Effects of maternal choline supplementation on performance and immunity of progeny from birth to weaning.

The objectives were to investigate whether supplementation with rumen-protected choline (RPC) during late pregnancy in Holstein cows affects offspring immunity and growth, and whether effects are utero-placental, colostrum dependent, or both. A total of 105 multiparous Holstein cows were assigned randomly to a prepartum diet (1.54 Mcal of NEL/kg of DM, and 15.8% CP) without (control) or with added RPC (12.9 g/d of choline ion). Calves (n = 111) were blocked by sex and assigned randomly to colostrum from control cows or colostrum from RPC cows, resulting in 4 treatments in a 2 × 2 factorial arrangement: (1) calves born and fed colostrum from non-supplemented dams (NN; n = 33); (2) calves from non-supplemented dams and fed colostrum from RPC-fed cows (NC; n = 25); (3) calves from RPC-supplemented dams and colostrum from non-supplemented cows (CN; n = 28); and (4) calves from RPC-supplemented dams and colostrum from RPC-fed cows (CC; n = 25). Growth, intakes, and immunity of females were evaluated up to 56 d of age. Growth and intake of male calves was evaluated up to 35 d of age, and physiological and immune responses to intravenous LPS challenge were evaluated from 21 to 35 d of age. Effects of prenatal and colostrum treatments and interactions between treatments were analyzed using mixed models. Calves fed colostrum from RPC-supplemented dams had a 17.4% increase in apparent efficiency of absorption of IgG compared with calves fed colostrum from control dams (27.4 vs. 23.3%). Incidence of fever in the first 21 d of age tended to be less in females born from RPC-supplemented dams compared with females born from control dams (31 vs. 58%). Prenatal RPC females had increased hematocrit and concentrations of red blood cells, leukocytes, neutrophils, and lymphocytes in blood compared with prenatal females born from control dams. Compared with prenatal control females, prenatal RPC females had greater intake of milk replacer (704 vs. 748 ± 9.9 g/d) and starter (45.4 vs. 60.2 ± 5.9 g/d) during the first 21 d of age. In male calves, mean intake of DM was greater (1,074 vs. 976 ± 45 g/d) after the LPS challenge (0 to 8 d) by calves born from dams fed RPC compared with males born from control dams. Calves born from RPC-fed dams also had lower mean rectal temperature (39.0 vs. 39.2°C) and mean respiration rate (35.6 vs. 39.3 breaths/min) compared with males born from control dams. Moreover, serum concentrations of metabolites (i.e., ß-hydroxybutyric acid, fatty acids, and glucose), cytokines (i.e., tumor necrosis factor-α) and acute phase proteins (i.e., serum amyloid A) were consistent with less-severe inflammatory response to LPS in males born from dams fed RPC compared with control. Source of colostrum and interaction between prenatal and colostrum treatments had minimal effects on calf responses to LPS. Overall, maternal RPC supplementation during late gestation suggests a positive effect on immunity, in that colostrum from RPC-fed dams increased efficiency of IgG absorption and maternal supplementation with RPC during late gestation, regardless of colostrum source, attenuated responses to LPS.

Lactational performance of dairy cows in response to supplementing N-acetyl-l-methionine as source of rumen-protected methionine.

The objective of this experiment was to evaluate the effects of supplementing a rumen-protected source of Met, N-acetyl-l-methionine (NALM), on lactational performance and nitrogen metabolism in early- to mid-lactation dairy cows. Sixty multiparous Holstein dairy cows in early lactation (27 ± 4.3 d in milk, SD) were assigned to 4 treatments in a randomized complete block design. Cows were blocked by actual milk yield. Treatments were as follows: (1) no NALM (control); (2) 15 g/d of NALM (NALM15); (3) 30 g/d of NALM (NALM30); and (4) 45 g/d of NALM (NALM45). Diets were formulated using a Cornell Net Carbohydrate and Protein System (CNCPS) v.6.5 model software to meet or exceed nutritional requirements of lactating dairy cows producing 42 kg/d of milk and to undersupply metabolizable Met (control) or supply incremental amounts of NALM. The digestible Met (dMet) supply for control, NALM15, NALM30, and NALM45 were 54.7, 59.8, 64.7, and 72.2 g/d, respectively. The supply of dMet was 88, 94, 104, and 115% of dMet requirement for control, NALM15, NALM30, and NALM45, respectively. Milk yield data were collected, dry matter intake (DMI) was measured daily, and milk samples were collected twice per week for 22 wk. Blood, ruminal fluid, urine, and fecal samples were collected during the covariate period and during wk 4, 8, and 16. Data were analyzed using the GLIMMIX procedure of SAS (SAS Institute) using covariates in the model for all variables except body weight. Linear, quadratic, and cubic contrasts were also tested. Treatments did not affect DMI, milk yield, and milk component concentration and yield; however, feed efficiency expressed as milk yield per DMI and 3.5% fat-corrected milk per DMI were quadratically affected, with greater response observed for NALM15 and NALM30 compared with control. Acetate proportion linearly increased, whereas propionate proportion linearly decreased with NALM supplementation. Blood urea nitrogen linearly decreased with NALM supplementation. Total plasma essential AA concentrations were quadratically affected, as greater values were observed for control and NALM45 than other treatments. Plasma Met concentration was quadratically affected as lower levels were observed with NALM15, whereas Met concentrations increased with NALM45 compared with control. Nitrogen utilization efficiency and apparent total-tract nutrient digestibility were not affected by treatment. Supplementation of NALM at 15 or 30 g/head per day resulted in the greatest improvements in feed efficiency without affecting N metabolism of early- to mid-lactation dairy cows.

Effects of a xylanase-rich enzyme on intake, milk production, and digestibility of dairy cows fed a diet containing a high proportion of bermudagrass silage.

