O. AlZahal

Rumen epithelial adaptation to high-grain diets involves the coordinated regulation of genes involved in cholesterol homeostasis

The molecular mechanisms underlying rumen epithelial adaption to high-grain (HG) diets are unknown. To gain insight into the metabolic mechanisms governing epithelial adaptation, mature nonlactating dairy cattle (n = 4) were transitioned from a high-forage diet (HF, 0% grain) to an HG diet (65% grain). After the cattle were fed the HG diet for 3 wk, they returned to the original HF diet, which they were fed for an additional 3 wk. Continuous ruminal pH, ruminal short chain fatty acids, and plasma β-hydroxybutyrate were measured on a weekly basis, and rumen papillae were biopsied from the ventral sac to assess alterations in mRNA expression profiles. The subacute form of ruminal acidosis was diagnosed during the first week of the HG period (4.6 ± 1.6 h/day <pH 5.6), but not during weeks 2 and 3, thereby indicating ruminal adaption to the HG diet. Changes in the mRNA expression profile of rumen papillae were initially examined using Bovine Affymetrix microarrays; a total of 521 differentially expressed genes (false discovery rate P < 0.08) were uncovered from the first to third week of the HG period. Ingenuity Pathway Analysis of microarray results revealed that enzymes involved in cholesterol synthesis were coordinately downregulated from the first to third week of the HG period. In addition, the LXR/RXR activation pathway was significant and included several genes involved in intracellular cholesterol homeostasis. The differential expression signature of eight genes representing the key regulatory points of cholesterol homeostasis was confirmed by quantitative real-time PCR. Based upon our pathway and network results we propose a model to explain cellular events during rumen epithelial adaptation to HG diets and thus provide molecular targets that may be useful in the treatment and prevention of ruminal acidosis.

Ruminal acidosis and the rapid onset of ruminal parakeratosis in a mature dairy cow: a case report.

A mature dairy cow was transitioned from a high forage (100% forage) to a high-grain (79% grain) diet over seven days. Continuous ruminal pH recordings were utilized to diagnose the severity of ruminal acidosis. Additionally, blood and rumen papillae biopsies were collected to describe the structural and functional adaptations of the rumen epithelium. On the final day of the grain challenge, the daily mean ruminal pH was 5.41 ± 0.09 with a minimum of 4.89 and a maximum of 6.31. Ruminal pH was under 5.0 for 130 minutes (2.17 hours) which is characterized as the acute form of ruminal acidosis in cattle. The grain challenge increased blood beta-hydroxybutyrate by 1.8 times and rumen papillae mRNA expression of 3-hydroxy-3-methylglutaryl-coenzyme A synthase by 1.6 times. Ultrastructural and histological adaptations of the rumen epithelium were imaged by scanning electron and light microscopy. Rumen papillae from the high grain diet displayed extensive sloughing of the stratum corneum and compromised cell adhesion as large gaps were apparent between cells throughout the strata. This case report represents a rare documentation of how the rumen epithelium alters its function and structure during the initial stage of acute acidosis.

Effects of Monensin and Dietary Soybean Oil on Milk Fat Percentage and Milk Fatty Acid Profile in Lactating Dairy Cows

