Performance, egg quality, and blood plasma chemistry of laying hens fed hempseed and hempseed oil.

The aim of this study was to compare the performance of hens (feed intake, rate of lay, egg weight, and BW gain), egg quality and blood biochemistry (enzymes, electrolytes, proteins, and other plasma constituents) of laying hens fed diets containing hemp products. Forty-eight Lohmann LSL-Classic (white-egg layers; 19 wk of age) were individually caged and fed 1 of 6 wheat-barley-soybean-based diets for a period of 12 wk. The diets consisted of hempseed (HS; 10, 20, or 30%), hempseed oil (HO; 4.5 or 9.0%), or a control diet (corn oil-based). All diets were formulated to contain similar levels of crude fat (11%), energy (2,800 kcal/kg), and CP (17%). Data were analyzed as a completely randomized design using the repeated measure analysis of the PROC MIXED procedure of SAS. The results indicated that the inclusion of up to 30 and 9.0% HS and HO, respectively, to diets of laying hens had no significant effects on hen performance, egg quality, or plasma level of metabolites (proteins, glucose, uric acid, and cholesterol) and electrolytes (Na, K, Cl, P, and Ca). Overall plasma enzyme concentrations, particularly gamma-glutamyl transferase, were significantly (P < 0.01) lowest at the 10 and 20% levels of HS inclusion, or at the 4.5% HO level of inclusion of the hempseed products compared with the higher levels or control fed hens. Similar effects were also observed for plasma aspartate aminotransferase levels but with the HS enriched diets only (P < 0.05), particularly being lowest at the inclusion levels of 10 and 20% HS compared with the control. The results may imply a possible protective effect of HS- and HO-containing diets, particularly at 10% HS, 20% HS, and 4.5% HO levels, on liver damage/injury. In summary, both HO and HS appear to be well tolerated by laying hens as judged by markers of plasma clinical chemistry supporting the safety and efficacy of hemp products for use in laying hen rations.

Performance, serum biochemical responses, and gene expression of intestinal folate transporters of young and older laying hens in response to dietary folic acid supplementation and challenge with Escherichia coli lipopolysaccharide.

The present study was conducted to investigate the effects of dietary folic acid (FA) supplementation on performance, serum biochemical indices, and mRNA abundance of intestinal folate transporters in young and older laying hens after acute lipopolysaccharide (LPS) challenge. Two experiments were conducted separately involving 48 Shaver White young laying hens (24 wk of age) in experiment 1 and 48 Shaver White older laying hens (58 wk of age) in experiment 2. Birds were fed 2 diets in a complete randomized design. The diets were wheat-soybean meal based, with or without supplemental 4 mg of FA/kg of diet. Birds were fed for 8 wk, during which time feed consumption and egg production were monitored. At the end of each feeding experiment, 6 hens from each dietary treatment were injected intravenously with 8 mg/kg of BW of either Escherichia coli LPS or sterile saline. Four hours after injection, blood and intestinal samples were collected for further analysis. Compared with the control, dietary FA supplementation increased egg weight and egg mass and decreased serum glucose levels in the young laying hens, and reduced serum uric acid in the older laying hens (P < 0.05). Relative to saline injection, plasma homocysteine, serum calcium, and phosphorus levels were found to be lower in both young and older laying hens after LPS challenge (P < 0.05). Other serum biochemical variables and the mRNA expression of 2 folate transport genes in the small and large intestine were differentially affected by LPS challenge, and some of those responses varied with the age of the birds. Additionally, interactions between diet and LPS challenge were specifically found in the older laying hens. In summary, in addition to improving production performance, there were effects of dietary FA supplementation and its interaction with LPS challenge on biochemical constituents, and age played a role in the development of responses to diet and bacterial LPS infections.

Response of older laying hens to an Escherichia coli lipopolysaccharide challenge when fed diets with or without supplemental folic acid.