We previously reported that milk production in dairy cows was increased by adding a specific xylanase-rich exogenous fibrolytic enzyme (XYL) to a total mixed ration (TMR) containing 10% bermudagrass silage (BMD). Two follow-up experiments were conducted to examine whether adding XYL would increase the performance of dairy cows consuming a TMR containing a higher (20%) proportion of BMD (Experiment 1) and to evaluate the effects of XYL on in vitro fermentation and degradability of the corn silage, BMD, and TMR (Experiment 2). In Experiment 1, 40 lactating Holstein cows in early lactation (16 multiparous and 24 primiparous; 21 ± 3 d in milk; 589 ± 73 kg of body weight) were blocked by milk yield and parity and randomly assigned to the Control and XYL treatments. The TMR contained 20% BMD, 25% corn silage, 8% wet brewer's grain, and 47% concentrate mixture in the dry matter (DM). Cows were fed the XYL-treated or untreated experimental TMR twice per day for 10 wk after a 9-d covariate period. In Experiment 2, ruminal fluid was collected from 3 cannulated lactating Holstein cows fed a diet containing 20% bermudagrass haylage, 25% corn silage and 55% concentrate. In Experiment 1, compared with Control, application of XYL did not affect DM intake (24.0 vs. 23.7 kg/d), milk yield (35.1 vs. 36.2 kg/d), fat-corrected milk yield (36.1 vs. 36.9 kg/d), or yields of milk fat (1.29 vs. 1.31 kg/d) or protein (1.07 vs. 1.08 kg/d). However, intake of neutral detergent fiber (4.67 vs. 4.41 kg/d) tended to increase with XYL; consequently, milk protein concentration was increased by XYL (3.02 vs. 2.95%). Feed efficiency tended to be lower in cows fed XYL (1.57 vs. 1.52 kg of fat-corrected milk/kg of DM intake) compared with Control. In Experiment 2, XYL tended to increase the rate of gas production in the TMR, the molar proportion of propionate for corn silage, and that of valerate for the TMR. In addition, XYL increased in vitro DM, neutral detergent fiber, and acid detergent fiber degradability of BMD and corn silage. Application of XYL to a diet with a relatively high proportion of BMD tended to increase digestible neutral detergent fiber intake, increased milk protein concentration, and in vitro degradability of DM, neutral detergent fiber, and acid detergent fiber. However, XYL did not affect milk production and tended to decrease feed efficiency in early lactation cows.

The effects of prepartum energy intake and peripartum rumen-protected choline supplementation on hepatic genes involved in glucose and lipid metabolism.

Nutritional interventions, either by controlling dietary energy (DE) or supplementing rumen-protected choline (RPC) or both, may mitigate negative postpartum metabolic health outcomes. A companion paper previously reported the effects of DE density and RPC supplementation on production and health outcomes. The objective of this study was to examine the effects of DE and RPC supplementation on the expression of hepatic oxidative, gluconeogenic, and lipid transport genes during the periparturient period. At 47 ± 6 d relative to calving (DRTC), 93 multiparous Holstein cows were randomly assigned in groups to dietary treatments in a 2 × 2 factorial of (1) excess energy (EXE) without RPC supplementation (1.63 Mcal of NEL/kg of dry matter; EXE-RPC); (2) maintenance energy (MNE) without RPC supplementation (1.40 Mcal of NEL/kg dry matter; MNE-RPC); (3) EXE with RPC supplementation (EXE+RPC); and (4) MNE with RPC supplementation (MNE+RPC). To achieve the objective of this research, liver biopsy samples were collected at -14, +7, +14, and +21 DRTC and analyzed for mRNA expression (n = 16/treatment). The interaction of DE × RPC decreased glucose-6-phosphatase and increased peroxisome proliferator-activated receptor α in MNE+RPC cows. Expression of cytosolic phosphoenolpyruvate carboxykinase was altered by the interaction of dietary treatments with reduced expression in EXE+RPC cows. A dietary treatment interaction was detected for expression of pyruvate carboxylase although means were not separated. Dietary treatment interactions did not alter expression of carnitine palmitoyltransferase 1A or microsomal triglyceride transfer protein. The 3-way interaction of DE × RPC × DRTC affected expression of carnitine palmitoyltransferase 1A, glucose-6-phosphatase, and peroxisome proliferator-activated receptor α and tended to affect cytosolic phosphoenolpyruvate carboxykinase. Despite previously reported independent effects of DE and RPC on production variables, treatments interacted to influence hepatic metabolism through altered gene expression.

Effects of choline on the phenotype of the cultured bovine preimplantation embryo.

Choline is a precursor of acetylcholine, phosphatidylcholine, and the methyl-donor betaine. Reports indicate that supplementation with rumen-protected choline improves postpartum reproductive function of dairy cows. The objective was to determine whether addition of choline to culture medium of in vitro-produced embryos alters the phenotype of the resultant blastocysts. Treatments were choline chloride (ChCl; 0.004, 1.3, 1.8, and 6.37 mM) and phosphatidylcholine (1.3 mM). Treatment with 0.004 mM ChCl improved development to the blastocyst stage, increased blastocyst cell number, and increased the percentage of blastocysts that were hatching or hatched. Development was not affected by higher concentrations of ChCl but was reduced by 1.3 mM phosphatidylcholine. Treatment of embryos with 1.3 mM ChCl (but not other concentrations) increased expression in blastocysts of 11 of 165 genes examined (AMOT, NANOG, HDAC8, HNF4A, STAT1, MBNL3, SOX2, STAT3, KDM2B, SAV1, and GPAM) and decreased expression of one gene (ASS1). Treatment with 1.3 mM ChCl decreased global DNA methylation at d 3.5 of development and increased DNA methylation at d 7.5 in blastocysts. Treatment with 1.8 mM ChCl also increased methylation in blastocysts. In conclusion, addition of choline to the culture medium alters the phenotype of preimplantation bovine embryos produced in vitro. Choline chloride can act in a concentration-dependent manner to alter development, expression of specific genes, and DNA methylation.

Timing of initiation and duration of feeding rumen-protected choline affects performance of lactating Holstein cows.

