energy intake in the HOL and HOL × RDC groups depending on feed ration treatment. The RDC heifers had similar feed intake irrespective of treatment. There were no significant differences in the feed conversion ratio among breeds and feed treatments. These results indicate the presence of a genotype by environment interaction in prepubertal HOL and RDC heifers in response to differences in feed ration treatment. Due to the influence of prepubertal growth on future milk production, reproduction, and health status, it is important to be aware of breed-specific requirements during the prepubertal period, particularly in mixed breed and crossbred groups, to optimize growth rates and production potential., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

The effects of high (2.5% of DM) versus normal dietary sodium chloride (NaCl) intake on renal urea-N kinetics and urea-N metabolism were investigated in 9 rumen-cannulated and multicatheterized lactating dairy cows in a crossover design with 21-d periods. It was hypothesized that urinary urea-N excretion would be greater, and BUN concentration lower in response to greater diuresis induced by high NaCl intake. Also, urea-N transport across ruminal and portal-drained viscera (PDV) tissues was hypothesized to be affected by dietary sodium intake. A second experiment was conducted using 8 lactating cows in a crossover design with 14-d periods to test high NaCl (2.5% of DM) versus high KCl (3.2% of DM) intake on milk yield and MUN concentrations. Experiment 1 showed that despite greater diuresis, high NaCl intake had no effect on urinary urea-N excretion or BUN concentration. The high NaCl intake did not affect rumen ammonia concentrations, total rumen VFA concentrations, ruminal venous - arterial concentration differences for ammonia, or ammonia absorption indicating that high NaCl did not adversely affect ruminal fermentation and microbial protein synthesis. High NaCl intake did not affect the total amount of urea-N transport from blood to gut, but ruminal venous - arterial concentration differences for urea-N were lower with high NaCl and ruminal extraction of arterial urea-N was numerically smaller, indicating that the ruminal epithelial urea-N transport was lower with high NaCl. Energy-corrected milk yield was greater with high NaCl (3.2 ± 1.5 kg/d); however, MUN concentrations were not affected by treatment. In experiment 2, ECM was greater with NaCl (1.4 ± 0.31 kg/d) compared with KCl (30.2 and 28.8 ± 0.91 kg ECM/d, respectively). Milk urea-N concentration was lower with KCl, suggesting a urea-N lowering effect in milk not evident with high NaCl intake. In conclusion, the present data show that dietary Na intake of 12 to 13 g/kg DM was followed by greater diuresis but did not affect urea-N excretion or BUN concentration. High NaCl intake did not affect the total amount of urea-N transfer across PDV tissues. Energy-corrected milk yield was greater with high NaCl compared with both control and feeding KCl; however, with KCl, MUN decreased., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Supplemental dietary rumen available fats show promise as enteric methane (eCH 4 ) mitigators for lactating dairy cows. However, concerns include variability in eCH 4 response and possible negative effects on dairy cow performance. Successful implementation of this mitigation option requires better prediction of responses specifically to rumen available fatty acids (FA) as well as understanding the modulating effects of other dietary and animal characteristics. Using meta-analytic and meta-regression techniques, 35 published studies with diet definition were used to assess changes in eCH 4 emissions and lactation performance associated with supplemental fat, specific supplemental rumen available FA types, and other dietary characteristics. Enteric CH 4 (g/d) was reduced by 3.77% per percentage unit of supplemental rumen available ether extract (EE). Supplemental rumen available PUFA (C18:2 and C18:3) and UFA (C18:1, C18:2, C18:3) mitigated eCH 4 (g/d) emissions in dairy cows by 6.88 and 4.65% per percentage unit increase, respectively. The anti-methanogenic effects of PUFA, MUFA and MCFA increased with correspondingly greater basal dietary levels of each FA type. Higher rumen-degradable starch (RDS; >18% DM) in the basal diet promoted greater reductions in eCH 4 yield (eCH 4 /DMI, g/kg) with supplemental rumen available PUFA and UFA. Both milk fat percentage and yield (kg/d) were reduced with rumen available fat supplementation with a reduction of 7.8% and 6.0%, respectively, relative to control diets. Our results highlight the importance of determining basal levels of the rumen available FA before providing supplemental rumen available FA as an option for enteric eCH 4 mitigation. Dairy nutritionists can use estimates generated from this analysis to predict changes in eCH 4 emissions and dairy cow performance associated with dietary supplementation of rumen available EE and