Folic acid plays a key role in nucleic acids and protein synthesis, and has been associated with anti-inflammatory effects in LPS-induced infections. To this end, a study was conducted to investigate the effects of dietary folic acid (FA) supplementation in older laying hens (58 to 66 wk of age) challenged with Escherichia coli lipopolysaccharide (LPS). A total of 24 Shaver White laying hens at 58 wk were fed 2 diets. The diets were wheat-soybean-based, with either 0 or 4 mg of supplemental FA per kg of diet. After 8 wk of feeding and at 66 wk, the hens were injected intravenously with 8 mg of LPS or saline per kg of BW. Four hours after injection, blood was collected and hens were euthanized to obtain spleen and cecal tonsils. The T cell subsets in the blood and the spleen (CD4+ and CD8+), total IgG, and biochemical constituents (total protein, albumin, globulin, and fibrinogen) were not influenced (P > 0.05) by dietary FA supplementation. However, LPS injection decreased (P < 0.05) biochemical constituents, CD4+, and CD8+ cells in the blood, whereas CD4+:CD8+ ratio and total IgG increased (P < 0.05), and fibrinogen was not influenced. Gene expression in the spleen and cecal tonsils was not influenced by dietary FA supplementation except a diet × challenge interaction for interleukin (IL)-8 in the spleen; IL-8 decreased in FA-fed hens that were treated with LPS. Also, FA supplementation decreased the expression of IL-8 in cecal tonsils. Relative to saline-injected hens, expression of IL-1ß, interferon-γ, and IL-10 increased in the LPS-injected hens in the spleen and cecal tonsils, IL-8 increased in LPS-injected hens only in the cecal tonsils, whereas Toll-like receptor 4, IL-4, IL-17, and IL-18 increased in the LPS-injected hens only in the spleen; however, LPS decreased expression of IL-13 in the cecal tonsils. In conclusion, FA did not affect inflammatory responses in older laying hens; more studies are required to investigate possible protective effects of FA in laying hens.

Characterization of the mechanical properties of high-moisture meat analogues using low-intensity ultrasound: Linking mechanical properties to textural and nutritional quality attributes.

Plant-based meat analogues offer possible alternatives to meat consumption. However, many challenges remain to produce a palatable meat analogue as well as to understand the roles of different processing steps and ingredients on both the texture and nutritional properties of the final product. The goal of this paper is to help with addressing these challenges by using a low-intensity ultrasonic transmission technique, both online and 24 h after production, to investigate high-moisture meat analogues made from a blend of soy and wheat proteins. To understand the ultrasonic data in the context of traditional characterization methods, physical properties (meat analogue thickness, density, peak cutting force) and protein nutritional quality attributes of the meat analogues were also characterized separately. The ultrasonic velocity was found to decrease with the feed moisture content and to be strongly correlated (r = 0.97) with peak cutting force. This strong correlation extends over a wide range of moisture contents from 58% to 70%, with the velocity decreasing from about 1730 m/s to 1660 m/s over this range. The protein quality was high for all moistures, with the highest amino acid score and in vitro protein digestibility being observed for the highest moisture content treatment. The accuracy of the ultrasonic measurements was enhanced by the development of an innovative non-contact method, suitable for materials exhibiting low ultrasonic attenuation, to measure the meat analogue thickness ultrasonically and in a sanitary fashion - an advance that is potentially useful for online monitoring of production problems (e.g., extruder barrel-fill and cooling-die temperature issues). This study demonstrates, for the first time, the feasibility of using ultrasonic transmission techniques to measure both velocity and sample thickness simultaneously and provide information in real time during production that is well correlated with some textural and nutritional attributes of meat analogues.

Effect of feeding hemp seed and hemp seed oil on laying hen performance and egg yolk fatty acid content: evidence of their safety and efficacy for laying hen diets.