Objectives were to evaluate the effects of altering timing of initiating and duration of supplementing rumen-protected choline (RPC) on lactation performance in dairy cows. The hypothesis was that RPC increases yields of milk and milk components, regardless of when supplementation is initiated, and that the effects of supplementing RPC starting prepartum and continuing post-transition would be additive. Cows at 241 ± 2.2 d of gestation were blocked by parity group (49 entering lactation 2, 50 entering lactation >2) and 305-d milk yield and, within block, assigned randomly to 1 of 4 treatments arranged as a 2 × 2 factorial with 2 levels of choline in transition, from 21 d pre- to 21 d postpartum, and 2 levels of choline in post-transition, from 22 to 105 d postpartum. The 2 levels of RPC supplemented were either 0 g/d or 12.9 g/d of choline ion fed as 60 g/d of an RPC product that was top-dressed onto the total mixed ration. Thus, treatments were as follows: NN (n = 25): no choline in transition or post-transition; NC (n = 25): no choline in transition and choline in post-transition; CN (n = 25): choline in transition and no choline in post-transition; CC (n = 24): choline in transition and in post-transition. Prepartum, treatments were supplemented (mean ± SD) for the last 18.8 ± 5.7 and 19.2 ± 5.0 d of gestation in treatments with 0 or 12.9 g/d of choline ion, respectively. Supplementing RPC prepartum did not affect dry matter intake (DMI), body weight (BW), or body condition score (BCS) in the last 3 weeks of gestation. Likewise, RPC did not affect the yield or the composition of colostrum. Supplementation with RPC during transition increased fat percent by 0.02 percentage units, fat yield by 0.16 kg/d, and energy-corrected milk (ECM) by 3.1 kg/d in the first 21 d postpartum, and increased fat yield by 0.10 kg/d and ECM by 2.4 kg/d from 22 to 105 d postpartum. Supplementing RPC during transition did not affect DMI postpartum, but it improved feed efficiency, and cows produced 0.11 kg/d more ECM per kg of DMI. Changes in BW and BCS during the first 21 d postpartum did not differ between treatments. Cows fed RPC during transition had more negative net energy balance and 0.1 unit smaller BCS in the first 105 d postpartum than non-supplemented cows. Supplementing RPC in post-transition did not influence productive performance in dairy cows, and choline supplementation during transition or post-transition did not affect measures of reproduction. Collectively, supplementing RPC to supply 12.9 g/d of choline ion benefited productive performance in dairy cows when supplementation occurred during the transition period, but no additional benefit was observed from supplementing RPC past 22 d postpartum.

Effect of prepartum energy intake and supplementation with ruminally protected choline on innate and adaptive immunity of multiparous Holstein cows.

Objectives were to evaluate the effect of prepartum energy intake and peripartal supplementation of ruminally protected choline (RPC) on select indicators of immune status in blood plasma and on lipopolysaccharide-stimulated blood cells ex vivo. At 47 ± 6 d before the expected calving date, 93 multiparous Holstein cows were assigned randomly to 1 of 4 dietary treatments in a 2 × 2 factorial arrangement. Cows were fed energy to excess [EXE; 1.63 Mcal of net energy for lactation (NEL)/kg of dietary dry matter (DM)] or to maintenance (MNE; 1.40 Mcal of NEL/kg of dietary DM) ad libitum throughout the nonlactating period. The RPC was fed at 0 or 60 g/d to supply 0 or 12.9 g/d of choline ions top-dressed for 17 ± 4.6 d prepartum through 21 d postpartum. After calving, cows were fed the same methionine-supplemented diet, apart from RPC supplementation. During the last 2 wk before calving and during the first 5 wk postpartum, blood was sampled repeatedly and analyzed for cell types, acute-phase proteins, tumor necrosis factor-α (TNFα), and neutrophil function. Samples of whole blood were collected at 3 and 14 DIM and stimulated with 1 µg/mL lipopolysaccharide (LPS) in vitro for 6 and 24 h. After 6 h of LPS exposure, peripheral blood leucocytes (PBL) were harvested, and relative transcript abundance for select cytokines were measured. Supernatant was analyzed for TNFα after 24 h of LPS exposure. The PBL from cows fed EXE diets during the whole dry period had increased transcripts for the proinflammatory cytokines CXCL8 and TNF, although the plasma concentrations of the acute-phase proteins haptoglobin and fibrinogen, and the killing activity of the blood neutrophils in the postpartum period, were not affected by feeding different energy levels prepartum. Feeding RPC to cows overfed energy prepartum modulated their inflammatory state, as evidenced by decreased IL6 in PBL and reduced mean fluorescence intensity of CD14 during the postpartum period, compared with cows not fed RPC. Feeding RPC also decreased TNFα protein production, abundances of IL1B, CXCL8, and TNF transcripts, and mean fluorescence intensity of CD80 of PBL stimulated by LPS, regardless of prepartum energy intake. In contrast, proportions of blood neutrophils undergoing phagocytosis and oxidative burst were increased at 17 d postpartum in cows supplemented with RPC. Collectively, these data indicate that transition cows supplemented with RPC experienced less inflammation, which may partially explain increased milk production in cows supplemented with RPC.

Effects of rumen-protected choline on the inflammatory and metabolic status and health of dairy cows during the transition period.