The objective of this study was to investigate the effect of monensin (MN) and dietary soybean oil (SBO) on milk fat percentage and milk fatty acid (FA) profile. The study was conducted as a randomized complete block design with a 2 x 3 factorial treatment arrangement using 72 lactating multiparous Holstein dairy cows (138 +/- 24 d in milk). Treatments were [dry matter (DM) basis] as follows: 1) control total mixed ration (TMR, no MN) with no supplemental SBO; 2) MN-treated TMR (22 g of MN/kg of DM) with no supplemental SBO; 3) control TMR including 1.7% SBO; 4) MN-treated TMR including 1.7% SBO; 5) control TMR including 3.4% SBO; and 6) MN-treated TMR including 3.4% SBO. The TMR (% of DM; corn silage, 31.6%; haylage, 21.2%; hay, 4.2%; high-moisture corn, 18.8%; soy hulls, 3.3%; and protein supplement, 20.9%) was offered ad libitum. The experiment consisted of a 2-wk baseline, a 3-wk adaptation, and a 2-wk collection period. Monensin, SBO, and their interaction linearly reduced milk fat percentage. Cows receiving SBO with no added MN (treatments 3 and 5) had 4.5 and 14.2% decreases in milk fat percentage, respectively. Cows receiving SBO with added MN (treatments 4 and 6) had 16.5 and 35.1% decreases in milk fat percentage, respectively. However, the interaction effect of MN and SBO on fat yield was not significant. Monensin reduced milk fat yield by 6.6%. Soybean oil linearly reduced milk fat yield and protein percentage and linearly increased milk yield and milk protein yield. Monensin and SBO reduced 4% fat-corrected milk and had no effect on DM intake. Monensin interacted with SBO to linearly increase milk fat concentration (g/100 g of FA) of total trans-18:1 in milk fat including trans-6 to 8, trans-9, trans-10, trans-11, trans-12 18:1 and the concentration of total conjugated linoleic acid isomers including cis-9, trans-11 18:2; trans-9, cis-11 18:2; and trans-10, cis-12 18:2. Also, the interaction increased milk concentration of polyunsaturated fatty acids. Monensin and SBO linearly reduced, with no significant interaction, milk concentration (g/100 g of FA) of short- and medium-chain fatty acids (

Rumen epithelial adaptation to ruminal acidosis in lactating cattle involves the coordinated expression of insulin-like growth factor-binding proteins and a cholesterolgenic enzyme

The objective of this study was to characterize the mRNA expression of metabolic and proliferative genes in the rumen epithelium during ruminal acidosis. To meet our objectives, 16 rumen-fistulated, lactating Holstein dairy cattle (618±35 kg of body weight, 221±32 d in milk) were used in a randomized complete block design. All cattle were fed a high-forage diet (HF; 88.9% of dry matter) for 5 wk before the experiment. After the baseline week (wk 0), half of the cattle were randomly assigned and transitioned to a high-concentrate diet (HC; 62.2% of dry matter) which was fed for 3 wk (wk 1, 2, and 3). For the last 48 h of each week, continuous ruminal pH, short-chain fatty acids, and plasma β-hydroxybutyrate were assessed, followed by a rumen papillae biopsy. Milk production was higher in HC cattle compared with HF during wk 1, 2, and 3 (17.4±0.5 vs. 23.4±0.9 kg/d, respectively); however, the mean ruminal pH was decreased (5.75±0.03 vs. 6.30±0.02). The HC cattle spent more time below pH 5.6 (594±54 vs. 3±3 min/d) and displayed greater concentrations of ruminal butyrate (15.8±0.9 vs. 10.2±0.4 mmol) and plasma β-hydroxybutyrate (1,036±63 vs. 778±20 μM) compared with the HF cattle. The mRNA expression of genes involved in ketogenesis (HMGCS2 and PPARA) and short-chain fatty acid transport (MCT1) was unchanged by treatment. However, a downregulation in HMGCS1 (0.72±0.09), one of the cholesterol biosynthesis genes, was observed in HC cattle during wk 1 of the grain challenge. In addition, the relative mRNA expression value of insulin-like growth factor-binding protein 3 was lower (0.78±0.06), whereas insulin-like growth factor-binding protein 5 was higher (1.79±0.15) in HC compared with HF cattle. These results suggest that grain-induced ruminal acidosis alters the mRNA expression of IGF-binding proteins and a cholesterolgenic enzyme in the rumen epithelium of lactating dairy cattle.

The effect of dietary fiber level on milk fat concentration and fatty acid profile of cows fed diets containing low levels of polyunsaturated fatty acids.