Forty-eight 19-wk-old Bovan White laying hens were fed 1 of 5 diets containing either hemp seed (HS) or hemp seed oil (HO). The level of HO was 4, 8, or 12%, whereas the level was 10 or 20% for the HS. A set of 8 birds fed wheat-, barley-, and corn oil-based diets served as the control. Performance was monitored over 12 wk. Average hen-day egg production was not affected upon feeding of either HS or HO diets. Egg weight was higher than that of the controls for hens consuming the 20% HS diet (P < 0.05). Feed intake was lower than that of the controls for birds consuming the 4% HO diet but similar across other treatments. Final BW were not affected by diet, with the exception of being lower than that of the controls (P < 0.05) in hens consuming the 12% HO diet. The total egg yolk n-3 fatty acid content increased linearly (P < 0.05) with increasing dietary α-linolenic acid provision with the HS- or HO-based diets. A quadratic response (P < 0.05) was observed for docosahexaenoic acid levels in egg yolk in response to increasing dietary α-linolenic acid supply. The expression of hepatic fatty acid desaturase 1 and 2, key genes for the desaturation of long-chain polyunsaturated fatty acids, was significantly decreased (50-60% of controls; P < 0.05) as a result of feeding HS or HO diets. Based on the results from the current study, the inclusion of the hemp products HS or HO in the diets of laying hens up to a maximum level of 20 and 12%, respectively, does not adversely effect the performance of laying hens and leads to the enrichment of the n-3 fatty acid content of eggs.

The adaptive transport of folic acid in the intestine of laying hens with increased supplementation of dietary folic acid.

Different aspects of folic acid (FA) transport in the intestine of the laying hen have been characterized. Less is known about the adaptive response of this process to a dietary challenge. To this end, a study was conducted to evaluate the effect of increased dietary FA supplementation on the rate of intestinal FA transport and the expression of the intestinal folate transporter genes, the proton-coupled folate transporters (PCFT) and the reduced folate carrier (RFC), in the laying hen. Twenty-four Shaver White hens at 34 wk of age were randomly assigned to receive 1 of 3 dietary treatments: 1) basal diet with no supplemental folate (n = 8), 2) basal diet + 10 mg/kg of crystalline FA (n = 8), and 3) basal diet + 100 mg/kg of crystalline FA (n = 8). A completely randomized design with 3 dietary treatments was used. Production performance was not affected by FA supplementation; however, egg and plasma folate concentrations increased (P < 0.001), whereas plasma homocysteine concentrations decreased (P < 0.011) in birds fed 10 or 100 mg of FA/kg of diet, relative to controls. Mucosal to serosal uptake of FA in the duodenum was decreased (P < 0.002), but the mRNA levels of the duodenal PCFT and RFC genes were not affected when birds were fed 10 or 100 mg of FA/kg of diet. In the jejunum, the mucosal to serosal uptake of FA, as well as the mRNA levels of the PCFT and RFC genes, were not influenced by increased FA supplementation. Overall, increased dietary levels of FA resulted in decreased transport of FA in the duodenum but not in the jejunum of laying hens. This decrease was not associated with decreased mRNA expression of the duodenal PCFT and RFC genes. Therefore, a posttranscriptional or translational adaptation of the intestinal folate transporters may be involved in the much lower transport of FA in the duodenum of laying hens during increased dietary supplementation of FA.

Immunomodulation in young laying hens by dietary folic acid and acute immune responses after challenge with Escherichia coli lipopolysaccharide.

We investigated the effects of dietary folic acid (FA) supplementation on immunological parameters in young laying hens under acute conditions of lipopolysaccharide (LPS) challenge. Twenty-four Shaver White laying hens at 24 wk were fed 2 diets in a completely randomized design. The diets were wheat-soybean based, with either 0 or 4 mg of supplemental FA per kilogram of diet. At 32 wk of age, 6 hens from each dietary treatment were injected intravenously with 8 mg/kg of BW of either LPS or saline. Four hours after injection, blood was collected and the hens were euthanized to obtain spleen and cecal tonsils. Heterophil:lymphocyte ratio, CD3+, CD4+, CD8+ T cells, and CD4+:CD8+ cells in the blood and spleen were not affected by dietary FA. Relative to saline-injected hens, LPS-injected hens had fewer (P < 0.05) CD3+, CD4+, CD8+, and CD4+:CD8+ cells in the blood, and no difference was found in the spleen. Total protein, albumin, and globulin were found to be higher (P < 0.05) in FA-supplemented hens compared with the control. However, total protein, albumin, and globulins decreased (P < 0.05) in the LPS-injected hens compared with the saline control. Expression of interleukin (IL)-1ß in cecal tonsils decreased (P < 0.05) in FA-supplemented hens, but no dietary influence was found on the expression of other genes in both the spleen and cecal tonsils. Lipopolysaccharide upregulated (P < 0.05) expression of IL-10 and interferon (IFN)-γ in the spleen, and IL-1ß, IL-10, and IFN-γ in the cecal tonsils, whereas the expression of Toll-like receptor (TLR)-4 and IL-8 was not influenced by LPS in the spleen and cecal tonsils. There was a diet × challenge interaction for total IgG, and cytokines IL-1ß and IL-18 in the spleen as well as IL-18 in the cecal tonsils. In conclusion, there were few interactions of dietary FA and LPS; however, FA increased biochemical constituents, enhanced generation of total IgG, as well as exhibiting pleoitropic effects in inflammatory responses.