The objectives of this study were to evaluate the effects of rumen-protected choline (RPC) supplementation from 21 d pre- to 21 d postpartum on markers of metabolic status and inflammatory response, concentrations of liposoluble vitamins, and plasma total Ca in parous Holstein cows. The hypotheses were that supplementing RPC during the transition period would reduce hepatic triacylglycerol accumulation postpartum and attenuate markers of inflammatory response following parturition, and collectively, such responses were expected to benefit health of dairy cows. Parous cows at 241 d of gestation were blocked by parity group and 305-d milk yield, and within block, they were assigned randomly to receive either 0 g/d [no choline in transition (NT), n = 55] or 12.9 g/d choline ion [choline in transition (CT), n = 58] from 21 d pre- to 21 postpartum. The RPC product was individually top-dressed onto the total mixed ration once daily. Prepartum, treatments were supplemented (mean ± standard deviation) for the last 18.8 ± 5.7 and 19.2 ± 5.0 d of gestation in NT and CT, respectively. Supplementing RPC prepartum did not affect concentrations of plasma metabolites and inflammatory markers during the last 3 wk of gestation. Postpartum, cows fed RPC had greater hepatic concentration of hepatic triacylglycerol (NT = 3.4 vs. CT = 4.4%) and tended to have increased concentration of ß-hydroxybutyrate (NT = 0.48 vs. CT = 0.53 mM) in plasma. In spite of the increased hepatic triacylglycerol in cows fed RPC, treatment did not affect the concentrations of the inflammatory marker tumor necrosis factor-α or of the positive acute phase proteins, haptoglobin and fibrinogen. Supplementing choline tended to increase the concentration of plasma triacylglycerol by 0.69 mg/dL in the first 21 d postpartum and reduced the incidence of subclinical hypocalcemia by 20.9 percentage units compared with NT. Supplementing transition cows with RPC did not affect the concentrations of liposoluble vitamins in the first 7 d postpartum or the incidence of individual diseases or morbidity in early lactation. The inability of supplemental choline to reduce hepatic triacylglycerol might have been a consequence of the increased productive performance without additional dry matter intake.

Meta-analysis of the effects of supplemental rumen-protected choline during the transition period on performance and health of parous dairy cows.

The objectives were to use meta-analytic methods to determine the effects of amount of supplemental choline ion as rumen-protected choline (RPC) starting prepartum on production and health of dairy cows. The literature was systematically reviewed and 21 experiments, with up to 66 treatment means and 1,313 prepartum parous cows, were included. All experiments had a treatment with no supplemental choline (0 g/d; n = 30 treatment means), and the amount of choline ion supplemented to treated cows ranged from 5.6 to 25.2 g/d (n = 36 treatment means). Duration of pre- and postpartum feeding of RPC averaged (±standard deviation) 22.0 ± 6.0 and 57.5 ± 42.2 d, respectively. Data collected included the ingredient composition and chemical analyses of pre- and postpartum diets, amount of choline ion supplemented, number of cows per treatment, frequency of health events, and the least squares means and respective standard error of the means for production responses, liver composition, and blood parameters. The concentrations of net energy for lactation and metabolizable amino acids and protein (MP) in pre- and postpartum diets were predicted for each treatment mean using National Research Council (2001). Mixed model meta-analysis was conducted including the random effect of experiment and weighting by the inverse of the standard error of the means squared. Increasing supplementation of choline ion during transition linearly increased pre- (ß = 0.0184 ± 0.00425) and postpartum dry matter intake (ß = 0.0378 ± 0.00974), and yields of milk (ß = 0.436 ± 0.112), energy-corrected milk (ECM; ß = 0.422 ± 0.0992), fat (ß = 0.00555 ± 0.000793), and protein (ß = 0.0138 ± 0.00378). Nevertheless, an interaction between choline and postpartum metabolizable methionine as a percent of MP (METMPPo) was observed for yields of milk, ECM, and protein because as METMPPo increased, the positive response to choline on yields of milk, ECM, and protein decreased. Supplementing choline during transition tended to reduce the risks of retained placenta and mastitis, but it had no effect on metritis, milk fever, displaced abomasum and ketosis, or the concentration of triacylglycerol in the hepatic tissue postpartum. The median amount of choline ion supplemented was 12.9 g/d and responses in postpartum dry matter intake and yields of milk, ECM, fat, and protein to that amount of supplementation were 0.5, 1.6, 1.7, 0.07, and 0.05 kg/d, respectively. No interactions were observed between supplemental choline and prepartum dietary net energy for lactation or metabolizable methionine as a percent of MP. Collectively, feeding RPC during the transition period improves performance in parous cows. Increases in yields of milk and milk components were observed in spite of pre- and postpartum diets, although the increments in milk, ECM, and protein yields with supplementing choline decreased as the concentration of methionine in postpartum diets increased. The optimum dose of choline ion was not detected, but likely it is more than the 12.9 g/d fed in most experiments evaluated in the current meta-analysis. Finally, the meta-analysis identified lack of sufficient data to understand the role of supplemental choline in nulliparous cows.

Responses to rumen-protected choline in transition cows do not depend on prepartum body condition.

It is often suggested that the benefits of supplemental rumen-protected choline (RPC) might be greater in cows predisposed to fatty liver, such as those that are overconditioned; however, limited data support this suggestion. Therefore, the hypothesis of this study was that responses to supplementing RPC to transition dairy cows is not dependent on the degree of fatness prepartum. Objectives were to evaluate the effects of supplementing RPC to transition dairy cows according to body condition score (BCS) prepartum on production and metabolic responses. Data from 2 randomized block experiments that evaluated the effects of RPC supplementation during the transition period were combined. Within each experiment, cows were assigned randomly to receive 0 (CTRL) or 12.9 g/d choline ion in an RPC form (CHOL) daily top-dressed onto the diet from 21 d prepartum to 21 d postpartum. Body condition was evaluated twice prepartum before enrollment and the mean value was used as an explanatory variable for statistical analyses. Data were collected for the last 21 d of gestation and the first 105 d postpartum. The BCS (mean ± standard deviation) prepartum were 3.51 ± 0.29 and 3.51 ± 0.32 for CTRL and CHOL, respectively, and ranged from 2.69 to 4.25. A total of 215 cows were enrolled in the respective experiments and contributed data for the incidence of diseases, whereas 192 cows contributed data for analyses of production responses, plasma metabolites, and liver composition. Irrespective of BCS, supplementing transition diets with CHOL increased yields of milk by 1.8 kg/d, fat by 0.08 kg/d, lactose by 0.08 kg/d, true protein by 0.04 kg/d, energy-corrected milk (ECM) by 1.9 kg/d, and fat-corrected milk by 2.1 kg/d. The improvements in productive performance were not followed by increased dry matter intake or measures of lipomobilization. Therefore, CHOL cows were more efficient in converting dry matter intake into ECM. Feeding CHOL increased concentration of hepatic triacylglycerol (CTRL = 3.23 vs. CHOL = 3.87% wet basis) in the first 21 d postpartum. Overconditioned cows were more prone to having exacerbated lipomobilization and increased prevalence and incidence of fatty liver, but no interactions between treatment and BCS were observed for body weight, BCS, or concentrations of metabolites in plasma or hepatic triacylglycerol. Treatment did not affect incidence of clinical diseases. Opposite to common suggestions, cows with increased hepatic triacylglycerol content also had increased yields of milk and ECM in the first 105 d postpartum. Collectively, these findings indicate that the effects of RPC supplementation during the transition period are independent of the degree of fatness of dairy cows prepartum. The findings also suggest that the effects on productive performance are not necessarily mediated by improvements in markers of metabolic health or reductions in hepatic triacylglycerol.

