Abstract Essential amino acids (EAA) may alter bovine mammary epithelial cell (BMEC) activities through regulation of gene transcription. For this study, we used pathway analysis of RNA-sequencing data to identify upstream transcription factors (TF) that potentially mediate the effects of single EAA deficiency on BMEC. Differentiated BMEC were cultured in 1 of 4 treatment media representing normal physiological concentrations of all amino acids (CTL), or ¼ the normal concentration of histidine (LH), lysine (LK), or methionine (LM). After 48 h, rates of DNA synthesis were reduced 30 to 40% by each of the deficiencies. Protein synthesis rate was 30 to 50% lower in LK and LM but was unaffected by LH. Up-regulated upstream TF included those related to nutrient stress, interferon signaling, and suppression of cell population growth. Down-regulated TF were primarily related to cell cycle progression. An array of classic ATF4 targets increased in response to EAA deficiencies including amino acid transporters, branched-chain aminotransferase, aminoacyl-tRNA synthetases, sestrin2 (a leucine sensor), and activators of apoptosis. Interferon signaling emerged as a novel pathway activated by EAA deficiencies, possibly contributing to reduced cell proliferation and protein synthesis. In contrast, the MAPK signaling pathway was unaffected. Expression of hypoxia-inducible factor 1α (HIF1A) increased, consistent with an anti-proliferative role. Downregulation of Forkhead box protein M1 (FOXM1) likely contributed to slower cell proliferation. These findings reveal a complex network of transcriptional regulators and signaling pathways influenced by EAA availability and shed light on the nutritional regulation of milk protein production by dairy cows.

Skeletal muscle is vital in maintaining metabolic homeostasis and adapting to the physiological needs of pregnancy and lactation. Despite advancements in understanding metabolic changes in dairy cows around calving and early lactation, there are still gaps in our knowledge, especially concerning muscle metabolism and the changes associated with drying off. This study aimed to characterize the skeletal muscle metabolome in the context of the dietary and metabolic changes occurring during the transition from the cessation of lactation to the resumption of lactation in dairy cows. Twelve Holstein dairy cows housed in tiestalls were dried off 6 wk before the expected calving date. Cows were individually fed ad libitum TMR composed of grass silage, corn silage, and concentrate during lactation and of corn silage, barley straw, and concentrate during the dry period. The metabolome was characterized in skeletal muscle samples (M. longissimus dorsi) collected on wk -7 (9 d before dry-off), -5 (6 d after dry-off), and wk -1, and wk 1 relative to calving. The targeted metabolomics approach was conducted using the MxP Quant 500 kit (Biocrates Life Sciences AG) with liquid chromatography, flow injection, and electrospray ionization triple quadrupole mass spectrometry. Statistical analysis on the muscle metabolite data was performed using MetaboAnalyst 5.0, which allowed us to conduct various multivariate analyses such as principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), informative heat map generation, and hierarchical clustering. The statistical analysis revealed a clear separation between pregnancy (wk -7, -5, and -1) and postcalving (wk 1). Starting 5 wk before calving and continuing through the first week thereafter, the concentration of 3-methylhistidine (3-MH) in the muscle increased. This coincided with an increase in the concentrations of 11 AA (Phe, His, Tyr, Trp, Arg, Asn, Leu, Ile, Gly, Ser, and Thr) in the first week after calving, whereas Gln decreased. l-Arginine pathway metabolites (homoarginine, ornithine, citrulline, and asymmetric dimethylarginine), betaine, and sarcosine followed a similar pattern, increasing from wk -7 to -5, but decreasing from wk -1 to 1. The transition from pregnancy to lactation was associated with an increase in concentrations of the long-chain acylcarnitine species C16, C16:1, C18, and C18:1 in the muscle, whereas the concentrations of phosphatidylcholine and sphingomyelin in the muscle remained stable. The significant changes observed in the metabolome mainly concerned the AA and AA-related metabolites, indicating muscle protein breakdown in the first week after calving. The metabolites produced by the l-Arg pathway might contribute to regulating skeletal muscle mass and function in periparturient dairy cows. The elevated concentrations of long-chain acylcarnitine species in the muscle in the first week after calving suggest incomplete fatty acid oxidation, likely due to insufficient metabolic adaptation in response to the fatty acid load around the time of calving.