specific rumen available FA types for the purpose of eCH 4 mitigation., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Fat supplements are well known for their multiple beneficial effects on ruminant health, reproduction and productivity, and as a source for certain bioactive compounds in ruminant products. On the other hand, numerous phytochemicals have demonstrated the potential to improve rumen fermentation through modifying the volatile fatty acid (VFA) pattern to favour those with greater energy efficiency, boosting microbial protein synthesis, and decreasing methane emission and ruminal ammonia concentration. Savory is an aromatic plant rich in various phytochemicals (mainly carvacrol and flavonoids) that can alter ruminal metabolism of dietary fatty acids, potentially increasing the production of some bioactive compounds such as conjugated linoleic acids (CLAs). This study aimed to investigate combined effects of oil type (fish oil (FO) versus soybean oil (SO)) and the inclusion of savory (Satureja khuzistanica) plant (SP) in the diet on total tract digestibility of nutrients, rumen fermentation, milk yield and milk fatty acid profile in dairy cattle. Eight multiparous lactating Holstein cows were used in a replicated 4 × 4 Latin square design experiment with four diets and four 21-d periods. During each experimental period consisted of 14 days of adaptation and a 7-day sampling period, cows were randomly assigned to one of the four dietary treatments: the diet supplemented with 2% (DM basis) fish oil (FOD) or soybean oil (SOD), the FOD or SOD plus 370 g DM/d/head SP (FODs, SODs, respectively). The experimental diets were arranged in a 2 × 2 factorial design, with the fat sources as the first and SP as the second factor. The FO-supplemented diets had lower dry matter intake (DMI) and higher total tract digestibility than SO-supplemented diets (P < 0.05), and including SP in the diet improved total tract digestibility of dry matter (DM), organic matter (OM), ether extract (EE), and non-fibrous carbohydrates (NFC) (P < 0.05) without negatively affecting DMI. Rumen pH was lower with SO than with FO diets (P < 0.01) and increased with SP inclusion in the diet (P < 0.05). Total protozoa count and ruminal ammonia concentration decreased, and the branched-chain VFA (BCVFA) proportion increased with SP inclusion in the diet (P < 0.05). Milk production, as well as the concentration and yield of milk components (except lactose concentration) were higher with SO than with FO diets (P < 0.05), but these variables remained unaffected by SP. The milk concentrations of both non-esterified fatty acids (NEFA) and beta-hydroxybutyrate (BHB) were lower with SO compared to FO diets, and these variables were reduced by SP (P < 0.01). The proportions of both mono- and polyunsaturated FA (MUFA and PUFA, respectively) in milk were higher with FO than with SO diets (P < 0.01), and their proportions increased by SP at the expense of saturated FA (SFA) (P < 0.01). Including SP in the diet increased the proportions of all the milk n-3 FA (C18:3c, C20:5, and C22:6) by 21%, 40%, and 97%, respectively, and those of conjugated linoleic acids (C18:2 (c9,t11-CLA) and C18:2 (t10,c12-CLA)) by 23% and 62%, respectively. There was no interaction between oil type and SP for the assessed variables. Fish oil, despite reducing milk production and milk components, was more effective than soybean oil in enriching milk with healthy FA. These findings also show promise for SP as a feed additive with the potential to improve total tract digestibility, rumen fermentation and milk FA composition., (© 2024. The Author(s).)

Cosmetically applicable soluble agonists for Toll-like receptor 2 (TLR2), which can strengthen skin barrier function, were produced by fermentation of asparagus ( Asparagus officinalis L.) extract supplemented with skimmed milk using Lactobacillus delbrueckii subsp. lactis TL24. Their molecular size was estimated to be >100 kDa. Their TLR2-stimulating activity was stable over 1 year at 4 °C, but it decreased by more than 95% within 10 and 4 months at 25 °C and 40 °C, respectively. The possibility of stabilization of TLR2-stimulating activity by powdering was tested, and we found that lyophilization with 10% or a higher amount of dextrin could stabilize the activity even at 40 °C. The powdered fermented product dose-dependently stimulated TLR2. It augmented the formation of tight junctions in normal human keratinocytes, as detected by fluorescence staining of occludin and ZO-1, whereas their protein and gene expression levels did not increase, suggesting that a change in subcellular localization of these proteins without significant changes in their amounts might be responsible. The powder nature has some benefits over the aqueous, besides stability, e.g., it can be dissolved just before application, allowing fresh material to be used each time, and it may widen a range of cosmetic applications in non-aqueous types of cosmetics.