The objective of this study was to investigate the effect of dietary fiber level on milk fat concentration, yield, and fatty acid (FA) profile of cows fed diets low in polyunsaturated fatty acid (PUFA). Six rumen-fistulated Holstein dairy cows (639 +/- 51 kg of body weight) were used in the study. Cows were randomly assigned to 1 of 2 dietary treatments, a high fiber (HF; % of dry matter, 40% corn silage, 27% alfalfa silage, 7% alfalfa hay, 18% protein supplement, 4% ground corn, and 4% wheat bran) or a low fiber (LF; % of dry matter, 31% corn silage, 20% alfalfa silage, 5% alfalfa hay, 15% protein supplement, 19% ground wheat, and 10% ground barley) total mixed ration. The diets contained similar levels of PUFA. The experiment was conducted over a period of 4 wk. Ruminal pH was continuously recorded and milk samples were collected 3 times a week. Milk yield and dry matter intake were recorded daily. The rumen fluid in cows receiving the LF diet was below pH 5.6 for a longer duration than in cows receiving the HF diet (357 vs. 103 min/d). Neither diet nor diet by week interaction had an effect on milk yield (kg/d), milk fat concentration and yield, or milk protein concentration and yield. During wk 4, milk fat concentration and milk fat yield were high and not different between treatments (4.30% and 1.36 kg/d for the HF treatment and 4.31% and 1.33 kg/d for the LF treatment, respectively). Cows receiving the LF diet had greater milk concentrations (g/100 g of FA) of 7:0; 9:0; 10:0; 11:0; 12:0; 12:1; 13:0; 15:0; linoleic acid; FA <C16; and PUFA; and lower concentrations of iso 15:0; 18:0; trans-9 18:1; cis-9, trans-11 conjugated linoleic acid (CLA); trans-9, cis-12 18:2; 20:0; and cis-9 20:1 compared with cows receiving the HF diet. Milk concentrations (g/100 g of FA) of total trans 18:1; trans-10 18:1; trans-11 18:1; trans-10, cis-12 CLA, and trans-9, cis-11 CLA were not different between treatments. The study demonstrated that cows fed a diet low in fiber and low in PUFA may exhibit subacute ruminal acidosis and moderate changes to milk fatty acid profile but without concomitant milk fat depression. The changes in FA profile may be useful for the diagnosis of SARA even in the absence of milk fat depression.

Technical note: The use of a telemetric system to continuously monitor ruminal temperature and to predict ruminal pH in cattle