Dairy cows undergo profound metabolic challenges as they transition from lactation cessation to lactation resumption. Adipose tissue (AT), serving as the primary energy reserve and an active endocrine organ, plays a crucial role in these adaptations. Thus, the objective of the current study was a comprehensive characterization of the metabolic changes in the AT metabolome of Holstein dairy cows as they transitioned from one lactation cycle to the next, providing key insights into the dynamic adaptations crucial for maintaining energy homeostasis and optimizing lactational performance. Twelve Holstein dairy cows (BW = 745 ± 71 kg, BCS = 3.43 ± 0.66), housed in tiestalls, were dried off 6 wk before their expected calving date (mean dry-off time = 42 d). Cows were individually fed ad libitum TMR, consisting of grass silage, corn silage, and concentrate during lactation and a mixture of corn silage, barley straw, and concentrate during the dry period. The metabolome was characterized in subcutaneous AT samples collected on wk -7 (before drying off), -5 (after drying off), and wk -1 and 1 relative to calving. A targeted metabolomics approach was employed using the MxP Quant 500 kit (Biocrates Life Sciences AG), integrating liquid chromatography, flow injection, and electrospray ionization triple quadrupole MS. Statistical analysis of the AT metabolite data was conducted using MetaboAnalyst 5.0, enabling various multivariate analyses, including principal component analysis, partial least squares discriminant analysis, hierarchical clustering, and the generation of informative heatmaps. Multivariate analyses revealed distinct and dynamic alterations in the AT metabolome, with minimal changes during the early dry period (wk -7 to -5) followed by pronounced metabolic reprogramming close to calving (wk -1 to 1). Amino acid profiles in AT remained stable during late gestation but declined significantly in Ala, Asp, and Gln between wk -1 and wk 1, likely due to increased utilization within AT, redirecting carbon skeletons from these AA toward glyceroneogenesis and the re-esterification of fatty acids (FA) into triglycerides. Such a shift in AA metabolism may also facilitate interorgan nutrient exchange, with Ala export through the glucose-Ala cycle providing essential gluconeogenic substrates to the liver during early lactation. Moreover, acylcarnitine profiles remained unchanged, reflecting the role of AT as a long-term lipid reservoir rather than a metabolically active site for FA oxidation. The data revealed a biphasic pattern in diglycerides and extensive remodeling of phosphatidylcholines, underscoring dynamic cellular membrane adaptations to heightened lipolytic activity and increased energy demands during the immediate postpartum period. Notably, sphingomyelin remained stable throughout the transition, suggesting potential mechanisms in preserving membrane integrity and ensuring cellular stability under fluctuating metabolic stress. Together, these data further support that AT functions not merely as a passive energy store but as a dynamic organ actively orchestrating metabolic homeostasis during the transition from lactation cessation to lactation resumption.

This study aimed to investigate the metabolic changes in the livers of dairy cows from 1 wk before dry-off to 1 wk after calving. Twelve high-yielding Holstein cows were included in a longitudinal study and housed in a tie-stall barn. The cows were dried off at 6 wk before the expected calving date (dry period length = 42 d). During the entire lactation, the cows were milked twice daily at 0600 and 1700 h. Liver biopsies were taken from each cow at 4 different times: wk −7 (before drying off), −5 (after drying off), −1 and +1 relative to calving. A targeted metabolomics approach was performed by liquid chromatography and flow injection with electrospray ionization triple quadrupole mass spectrometry using the MxP® Quant 500 kit. A total of 185 metabolites in the liver were used for the final data analysis. Principal component analysis (PCA) revealed a clear separation by days of sampling, indicating a notable shift in metabolic phenotype from late lactation to dry period and further changes after calving. Changes were observed in several classes of compounds, including amino acids and biogenic amines. In particular, the changes in acylcarnitines (AcylCN), phosphatidylcholines (PC), sphingomyelins (SM) and bile acids (BA) indicated extensive remodeling of the hepatic lipidome. The changes in AcylCN concentrations in early lactation suggest incomplete fatty acid oxidation in the liver, possibly indicating mitochondrial dysfunction or enzymatic imbalance. In addition, the changes in PC and SM species in early lactation indicate altered cell membrane composition, which may affect cell signaling and functionality. In addition, changes in BA concentrations and profiles indicate dynamic adaptations in BA synthesis, lipid digestion and absorption during the observation period. In particular, PCA analysis showed an overlapping distribution of liver metabolites in primiparous and multiparous cows, indicating no significant difference between these groups. In addition, Volcano plots showed similar liver metabolism between primiparous and multiparous cows, with no significant fold changes (>1.5) in any metabolite at significant P-values (false discovery rate <0.05). These results provide valuable insight into the physiological ranges of liver metabolites during dry period and calving in healthy dairy cows and should contribute to the design and interpretation of future metabolite-based studies of the transition dairy cow.