Effects of altering the ratio of dietary n-6 to n-3 fatty acids on spontaneous luteolysis in lactating dairy cows.

Objectives were to evaluate the effects of altering the dietary ratio of omega-6 (n-6) to omega-3 (n-3) fatty acids on the profile of fatty acids and expression of genes related to the prostaglandin biosynthesis on endometrial tissue, uterine secretion of PGF2α, and timing of spontaneous luteolysis in dairy cows. Multiparous lactating Holstein cows (n = 45) were blocked based on milk yield and, within each block, assigned randomly to 1 of 3 dietary treatments at 14 d postpartum for 90 d. Diets were supplemented with a mixture of Ca salts of fish, safflower, and palm oils to create 3 different ratios of n-6 to n-3 fatty acids, namely R4, R5, and R6, which resulted in 3.9, 4.9, and 5.9 parts of n-6 to 1 part of n-3 fatty acids, respectively. Blood was sampled every 2 h from d 16 to 23 of the estrous cycle and assayed for concentrations of progesterone and the PGF2α metabolite 13,14-dihydro-15-keto-PGF2α (PGFM). In a subsequent estrous cycle, endometrial tissue was collected for biopsy on d 8 and endometrial fatty acids profile and gene expression were quantified. The proportion of arachidonic acid of the endometrial fatty acids increased as the dietary ratio n-6 to n-3 fatty acids increased (R4 = 9.05, R5 = 11.64, and R6 = 13.41%). On the other hand, proportions of eicosapentaenoic (R4 = 2.85, R5 = 2.14, and R6 = 2.02%) and docosahexaenoic (R4 = 3.30, R5 = 1.57, and R6 = 1.08%) decreased as the ratio of n-6 to n-3 fatty acids in the diet increased. Increasing the ratio of dietary n-6 to n-3 fatty acids increased mRNA expression of estrogen receptor 1, oxytocin receptor, cyclooxygenase 2, prostaglandin E and F synthases, and steroidogenic acute regulatory protein in endometrium, but decreased expression of peroxisome proliferator-activated receptor gamma and insulin-like growth factor-1. The changes in endometrium gene expression caused by dietary treatments were associated with changes in the ratio of n-6 to n-3 fatty acids in the endometrium. As the ratio increased from R4 to R6, the number of PGFM pulses (R4 = 5.6, R5 = 4.3, and R6 = 3.8 ± 0.6 pulses; least squares means ± standard error of the means) decreased, but the amplitude of the greatest PGFM pulse increased (R4 = 226, R5 = 267, and R6 = 369 ± 38 pg/mL). Luteolysis by d 23 of the estrous cycle was observed in 79.6% of the cows (R4 = 11/14; R5 = 13/15; and R6 = 11/15) and day of spontaneous luteolysis did not differ among treatments (R4 = 20.8; R5 = 21.1; and R6 = 21.0 ± 0.4). Three pulses of PGFM was the best predictor of luteolysis in dairy cows. Collectively, supplying the same quantity of fatty acids in the diet of lactating dairy cows, but altering the ratio of n-6 to n-3 fatty acids, influenced the endometrial fatty acids profile and gene expression and altered the pattern of prostaglandin synthesis; however, the changes were not sufficient to alter the length of the estrous cycle.

Feeding increasing amounts of ruminally protected choline decreased fatty liver in nonlactating, pregnant Holstein cows in negative energy status.

The objectives were to determine the optimal feeding amount of choline in a ruminally protected form to reduce the triacylglycerol (TAG) concentration in liver and to increase TAG in blood plasma of dairy cows. Pregnant, nonlactating multiparous Holstein cows (n = 77) were blocked by body condition score (3.59 ± 0.33) and assigned to treatment at 64 ± 10 d before calculated calving date. Dietary treatments were top-dressing of 0, 30, 60, 90, or 120 g/d of ruminally protected choline (RPC; Balchem Corp., New Hampton, NY) ions to supply the equivalent of 0, 6.5, 12.9, 19.4, and 25.8 g/d of choline ions. Diets were formulated to exceed nutrient requirements for maintenance and pregnancy and fed in ad libitum amounts for the first 5 d. From d 6 to 15, cows were restricted to consume approximately 31% of their net energy requirements to simulate early lactating cows in negative energy balance. Methionine intake was maintained throughout each 15-d period. Liver was biopsied at 5 and 14 d and analyzed for TAG and glycogen. Blood was sampled on d 5 and 14 and plasma analyzed for glucose, insulin, cholesterol, ß-hydroxybutyrate, long-chain fatty acids, and haptoglobin. On d 14, a mixture of saturated long-chain fatty acids, ground corn, and dried molasses (50:37:13) was offered (908 g, as-is basis) 10 h after the single daily feeding. Blood samples were collected for 19 h and plasma analyzed for TAG and cholesterol to assess apparent absorption of dietary fat. Mean dry matter intake and energy balance decreased from means of 9.5 to 3.3 kg/d and from 0.6 to -9.2 Mcal of net energy for lactation/d during the ad libitum and restricted feeding periods, respectively. Plasma concentrations of the lipid-soluble choline biomolecules, namely total phosphatidylcholines, total lysophosphatidylcholines, and sphingomyelin, increased with choline supplementation. Feed restriction increased plasma concentrations of ß-hydroxybutyrate and free long-chain fatty acids, whereas those of glucose, insulin, and total cholesterol decreased. During feed restriction, concentration of hepatic TAG and plasma haptoglobin decreased linearly, whereas concentration of hepatic glycogen tended to increase quadratically with increasing intake of RPC. After fat supplementation, mean plasma concentration of TAG increased by an average of 21% with intake of RPC ions, peaking at intakes of ≥6.5 g/d of RPC ion. In summary, feeding RPC ions to cows in negative energy balance had increasing lipotropic effects on the liver when consumed up to 25.8 g/d, whereas feeding only 6.5 g/d increased concentrations of hepatic glycogen and TAG in the blood.

