Incremental supply of fat, lactose, or protein influences the diurnal pattern of heat production and substrate oxidation in pre-weaning calves.

Increasing nutrient supply to dairy calves has well known benefits; however, the effects of milk replacer (MR) composition when supplied in higher amounts are not fully understood, particularly in the first weeks of life. To better understand the metabolism of macronutrient supply in young calves (21 d old), we investigated diurnal patterns of heat production and substrate oxidation in young calves fed MR with an incremental supply of fat, lactose, or protein. Thirty-two groups of 3 mixed-sex Holstein-Friesian newborn calves (3.4 ± 1.6 d of age), were randomly assigned to one of 4 dietary treatments and studied for 21 d. Diets consisted of a basal MR (23.3% CP, 21.2% EE, and 48.8% lactose of DM) fed at 550 kJ/kg BW0.85 per day (CON; n = 24), or the basal MR incrementally supplied with 126 kJ of DE/BW0.85 per day as milk fat (+FAT; n = 23), lactose (+LAC; n = 24), or milk protein (+PRO; n = 23). Calves were fed MR in 2 daily meals and had ad libitum access to water, but were not supplied with any calf starter nor forage. After 2 weeks of adaptation to the diets, groups of 3 calves were placed for 1 week in an open-circuit respiration chamber for nitrogen and energy balance measurements (lasting 7 d). On d 3, glucose oxidation kinetics was estimated by using [U-13C]glucose. Measurements included total heat production (total energy [HP], activity [Hact] expenditure, resting metabolic rate [RMR]), respiration quotient (RQ), carbohydrate (COX) and fat oxidation (FOX) in 10 min. intervals and averaging these values per hour over days. Incremental supply of lactose and fat increased body fat deposition, with observed patterns in RMR indicating that this increase occurred primarily after the meals. Specifically, the average daily RMR was highest in the +PRO group and lowest in the CON treatment. The HP was higher in the +PRO group and throughout the day, hourly means of HP were higher in this treatment mainly caused by an increase in Hact. The recovery of 13CO2 from oral pulse-dosed [U-13C]glucose was high (77%), and not significantly different between treatments, indicating that ingested lactose was oxidized to a similar extent across treatments. Increasing lactose supply in young calves increased fat retention by reduction in fatty oxidation. Calves fed a MR with additional protein or fat raised RMR persistently throughout the day, while extra lactose supply only affects RMR after the meal. Dietary glucose was almost completely oxidized (77% based on (13C) glucose measurement) regardless of nutrient supplementation. Extra protein supply increased HP and FOX compared with similar intakes of fat and lactose. Fasting heat production (FHP) of young, group-housed calves is comparable to literature values and unaffected by energy intake. Overall, these findings deepen our understanding of how different nutrients impact metabolic processes, fat retention, and energy expenditure in young dairy calves.

Particle size of insoluble fibres and gelation of soluble fibres influence digesta passage rate throughout the gastrointestinal tract of finishing pigs.

Fibres, as abundant in agricultural by-products, exhibit a large range of physicochemical properties that can influence digestive processes such as digesta mean retention time (MRT), thereby affecting nutrient digestion kinetics. In this study, we investigated the effects of particle size of insoluble fibres, and gelation of soluble fibres on MRT of liquids, fine solids, and fibrous particles in the different segments of the gastrointestinal tract (GIT) of pigs. Twenty-four boars (51.6 ± 4.90 kg) were allocated to four diets; two diets contained 15% wheat straw, either coarsely chopped or finely ground (1-mm screen), two diets contained 27% wheat bran without or with the addition of 10% low-methylated pectin. After 14 days of adaptation to the diet, a total collection of faeces was performed to determine the total tract digestibility of nutrients. Thereafter, pigs were fed diets supplemented with tracers for at least 5 days and dissected following a frequent feeding procedure to approach steady-state passage of digesta. The MRT of liquids (Co-EDTA), fine solids (TiO2), and fibrous particles (Chromium-mordanted fibres) in the different segments of the GIT were quantified. In the stomach, particle size reduction of straw decreased the MRT of fine solids by 02:39 h, and fibrous particles by 07:21 h (P < 0.10). Pectin addition to the wheat bran diet reduced the MRT of fine solids by 03:09 h, and fibrous particles by 07:10 h (P < 0.10), but not of liquids, resulting in less separation between digesta phases in the stomach compared with the bran diet (P < 0.05). In the mid-small intestine (SI), pectin addition reduced the MRT of fibrous particles and the separation between fibrous particles and fine solids. No further effects of particle size reduction of straw nor pectin addition on MRT and digestibility of starch, nitrogen, or fat were observed in the SI. In the large intestine (LI), particle size reduction of straw reduced separation between fibrous particles and fine solids (P < 0.10), while pectin addition had no effects. Total tract, non-starch polysaccharide degradation of straw was poor (∼31%), and unaffected by particle size reduction (P > 0.10). The complete fermentation of pectin did not influence the degradation of wheat bran fibres (∼51%). In conclusion, the effects of particle size of insoluble fibres and gelling properties of soluble fibres on the passage of digesta phases were most pronounced in the stomach, but less prominent in distal segments of the GIT.

High variation in the response of calves to a low-dose lipopolysaccharide challenge is associated with early-life measurements.