While there is generally no consensus about how nutrients determine milk synthesis in the mammary gland, it is likely that the mechanistic target of rapamycin (mTOR) complex 1 (mTORC1) plays a role as a key integrator of nutritional and mitogenic signals that can influence a multitude of catabolic and anabolic pathways. The objectives of this study were to evaluate acute changes (<24 h) in translational signaling, in addition to chronic changes (14 d) in mammary gland structure and composition, in response to a severe feed restriction. Fourteen lactating Holstein dairy cows were assigned to either ad libitum feeding (n = 7) or a restricted feeding program (n = 7). Feed-restricted cows had feed removed after the evening milking on d 0. Mammary biopsies and blood samples were collected 16 h after feed removal, after which cows in the restricted group were fed 60% of their previously observed ad libitum intake for the remainder of the study. On d 14, animals were slaughtered and their mammary glands dissected. In response to feed removal, an acute increase in plasma nonesterified fatty acid concentration was observed, concurrent to a decrease in milk yield. In mammary tissue, we observed downregulation of the mTORC1-S6K1 signaling cascade, in addition to reductions in mRNA expression of markers of protein synthesis, endoplasmic reticulum biogenesis, and cell turnover (i.e., transcripts associated with apoptosis or cell proliferation). During the 14 d of restricted feeding, animals underwent homeorhetic adaptation to 40% lower nutrient intake, achieving a new setpoint of 14% reduced milk yield with 18% and 29% smaller mammary secretory tissue DM and CP masses, respectively. On d 14, no treatment differences were observed in markers of protein synthesis or mammary cell turnover evaluated using gene transcripts and immunohisto chemical staining. These findings implicate mTORC1-S6K1 in the early phase of the adaptation of the mammary gland's capacity for milk synthesis in response to changes in nutrient supply. Additionally, changes in rates of mammary cell turnover may be transient in nature, returning to basal levels following brief alterations that have sustained effects.

Twenty-one beef heifers were used in a randomized complete block design lasting 206 ± 8 day (mean ± SD) to evaluate dry matter intake (DMI), growth, and gastrointestinal tract and carcass characteristics with increasing dose of dietary hydrogenated fat-embedded calcium gluconate (HFCG). Dietary treatments included HFCG included at: 0.0% (CON), 0.09% (CG09), or 0.18% (CG18) DM. Heifers were fed individually and fecal samples were collected on d 171 and 192. Gastrointestinal tissue, digesta, carcass merit, and meat quality and sensory characteristics were assessed after slaughter. Data were analyzed to determine the linear and quadratic effects of HFCG dose. DMI, average daily gain (ADG), and G:F were not affected ( P ≥ 0.08) by dose of HFCG. Gastrointestinal fermentation was minimally affected, while quadratic responses ( P = 0.03 and 0.01, respectively) were observed for jejunal digesta pH and the molar proportion of isovalerate in colonic digesta where both increased and then decreased as HFCG dose increased. Carcass ( P ≥ 0.30), meat quality ( P ≥ 0.06), and sensory characteristics ( P ≥ 0.21) were not affected by dose of HFCG. We conclude that feeding HFCG to finishing beef heifers did not affect DMI, growth, gastrointestinal fermentation parameters, or carcass merit, meat quality, and palatability.