Effect of adding clay with or without a Saccharomyces cerevisiae fermentation product on the health and performance of lactating dairy cows challenged with dietary aflatoxin B1.

The study was conducted to examine the effect of supplementing bentonite clay with or without a Saccharomyces cerevisiae fermentation product (SCFP; 19 g of NutriTek + 16 g of MetaShield, both from Diamond V, Cedar Rapids, IA) on the performance and health of dairy cows challenged with aflatoxin B1 (AFB1). Twenty-four lactating Holstein cows (64 ± 11 d in milk) were stratified by parity and milk production and randomly assigned to 1 of 4 treatment sequences. The experiment had a balanced 4 × 4 Latin square design with 6 replicate squares, four 33-d periods, and a 5-d washout interval between periods. Cows were fed a total mixed ration containing 36.1% corn silage, 8.3% alfalfa hay, and 55.6% concentrate (dry matter basis). Treatments were (1) control (no additives), (2) toxin (T; 1,725 µg of AFB1/head per day), (3) T + clay (CL; 200 g/head per day; top-dressed), and (4) CL+SCFP (CL+SCFP; 35 g/head per day; top-dressed). Cows were adapted to diets from d 1 to 25 (predosing period) and then orally dosed with AFB1 from d 26 to 30 (dosing period), and AFB1 was withdrawn from d 31 to 33 (withdrawal period). Milk samples were collected twice daily from d 21 to 33, and plasma was sampled on d 25 and 30 before the morning feeding. Transfer of ingested AFB1 into milk aflatoxin M1 (AFM1) was greater in T than in CL or CL+SCFP (1.65 vs. 1.01 and 0.94%, respectively) from d 26 to 30. The CL and CL+SCFP treatments reduced milk AFM1 concentration compared with T (0.45 and 0.40 vs. 0.75 µg/kg, respectively), and, unlike T, both CL and CL+SCFP lowered AFM1 concentrations below the US Food and Drug Administration action level (0.5 µg/kg). Milk yield tended to be greater during the dosing period in cows fed CL+SCFP compared with T (39.7 vs. 37.7 kg/d). Compared with that for T, plasma glutamic oxaloacetic transaminase concentration, indicative of aflatoxicosis and liver damage, was reduced by CL (85.9 vs. 95.2 U/L) and numerically reduced by CL+SCFP (87.9 vs. 95.2 U/L). Dietary CL and CL+SCFP reduced transfer of dietary AFB1 to milk and milk AFM1 concentration. Only CL prevented the increase in glutamic oxaloacetic transaminase concentration, and only CL+SCFP prevented the decrease in milk yield caused by AFB1 ingestion.

Effects of supplementation with ruminally protected choline on performance of multiparous Holstein cows did not depend upon prepartum caloric intake.

Objectives were to evaluate the effect of prepartum energy intake on performance of dairy cows supplemented with or without ruminally protected choline (RPC; 0 or 17.3 g/d of choline chloride; 0 or 60 g/d of ReaShure, Balchem Corp., New Hampton, NY). At 47 ± 6 d before the expected calving date, 93 multiparous Holstein cows were assigned randomly to 1 of 4 dietary treatments in a 2 × 2 factorial arrangement. Cows were fed energy to excess [EXE; 1.63 Mcal of net energy for lactation/kg of dry matter (DM)] or to maintenance (MNE; 1.40 Mcal of net energy for lactation/kg of DM) in ad libitum amounts throughout the nonlactating period. The RPC was top-dressed for 17 ± 4.6 d prepartum through 21 d postpartum (PP). After calving, cows were fed the same methionine-balanced diet, apart from RPC supplementation, through 15 wk PP. Liver was biopsied at -14, 7, 14, and 21 d relative to parturition. Cows fed EXE or MNE diets, respectively, consumed 40 or 10% more Mcal/d than required at 15 d before parturition. Cows fed the MNE compared with the EXE diet prepartum consumed 1.2 kg/d more DM postpartum but did not produce more milk (41.6 vs. 43.1 kg/d). Thus, PP cows fed the EXE diet prepartum were in greater mean negative energy balance, tended to have greater mean concentrations of circulating insulin, fatty acids, and ß-hydroxybutyrate, and had greater triacylglycerol in liver tissue (8.3 vs. 10.7% of DM) compared with cows fed the MNE diet prepartum. Cows fed RPC in transition tended to produce more milk (43.5 vs. 41.3 kg/d) and energy-corrected milk (44.2 vs. 42.0 kg/d) without increasing DM intake (23.8 vs. 23.2 kg/d) during the first 15 wk PP, and tended to produce more milk over the first 40 wk PP (37.1 vs. 35.0 kg/d). Energy balance of cows fed RPC was more negative at wk 2, 3, and 6 PP, but mean circulating concentrations of fatty acids and ß-hydroxybutyrate did not differ from those of cows not fed RPC. Despite differences in energy balance at 2 and 3 wk PP, mean concentration of hepatic triacylglycerol did not differ between RPC treatments. Feeding RPC reduced the daily prevalence of subclinical hypocalcemia from 25.5 to 10.5%, as defined by concentrations of total Ca of <8.0 mg/dL in serum in the first 7 d PP. Pregnancy at first artificial insemination tended to be greater for cows fed RPC (41.3 vs. 23.6%), but the proportion of pregnant cows did not differ by 40 wk PP. Heifers born from singleton calvings from cows fed RPC tended to experience greater daily gain between birth and 50 wk of age than heifers from cows not supplemented with RPC. Feeding RPC for approximately 38 d during the transition period tended to increase yield of milk for 40 wk regardless of amount of energy consumed during the pregnant, nonlactating period.