The objective of this study was to compare a telemetric monitoring system to an existing in situ methodology (conventional system) of monitoring ruminal temperature and to validate its use to detect changes in ruminal pH (RpH). Four nonlactating, ruminally cannulated Holstein dairy cows (760 +/- 30 kg of body weight, mean +/- standard deviation) housed in a tie-stall facility were used in the study. The experiment was conducted during the month of May and the recorded ambient temperature was 8.0 +/- 2.0 degrees C (mean +/- SD). The cows were fed a diet consisting of chopped mixed hay (MH; 11.3% crude protein, 59.7% neutral detergent fiber, 17.3% nonfiber carbohydrate, 3.1% ether extract, and 11.3% ash; dry matter basis) during wk 1 and were gradually switched to a high-grain (HG) diet (11.6% crude protein, 30.2% neutral detergent fiber, 50.7% nonfiber carbohydrate, 3.0% ether extract, and 6.0% ash; dry matter basis) during wk 2. A conventional system that utilized an indwelling electrode was used to monitor RpH and ruminal temperature (RT(C)) during d 6 and 7 of each week. The indwelling electrode was attached to a telemetric bolus and ruminal temperature (RT(T)) was logged into a personal computer. The daily mean, minimum, and maximum RpH and duration (min/d) RpH <6.2 were 6.39 +/- 0.04, 6.10 +/- 0.05, 6.66 +/- 0.03, and 107 +/- 50 during MH feeding (wk 1) and 5.84 +/- 0.03, 5.35 +/- 0.05, 6.35 +/- 0.03, and 1,257 +/- 40 during HG feeding (wk 2), respectively, and were different across diets (week effect). Ruminal pH did not decrease below 5.6, 5.8, and 6.0 during MH feeding; mean duration of RpH <5.6, <5.8, and <6.0 during HG feeding was 279 +/- 149, 611 +/- 139, and 894 +/- 101, respectively. Mean daily RT(C) increased from 37.5 degrees C +/- 0.1 in wk 1 to 38.6 degrees C +/- 0.1 in wk 2; there was also an increase from wk 1 to wk 2 in minimum and maximum daily RT(C) and durations (min/d) of RT(C) >38.0, >38.2, >38.4, and >38.6 degrees C. These increases were not detectable with the telemetric system. Ruminal temperature obtained by the conventional system was 0.68 degrees C +/- 0.005 lower than RT(T) during MH feeding (wk 1), whereas RT(C) was 0.04 degrees C +/- 0.004 higher than RT(T) during HG feeding (wk 2). Daily minimum RpH was associated with maximum daily RT(C) and RT(T) during MH and HG feeding (R(2) = 0.88 and 0.43, respectively). There was a high association between low RpH and high ruminal temperature, with the highest associations being between duration (min/d) of RpH <6.0 and duration of RT(C) >39.0 degrees C (R(2) = 0.68) and RT(T) >39.2 degrees C (R(2) = 0.72). Unlike the telemetric system, the conventional system requires cow cannulation; therefore, the current study provided a noninvasive alternative for measuring ruminal temperature and the prediction of RpH. Additional studies are needed to develop an algorithm that accounts for diet type, seasonal variation in temperature, and core body temperature to predict subacute ruminal acidosis effectively on farm.

The periparturient period is associated with structural and transcriptomic adaptations of rumen papillae in dairy cattle

The structural and functional adaption of the rumen epithelium during the transition period is largely undescribed. To characterize the adaptation of the rumen epithelium during transition, multiparous dairy cattle (n=12) fitted with rumen fistulas and fed a low-energy dry cow diet (1.37 Mcal/kg, net energy for lactation) were transitioned abruptly to a high-energy lactating cow diet (1.68 Mcal/kg, net energy for lactation) immediately after parturition. Rumen papillae were biopsied at -3, +1, and +6 wk relative to calving. The histology of morphology of the rumen papillae was evaluated under the light microscope and electron microscope, and mRNA profiling was performed using an Affymetrix GeneChip Bovine Gene 1.0 ST Array (Affymetrix, Santa Clara, CA). Data preprocessing was conducted using the robust multi-array average method, and detection of significant genes was conducted using ANOVA. Also, the Benjamini-Hochberg false discovery rate of 0.1 was applied. Microscopic examination of rumen papillae revealed an increase in epithelial desquamation during early lactation as sloughing scores increased from 1.7 ± 0.2 at -3 wk to 4.1 ± 0.3 and 3.4 ± 0.2 at +1 and + 6 wk, respectively. A total of 1,011 (-3 vs. +1 wk) and 729 (-3 vs. +6 wk) differentially expressed genes were identified (false discovery rate of 0.10, P<10(-3), fold-change ± 1.2 cut-off). A group of differentially expressed genes involved in desmosome assembly (DSG1, CDSN), epidermal growth factor signaling (EGFR, EREG), transforming growth factor β signaling (TGFB1), and the insulin-like growth factor-axis (GHR, IGFBP2, IGFBP3, CTGF) was also validated using PCR. Gene network analysis found that EGFR, GHR, and TGFB1 were focal points of the top pathways, thereby supporting the importance of these regulatory genes to the adaptive response of rumen papillae in early lactation. The microscopic and transcriptomic changes in this study provide insight into the mechanisms responsible for the rapid rate of cellular and molecular adaptations of rumen papillae tissue during the transition period in dairy cattle. In conclusion, the experimental data support the hypothesis that rumen papillae adapt in early lactation by altering their gene expression patterns and, thus, their epithelial structure.