Commercial dairy production occurs in a complex management environment, but increasingly, the dairy manager is expected to provide detailed reporting of productivity and environmental outcomes, for which conventional research methods double-blind crossover or case:control trials are inappropriate. This paper demonstrates the development of a milk protein production monitoring tool using a temporal (baseline) control in longitudinal, census-type investigations of modulation of system performance in response to factor change. It utilises farm-derived current and historical data, and contrasts seasonal responses with those achieved on neighbouring farms in a 2 × 2 contingency table. The approach is then shown to be useful in assessing the effect of two approaches to moderating milk urea concentration. Firstly, milk urea content can be monitored as it falls due to reduced feed protein content, and this fall can be arrested when milk protein content starts to decline relative to the value expected for the herd at any lactation stage. Secondly, by providing a dietary intervention aimed at increasing the availability of metabolic energy in the last month before calving, udder development can be augmented, leading to greater protein secretion capacity, meaning greater utilisation of circulating amino acids, and thus more limited substrate for urea synthesis. Thus, the changing impact of differing nutrition practices on dairy herd nitrogen excretion to environment can be followed with daily precision. In principle this approach can provide useful insights into a wide range of practical management interventions., Competing Interests: Graeme Coles is a director of Knewe Biosystems Ltd, the beneficial owner of Rich Technology Solutions Ltd. Jacqueline Rowarth is a farmer-elected director of Dairy NZ and of Ravendown Ltd. None of these entities have had any input into this submission., (©2024 Coles and Rowarth.)

Background: Despite the widespread adoption of TMR or PMR and the formulas designed to sufficiently cover the cows' requirements, individual dairy cows' milk production varies significantly. The liver is one of the most important organs in cow lactation metabolism and plays an essential role in the initiation of lactation., Objectives: This study aimed to investigate the potential key genes in the liver contributing to the different milk production., Methods: We enrolled 64 cows and assigned them to high or low milk yield (MY) groups according to their first 3 weeks of milk production. We performed RNAseq for 35 liver samples with 18 from prepartum and 17 from postpartum cows., Results: The continuous milk yield observation showed a persistently higher milk yield in high MY cows than low MY cows in the first 3 weeks. High MY cows showed better feed conversion efficiency. We identified 795 differentially expressed genes (DGEs) in the liver of high MY cows compared with low MY cows, with up-regulated genes linked to morphogenesis and development pathways. Weighted gene co-expression network analysis (WGCNA) revealed four gene modules positively correlating with milk yield, and protein and lactose yield ( p < 0.05). Using the intersected genes between the four gene modules and DEGs, we constructed the linear mixed-effects models and identified six hub genes positively associated and two hub genes negatively associated with milk yield (Coefficients > 0.25, p < 0.05). Random forest machine learning model training based on these eight hub genes could efficiently predict the milk yield ( p < 0.001, R 2 = 0.946). Interestingly, the expression patterns of these eight hub genes remained remarkably similar before and after parturition., Conclusions: The present study indicated the critical role of liver in milk production. Activated processes involved in morphogenesis and development in liver may contribute to the higher milk production. Eight hub genes identified in this study may provide genetic research materials for dairy cow breeding., Competing Interests: The authors declare no conflicts of interest.

MicroRNA (miRNA) is a type of endogenous non-coding small RNA, which is abundant in living organisms. miRNAs play an important role in regulating gene expression and myriad cellular processes by binding to target messenger RNAs through complementary base pairing, and cross-species regulation mammalian cells by plant-derived xeno-miRNAs has been described. Here, we examined the miRNA species in two alfalfa (Medicago sativa, lucerne) cultivars commonly grown in Ningxia, China: cv. Zhongmu 1 and cv. Xinyan 52. Both cultivars have good salt and drought resistance. We found that the miRNA profiles were similar between the cultivars, with a slightly higher number of miRNAs present in the newer cv. Xinyan 52, which may contribute to its improved salt and drought tolerance. miRNAs were stable during drying, and some miRNAs were increased in dry versus fresh alfalfa, suggesting some miRNAs may be upregulated during drying. Alfalfa-derived miRNAs could be detected in exosomes from serum and whey collected from dairy cows, confirming the ability of the exogenous miRNAs (xeno-miRNAs) to enter the circulation and reach the mammary epithelium. In vitro studies confirmed that overexpression of mtr-miR156a could downregulate expression of Phosphatase 2 Regulatory Subunit B'gamma ( PPP2R5D) and Phosphoinositide-3-kinase Regulatory Subunit 2 (PIK3R2). Overexpression of mtr-miR156a also modulated PI3K-AKT-mTOR signaling as well as the casein content of milk produced by bovine mammary epithelial cells. Based on the known roles of PPP2R5D and PIK3R2 in regulating the PI3K-AKT-mTOR pathway as well as the effect of PI3K-AKT-mTOR on milk protein content, our findings implicate alfalfa-derived miR156a as a new cross-species regulator of milk quality in dairy cows., (© 2024. The Author(s).)