Lipopolysaccharide (LPS) challenges are commonly used in animal studies as a model for infection with gram-negative bacteria and innate immune activation. We used a low-dose LPS challenge for evaluating interindividual variation in innate immune responses in calves. This was part of a larger study aimed at predicting interindividual variation in feed efficiency in veal calves by variation in feeding motivation, digestion, metabolism, immunology, and behavioral traits. However, due to unexpected high mortality, this LPS challenge was performed in 32 calves rather than in 130 calves, which was initially intended in that larger study, and the 32 calves subjected to the LPS challenge were removed from that larger study. The objective of this short communication is to report the effects of a low-dose LPS challenge in those 32 calves and to examine whether the high variation in calves' responses to LPS could be explained by parameters related to feeding motivation, digestion, behavior, and immunology measured in early life. Thirty-two male Holstein-Friesian calves of Dutch origin were intravenously injected with LPS (0.05 µg/kg of body weight) at an age of 72 ± 0.6 d. Rectal temperature and respiratory frequency were recorded before injection and every hour after injection up to 6 h. In the 8 wk before the LPS challenge, measurements were performed related to general health, feeding motivation, digestion, behavior, and immunity. Following LPS administration, 3 calves died of shock, a fourth calf was euthanized because of severe symptoms of shock and 3 other calves were treated with corticosteroids to counteract observed symptoms of shock. Within the group of 25 relatively mild-responding calves, large interindividual variation in clinical responses to LPS was observed. The maximum increase in rectal temperature varied from 0.6 to 1.9°C and averaged 1.2 ± 0.39°C (coefficient of variation was 32%). The maximum increase in respiratory frequency varied from 16 to 132 bouts/min and averaged 60 ± 28 bouts/min (coefficient of variation was 48%). Little differences were found in early-life measurements between the 7 heavy and 25 mild responders, although heavy responders tended to have a better umbilical hernia score, and had a lower score in a human approach test (i.e., were less reactive) and lower presence of fecal pathogens. The maximum increase in rectal temperature correlated negatively with blood hemoglobin concentration at arrival of the calves at the facilities (r = -0.59) and in wk 4 (r = -0.53). The maximum increase in respiratory frequency correlated negatively with fecal color score (r = -0.43) and positively with fur score in wk 5 (r = 0.50). Overall, mortality (12.5%) and variation in clinical response was high after a low-dose LPS challenge in clinically healthy calves and some hematological and health measurements in early life were related to the clinical response of calves to LPS.

Responses to incremental nutrient supply on energy and protein metabolism in pre-weaning dairy calves.

Recently reviewed development objectives and feeding practices in young dairy calves require an adaptation of nutrient recommendations set for milk replacer (MR) composition. Nutrient requirements of calves younger than 21 d of age, and those of calves fed with high levels of milk replacer are insufficiently quantified. The efficiency at which macronutrients are utilized, particularly protein, substantially diminishes with age, and there is little data for the first weeks of life. In addition, in older (pre-)ruminants, protein and energy can be simultaneously limiting for protein gain. Whether this also applies to calves in the first weeks of life is unknown. Therefore, this study aimed to quantify the responses in protein and fat gain to incremental supply of protein, fat, or lactose to MR in very young calves. Thirty-two groups of 3 mixed-sex Holstein-Frisian newborn calves (3.4 ± 1.6 d of age), were randomly assigned to one of 4 dietary treatments applied for 19 d: a basal MR (23.3% CP, 21.2% CF and 48.8% lactose of DM), provided at 550 kJ/kg BW0.85 per day (CON; n = 24), or the basal MR incrementally supplied with 126 kJ of DE/BW0.85 per day as milk fat (+FAT; n = 23), lactose (+LAC; n = 24) or milk protein (+PRO; n = 23). Calves were fed MR in 2 daily meals and had ad libitum access to water, but did not have access to calf starter nor any other solid feed. After 2 weeks of adaptation to their respective diets, groups of calves were placed for one week in an open-circuit respiration chamber for nitrogen and energy balance measurements (5 d). The incremental nutrient efficiencies indicate what percentage of extra intake of nutrients is retained. In this study, we observed that with every 100 g increase in protein intake, 52% was converted into protein deposition, while 44% contributed to heat production. Similarly, a 100 g increase in fat intake resulted in 67% being stored as fat, 22% being released as heat, and only 5% being retained as protein. Likewise, a 100 g increment in lactose intake led to 49% being stored as fat, with 38% being released as heat. Additional protein intake was not deposited as fat, extra energy intake (fat, and additional lactose) increased post absorptive N efficiency in young calves.

Effects of nondigestible oligosaccharides on inflammation, lung health, and performance of calves.