Effects of maturity at ensiling of bermudagrass and fibrolytic enzyme application on the performance of early-lactation dairy cows.

The objective of this study was to examine effects of adding fibrolytic enzymes to diets containing bermudagrass ensiled after 4 or 7wk of regrowth on the diet digestibility, ruminal fermentation and performance of lactating cows, and the interaction of the treatments. In experiment 1, 64 Holstein cows (22±4d in milk) were assigned to an experiment with a 2×2 factorial treatment arrangement and a 56-d duration. Treatments were diets containing 4 or 7wk regrowth bermudagrass silage without or with an exogenous fibrolytic enzyme cocktail. The cellulase-xylanase enzyme was applied at 2.33g/kg of total mixed ration dry matter (DM) during mixing immediately before feeding. Experiment 2 was aimed at examining treatment effects on the ruminal fermentation profile. Four ruminally cannulated cows were assigned to the 4 treatments using a 4×4 Latin square design with 14-d periods. No enzyme by maturity interaction was detected for any measurement. Regardless of forage maturity, applying the fibrolytic enzyme did not affect DM intake, milk yield, apparent digestibility, feed efficiency, energy balance, and ruminal fermentation though it tended to increase milk lactose concentration (4.88 vs. 4.81%). Feeding the 4-wk diet instead of the 7-wk diet increased DM intake (22.4 vs. 21.3kg/d), digestibility of DM, neutral detergent fiber, and acid detergent fiber, and tended to increase 3.5%-fat corrected milk yield (47.2 vs. 44.3kg/d) and milk fat yield (1.88 vs. 1.73kg/d). Therefore, daily intake of net energy and secretion of milk energy were greater for the 4-wk diet. In addition, the 4-wk diet increased the ruminal concentrations of acetate, propionate, valerate, lactate, and total volatile fatty acids, and decreased ruminal pH, without affecting the acetate:propionate ratio. Feeding fibrolytic enzymes did not improve the performance of early-lactation dairy cows, but harvesting the forage earlier tended to improve milk production.

Supplementation of essential fatty acids to Holstein calves during late uterine life and first month of life alters hepatic fatty acid profile and gene expression.

Linoleic acid is an essential dietary fatty acid (FA). However, how the supplementation of linoleic acid during uterine and early life may modify the FA profile and transcriptome regulation of the liver, and performance of preweaned dairy calves is unknown. Our objective was to evaluate the effect of supplementation of essential FA to Holstein calves during late uterine and early life on their hepatic FA profile and global gene expression at 30 d of age. During the last 8 wk of pregnancy, Holstein cattle (n=96) were fed either no fat supplement (control), a saturated FA supplement enriched with C18:0, or an unsaturated FA supplement enriched with linoleic acid. Male calves (n=40) born from these dams were fed a milk replacer (MR) with either low (LLA) or high linoleic acid (HLA) concentration as the sole feedstuff during the first 30 d. Liver biopsy was performed at 30 d of age, and microarray analysis was performed on 18 liver samples. Total concentration of FA in liver were greater in calves fed LLA compared with those fed HLA MR (8.2 vs. 7.1%), but plasma concentrations of total FA did not differ due to MR diets. The FA profiles of plasma and liver of calves were affected differently by the prepartum diets. Specifically, the FA profile in liver was affected moderately by the feeding of fat prepartum, but the profiles did not differ due to the type of FA fed prepartum. The type of MR fed during the first 30 d of life had major effects on both plasma and liver FA profiles, resembling the type of fat fed. Plasma and liver of calves fed LLA MR had greater percentage of medium-chain FA (C12:0 and C14:0), whereas plasma and liver from calves fed HLA MR had greater percentages of linoleic and α-linolenic acids. Dams fed fat or a specific type of FA modified the expression of some genes in liver of calves, particularly those genes involved in biological functions and pathways related to upregulation of lipid metabolism and downregulation of inflammatory responses. Feeding HLA instead of LLA MR modified the expression of hepatic genes, including genes predicted to decrease infections and to increase lipid utilization and protein synthesis. Research evaluating the effect of FA supplementation during uterine and neonatal life on the future productivity of the neonate is warranted.

Improving the performance of dairy cattle with a xylanase-rich exogenous enzyme preparation.

The objective of this experiment was to examine effects of adding 2 exogenous fibrolytic enzymes (EFE) to the total mixed ration (TMR) on the performance of lactating dairy cows (experiment 1) and the kinetics of ruminal degradation of the diet (experiment 2). Twelve EFE had been screened in a series of in vitro assays that identified the most potent EFE and their optimal doses for increasing the digestibility of bermudagrass. In experiment 1, 66 Holstein cows (21±5 d in milk) were grouped by previous milk production and parity (45 multiparous and 21 primiparous) and assigned randomly to 1 of the following 3 treatments: (1) control (CON, untreated), (2) Xylanase Plus [2A, 1mL/kg of TMR dry matter (DM); Dyadic International, Jupiter, FL], and (3) a 75:25 (vol/vol) mixture of Cellulase Plus and Xylanase Plus EFE (3A, 3.4mL/kg of TMR DM; Dyadic International). The EFE were sprayed twice daily onto a TMR (10% bermudagrass silage, 35% corn silage, 5% alfalfa-orchardgrass hay mixture, and 50% concentrates; DM basis) and fed for a 14-d training and covariate period and a 70-d measurement period. Experiment 2 aimed to examine the in situ DM ruminal degradability and ruminal fermentation measurements of the diets fed in experiment 1. Three ruminally fistulated lactating Holstein cows were assigned to the diets. The experiment had a 3×3 Latin square design with 23-d periods. In experiment 1, application of 2A increased intakes (kg/d) of DM (23.5 vs. 22.6), organic matter (21.9 vs. 20.9), and crude protein (3.9 vs. 3.7) and tended to increase yields (kg/d) of fat-corrected milk (41.8 vs. 40.7) and milk fat (1.48 vs. 1.44). In particular, 2A increased milk yield (kg/d) during wk 3 (41.2 vs. 39.8, tendency), 6 (41.9 vs. 40.1), and 7 (42.1 vs. 40.4), whereas 3A increased milk yield (kg/d) during wk 6 (41.5 vs. 40.1, tendency), 8 (41.8 vs. 40.0), and 9 (40.9 vs. 39.5, tendency). In experiment 2, EFE treatment did not affect ruminal DM degradation kinetics or ruminal pH, ammonia-N, and volatile fatty acid concentration. Application of 2A to the bermudagrass-based TMR increased DM intake and milk production, implying that this EFE could be used to increase the performance of lactating dairy cows fed diets containing up to 10% bermudagrass.