It has been established that essential amino acids (EAA) regulate protein synthesis in mammary epithelial cells by rapidly altering the phosphorylation state of translation factors. However, the long-term transcriptional response to EAA supply has been investigated much less. Eight transcription factors were selected as candidate mediators of EAA effects on mammary cell function via the amino acid response ( ATF4 , ATF6 ), mitogen-activated protein kinase ( JUN , FOS , EGR1 ), and mechanistic target of rapamycin complex 1 ( MYC , HIF1A , SREBF1 ). The objective was to determine if and when expression of these candidate genes was affected in primary cultures of bovine mammary epithelial cells more than 24 h after imposing an EAA deficiency, and to evaluate effects of EAA deficiency on protein synthesis, endoplasmic reticulum size, cell proliferation, and lipogenesis. Differentiated cells were cultured in 1 of 3 treatment media representing normal physiological concentrations of all amino acids (CTL), low lysine (LK), or low methionine (LM) for 24, 40, 48, or 60 h. Both LK and LM suppressed protein synthesis and activated ATF4 expression, indicating the classic amino acid response pathway had been triggered. However, there was no effect of LK or LM on endoplasmic reticulum size, possibly related to elevated ATF6 expression on LM. Expression of early response genes JUN , FOS , EGR1 and MYC was not elevated by EAA deficiency but LM decreased EGR1 expression. LM also increased expression of HIF1A . The EGR1 and HIF1A expression results are consistent with the decrease in cell proliferation rate observed. Variable responses in SREBF1 expression to LK and LM at different timepoints may have contributed to a lack of effect on lipogenesis rates. These findings indicate that EAA deficiency may inhibit mammary protein synthesis and cell proliferation through transcription factors.

The objective of this pilot study was to generate data to support the development of an experimental model of hindgut acidosis to further understand its systemic consequences independently of rumen acidosis. Four ruminally fistulated multiparous Holstein cows (213 +/- 11 d in milk) were subjected to 2 consecutive experimental periods (P1 and P2), separated by a 3-d washout. Experimental periods were 96 h long from the baseline to the final measurements but expanded over 5 calendar days (d 0-4). Abomasal infusions of saline and corn starch (2.8 kg/d) were performed for the first 72 h (d 0-3) of P1 and P2, respectively. Final measurements were performed 24 h after the end of the infusions (d 4). Each cow was used as its own control by comparing P2 to P1. Postruminal-intestinal permeability was assessed by Cr appearance in blood after a pulse dose administration of Cr-EDTA into the abomasum on d 2 (48 h after infusion initiation) of each period. Starch infusion during P2 was associated with a milk protein yield increase (3.3%) and a decrease in milk urea nitrogen (11%). Fecal dry matter increased (8.8%), and starch content tended to increase (similar to 2 fold) during P2. There was a period-by-day interaction for fecal pH as it decreased during starch infusion (1.3 pH points) but remained constant during P1. Although fecal lactate was not detectable during P1, it consistently increased during starch infusion. Fecal alkaline phosphatase activity also increased (similar to 17 fold) in association with starch infusion. Two hours after Cr-EDTA administration, blood Cr concentration was higher during starch infusion, resulting in a tendency for a treatment-by-hour interaction. Furthermore, blood d-lactate increased (similar to 2.5 fold), serum Cu decreased (18%), and blood urea nitrogen, cholesterol, and Ca tended to decrease (9.4%, 1.2%, and 2.4%, respectively), relative to P1. The current results suggest that hindgut acidosis was successfully induced by postruminal starch infusion, leading to gut damage and increased intestinal permeability. However, indications of systemic inflammation were not observed. The herein described preliminary results will require confirmation in a properly powered study. Key words: leaky gut, hindgut acidosis, transition period