Our objective was to determine the effects of nondigestible oligosaccharides (NDO) on lung health and performance. Three hundred male Holstein-Friesian calves aged 18.0 ± 3.6 d received 1 of 6 treatments for 8.5 wk (period 1). Treatments included a negative control (CON), galacto-oligosaccharides (GOS) administered as a spray via the nose once daily (SPR), GOS administered via the milk replacer (MR) at 1% (GOS-L) and 2% (GOS-H), fructo-oligosaccharides administered via the MR at 0.25% (FOS) and a combination of GOS and fructo-oligosaccharides administered via the MR at 1% and 0.25%, respectively (GOS-FOS). Milk replacer was fed twice daily. Feeding levels were equal between calves and increased progressively in time. Body weight was measured every 4 wk and clinical health was scored weekly. Blood and broncho-alveolar lavage fluid (BALF) samples were collected bi-weekly from a subset of calves (n = 120). After period 1, all calves received the same control MR for 18 wk until slaughter (period 2), during which general performance and clinical health were measured. Generally, infection pressure was high, with clinical scores and BALF proinflammatory TNFα concentrations increasing with time in period 1, which resulted in a high number of required group antimicrobial treatments (6 group antimicrobial treatments in 13 wk, supplied to all calves). Average daily gain adjusted to equal solid feed intake was increased for GOS-L (+61 g/d) compared with CON calves from experimental wk 1 to 5. Plasma white blood cell concentration tended to be lowered by GOS-L, plasma IL-8 concentration was reduced by all orally supplemented NDO, plasma IL-6 was reduced by all NDO treatments except GOS-FOS and plasma IL-1ß was reduced by all NDO treatments compared with CON, although this differed per time point for SPR. The neutrophil percentage in BALF was reduced by GOS-L in wk 6, which was associated with a relative increase in macrophages. The BALF concentration of TNFα and IL-8 was reduced or tended to be reduced by GOS-L and GOS-H, while IL-6 was or tended to be reduced by SPR, GOS-L, GOS-H, and GOS-FOS, and IL-1ß was reduced by SPR, GOS-L, GOS-H, and FOS. Generally, feeding the combination of GOS and FOS was not more effective than feeding GOS or FOS alone, because feeding GOS-FOS resulted in higher concentrations of plasma and BALF cytokine and chemokine concentrations compared with feeding GOS-L alone, and resulted in higher plasma cytokine concentrations compared with feeding FOS alone. None of the BALF and plasma cytokine or chemokine concentrations differed between the GOS-L and GOS-H treatment. Performance and clinical scores in period 2 did not differ among treatments. Altogether, all tested NDO reduced systemic and lung inflammation in calves under high natural infection pressure and for GOS-fed calves, this increased performance during the first 4 wk. Combining GOS and FOS did not have a synergistic effect. The intranasal administration of GOS also lowered systemic and lung inflammation, but tended to negatively affect performance. Overall, this study demonstrates the potential of NDO to alleviate systemic and respiratory inflammation in calves.

Short communication: Estimation of ileal digestibility in chickens using single- and dual-tracer methods.

Fractionation of digesta, as occurs during gastrointestinal transit in chickens, complicates accurate measurements of ileal digestibility using tracers. Dual-tracer methods using separate tracers for solid and fluid digesta phases may improve the accuracy of digestibility measurements when assumptions of the single tracer method are violated. The aim of the present study was to compare the apparent ileal digestibility (AID) of nutrients calculated with single- and dual-tracer methods in chickens fed diets varying in particle size, anticipating digesta phase separation in the proximal gastrointestinal tract. A total of 112 Dekalb White (BW: 1.53 ± 0.107 kg) and 112 Bovans Black (BW: 1.79 ± 0.127 kg) 29-week-old laying hens were distributed over 32 pens (seven birds/pen). Within breed, pens were randomly assigned to one of two experimental diets (coarse vs fine oat hulls; n = 8 replicate pens per diet/breed combination). Diets were supplemented with TiO2 (3 g/kg) and Co-EDTA (2 g/kg). On days 34, 35, or 36, birds were euthanised and digesta from the ileum was collected for tracer and nutrient analyses. Apparent ileal digestibility was subsequently calculated by single- and dual-tracer methods. Although coarse oat hulls were hypothesised to increase the fractionation of solid and fluid digesta phases, no breed or diet × method interactions were found. Using a single tracer method based on TiO2, AID of nitrogen (N) was overestimated by 3%-units (P < 0.01) compared with the dual-tracer method, whereas AID estimates of DM, starch, fat, and non-starch polysaccharides did not differ (P > 0.09) and precision of all AID estimates was improved. In conclusion, these results show that although from a conceptual perspective, dual-tracer methods are presumed to better account for the variation in flow behaviour of different digesta phases, AID estimates obtained by the commonly used single tracer method using solid-phase tracer TiO2 were more precise and only marginally differed from estimates obtained by a dual-tracer method using distinct tracers for solid (TiO2) and liquid (Co-EDTA) digesta phases. Considering technical and economical constraints, the single tracer method may thus be the method of choice in many situations. Only when digestibility of proteins or amino acids is of specific interest, single tracer methods using a solid-phase tracer may not suffice. Nevertheless, for both single- and dual-tracer methods, tracer selection is critical, and the choice of tracers should depend on the nutrient(s) of interest.

Standardisation of the C:N ratio in ileal digesta changes relationships among fermentation end-products during in vitro hindgut fermentation in pigs.

Undigested proteins that become available for the microbiota in the hindgut can be used as building blocks for bacterial cells, or can enter various catabolic pathways. Degradation via protein fermentation pathways is least preferred, as several fermentation end-products released can be toxic for the host. Directing microbial protein metabolism towards protein synthesis or degradative pathways that result in less toxic end-products, for example through nutritional interventions, is an interesting strategy for improving health. We studied variation in protein fermentation patterns, resulting from variation in substrate composition. Ileal digesta, obtained from cannulated pigs fed different protein sources, were subjected to fermentation in vitro under different conditions; (1) ileal digesta were fermented as-is, (2) ileal digesta were fermented after standardisation to a constant high C:N ratio, by addition of high fermentable carbohydrates and (3) ileal digesta samples were incubated under limiting N concentrations. Gas production was monitored as an indirect measure of microbial activity, and fermentation end-products at different points in time were analysed by gas chromatography and high resolution mass spectrometry. Using principal component analysis, we identified patterns in protein fermentation end-products and related them to the composition of ileal digesta. Protein-associated fermentation end-product concentrations of e.g. isovaleric-, isobutyric-, phenylacetic acid and p-cresol were negatively affected by the available amount of high fermentable carbohydrates combined with a high C:N ratio. The aforementioned fermentation end-products positively correlated with NH3 concentrations and negatively with short-chain fatty acid (SCFA) concentrations. Standardisation to a constant high C:N ratio changed their relationship; isovaleric-, isobutyric-, phenylacetic acid and p-cresol lost their correlation with NH3 concentrations, became positively correlated with SCFA concentrations, and now showed a positive correlation with available amounts of high fermentable carbohydrates. Our observations demonstrate an important role of the C:N ratio in the relationship between fermentation end-products. At constant C:N, protein fermentation end-products correlate with end-products of carbohydrate fermentation and NH3, often considered as a proxy for protein fermentation, loses its predictive power.