Background: Goat milk is gaining popularity as a superior alternative to bovine milk due to its closer resemblance to human milk. Understanding the molecular processes underlying lactation is crucial for improving milk quality and production in goats. However, the genetic mechanisms governing lactation in goats, particularly in indigenous breeds like the Jakhrana, remain largely unexplored., Results: In this study, we performed a comprehensive transcriptomic analysis of Jakhrana goat mammary glands during early and late lactation stages. We isolated milk somatic cells and conducted RNA sequencing, followed by transcript quantification and mapping against the ARS1.2 Capra hircus reference assembly. Our analysis identified differentially expressed genes (DEGs) and commonly expressed genes (CEGs) across the lactation phases. Early lactation showed enrichment of genes encoding antimicrobial peptides and lubrication proteins, while late lactation exhibited heightened expression of genes encoding major milk proteins. Additionally, DEG analysis revealed upregulation of pivotal genes, such as the ABC transporter gene MRP4, implicated in modulating milk composition and quality., Conclusion: Our findings provide insights into the genetic mechanisms underlying lactation dynamics in the Jakhrana goat. Understanding these mechanisms could help in improving milk production and quality in goats, benefiting both the dairy industry and consumers., (© 2024. The Author(s).)

Obesity is a major global health concern. Studies suggest that the gut microflora may play a role in protecting against obesity. Probiotics, including lactic acid bacteria and Bifidobacterium , have garnered attention for their potential in obesity prevention. However, the effects of Bifidobacterium -fermented products on obesity have not been thoroughly elucidated. Bifidobacterium , which exists in the gut of animals, is known to enhance lipid metabolism. During fermentation, it produces acetic acid, which has been reported to improve glucose tolerance and insulin resistance, and exhibit anti-obesity and anti-diabetic effects. Functional foods have been very popular around the world, and fermented milk is a good candidate for enrichment with probiotics. In this study, we aim to evaluate the beneficial effects of milks fermented with Bifidobacterium strains on energy metabolism and obesity prevention. Three Bifidobacterium strains (Bif-15, Bif-30, and Bif-39), isolated from newborn human feces, were assessed for their acetic acid production and viability in milk. These strains were used to ferment milk. Otsuka-Long-Evans Tokushima Fatty (OLETF) rats administered Bif-15-fermented milk showed significantly lower weight gain compared to those in the water group. The phosphorylation of AMPK was increased and the expression of lipogenic genes was suppressed in the liver of rats given Bif-15-fermented milk. Additionally, gene expression related to respiratory metabolism was significantly increased in the soleus muscle of rats given Bif-15-fermented milk. These findings suggest that milk fermented with the Bifidobacterium strain Bif-15 can improve lipid metabolism and suppress obesity.

Background: Milk is essential for mammalian nutrition because it provides vital nutrients for growth and development. Milk composition, which is influenced by genetic and environmental factors, supports lactation, a complex process crucial for milk production and quality. Recent research has focused on noncoding RNAs, particularly microRNAs (miRNAs), which are present in body fluids and regulate gene expression post-transcriptionally. This study comprehensively characterizes miRNAs in milk of four livestock species, namely Bubalus bubalis, Capra hircus, Equus asinus, and Ovis aries and identifies potential target genes., Results: High-throughput sequencing of milk RNA resulted in distinct read counts across species: B. bubalis (8,790,441 reads), C. hircus (12,976,275 reads), E. asinus (9,385,067 reads), and O. aries (7,295,297 reads). E. asinus had the highest RNA mapping rate (94.6%) and O. aries the lowest (84.8%). A substantially greater proportion of miRNAs over other small RNAs was observed for the donkey milk sample (7.74%) compared to buffalo (0.87%), goat (1.57%), and sheep (1.12%). Shared miRNAs, which included miR-200a, miR-200b, miR-200c, and miR-23a among others, showed varying expression levels across species, confirmed by qPCR analysis. Functional annotation of predicted miRNA target genes highlighted diverse roles, with an enrichment in functions linked to metabolism and immunity. Pathway analysis identified immune response pathways as significant, with several miRNAs targeting specific genes across species, suggesting their regulatory function in milk., Conclusions: Both conserved and species-specific miRNAs were detected in milk of the investigated species. The identified target genes of these miRNAs have important roles in neonatal development, adaptation, growth, and immune response. Furthermore, they influence milk and meat production traits in livestock., (© 2024. The Author(s).)