Streptococcus suis (S. suis) is an endemic zoonotic pathogen still lacking adequate prevention in pigs. The present case study looked back to the occurrence and consequences of S. suis outbreaks in our swine research facilities in search of new metabolic and physiological insight. From a series of outbreaks, a dataset was created including 56 pigs sampled during disease detection based on clinical signs. Pigs suspected with S. suis infection were defined as diseased (n = 28) and included pigs defined as neurologically diseased (n = 20) when severe neurological signs (central nervous system dysfunctions, i.e., opisthotonos, ataxia, and generalized tremor) were observed. Another set of 28 pigs included respective pen mates from each case and were defined as control. Representative deaths were confirmed to be caused by S. suis. Tonsillar swabs were collected and analyzed by quantitative polymerase chain reaction (qPCR) for total bacteria, total S. suis, and S. suis serotypes (SS) 2 (and/or 1/2) and 9. Blood and sera were analyzed to quantify blood gases, minerals, and S. suis reactive immunoglobulins against current isolates. Data collected included litter sibling associations, birth and weaning body weight (BW), and average daily gain (ADG) 7 d after the disease detection. In general, the disease increased pH, sO2 and the incidence of alkalosis, but reduced pCO2, glucose, Ca, P, Mg, K, and Na in blood/serum compared to control. The SS2 (and/or SS1/2) prevalence was significantly (P < 0.05) increased in neurologically diseased pigs and its relative abundance tended (P < 0.10) to increase in tonsils. In contrast, the relative abundance of total S. suis was lower (P > 0.05) in diseased pigs than control pigs. Levels of S. suis reactive IgG2 were lower, but IgM were higher (P < 0.03) in neurologically affected pigs compared to control. Furthermore, there was an increased proportion of sibling pigs that were diseased compared to control. In conclusion, our results evidence that naturally affected pigs were associated to average performing pigs without any predisease trait to highlight but a sow/litter effect. Besides, neurologically affected pigs had increased S. suis (SS2 and/or 1/2) prevalence and relative abundance, a respiratory alkalosis profile, and mineral loss.

The objective of this study was to compare the effects of post-ruminal provision of Ca-butyrate (CaB) when delivered via abomasal dosing, and Ca-gluconate (CaG) when provided ruminally using a rumen protected form or using an unprotected form via abomasal dosing on short-chain fatty acid (SCFA) concentration throughout the GIT, nutrient digestibility, GIT barrier function, ruminal SCFA absorption, ruminal morphometrics, intestinal brush border enzyme activity, and blood parameters for beef heifers. Thirty-two beef heifers fitted with ruminal cannulas were used in a randomized complete block design and assigned to one of four treatments: 1) negative control (ruminal infusion of double-distilled water; CON); 2) abomasal infusion of CaB (AB; 0.0029% of BW); 3) abomasal infusion of CaG (AG; 0.0077% of BW); and 4) ruminal infusion of a hydrogenated fat-embedded CaG (RG; 0.0192% of BW) to provide ruminal protection. Excluding CON, treatments were designed to deliver the same amount of butyrate in the small intestine. Heifers were housed in individual pens and DMI was limited to 95% of voluntary intake to minimize a potential confounding effect of DMI on treatment responses. Total GIT barrier function was assessed on day 17 and SCFA disappearance was evaluated on day 21 using the temporarily isolated and washed reticulo-rumen technique. On day 28, heifers were slaughtered, and ruminal and colonic digesta were collected to assess SCFA concentration. Additionally, ruminal, jejunal, and colonic tissues were collected to assess SCFA fluxes and regional barrier function ex vivo using the Ussing chamber technique. For colonic digesta, both AB and CaG treatments reduced the proportion of acetate (P < 0.05) and increased the proportion on propionate (P < 0.05) compared to CON. Relative to CON, AB but not CaG treatments increased in vivo ruminal disappearance of total SCFA (P = 0.01), acetate (P = 0.03), propionate (P = 0.01), and butyrate (P > 0.01). Treatments did not affect (P ≥ 0.10) acetate and butyrate fluxes in the ruminal and colonic tissues when measured ex vivo; however, when compared with CON, AB tended to decrease (P = 0.09) mannitol flux across ruminal tissue. In addition, mannitol flux was affected (P < 0.01) by region, with greater mannitol flux across the jejunum than rumen and colon. We conclude that while both abomasal infusion of CaB and CaG affect the molar proportion of acetate and propionate in the colon, only abomasal CaB stimulated ruminal SCFA absorption for growing beef heifers.