Screening exogenous fibrolytic enzyme preparations for improved in vitro digestibility of bermudagrass haylage.

Our objectives were to evaluate the effects of 12 exogenous fibrolytic enzyme products (EFE) on ruminal in vitro neutral detergent fiber digestibility (NDFD) and preingestive hydrolysis of a 4-wk regrowth of bermudagrass haylage (BH), to examine the accuracy of predicting NDFD with EFE activity measures, and to examine the protein composition of the most and least effective EFE at increasing NDFD. In experiment 1, effects of 12 EFE on NDFD of BH were tested. Enzymes were applied in quadruplicate to culture tubes containing ground BH. The suspension was incubated for 24 h at 25 °C before addition of rumen fluid media and further incubation for 24 h at 39 °C. The experiment was repeated twice. In addition, regression relationships between EFE activity measures and NDFD were examined. Compared with the values for the control, 9 EFE-treated substrates had greater NDFD (37.8 to 40.4 vs. 35.6%), 6 had greater total VFA concentration (59.1 to 61.2 vs. 55.4 mM), and 4 had lower acetate-to-propionate ratios (3.03 to 3.16 vs. 3.24). In experiment 2, EFE effects on preingestive fiber hydrolysis were evaluated by incubating enzyme-treated and untreated bermudagrass suspensions in quadruplicate for 24 h at 25 °C and examining fiber hydrolysis measures. Compared with values for the control, 3 EFE reduced neutral detergent fiber concentration (62.8 to 63.7 vs. 67.3%), 10 increased release of water-soluble carbohydrates (26.8 to 58.5 vs. 22.8 mg/g), and 8 increased release of ferulic acid (210 to 391 vs. 198 µg/g). Regression analyses revealed that enzyme activities accurately [coefficient of determination (R(2)) = 0.98] predicted preingestive hydrolysis measures (water-soluble carbohydrates, ferulic acid), moderately (R(2) = 0.47) predicted neutral detergent fiber hydrolysis, but poorly (R(2) ≤ 0.1) predicted dry matter and NDFD. In experiment 3, proteomic tools were used to examine the protein composition of the most and least effective EFE at improving NDFD. Relative to the least effective, the most effective EFE at increasing NDFD contained 10 times more endoglucanase III, 17 times more acetylxylan esterase with a cellulose-binding domain 1, 33 times more xylanase III, 25 times more ß-xylosidase, and 7.7 times more polysaccharide monooxygenase with cellulose-binding domain 1 and 3 times more swollenin. The most effective EFE had a much greater quantity of fibrolytic enzymes and key proteins necessary for hemicellulose and lignocellulase deconstruction. This study identified several EFE that increased the NDFD and in vitro fermentation of 4-wk BH and revealed why some EFE are more effective than others.

Effects of altering the ratio of dietary n-6 to n-3 fatty acids on performance and inflammatory responses to a lipopolysaccharide challenge in lactating Holstein cows.

The study was designed to evaluate the effects of altering the ratio between n-6 and n-3 fatty acids (FA) in the diet and the intake of these FA by lactating dairy cows on lactation performance and inflammatory acute phase responses to a challenge with lipopolysaccharide (LPS). Multiparous Holstein cows (n=45) were blocked based on milk yield from d 6 to d 10 postpartum and, within each block, assigned randomly to 1 of 3 dietary treatments at 14d postpartum; treatments lasted for 90d. Diets were supplemented with a mixture of Ca salts of fish, safflower, and palm oils to create 3 different ratios of n-6 to n-3 FA; namely, 3.9, 4.9, or 5.9 parts of n-6 to 1 part of n-3 FA (R4, R5, and R6, respectively). During the first 5 wk of the study, blood was sampled weekly and analyzed for concentrations of metabolites and hormones. On d 75 postpartum, cows received an infusion of 10µg of LPS into one quarter of the mammary gland to evaluate inflammatory acute phase responses. Altering the ratio of dietary n-6 to n-3 FA was reflected in changes in the FA composition of plasma and milk fat. Reducing the ratio of n-6 to n-3 FA from R6 to R4 increased dry matter intake (24.7, 24.6, and 26.1±0.5kg/d for R6, R5, and R4, respectively), with concurrent increases in yields of 3.5% fat-corrected milk (43.4, 45.4, and 48.0±0.8kg/d), milk fat (1.53, 1.60, and 1.71±0.03kg/d), milk true protein (1.24, 1.28, and 1.32±0.02kg/d), and milk lactose (2.12, 2.19, and 2.29±0.04kg/d). After the LPS challenge, concentrations of IL-6 in plasma increased as the ratio of n-6 to n-3 FA increased (112.5, 353.4, and 365.1±86.6pg/mL for R4, R5, and R6, respectively). Elevations of body temperature and somatic cell count were greater for cows fed R5 compared with those fed R4 or R6 (41.3, 40.8, and 40.8±0.2°C; 4.33, 3.68, and 3.58±0.25×10(6)/mL, for R5, R4, and R6, respectively). Haptoglobin concentration was greatest at 24h after LPS challenge for cows fed R6. Phagocytosis and oxidative burst by neutrophils collected from circulation were unaffected by dietary treatment in the first 48h after intramammary LPS infusion. In conclusion, supplying the same quantity of FA in the diet of early lactation dairy cows but altering the ratio of the polyunsaturated FA of the n-6 to n-3 families influenced lactation performance and inflammatory responses to an LPS challenge.