Local and systemic innate immunity in broiler chickens supplemented with yeast-derived carbohydrates.

A study was conducted to assess the effect of yeast-derived carbohydrates (YDC) on performance and innate immune responses of broiler chickens. In total, 1,080 one-day-old birds were randomly assigned to one of 3 dietary treatments (n = 360): a standard broiler diet containing monensin (control), control + bacitracin methylene disalycylate (BMD), and YDC treatment (control + YDC at 0.02%, 0.01%, and 0.005% for starter, grower, and finisher, respectively). Weekly BW, feed intake (FI), and feed conversion ratio (FCR) were recorded. Immune organ weights, gut morphology, gene expression, heterophil:lymphocyte (H:L), and serum IgG were determined at d 42. No significant difference in FCR, FI, and mortality was observed among treatments. However, BW gain in starter phase was higher in control and YDC treatments compared with BMD treatment. Ileal villi height, crypt depth, and their ratio were not significantly different among treatments, whereas villi width was lower in control and YDC treatments compared with BMD treatment. The number of goblet cells per unit area in the ileum was lower in BMD treatment compared with control and YDC treatments. Expression of TLR2b and IL-6 in the ileum and cecal tonsils was not significantly different among treatments (P > 0.05). Expression of TLR4 was downregulated in YDC treatment compared with control in the ileum. Expression of IL-12p35 and IFN-γ were downregulated in the YDC treatment only in the cecal tonsils. Compared with the control, the expression of IL-10 in both the ileum and the cecal tonsils was downregulated in YDC treatment. Serum IgG and H:L ratio were lower and higher, respectively, in the YDC treatment compared with control and BMD treatments. In conclusion, dietary inclusion of YDC affected intestinal cytokines anti-inflammatory profile on a gut location associated immune pathways manner, suggesting different immune pathways that require further studies in this field.

Innate immune response to yeast-derived carbohydrates in broiler chickens fed organic diets and challenged with Clostridium perfringens.

Necrotic enteritis (NE) caused by Clostridium perfringens is a reemerging disease of economic importance in areas of the world where antibiotic growth promoters have been banned. The effect of mannan-oligosaccharide (MOS) supplementation in organic diets of broilers challenged with C. perfringens on performance, gut morphology, and innate immunity was investigated. Three hundred Ross-308 broilers were fed antibiotic-free certified organic starter and grower diets. On d 14, birds were orally challenged with 1 mL of C. perfringens culture at 3 × 10(10) cfu/bird. Treatments consisted of a control no-challenge (CO; 0 g/kg of MOS in the basal diet), control challenge (COC, 0 g/kg of MOS in the basal diet), and MOS challenge (2 g/kg of MOS in the basal diet). Challenge of birds resulted in decreased feed intake and BW gain (P = 0.048 and P = 0.026, respectively). Even though supplementation of diet with MOS improved feed intake (P = 0.985), BW gain and G:F were not improved compared with those of the CO group (P = 0.026 and P = <0.001, respectively). There was no significant difference among treatments in jejunal and ileal villus height, crypt depth, and goblet cells/mm(2) (P > 0.05). Quantitative real-time PCR showed that, in the ileum, the MOS diet resulted in an upregulation of toll-like receptor (TLR)2b, TLR4, interleukin (IL)-12p35, and interferon (IFN)-γ compared with CO (P = 0.003, P = 0.018, and P = 0.024, respectively). In the cecal tonsil, challenging birds with C. perfringens resulted in an upregulation of TLR2b compared with CO (P = 0.036), and MOS resulted in an upregulation of TLR4 (P = 0.018). In conclusion, feeding a MOS-supplemented diet to C. perfringens-challenged broiler chickens did not improve performance and gut morphology-associated responses. However, MOS was capable of altering TLR and cytokine profiles, where dual TLR2 and TLR4 pathways were associated with MOS supplementation with subsequent upregulation of ileal IL-12p35 and IFN-γ, implying that MOS supplementation in C. perfringens-challenged chickens supports a proinflammatory effect via T-helper cell-1 associated pathways.