Prebiotic compounds may be supplemented in the diet to improve animal health and performance in a variety of ways. In dairy cattle, the transition from pregnancy through parturition and lactation represents a critical life stage with many concurrent stressors. The objectives of this study were to evaluate responses to the provision of a hindgut-targeted prebiotic compound (calcium gluconate; HFCG) when supplemented prepartum and/or postpartum in a 2 x 2 factorial design. One hundred and sixty-four multiparous Holstein cattle were enrolled and followed from approximately 21 d prior to calving until 100 d of lactation. Treatments were administered as a pelleted compound feed offered in the rotary milking parlor once daily prepartum and thrice daily postpartum. Information pertaining to milk production and body weight were automatically recorded by the milking equipment, and information pertaining to reproductive and health performance was recorded by farm staff. Cattle that received HFCG prepartum were confirmed pregnant approximately 21 d earlier (P = 0.024). Cattle that received HFCG both pre- and postpartum had 9% to 10% higher yields of milk protein, fat, and energy-corrected milk (P & LE; 0.037) from weeks 4 to 9 of lactation relative to those that received HFCG exclusively prepartum. Conversely, cattle that received HFCG exclusively postpartum had 9% to 10% higher yields of milk protein, fat, and energy-corrected milk (P & LE; 0.037) from weeks 9 to 14 of lactation relative to those that received exclusively the negative control in both periods. The mechanism underlying these responses remains unclear, however, we hypothesize that these responses are due to localized reductions in inflammation in the gut and/or signaling to extragastrointestinal tissues altering energy partitioning and balance.

A 305-d lactation followed by a 60-d dry period has traditionally been considered economically optimal, yet dairy cows in modern intensive dairy systems are frequently dried off while still producing significant quantities of milk. Managing cows for an extended lactation has reported production, welfare and economic benefits, but not all cows are suitable for an extended lactation. Implementation of an extended lactation strategy on-farm could benefit from use of a decision support system, based on a mathematical lactation model, that can identify suitable cows during early lactation that have a high likelihood of producing above a target milk yield (MY) at 305 d in milk (DIM). Therefore, our objectives were 1) to compare the suitability of 3 commonly used lactation models for modeling extended lactations (Dijkstra, Wood, and Wilmink) in primiparous and multiparous cows under a variety of lactation lengths, and 2) to determine the amount of early lactation daily MY data needed to accurately forecast MY at d 305 by using the most suitable model and determine if this is sufficient for identifying cows suitable for an extended lactation before the end of a typical voluntary waiting period (50 to 90 d). Daily MY data from 467 individual Holstein-Friesian lactations (DIM >305 d; 379 ± 65 d lactation length [mean ± SD]) were fitted by the 3 lactation models using a nonlinear regression procedure. The parameter estimates of these models, lactation characteristics (peak yield, time to peak yield, and persistency), and goodness-of-fit were compared between parity and different lactation lengths. The models had similar performance and differences between parity groups were consistent with previous literature. Then, data from only the first i DIM for each individual lactation, where i was incremented by 30 d from 30 to 150 DIM and by 50 d from 150 to 300 DIM, were fitted by each model to forecast MY at d 305. The Dijkstra model was selected for further analysis as it had superior goodness-of-fit statistics for i = 30 and 60. The data set was fit twice by the Dijkstra model, with parameter bounds either unconstrained or constrained. The quality of predictions of MY at d 305 improved with increasing data availability for both models and assisting the model fitting procedure with more biologically relevant constraints on parameters improved the predictions, but neither was reliable enough for practical use on-farm due to the high uncertainty of forecasted predictions. Using 90 d of data, the constrained model correctly classified 66% of lactations as being above or below a target MY at d 305 of 25 kg/d, with a probability threshold of 0.95. The proportion of correct classifications became smaller at lower targets of MY at d 305 and became greater when using more lactation days. Overall, further work is required to develop a model that can forecast late lactation MY with sufficient accuracy for practical use. We envisage that a hybridized machine learning and mechanistic model that incorporates additional historical and genetic information with early lactation MY could produce meaningful lactation curve forecasts.