The ABCG2 membrane transporter affects bioavailability and milk secretion of xenobiotics and natural compounds, including vitamins such as riboflavin. We aimed to characterize the in vitro and in vivo interaction of ABCG2 with lumichrome, the main photodegradation product of riboflavin, which has proven in vitro anti-cancer activity and a therapeutical role in antibacterial photodynamic therapy as an efficient photosensitizer. Using MDCK-II polarized cells overexpressing murine Abcg2 and human ABCG2 we found that lumichrome was efficiently transported by both variants. After lumichrome administration to wild-type and Abcg2 -/- mice, plasma AUC 20-120 min was 1.8-fold higher in Abcg2 -/- mice compared with wild-type mice. The liver and testis from Abcg2 -/- mice showed significantly higher lumichrome levels compared with wild-type, whereas lumichrome accumulation in small intestine content of wild-type mice was 2.7-fold higher than in Abcg2 -/- counterparts. Finally, a 4.1-fold-higher lumichrome accumulation in milk of wild-type versus Abcg2 -/- mice was found. Globally, our results show that ABCG2 plays a crucial role in plasma levels, tissue distribution and milk secretion of lumichrome potentially conditioning its biological activity.

Background: Mitochondrial genomes differ from the nuclear genome and in humans it is known that mitochondrial variants contribute to genetic disorders. Prior to genomics, some livestock studies assessed the role of the mitochondrial genome but these were limited and inconclusive. Modern genome sequencing provides an opportunity to re-evaluate the potential impact of mitochondrial variation on livestock traits. This study first evaluated the empirical accuracy of mitochondrial sequence imputation and then used real and imputed mitochondrial sequence genotypes to study the role of mitochondrial variants on milk production traits of dairy cattle., Results: The empirical accuracy of imputation from Single Nucleotide Polymorphism (SNP) panels to mitochondrial sequence genotypes was assessed in 516 test animals of Holstein, Jersey and Red breeds using Beagle software and a sequence reference of 1883 animals. The overall accuracy estimated as the Pearson's correlation squared (R 2 ) between all imputed and real genotypes across all animals was 0.454. The low accuracy was attributed partly to the majority of variants having low minor allele frequency (MAF < 0.005) but also due to variants in the hypervariable D-loop region showing poor imputation accuracy. Beagle software provides an internal estimate of imputation accuracy (DR2), and 10 percent of the total 1927 imputed positions showed DR2 greater than 0.9 (N = 201). There were 151 sites with empirical R 2  > 0.9 (of 954 variants segregating in the test animals) and 138 of these overlapped the sites with DR2 > 0.9. This suggests that the DR2 statistic is a reasonable proxy to select sites that are imputed with higher accuracy for downstream analyses. Accordingly, in the second part of the study mitochondrial sequence variants were imputed from real mitochondrial SNP panel genotypes of 9515 Australian Holstein, Jersey and Red dairy cattle. Then, using only sites with DR2 > 0.900 and real genotypes, we undertook a genome-wide association study (GWAS) for milk, fat and protein yields. The GWAS mitochondrial SNP effects were not significant., Conclusion: The accuracy of imputation of mitochondrial genotypes from the SNP panel to sequence was generally low. The Beagle DR2 statistic enabled selection of sites imputed with higher empirical accuracy. We recommend building larger reference populations with mitochondrial sequence to improve the accuracy of imputing less common variants and ensuring that SNP panels include common variants in the D-loop region., (© 2024. Crown.)

This study was performed to determine the influence of fish oil, an omega-3 fatty acids source, supplemented to diets of goats throughout all stages of gestation on the growth and milk production of weaned female kids. Eighty German Fawn (75%) x Hair (25%) crossbred goats were randomly assigned to treatment (fish oil, FiO group) and control (Rumen protected fat, RPF group) groups during the first half of pregnancy. Subsequently, the FiO group was further allocated into FiO-FiO and FiO-RPF subgroups and RPF group was further divided into RPF-FiO and RPF-RPF subgroups containing 20 goats in each during the second half of pregnancy. The growth and feed intake of 41 female kids (aged 75.1 ± 6.73 days, with a mean live weight of 11.6 ± 3.00 kg) were recorded for a 98 day post-weaning, In the continuation of the study, live weight changes, milk yield and composition of young female goats from mating to the second month of lactation and the growth of female kids until weaning were studied for a total of 210 days. Maternal nutrition slightly influenced the live weight gain of female kids over a 98-day investigation period (p = 0.070). When growth performance was considered, a higher feed conversion efficiency of female offspring was determined in RPF-FiO (5.52) treatment group compare to female kids in other treatment groups (p = 0.086). However, the maternal feeding system significantly affected live weight in the RPF-FiO treatment group during the mating period (P = 0.054). Concerning the feed intake, maternal nutrition significantly affected the feed intake of female kids (p < 0.01) with the highest feed consumption in the FiO-RPF group. The findings of this study have shown that fish oil enriched diet given to goats during gestation improved daily live weight changes and total live weight gain of female kids despite the initial disadvantage after weaning. At mating time, the live weight of young female goats in the RPF-FiO treatment group, which exhibited the highest feed conversion ratio during the 98-day study, was higher than the remaining treatment groups. Maternal nutrition had no effect on milk yield or milk components in young goats during lactation. Young female goats born to dams in the FiO-RPF group showed better performance than the other groups regarding live weight performance of their offspring on 56th day postpartum., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Erez, Serbester. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Selecting and raising dairy animals that are more likely to reach their potential is a strategy to increase milk production efficiency and overall profitability. However, indicators are necessary for the early identification of animals that are less likely to perform well, allowing for their early culling and ensuring that resources are allocated to those with the highest potential. The objective of this study was to analyze the association between early-life animal health and performance with longevity, production, and profitability. After data cleaning, the following early-life measures (i.e., predictors) were available for 363 female calves born between June 2014 and November 2015 in eight dairy herds from New Brunswick, Canada (average: 45 calves/farm; SD: 26.1 calves/farm; median: 42 calves/farm; range: 15-95 calves/farm): birth weight, weaning weight, weaning age, weaning average daily gain (weaning ADG), immunoglobulin G (IgG) serum concentration, the occurrence of navel infection, diarrhea, and pneumonia, and if animals received antibiotic treatment between birth and weaning. Their subsequent length of life (LL), length of productive life (LPL), lifetime cumulative energy-corrected milk (ECM), and lifetime cumulative milk value (i.e., response variables) were provided by the Canadian dairy herd improvement agency. Bayesian Additive Regression Tree models were trained for each response variable using 5-fold cross-validation. Models were evaluated using the RMSE and R 2 . The three most important predictors were identified using permutation, and the relationship between response variables and important predictors was assessed using accumulated local effect plots. The RMSE for LL, LPL, ECM, and milk value were 1.43 years, 1.37 years, 16 314.94 kg, and $CAD 11 525.68, respectively, whereas the R 2 values were 0.30, 0.25, 0.29, and 0.29, respectively, indicating a moderate relationship between predictors and response variables. Non-linear relationships were found between the response variables and important predictors. Animals born with low or high birth weights were associated with decreased LL, LPL, ECM, and milk value. The highest LL, LPL, and milk value was observed for calves weaned between 1.9 and 2.0 months old, followed by a decline for calves weaned at older ages. The lowest LL and ECM were associated with weaning ADG of 0.786 kg/day, while 0.787 kg/day was associated with the lowest LPL. Lastly, both ECM and milk value were highest when serum IgG values were 1 659 mg/dL. These findings provide valuable insights for optimizing early culling decisions and enhancing the productivity and profitability of dairy farms., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Reduced piglet feed intake immediately postweaning (pw) leads to disruption of small intestine structure and function and reduced growth. Our objective was to evaluate the effect of providing supplemental milk or liquid starter diet for either 4 or 11 days pw, on intestinal parameters of newly weaned piglets and growth to slaughter. At weaning (28 ± 0.6 days old), five hundred and eighty-seven piglets ((Large White × Landrace) × Duroc) were divided into 59 pen groups, each containing 9-10 same sex (entire male or female) piglets. The pen groups were blocked by sex and weaning weight and provided with ad-libitum access to one of five dietary treatments: (1) Dry pelleted starter diet (control; CON); (2) CON+liquid milk replacer for 4 days pw (M4); (3) CON+liquid milk replacer for 11 days pw (M11); (4) CON+liquid starter diet for 4 days pw (S4) and (5) CON+liquid starter diet for 11 days pw (S11). Pen groups were weighed at weaning, days 11, 20, 28, and 47 pw and at target sale weight. Feed disappearance (on a DM basis) was recorded on each weighing day. On day 7 pw, 10 piglets per treatment were euthanised to collect small intestine tissue samples for determination of villus height (VH), crypt depth and brush-border membrane enzyme activity. Data were analysed using SAS-version 9.4. Between days 0 and 11 pw, M11 increased average daily feed intake by 48% and average daily gain (ADG) by 57% compared to CON (P < 0.05), and increased ADG by 54% (P < 0.05) compared to S4. Piglets on M11 also had improved feed conversion efficiency compared with CON piglets between days 0 and 11 pw. Treatment did not affect growth performance after day 28 pw, or carcass parameters at slaughter. At day 7 pw, M11 piglets had 37% higher jejunal VH than CON piglets (P < 0.05) and S11 piglets had 28% higher ileal VH than S4 piglets (P < 0.05). M11 piglets had up to 150% higher ileal sucrase activity than M4, S4 and S11 piglets (P < 