SNAPIG: a model to study nutrient digestion and absorption kinetics in growing pigs based on diet and ingredient properties.

Current feed formulation and evaluation practices rely on static values for the nutritional value of feed ingredients and assume additivity. Hereby, the complex interplay among nutrients in the diet and the highly dynamic digestive processes are ignored. Nutrient digestion kinetics and diet × animal interactions should be acknowledged to improve future predictions of the nutritional value of complex diets. Therefore, an in silico nutrient-based mechanistic digestion model for growing pigs was developed: "SNAPIG" (Simulating Nutrient digestion and Absorption kinetics in PIGs). Aiming to predict the rate and extent of nutrient absorption from diets varying in ingredient composition and physicochemical properties, the model represents digestion kinetics of ingested protein, starch, fat, and non-starch polysaccharides, through passage, hydrolysis, absorption, and endogenous secretions of nutrients along the stomach, proximal small intestine, distal small intestine, and caecum + colon. Input variables are nutrient intake and the physicochemical properties (i.e. solubility, and rate and extent of degradability). Data on the rate and extent of starch and protein hydrolysis of different ingredients per digestive segment were derived from in vitro assays. Passage of digesta from the stomach was modelled as a function of feed intake level, dietary nutrient solubility and diet viscosity. Model evaluation included testing against independent data from in vivo studies on nutrient appearance in (portal) blood of growing pigs. When simulating diets varying in physicochemical properties and nutrient source, SNAPIG can explain variation in glucose absorption kinetics (postprandial time of peak, TOP: 20-100 min observed vs 25-98 min predicted), and predict variation in the extent of ileal protein and fat digestion (root mean square prediction errors (RMSPE) = 12 and 16%, disturbance error = 12 and 86%, and concordance correlation coefficient = 0.34 and 0.27). For amino acid absorption, the observed variation in postprandial TOP (61 ± 11 min) was poorly predicted despite accurate mean predictions (58 ± 34 min). Recalibrating protein digestion and amino acid absorption kinetics require data on net-portal nutrient appearance, combined with observations on digestion kinetics, in pigs fed diets varying in ingredient composition. Currently, SNAPIG can be used to forecast the time and extent of nutrient digestion and absorption when simulating diets varying in ingredient and nutrient composition. It enhances our quantitative understanding of nutrient digestion kinetics and identifies knowledge gaps in this field of research. Already useful as research tool, SNAPIG can be coupled with a postabsorptive metabolism model to predict the effects of dietary and feeding-strategies on the pig's growth response.

Short communication: Quantifying postruminal starch fermentation in early-lactation Holstein-Friesian cows.

It has previously been shown that fermentation may contribute substantially to small intestinal carbohydrate disappearance. The fact that the energetic efficiency of starch fermentation is considerably less than that of enzymatic digestion of starch, makes it of nutritional importance to quantify the level of postruminal starch fermentation for dairy cows. Hence, we subjected six rumen-fistulated Holstein-Friesian dairy cows (48 ± 17 days in milk) to 5 d of continuous abomasal infusions of 0.0, 2.5, and 5.0 mol NH4Cl/d, with and without 3 kg ground maize/d, followed by 2 d of rest in a 6 × 6 Latin square design. A total mixed ration (TMR) consisting of (DM basis) 70% grass silage and 30% concentrate was fed at 95% of ad libitum intake. Separation of postruminal starch disappearance into enzymatically digested starch and fermented starch was based on the measurement of natural 13C enrichment of the TMR, abomasally infused ground maize, and resulting 13C enrichment of faeces. Within each cow, 0.0, 2.5, and 5.0 mol NH4Cl/d without ground maize served as control for the same levels of NH4Cl with 3 kg ground maize/d. Abomasal infusion of ground maize was associated with increased total DM and starch intake, faecal starch excretion, and digestibility of starch, and with decreased digestibility of DM and N. The increased faecal volatile fatty acid (VFA) output and 13C enrichment of the individual VFA indicate increased starch fermentation with abomasally infused ground maize. On average, 1 311 g starch/d was postruminally fermented, representing 60.8% of total starch intake. Overall, postruminal starch fermentation of early-lactation dairy cows abomasally infused with 3 kg ground maize/d is considerable and may result in substantial amounts of VFA rather than glucose production.

Changes in body composition and energetic efficiency in response to growth curve and dietary energy-to-protein ratio in broiler breeders.