Active dry Saccharomyces cerevisiae can alleviate the effect of subacute ruminal acidosis in lactating dairy cows

The objective of the study was to determine the effect of active dry Saccharomyces cerevisiae (ADSC) supplementation on dry matter intake, milk yield, milk components, ruminal pH, and microbial community during a dietary regimen that leads to subacute ruminal acidosis (SARA). Sixteen multiparous, rumen-cannulated lactating Holstein cows were randomly assigned to 1 of 2 dietary treatments that included ADSC (Biomate; AB Vista, Marlborough, UK; 8 × 10(10) cfu/head per day) or control. During wk 1 to 6, all cows received a high-forage (HF) diet (77:23, forage:concentrate). Cows were then abruptly switched during wk 7 to a high-grain (HG) diet (49:51, forage:concentrate) and remained on the HG until the end of wk 10. Feed intake and milk yields were recorded daily. Ruminal pH was recorded continuously using an indwelling system for 1 to 2 d per week during the pre-experimental phase, and wk 6, 7, and 10. Ruminal digesta samples were collected at the end of the experiment and analyzed for relative change in microbial communities using real-time quantitative PCR. Cows were considered to have SARA if the duration below pH 5.6 was ≥300 min/d. Ruminal pH during wk 6 (HF plateau) was not different across treatments (15 ± 46 min/d at pH <5.6). The dietary regimen successfully induced SARA during wk 7 (transition from HF to HG diet), and ruminal pH (551 ± 46 min/d at pH <5.6) was not different across treatments. However, cows receiving ADSC had an improved ruminal pH (122 ± 57 vs. 321 ± 53 min/d at pH <5.6) during wk 10 (HG plateau) compared with control. Additionally, cows receiving ADSC had a better dry matter intake (23.3 ± 0.66 vs. 21.6 ± 0.61 kg/d) and 4% fat-corrected milk yield (29.6 ± 1.2 vs. 26.5 ± 1.2 kg/d) than control cows during the HG phase (wk 8 to 10). During HG feeding, cows receiving ADSC had greater total volatile fatty acid and propionate concentrations (175 ± 7.5 vs. 154 ± 7.5 and 117 ± 6.1 vs. 94 ± 5.7 mM for ADSC and control, respectively) and lower acetate:propionate ratio (0.26 ± 0.5 vs. 0.36 ± 0.05 for ADSC and control, respectively). Microbial analyses conducted on samples collected during wk 10 showed that cows supplemented with S. cerevisiae had a 9-fold, 2-fold, 6-fold, 1.3-fold, and 8-fold increase in S. cerevisiae, Fibrobacter succinogenes, Anaerovibrio lipolytica, Ruminococcus albus, and anaerobic fungi, respectively, which suggested an increase in cellulolytic microbes within the rumen. Cows supplemented with ADSC had 2.2-fold reduction in Prevotella albensis, which is a gram-negative bacterium predominant during SARA. Prevotella spp. are suggested to be an important source of lipopolysaccharide responsible for inflammation within the rumen. Cows supplemented with ADSC had a 2.3-fold increase in Streptococcus bovis and a 12-fold reduction in Megasphaera elsdenii. The reduction in M. elsdenii may reflect lower concentration of lactic acid within the rumen for ADSC cows. In conclusion, ADSC supplementation to dairy cows was demonstrated to alleviate the condition of SARA caused by abrupt dietary changes from HF to HG, and can potentially improve rumen function, as indicated by greater numbers of cellulolytic microorganisms within the rumen.