This research communication reports the responses to supplementing dairy cattle with a hydrogenated fat-embedded calcium gluconate feed additive. The role of hindgut health in ruminant performance and wellbeing is an area of growing interest. Various prebiotic compounds have been used to promote lower gut health in various non-ruminant species. Calcium gluconate, a prebiotic compound, has previously been observed to increase milk fat yield when fed to ruminants in a form capable of resisting fermentation in the rumen, though the mechanism(s) behind this response remain unclear. The objective of this study was to compare the responses of lactating cattle to two different supplementation levels of a hydrogenated fat-embedded calcium gluconate (HFCG) product to evaluate a potential linear dose response. Forty-six lactating Holstein dairy cattle were used in a 3 × 3 replicated Latin square design with 28 d periods to evaluate a previously used dose of HFCG (approximately 16 g/d) with both a negative control and a dose of 25 g/d. Supplementation of multiparous animals with 16 g/d HFCG significantly (P < 0.05) increased milk fat yield and content relative to the negative control, and subsequently improved gross feed efficiency (P < 0.05); additionally, the presence of a potential non-linear dose response was observed for these parameters. Responses when supplemented with 25 g/d HFCG did not differ from the negative control. No production responses were observed in primiparous animals. The mode of action of HFCG, in addition to the potential differential response in primiparous animals remains unclear and warrants further investigation.

BACKGROUND:Trace metals are supplemented in cattle to prevent nutrient deficiencies. Levels supplemented to mitigate worst-case basal supply and availability scenarios can, however, result in trace metal intakes far above the nutritional requirements of dairy cows with high feed intakes. OBJECTIVES:We evaluated Zn, Mn, and Cu balance in dairy cows from late lactation through the subsequent mid-lactation, a period of 24 wk characterized by large changes in dry matter intake. METHODS:Twelve Holstein dairy cows were housed in a tie-stall from 10 wk before to 16 wk after parturition and fed 1 unique lactation diet when lactating and a dry cow diet otherwise. After 2 wk of adaptation to the facility and diet, Zn, Mn, and Cu balances were determined at weekly intervals, by calculating the difference between total intakes and complete fecal, urinary, and milk outputs, with the latter 3 fluxes quantified over a 48-h period. Repeated measure mixed models were used to evaluate the effects on trace mineral balances over time. RESULTS:The Mn and Cu balances of cows were not significantly different from 0 mg/d between 8 wk prepartum and calving (P ≥ 0.54), when dietary intake was the lowest of the period evaluated. However, when dietary intake was highest, between wk 6 and 16 postpartum, positive Mn and Cu balances were observed (80 and 20 mg/d, respectively, P ≤ 0.05). Cows were in positive Zn balance throughout the study except during the first 3 wk after calving during which the Zn balance was negative. CONCLUSIONS:Large adaptations occur in trace metal homeostasis in transition cows in response to changes in dietary intake. High dry matter intakes, associated with high milk production of dairy cows, combined with current Zn, Mn, and Cu supplementation practices may exceed regulatory homeostatic mechanisms resulting in potential body accumulation of Zn, Mn, and Cu.

The objective of this study was to characterize total-tract nutrient digestibility, energy balance, and N balance in the critical dietary and metabolic transitions of the lactation cycle. Twelve dairy cows were housed in tiestalls from 10 wk before to 16 wk after parturition. After 2 wk of adaptation to the facility and diet, digestibility of organic matter (OM), neutral detergent fiber (NDF), starch, and N were measured, and energy and N balances determined at weekly intervals by total collection of feces, urine, and milk over 48 h. Cows were individually fed ad libitum a grass silage- and corn silage-based total mixed ration during lactation and a corn silage- and barley straw-based total mixed ration during the dry period. Effects of stage of lactation were evaluated by clustering week in 5 groups: late lactation (wk -8 to -7), dry period (wk -6 to -1), and 3 early lactation periods (wk 1 to 5, wk 6 to 10, and wk 11 to 16). In lactation, apparent total-tract digestibility of OM, NDF, and starch was lowest in the first 5 wk of lactation. From wk 2 to 16 after parturition, apparent nutrient digestibility of all nutrients increased linearly, but with a negative quadratic component for dry matter, OM, and NDF, to levels comparable to those reported in last 2 wk of the previous lactation. However, differences in digestibility across lactation stage were moderate, illustrated by the difference between OM digestibility in late lactation (last 2 wk, 74.8%) and early lactation (first 5 wk, 72.5%). Cows were in negative energy balance for the first 8 wk after calving, and in negative N balance for the first 4 wk after calving. Based on energy and N balance, we predicted that 36.5 kg of body fat and 3.5 kg of body protein were gained in the last 8 wk before calving, and that 47.5 kg of body fat and 7.6 kg of body protein were mobilized in the first weeks of lactation. These predicted changes in body mass, both the gain before calving and loss after calving, were greater by 37% and 10%, respectively, than fluctuations in measured body weight (corrected for predicted gut fill and fetus weights). At wk 1 and 2 postpartum, body N loss corresponded to 25 and 29%, respectively, of total N excretion in milk, and body energy loss corresponded to 64% and 44%, respectively, of the energy exported to milk, illustrating the important contribution of N and energy from body stores to milk production in early lactation. Metabolic N efficiency, measured as total N output (milk and body) over digestible N input (from diet and body), averaged 54.4% in the last 2 wk of lactation, increased to 65.9% 2 wk after calving, and decreased linearly as lactation advanced to 61.9% by wk 16. Short (48 h) but weekly repetition of total collection of feces and urine appears to be a suitable approach to evaluate temporal changes in nutrient digestibility, energy balance, and N balance across lactation and the dry period.