Body composition plays an important role in reproduction in broiler breeders. The aim of this study was to evaluate the dynamics in body composition and energetic efficiency in broiler breeders, using different dietary strategies. About 1,536-day-old pullets were randomly allotted to 24 pens in a 2 × 4 factorial design with 2 growth curves (standard or elevated (+15%)) and 4 diets, with a step-wise increment in energy (96, 100, 104, and 108% apparent metabolizable energy nitrogen corrected [AMEn]) fed on a pair-gain basis. Body composition was determined at 10 time points from 0 to 60 wk of age. Body protein mass was linearly related to body weight (BW) in growing breeders, which can be expressed as -6.4+0.184*BW (R2 = 0.99; P < 0.001). Body fat mass was exponentially related to BW in growing breeders, which can be expressed as -42.2+50.8*1.0006BW (R2 = 0.98; P < 0.001). A higher energy-to-protein ratio resulted in higher body fat mass at the same BW (P < 0.001). Sexual maturation was related to body protein mass at 21 wk of age, where each 100 g of body protein mass extra advanced sexual maturation by 5.4 d (R2 = 0.83). Estimates of energetic efficiency for growth (kg) and egg production (ke) appeared not constant, but varied with age in a quadratic manner between 0.27 and 0.54 for kg and between 0.28 and 0.56 for ke. The quadratic relationship could be expressed as kg=0.408-0.0319*Age+0.00181*Age2 (R2 = 0.72; P < 0.001) and ke=-0.211+0.034*Age-0.00042*Age2 (R2 = 0.46; P < 0.001). Body protein mass in broiler breeders is tightly regulated and mainly depended on BW and seems to be the main determinant for sexual maturation. Body fat mass is exponentially related to BW, where an increase in dietary energy-to-protein ratio results in a higher body fat mass. Treatments had minimal effects on estimated energetic efficiencies in breeders.

Functional metabolic capacity of pig colonocytes is differentially modulated by fermentable fibre and poorly digestible protein.

The intestine is a highly metabolic organ that relies on energy production within the intestinal cells to sustain its functions. In the colon, intestinal cell metabolic function could be affected positively and negatively by microbiota-derived metabolites. Protein fermentation metabolites are known to negatively impact intestinal metabolic function, while fibre fermentation metabolites are generally thought beneficial. We aimed to investigate whether proteins of different digestibility in the absence and presence of fibres impact the energy metabolism of colonocytes, with potentially adverse health effects. We fed 32, 9-week-old boars one of four experimental diets for 14 days in a 2 × 2 factorial arrangement. Whey and collagen were added as a well and a poorly digestible protein source, respectively, and fibre was either included at 5% or 23%. We examined the effects of the diets on the flux of fermentation metabolites in colon digesta and assessed the impact of the diets on functional metabolic capacity of isolated colonocytes using the Seahorse XF analyzer. Feeding the poorly digestible protein source collagen indeed increased nitrogen flow into the colon by 135% compared to the well-digestible whey-protein source. Feeding high fermentable fibre increased colonic fluxes of both fibre-derived metabolites acetate, propionate, butyrate and caproate, but also increased flux of protein-derived metabolites ammonia, isobutyrate, isovalerate, valerate and isocaproate. To analyse the impact of the diets and the induced differential metabolic composition of the intestinal lumen on functional metabolic capacity of the intestine, we used extracellular flux analysis on freshly isolated pig colonocytes. Colonocytes isolated from high fermentable fibre-fed pigs in the whey-protein diet, but not in the collagen-protein diet, had a reduced mitochondrial capacity, as indicated by a 35% reduction of maximal respiration (interaction P < 0.05) and a 20% reduction of spare respiratory capacity (interaction P < 0.05). Colonocytes from high fermentable fibre-fed pigs had a 37% decreased glycolytic activity compared to the colonocytes isolated from the low fermentable fibre-fed pigs (P < 0.001). This indicated that different diets, and in particular different protein sources and fibre levels, differentially affect colonic epithelial cell metabolism in pigs. Especially, high fermentable fibre lowered both colonocyte mitochondrial and glycolytic metabolism, indicating that high-fibre intake in pigs could lower colonocyte energetic status. Because the metabolic capacity of colonocytes is tightly linked with their functionality, assessment of intestinal cell metabolic capacity may be a valuable tool for future research.

Effects of mixing a high-fat extruded pellet with a dairy calf starter on performance, feed intake, and digestibility.

During weaning, withdrawal of milk replacer is not directly compensated for by an increase in solid feed intake. Therefore, greater fat inclusion in the starter might mitigate this temporary dietary energy decline. However, fat inclusion in solid feeds may generally limit rumen fermentability and development. To address these potentially conflicting outcomes, we conducted 2 experiments to evaluate the effect of supplementing a high-fat extruded pellet mixed with a calf starter on feed intake, performance, and nutrient digestibility in calves. In experiment 1, 60 Holstein bull calves were blocked by serum IgG (2,449 ± 176 mg/dL) and date of arrival (2.5 ± 0.5 d of age). Within each block, calves were randomly assigned to 1 of 3 treatments: a standard control calf starter (CON; 3.1% fat) and mixtures of CON with 10% inclusion of 1 of 2 different high-fat extruded pellets containing 85% of either hydrogenated free palm fatty acids (PFA, 7.1% fat) or hydrogenated rapeseed triglycerides (RFT, 6.7% fat). Calves were offered milk replacer up to 920 g/d until 42 d of age, followed by a gradual weaning period of 7 d. Calves had ad libitum access to the starter diets, straw, and water. No differences were observed between CON, PFA, and RFT calves on body weight (BW) or average daily gain (ADG) until 49 d of age. From weaning (50 d) until 112 d, PFA calves had a greater BW and ADG than RFT and CON animals. Moreover, PFA calves had the highest intakes of starter, straw, calculated metabolizable energy, and crude protein after weaning. Overall, no differences were present in blood ß-hydroxybutyrate and glucose concentrations between treatments; however, calves in the RFT treatment had a higher concentration of insulin-like growth factor-1. In experiment 2, 24 Holstein bull calves at 3 mo of age were assigned to 1 of 8 blocks based on arrival BW and age. Within each block, calves were randomly assigned to 1 of the 3 treatments previously described for experiment 1. Calves on the RFT treatment had the lowest total-tract apparent dry matter and fat digestibility, potentially explaining the differences in performance observed between PFA and RFT calves. Inclusion of the PFA pellet at 10% with a calf starter improved BW, solid feed, and energy intake after weaning. However, these benefits were conditioned by fat source and its digestibility.