0.05) and 180% higher ileal maltase activity than S4 piglets (P < 0.05). In conclusion, M11 reduced the immediate negative effects of weaning, as it was associated with increased feed intake, growth, brush-border membrane enzyme activity and improved intestinal structure early pw. However, there were no carryover effects of any of the liquid supplements on growth or feed efficiency or carcass weight at slaughter., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Grazing management significantly contributes to low beef production in cow-calf systems within the Rio de la Plata native grasslands. An herbage allowance (HA) of 4 kg DM/kg BW increased the productive response of primiparous cows grazing shallow soils compared to 2.5. However, the impact of HA on metabolic changes and its association with productive response were not studied. We studied two levels of native grassland HA from -150 days relative to calving (DC) to weaning (195 DC) in spring-calving primiparous beef cows undergoing temporary weaning (TW) and flushing at 86 ± 12 DC on herbage intake (HI), body condition score (BCS), BW, milk yield, calf weight, concentrations of metabolic hormones, and the probability of ovulation and pregnancy. Thirty-one heifers were assigned to HA treatments that fluctuated throughout seasons: autumn (-150 to -90 DC) at 5 and 3 kg DM/kg BW, winter (-90 to 0 DC) at 3 and 3 kg DM/kg BW, and spring-summer (0 to 195 DC) at 4 and 2 kg DM/kg BW for High and Low HA, respectively. Data were analysed using linear models and generalised linear models for continuous and categorical variables, respectively. During the autumn period, HI, insulin, IGF-I, BCS, and BW were higher in High HA than Low, despite small differences in herbage mass between HA. Throughout the winter, spring, and summer, HI, insulin, leptin, and BCS changes did not differ between HA. However, IGF-I concentrations were greater at -65 and -40 DC (84 vs 55 ± 8.6 ng/mL; P < 0.05) and tended to be greater after TW in High HA than Low. The probability of ovulation did not reach significance (0.94 vs 0.75 ± 0.11 for High and Low HA, respectively; P = 0.125), while the probability of pregnancy was greater in High HA than in Low HA (0.9 vs 0.61 ± 0.10; P = 0.07). Ovulation probability exhibited a positive association with IGF-I concentrations at -90 and -40 DC (P < 0.05), but not postpartum. Milk yield did not differ between treatments, while calf weight was heavier at weaning in High HA cows (194 vs 178 ± 3.3 kg; P < 0.05). High HA enhances autumn HI and BCS and generates a carry-over effect on IGF-I concentrations throughout winter and after TW ("metabolic memory"), explaining the better reproductive response. Moderate changes in cows' nutrition during autumn contribute to changes in metabolic status and reproductive outcomes in primiparous cows grazing moderate herbage production native grasslands., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Increasing nutrient supply to dairy calves has well known benefits; however, the effects of milk replacer (MR) composition when supplied in higher amounts are not fully understood, particularly in the first weeks of life. To better understand the metabolism of macronutrient supply in young calves (21 d old), we investigated diurnal patterns of heat production and substrate oxidation in young calves fed MR with an incremental supply of fat, lactose, or protein. Thirty-two groups of 3 mixed-sex Holstein-Friesian newborn calves (3.4 ± 1.6 d of age), were randomly assigned to one of 4 dietary treatments and studied for 21 d. Diets consisted of a basal MR (23.3% CP, 21.2% EE, and 48.8% lactose of DM) fed at 550 kJ/kg BW 0.85 per day (CON; n = 24), or the basal MR incrementally supplied with 126 kJ of digestible energy/BW 0.85 per day as milk fat (+FAT; n = 23), lactose (+LAC; n = 24), or milk protein (+PRO; n = 23). Calves were fed MR in 2 daily meals and had ad libitum access to water, but were not supplied with any calf starter nor forage. After 2 weeks of adaptation to the diets, groups of 3 calves were placed for 1 wk in an open-circuit respiration chamber for nitrogen and energy balance measurements (lasting 7 d). On d 3, glucose oxidation kinetics was estimated by using [U- 13 C]glucose. Measurements included total heat production (total energy [HP], activity [H act ] expenditure, resting metabolic rate [RMR]), respiration quotient (RQ), carbohydrate (COX) and fat oxidation (FOX) in 10 min. intervals and averaging these values per hour over days. Incremental supply of lactose and fat increased body fat deposition, with observed patterns in RMR indicating that this increase occurred primarily after the meals. Specifically, the average daily RMR was highest in the +PRO group and lowest in the CON treatment. The HP was higher in the +PRO group and throughout the day, hourly means of HP were higher in this treatment mainly caused by an increase in H act . The recovery of 13 CO 2 from oral pulse-dosed [U- 13 C]glucose was high (77%), and not significantly different between treatments, indicating that ingested lactose was oxidized to a similar extent across treatments. Increasing lactose supply in young calves increased fat retention by reduction in fatty oxidation. Calves fed a MR with additional protein or fat raised RMR persistently throughout the day, whereas the extra lactose supply only affects RMR after the meal. Dietary glucose was almost completely oxidized (77% based on ( 13 C) glucose measurement) regardless of nutrient supplementation. Extra protein supply increased HP and FOX compared with similar intakes of fat and lactose. Fasting heat production (FHP) of young, group-housed calves is comparable to literature values and unaffected by energy intake. Overall, these findings deepen our understanding of how different nutrients affect metabolic processes, fat retention, and energy expenditure in young dairy calves., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