The use of a radiotelemetric ruminal bolus to detect body temperature changes in lactating dairy cattle

The objective of this study was to validate the efficacy of a radiotelemetric bolus (RTB) to detect changes in ruminal temperature resulting from (1) systemic illnesses that are associated with febrile responses and (2) subacute ruminal acidosis (SARA). Eight rumen-fistulated, lactating Holstein cows (586±37 kg of body weight, 106±18 d in milk) were used in a replicated 4 × 4 Latin square design with a 2 × 2 factorial arrangement. Each period consisted of 21 d. The factors were 2 diets, a moderate forage:concentrate [MFC; 52:48; % of dry matter (DM)] or a high forage:concentrate (HFC; 65:35, % of DM) total mixed ration, and a challenge with a single intramammary injection of lipopolysaccharide (LPS; 100 μg derived from Escherichia coli 0111:B4) or no LPS (sterile saline). Thus, the 4 resulting treatments were (1) MFC with LPS challenge, (2) MFC with saline, (3) HFC with LPS challenge, and (4) HFC with saline. Cows were fed at 0800 and 1400 h daily. Cows received the intramammary injections at 0900 h of d 21. Ruminal pH and ruminal temperature were also measured on d 21 every minute via an indwelling logging system that resided in the ventral sac of the rumen and via a radiotelemetric bolus that resided in the reticulum. Vaginal temperature was also recorded every minute via temperature loggers. Prior to LPS injection, the duration of rumen pH below 5.6 (indicative of SARA) was higher in cows receiving MFC than cows receiving HFC (148±24 and 62±24 min/d, respectively). The temperature measured at the same time via RTB was higher for MFC than HFC cows (167±21 vs. 104 vs. 21 min/d above 38.8°C, respectively). The following day, cows challenged with LPS showed signs of mastitis within the injected quarters, depressed DM intake, decreased milk yield, and a peak vaginal temperature of 41.3±0.1°C 5.5h after the LPS injection. The RTB system successfully detected a fever response parallel to that measured by the vaginal loggers but temperature peak detected by RTB was, on average, 0.5°C lower than that detected by the vaginal logger. Although the RTB system was able to detect a temperature response to the diet effect before LPS challenge, it was unable to detect this effect during the LPS challenge, likely because cows receiving the LPS challenge had decreased feed consumption. In conclusion, radiotelemetry has the potential to improve the detection of SARA and fever on farm.

Effect of subacute ruminal acidosis on milk fat concentration, yield and fatty acid profile of dairy cows receiving soybean oil

The objective of this study was to investigate the effect of ruminal infusion of soybean oil (SBO) with either a moderate- or high-forage diet on fat concentration, yield and composition in milk from dairy cows. Six rumen-fistulated Holstein dairy cows (639+/-51 kg body weight, 140+/-59 days in milk) were used in the study. Cows were randomly assigned to one of two dietary treatments, a high forage:concentrate (HFC, 74:26) or a moderate forage:concentrate (MFC, 56:44) total mixed ration. Cows were fed at 08.00 and 13.00 h and pulse-dosed ruminally at 13.00 h over a 10-min duration with 2% of diet dry matter of SBO. Ruminal pH was recorded continuously. Cows receiving the MFC treatment had lower daily mean ruminal pH and ruminal pH was below 6.0 for a longer duration compared with the HFC treatment (640 vs. 262 min/d, P<0.05). Cows receiving the MFC treatment had a greater reduction (diet by week interaction, P<0.05) in milk fat concentration and yield than cows receiving the HFC treatment (42 vs. 22% and 45 vs. 21%, respectively). Additionally, cows receiving the MFC diet had a greater reduction in milk fat concentration (g/100 g FA) of FA C16 (17 vs. 9%), trans-10 18:1 (159 vs. 21%) and trans-9, cis-11 conjugated linoleic acid (121 vs. 55%) (P<0.05) compared with cows receiving the HFC diet. This study demonstrated that cows fed the MFC diet had lower ruminal pH and showed a greater rate of milk fat depression when infused with SBO.