Impact of growth curve and dietary energy-to-protein ratio of broiler breeders on offspring quality and performance.

The impact of growth curve (GC) and dietary energy-to-protein ratio of broiler breeder hens on chick quality and broiler performance was investigated. Pullets (n = 1,536) were randomly allotted to 24 pens and assigned to 1 of 8 treatments from hatch onwards, according to a 2 × 4 factorial arrangement with 2 GC (standard growth curve = SGC or elevated growth curve = EGC, +15%) and 4 diets, differing in energy-to-protein ratio (96%, 100%, 104%, and 108% AMEn diet). At 28 and 36 wk of age, 60 hatching eggs per maternal pen were selected for incubation and 768-day-old broilers were assigned to 32 pens according to maternal treatment. Broilers from EGC breeders were 1.9 g heavier at hatch (P < 0.001) and 36 g heavier at slaughter (P = 0.001) than broilers from SGC breeders due to a 1.0 g/d higher growth rate (P = 0.003) and 1.5 g/d higher feed intake (P = 0.006) from hatch to 32 d of age. An increase in breeder dietary energy-to-protein ratio resulted in a linear decrease in embryonic mortality in the first 3 d of incubation (ß = -0.2% per % AMEn; P = 0.05). At hatch, broiler BW decreased with an increasing breeder dietary energy-to-protein ratio (ß = -0.1 g per % AMEn; P = 0.001), whereas at slaughter broiler BW increased with an increasing breeder dietary energy-to-protein ratio (ß = 3.2 g per % AMEn; P = 0.02). This was due to a linear increase in growth rate (ß = 0.1 g/d per % AMEn; P = 0.004) and feed intake (ß = 0.1 g/d per % AMEn; P = 0.02). Additionally, an increase in breeder dietary energy-to-protein ratio resulted in a linear decrease in body weight corrected feed conversion ratio (ß = -0.002 per % AMEn; P = 0.002). Overall, it can be concluded that a higher GC of breeders and an increase in breeder dietary energy-to-protein ratio enhances offspring performance.

Essential amino acid profile of supplemental metabolizable protein affects mammary gland metabolism and whole-body glucose kinetics in dairy cattle.

This study investigated mammary gland metabolism and whole-body (WB) rate of appearance (Ra) of glucose in dairy cattle in response to a constant supplemental level of metabolizable protein (MP) composed of different essential AA (EAA) profiles. Five multiparous rumen-fistulated Holstein-Friesian dairy cows (2.8 ± 0.4 lactations; 81 ± 11 d in milk; mean ± standard deviation) were abomasally infused according to a 5 × 5 Latin square design with saline (SAL) or 562 g/d of EAA delivered in different profiles where individual AA content corresponded to their relative content in casein. The profiles consisted of (1) a complete EAA mixture (EAAC), (2) Ile, Leu, and Val (ILV), (3) His, Ile, Leu, Met, Phe, Trp, Val (GR1+ILV), and (4) Arg, His, Lys, Met, Phe, Thr, Trp (GR1+ALT). A total mixed ration (58% corn silage, 16% alfalfa hay, and 26% concentrate on a dry matter basis) was formulated to meet 100 and 83% of net energy and MP requirements, respectively, and was fed at 90% of ad libitum intake on an individual cow basis. Each experimental period consisted of 5 d of continuous abomasal infusion followed by 2 d of no infusion. Arterial and venous blood samples were collected on d 4 of each period for determination of mammary gland AA and glucose metabolism. On d 5 of each period, D-[U-13C]glucose (13 mmol priming dose; continuous 3.5 mmol/h for 520 min) was infused into a jugular vein and arterial blood samples were collected before and during infusion to determine WB Ra of glucose. Milk protein yield did not differ between EAAC, GR1+ILV, and GR1+ALT, or between SAL and ILV, and increased over SAL and ILV with EAAC and GR1+ILV. Mammary plasma flow increased with ILV infusion compared with EAAC and GR1+ILV. Infusion of EAAC tended to increase mammary gland net uptake of total EAA and decreased the mammary uptake to milk protein output ratio (U:O) of non-EAA compared with SAL. Infusion of ILV increased mammary net uptake and U:O of Ile, Leu, and Val markedly over all treatments. The U:O of total Ile, Leu, and Val increased numerically (25%) with GR1+ILV infusion compared with EAAC, and the U:O of total Arg, Lys, and Thr tended to decrease, primarily from decreased U:O of Lys. During GR1+ALT infusion, U:O of total Arg, Lys, and Thr was greater than that during EAAC infusion, whereas U:O of Ile, Leu, and Val did not differ from EAAC. Glucose WB Ra increased 16% with GR1+ALT over SAL, and increased numerically 8 and 12% over SAL with EAAC and GR1+ILV, respectively. The average proportion of lactose yield relative to glucose WB Ra did not differ across treatments and averaged 0.53. On average, 28% of milk galactose arose from nonglucose precursors, regardless of treatment. In conclusion, intramammary catabolism of group 2 AA increased to support milk component synthesis when the EAA profile of MP was incomplete with respect to casein. Further, WB and mammary gland glucose metabolism was flexible in support of milk component synthesis, regardless of absorptive EAA profile.

Impact of growth curve and dietary energy-to-protein ratio of broiler breeders on egg quality and egg composition.