Calcium and phosphorus dynamics in commercial laying hens housed in conventional or enriched cage systems.

Calcium (Ca) and phosphorus (P) dynamics in Shaver White hens (19-63 wk of age) were compared between enriched (EC) and conventional cage (CC) systems. Calcium and P intake and their levels in egg components and excreta were considered. Using commercial levels of production (4,836 hens), 10 test cages/system (24 hens/test cage) were used as replicate units. Enriched cages provided a nesting area, scratch pad, perches, and more floor space (643 cm(2)/hen ) than CC (468 cm(2)/hen). All birds were offered similar phase-fed diets based on wheat-soybean formulation and housed under semicontrolled environmental conditions for 11 periods (28 d each). Egg weight, production, and shell quality indices (egg specific gravity, shell weight, thickness, and percentage shell) were also measured. Data were analyzed as a repeated measures design using the MIXED procedure of SAS. Egg production, weight, and shell quality measurements were not significantly different between the 2 systems. On a DM basis, EC hens exhibited lower feed disappearance than CC hens (92.5 vs. 95.0 ± 0.61 g/hen per day, respectively; P < 0.01), and lower Ca and P excretions in manure compared with CC hens (Ca: 2.11 vs. 2.29 ± 0.04 g/hen per day, respectively; P: 0.619 vs. 0.643 ± 0.005 g/hen per day, respectively; P < 0.01). Even though eggs from EC compared with CC had lower Ca deposition (2.07 vs. 2.13 ± 0.01 g/hen per day, respectively; P < 0.0001) and output (38.3 vs. 38.8 ± 0.15 mg/g of egg, respectively; P < 0.05), both EC and CC systems exhibited similar Ca outputs in eggs when expressed as a proportion of Ca intake (56.5 vs. 56.6 ± 0.51% Ca intake, respectively). The overall mean P retention between EC and CC hens was not significantly different (-7.22 vs. -7.45 ± 0.71% P intake, respectively), but Ca retention was higher in EC than CC hens (-1.37 vs. -4.76 ± 0.89% Ca intake, respectively; P < 0.05). In addition to providing environmental enrichment, EC systems may help to reduce Ca and P excretions when compared with CC systems, thereby improving the utilization of these nutrients.

Production performance and nitrogen flow of Shaver White layers housed in enriched or conventional cage systems.

Despite the large number of studies examining the impact of cage systems on Ca and P nutrition, data are limited on the N balance of hens when housed under different systems. To this end, an experiment was conducted to assess N balance, manure characteristics, and indices of the performance of laying hens housed in 2 distinct caging systems. A total of 4,836 commercial Shaver White hens were housed in either enriched (EC) or conventional (CC) cages (average floor space per bird of 643 and 468 cm(2), respectively) under semicontrolled environmental conditions. Enriched cages provided hens with a curtained nesting area, scratch pad, and perches. Birds in both systems were phase fed similar layer diets for 11 periods (4 wk each). Data, expressed on a hen basis, were analyzed as repeated measures using the mixed model procedure of SAS. Lower feed disappearance (P < 0.01; 92.5 vs. 95.0 ± 0.6 g/d, DM basis) and manure output (P < 0.01; 79.8 vs. 91.3 ± 1.2 g/d, as-is basis, and 27.0 vs. 28.1 ± 0.2 g/d, DM basis) were observed in birds housed in EC compared with CC, respectively. Manure DM was 34.1 and 31.0 ± 0.3% for EC and CC, respectively. Egg production, feed conversion ratio, BW, egg weight, and egg mass were not significantly different between the 2 systems. Overall egg N output decreased with age for both cage systems and was not significantly different between the systems. Although no difference was observed in the overall manure N excretion (1.94 and 1.96 ± 0.02 g/d for EC and CC, respectively), hens housed in CC had a significantly (P < 0.05) higher N balance compared with those in the EC system (85.0 vs. 30.2 ± 13.6 mg/d, respectively), which could potentially be explained by a higher (P < 0.05) manure N excretion in the EC at the later stages of production. The current data provide estimates of the efficiency of N utilization in laying hens housed under different housing conditions.