Hydrogenated fat-embedded calcium gluconate (HFCG), a prebiotic mixture designed to target the hindgut, has improved milk and component yields when supplemented in mid-lactation cows, likely due to improved hindgut health. The objective of this study was to evaluate production responses to HFCG when fed to dairy cattle over a full lactation. Seventy-four Holstein cows (21 primiparous, 53 multiparous) were used in a randomized complete block design comparing supplementation with HFCG (approximately 16 g/day supplement delivering approximately 6.4 g active ingredient) to a negative control from approximately 21 days prior to calving until end of lactation. In multiparous cattle supplemented with HFCG, average daily milk protein yield ( P = 0.037) was increased during the first 8 weeks of lactation, while average daily yields of milk fat, and fat- and energy-corrected milk tended ( P ≤ 0.075) to increase over the same period of time. Increased yields were likely supported by the concurrent increase in dry matter intake ( P = 0.036). Future work is needed to characterize the mode of action of this product within both the hindgut lumen and host, as well as investigate the potential differential responses between primiparous and multiparous animals over the course of lactation.

Gluconate salts have been identified as a butyrate precursor when fed to non-ruminant species and may increase the butyrate concentration in the large intestine supporting gastrointestinal health and development. The objective of this study was to evaluate the dose response of hydrogenated fat-embedded calcium gluconate (HFCG) on performance and gastrointestinal tract (GIT) development in growing lambs. Thirty-two wether lambs were used in a randomized complete block design and assigned to 1 of 4 treatments differing in the inclusion of HFCG: 0.0% (CON), 0.075% (LOW), 0.30% (MED), and 0.60% of the diet (HIGH). Lambs were allocated into individual pens and fed ad libitum with feed delivered twice daily. Feed intake was recorded daily, and body weight (BW) was assessed at the beginning and the end of the 29-d period. Blood was sampled on day 21, prior to feeding and 6 h post-feeding to evaluate changes in β-hydroxybutyrate, glucose, and insulin concentrations. Total fecal collection was conducted during days 25 to 28 to assess apparent total tract digestibility. On day 29, lambs were slaughtered, and the entire GIT was separated by region to enable sampling of tissue and digesta. Data were analyzed to assess linear, quadratic, and cubic effects of HFCG dose. Final BW, average daily gain, and dry matter intake decreased linearly (P ≤ 0.02) with increasing HFCG. Increasing inclusion of HFCG linearly decreased (P = 0.01) the thickness of the stratum corneum in ruminal papillae but did not affect other strata (P ≥ 0.34). Omasal digesta weight linearly decreased (P = 0.01) as the concentration of HFCG increased and abomasal digesta weight was cubically affected (P = 0.03) the increasing dose of HFCG. Short-chain fatty acid concentration in the cecum was cubically affected (P < 0.01) with increasing dose of HFCG where low dose had the greatest concentration. Moreover, increasing the dietary supply of HFCG linearly increased the proportion of acetate (P = 0.04) in the cecum and linearly decreased the proportion of propionate in the digesta of both the cecum (P < 0.01) and colon (P = 0.01). Colon crypt depth was quadratically (P = 0.03) affected with the increasing dose of HFCG, where lambs fed MED had greatest crypt depth. We conclude that feeding HFCG to growing lambs did not increase butyrate concentration in the large intestine and consequently does not increase the absorptive surface area of the whole tract, the size of the GIT, or the functionality of the intestine.