Egg characteristics have an impact on embryonic development and post-hatch performance of broilers. The impact of growth curve (GC) and dietary energy-to-protein ratio of broiler breeder hens on egg characteristics was investigated. At hatch, 1,536 pullets were randomly allotted to 24 pens in a 2 × 4 factorial dose-response design with 2 GC (standard growth curve = SGC or elevated growth curve = EGC (+ 15%)) and 4 diets, differing in energy-to-protein ratio (defined as 96%, 100%, 104% and 108% AMEn diet). Feed allocation per treatment was adapted weekly to achieve the targeted GC and to achieve pair-gain of breeders within each GC. Breeders on an EGC produced larger eggs (∆ = 2.3 g; P < 0.001) compared to breeders on a SGC. An exponential regression curve, with age (wk) of the breeders, was fitted to describe the impact of GC and dietary energy-to-protein ratio on egg composition. Yolk weight was 0.8 g higher for eggs from EGC breeders than from SGC breeders (a-108.1*0.907Age, where a was 22.1 and 22.9 for SGC and EGC, respectively; R2 = 0.97; P<0.001). An interaction between GC and dietary energy-to-protein ratio on albumen weight was observed (P = 0.04). Dietary energy-to-protein ratio did not affect albumen weight in SGC breeders (42.7-56.2*0.934Age; R2 = 0.89), but for EGC breeders, a higher dietary energy-to-protein ratio resulted in a 0.9 g lower albumen weight from 96% AMEn to 108% AMEn (a-62.9*0.926Age, where a was 43.4, 43.2, 42.8, and 42.5 for 96% AMEn, 100% AMEn, 104% AMEn, and 108% AMEn, respectively; R2 = 0.86). Albumen DM content decreased linearly with an increased dietary energy-to-protein ratio, but this was more profound in EGC breeders (ß = -0.03 %/% AMEn) than in SGC breeders (ß = -0.01 %/% AMEn; P = 0.03). Overall, it can be concluded that an EGC for breeders led to larger eggs with a more yolk and albumen, whereas dietary energy-to-protein ratio had minor effects on egg composition.

Interactive effects of protein and energy intake on nutrient partitioning and growth in Nile tilapia.

Studies of fish growth response to changes in dietary protein and energy content are often conducted with fish fed to apparent satiation or at fixed percentages of their body mass. Such designs result in simultaneous changes in protein and non-protein energy intake, thereby failing to distinguish their separate effects on nutrient partitioning and growth. The present study was designed to address this limitation and test the existence of distinct protein- and non-protein energy-dependent growth phases in Nile tilapia (Oreochromis niloticus). All-male Nile tilapia (63 g, SD = 1.3) were subjected to an 8 × 2 factorial design consisting of eight levels of digestible protein (DP) intake (0.44-1.25 g/day) and two levels of non-protein digestible energy (NPDE) intake (16.0 and 22.4 kJ/day). Fish (n = 960) were housed in 60-litre tanks with two replicates per treatment and hand-fed twice a day for 42 days. Nutrient balances were calculated from changes in body mass, analysed body composition and digestible nutrient intake. Linear regression models were compared to linear-plateau regression models to determine whether protein gain followed distinct protein- and non-protein energy-dependent phases or not. Body mass gain increased linearly with increasing DP intake and was significantly higher (2.6 vs 2.3 g/d, P < 0.05) in fish receiving a high NPDE intake. This increase mainly reflected a higher mean fat gain (0.29 vs 0.20 g/d) rather than a higher protein gain (0.42 vs 0.39 g/d) in fish fed a high vs low level of NPDE intake. The comparison of linear and linear-plateau models did not give clear support for the presence of distinct protein and non-protein energy-dependent phases in protein gain. These results indicate that non-protein energy intake has a modest protein-sparing potential, and that protein gain is simultaneously limited by protein and energy intake in Nile tilapia.

A dual marker technique to estimate individual feed intake in young pigs.

Accurate estimation of individual feed intake (FI) of pigs could help better understand the variation in performance between individual animals. We studied dual marker methods to estimate individual FI in pigs. This method is based on the measurement of the ratio between two indigestible markers in faeces. Twelve 6.5-week-old individually housed male pigs were assigned to one of three oral dosing treatments supplying 180 mg of ytterbium chloride (YbCl3)/day and 111 mg of dotriacontane (C32)/day as reference markers, either once (R1), three times (R3) or five times (R5) daily. Pigs were offered a diet containing 0.46 g/kg of chromium chloride (CrCl3) and 0.15 g/kg of hexatriacontane (C36) as in-feed markers. The experiment lasted for 10 days: days -5 to 0: adaptation; days 1-3: dosing of reference marker; days 2-4: total faecal collection. Spot faecal samples were taken on day 3 at 1200 h, 1700 h and on day 4 at 0700 h. Pigs were fed restrictedly three times daily, at 133.6 g/kg BW0.60. Individual measured FI was recorded daily and was compared to predicted FI using the ratio of the dual marker pairs (Yb:Cr and C32:C36), both in total faecal collection and spot samples. Due to unequal variance, R1 pigs were omitted from the statistical treatment comparison. When using total faecal collection samples, the absolute prediction error (APE) (predicted FI minus measured FI) in R3 and R5 pigs was numerically lower than in R1 pigs, regardless of the marker pair used. The APE measured by C32:C36 was numerically lower than measured by Yb:Cr at all frequencies, and significantly (P = 0.039) in R3 pigs (C32:C36: 0.15 ± 0.02 kg/day; Yb:Cr: 0.29 ± 0.04 kg/day). This was related to a larger difference in faecal recovery between Yb and Cr compared with C32 and C36. Daily total faecal collection revealed that for R3 pigs, starting faecal collections 2 days after the onset of provision of the reference marker improved the APE when compared with starting after 1 day. When using C32:C36 to predict feed intake, pooled, but not single spot samples gave similar APEs compared with total faecal collections. Therefore, we recommend dosing the reference marker three times per day for 2 days on days 1 and 2, combined with pooled spot faecal sampling collected on days 3 and 4. In this way, absolute prediction errors of 10%-15% of simultaneously measured intakes of multiple nutrient resources in a complex housing system are feasible using the dual marker technique.