We investigated the short- and long-term effects of different forage types supplemented in preweaning dairy calves on growth performance, blood metabolites, rumen fermentation, bacterial community, and milk production during first lactation. A total of 60 healthy 30-d-old female Holstein calves were blocked by birth date and body weight and randomly assigned to 1 of 3 groups (n = 20): normal milk and pelleted starter feeding (CON), supplemented with chopped oat hay (OAH; 75.0 g/d per calf [DM basis]), or alfalfa hay (ALF; 75.0 g/d per calf [DM basis]). The forage supplementation started when calves were 30 d old (d 1 of the experimental period) and ended when they were 73 d old (d 44 of the experimental period, when calves were weaned). Milk and feed intakes and fecal consistency scores were recorded daily. Growth performance, rumen fluid, and blood samples were collected biweekly. After weaning, all the calves were integrated with the same barn and diets. After calving, the milk production was recorded daily. During the experimental period, the OAH group had greater solid feed and total DM intakes and greater rumen pH than the CON group (P ≤ 0.04), but had lower forage intake and CP digestibility than the ALF group (P ≤ 0.04). The ALF group had higher rumen pH and blood BHB concentration (P ≤ 0.04), lower fecal score (P = 0.02), and greater ether extract digestibility (P = 0.02) than the CON group. The ALF and OAH groups had lower concentrations of ruminal total VFA (P = 0.01). Still, the ALF group had a greater proportion of acetate and a relative abundance of cellulose degradation-related bacteria (Lachnoclostridium&#95;1 and Oribacterium) and a lower relative abundance of inflammation-related bacteria (Erysipelotrichaceae&#95;UCG-009) in the rumen compared with CON. Interestingly, the average milk production from 6 to 200 DIM was greater in the ALF group (P < 0.01), even though no significant effects were found on the rumen fermentation parameters and blood metabolites at 200 DIM. Generally, alfalfa hay supplementation in preweaning dairy calves had positive effects in the short- and long-term for rumen development, health status, and future milk production., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)

Reduced feed intake in early lactation prompts increased fat mobilization to meet dairy cow energy needs for milk production. The increased lipolysis in cows presents significant health risks with unclear mechanisms. The objectives of our study were to compare the longitudinal profiles of metabolites and lipids of serum from high- and low-lipolysis cows. Forty multiparous Holstein dairy cows were enrolled in the retrospective study. Serum samples were collected on d 7 before expected calving, as well as on d 5, 7, 14, and 21 postpartum. Dairy cows were grouped according to mean serum nonesterified fatty acids on d 5 and 7 after parturition as low (<0.600 mmol/L; n = 8; LFM) and high (>0.750 mmol/L; n = 8; HFM), indicating fat mobilization during early lactation. Lactational performance and serum metabolic parameters related to glucose and lipid metabolism, liver functions, oxidative status, and inflammatory responses were determined. Serum samples were subjected to liquid chromatography-MS-based metabolomics and lipidomics. Despite differences in postpartum BW change, there were no observed variations in milk yield and composition between the 2 groups. Serum β-hydroxybutyric acid, glucose, leptin, aspartate aminotransferase, IL-6, and tumor necrosis factor alpha were greater in cows with HFM than in cows with LFM. Serum adiponectin, revised quantitative insulin sensitivity check index, and albumin were lower in cows with HFM than in cows with LFM. Intensified fat mobilization in the HFM cows came along with reduced estimated insulin sensitivity, impaired liver functions, and increased oxidative stress and inflammatory responses. Differences in metabolic patterns were observed across the transition period when comparing serum blood matrixes (e.g., in different amino acids, acylcarnitines, and sphingolipids). The serum metabolome of the HFM cows was characterized by higher concentrations of glycine, acylcarnitines, carnosine, Cer(d20:0/18:0), Cer(d18:1/16:0), and Cer(t18:0/24:0) compared with LFM cows. The differential serum metabolites and lipids at different sampling times during the peripartum period were enriched in the sphingolipid metabolism. Differences in serum metabolic status parameters suggest that cows adopt varied metabolic adaptation strategies to cope with energy deficits postpartum. Our investigation found a comprehensive remodeling of the serum metabolic profiles in transition dairy cattle, highlighting the significance of alterations in sphingolipid species, as they play a crucial role in insulin resistance and metabolic disorders., (The Authors. Published by Elsevier Inc. on behalf of the American Dairy Science Association®. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).)