Early life feeding accelerates gut microbiome maturation and suppresses acute post-weaning stress in piglets.

Early life microbiome perturbations can have important effects on host development, physiology and behaviour. In this longitudinal study, we evaluated the impact of early feeding on gut microbiome colonization in neonatal piglets. Early-fed (EF) piglets had access to a customized fibrous diet from 2 days after birth until weaning in addition to mother's milk, whereas control piglets suckled mother's milk only. Rectal swabs were collected at multiple time points until 6 weeks of age to investigate microbiota development using 16S rRNA gene profiling. The dynamic pre-weaning microbiota colonization was followed by a relatively stable post-weaning microbiota, represented by Prevotella, Roseburia, Faecalibacterium, Ruminococcus, Megasphaera, Catenibacterium and Subdoligranulum. EF piglets showed an accelerated microbiota maturation, characterized by increased microbial diversity, pre-weaning emergence of post-weaning-associated microbes and a more rapid decline of typical pre-weaning microbes. Furthermore, the individual eating behaviour scores of piglets quantitatively correlated with their accelerated microbiome. Importantly, EF piglets displayed a smoother relative weight gain and tended to reach a higher relative weight gain, in addition to reduced diarrhoea scores in the first week post-weaning. Overall, these findings demonstrate the beneficial impact of early feeding on microbiota development as well as pig health and performance during the weaning transition.

Mitochondrial and glycolytic extracellular flux analysis optimization for isolated pig intestinal epithelial cells.

Intestinal epithelial cells (IECs) are crucial to maintain intestinal function and the barrier against the outside world. To support their function they rely on energy production, and failure to produce enough energy can lead to IEC malfunction and thus decrease intestinal barrier function. However, IEC metabolic function is not often used as an outcome parameter in intervention studies, perhaps because of the lack of available methods. We therefore developed a method to isolate viable IECs, suitable to faithfully measure their metabolic function by determining extracellular glycolytic and mitochondrial flux. First, various methods were assessed to obtain viable IECs. We then adapted a previously in-house generated image-analysis algorithm to quantify the amount of seeded IECs. Correcting basal respiration data of a group of piglets using this algorithm reduced the variation, showing that this algorithm allows for more accurate analysis of metabolic function. We found that delay in metabolic analysis after IEC isolation decreases their metabolic function and should therefore be prevented. The presence of antibiotics during isolation and metabolic assessment also decreased the metabolic function of IECs. Finally, we found that primary pig IECs did not respond to Oligomycin, a drug that inhibits complex V of the electron transport chain, which may be because of the presence of drug exporters. A method was established to faithfully measure extracellular glycolytic and mitochondrial flux of pig primary IECs. This tool is suitable to gain a better understanding of how interventions affect IEC metabolic function.

Impact of growth curve and dietary energy-to-protein ratio on productive performance of broiler breeders.

The impact of growth curve (GC) and dietary energy-to-protein ratio on productive performance of broiler breeder females was investigated from 0 to 60 wk of age. One-day-old pullets (n = 1,536) were randomly allotted to 24 pens according to a 2 × 4 factorial arrangement, with 2 GC (standard growth curve = SGC or elevated growth curve = EGC, +15%) and 4 diets, differing in energy-to-protein ratio (96%, 100%, 104%, or 108% AMEn). Feed allocation per treatment was adapted weekly based on the desired GC, meaning that breeders fed the different diets within each GC were fed according to a paired-gain strategy. Linear and quadratic contrasts for energy-to-protein ratio for each GC were evaluated. Elevated growth curve breeders had an earlier sexual maturity (∆ = 4.1 d) than SGC breeders. Egg weight was higher for EGC breeders (∆ = 2.3 g) than for SGC breeders over the whole laying phase (22-60 wk). No differences between EGC and SGC breeders were observed on settable egg production. An increase in dietary energy-to-protein, at a similar BW, led to a linear increase in age at sexual maturity (ß = 0.14 d/% AMEn). From 22 to 40 wk of age, an increase in dietary energy-to-protein ratio led to a linear decrease in egg weight (ß = -0.06 g/% AMEn), regardless of GC. An interaction between GC and dietary energy-to-protein ratio was observed on settable egg production in this phase. An increase in dietary energy-to-protein ratio led to a linear decrease on settable egg production, which was more profound in EGC breeders (ß = -0.70 eggs/% AMEn) than in SGC breeders (ß = -0.19 eggs/% AMEn). From 41 to 60 wk of age, an interaction between GC and dietary energy-to-protein ratio was observed on egg weight. In the EGC, an increase in dietary energy-to-protein ratio led to a linear decrease in egg weight (ß = -0.13 g/% AMEn), whereas in the SGC, a linear increase in egg weight was observed (ß = 0.03 g/% AMEn). From 41 to 60 wk of age, no differences between diets were observed on settable egg production. It can be concluded that a higher GC of breeders has beneficial effects on egg weight, while maintaining settable egg production. Feeding breeders a lower dietary energy-to-protein ratio stimulated productive performance of broiler breeder hens, mainly during the first phase of lay. This effect was more profound when breeders were fed according to a higher GC.