Functional characterization of folic acid transport in the intestine of the laying hen using the everted intestinal sac model.

Absorption at the level of the intestine is likely a primary regulatory mechanism for the deposition of dietary supplemented folic acid into the chicken egg. Therefore, factors affecting the intestinal transport of folic acid in the laying hen may influence the level of egg folate concentrations. To this end, a series of experiments using intestinal everted sacs were conducted to characterize intestinal folic acid absorption processes in laying hens. Effects of naturally occurring folate derivatives (5-methyl and 10-formyltetrahydrofolate) as well as heme on folic acid absorption were also investigated. Folic acid absorption was measured based on the rate of uptake of (3)H-labeled folic acid in the everted sac from various segments of the small and large intestines. Folic acid concentration, incubation length, and pH condition were optimized before the performance of uptake experiments. The distribution profile of folic acid transport along the intestine was highest in the upper half of the small intestine. Maximum uptake rate (nmol·100 g tissue(-1)·min(-1)) was observed in the duodenum (20.6 ± 1.9) and jejunum (22.3 ± 2.0) and decreased significantly in the ileum (15.3 ± 1.1) and cecum (9.3 ± 0.9). Transport increased proportionately (P < 0.05) between 0.0001 and 0.1 µM folic acid. Above 0.1 µM, the slope of the regression line was not significantly different from zero (P < 0.137). Folic acid uptake in the jejunum showed a maximum rate of transport at pH 6.0, but was lowest at pH 7.5. The presence of 5-methyl and 10-formyltetrahydrofolate as well as heme impeded folic acid uptake, reducing intestinal folic acid absorption when added at concentrations ranging from 0 to 100 µM. Overall, these data indicated the presence of a folic acid transport system in the entire intestine of the laying hen. Uptake of folic acid in the cecum raises the likelihood of absorption of bacterial-derived folate.

Effects of phytase supplementation on production performance, egg and bone quality, plasma biochemistry and mineral excretion of layers fed varying levels of phosphorus.

Excessive fecal excretion of phosphorus (P) has increasingly become an environmental issue due to oversupply of P in layer rations, and thus it is imperative to minimize safety margins for P to ensure the sustainability of the egg industry. In this study, a 12-week feeding trial (22 to 34 weeks of age) was conducted to evaluate the effects of phytase supplementation on production performance, plasma biochemistry, egg and bone quality and P excretion of laying hens fed various levels of non-phytate P (NPP). Forty-eight Lohmann white laying hens were randomly allocated to one of six corn-soybean meal-oat-based diets: diets containing 2.0, 2.5 or 3.0 g/kg NPP without phytase, and diets containing 1.0, 1.5 or 2.0 g/kg NPP with phytase (1 000 U/kg diet) where phytase inclusion was expected to provide 1.0 g/kg NPP to laying hens, thus making the phytase-unsupplemented treatment served as a control for the phytase-supplemented treatment accordingly. Productive performance was recorded during the experimental period. Blood and egg samples were collected, and digestibility studies were conducted at weeks 6 and 12 of the experiment. Bone mineralization was evaluated at the end of the experiment. Egg weight and egg production, feed consumption, BW and feed conversion ratio of laying hens fed lower NPP diets supplemented with phytase were comparable to those of hens fed high NPP phytase-unsupplemented controls. Eggshell thickness, specific gravity, Haugh units, tibia bone mineral density, tibia ash percent, plasma P and other biochemical parameters were not significantly different among dietary treatments. Total P intake, excretion and retention were affected by diet (P < 0.001), but its deposition in eggs was not significantly different. Contrast analysis further showed that total P excretion of phytase present vs phytase absent was averagely reduced by 40.4 mg/hen per day (P < 0.01). Moreover, total P excretion was linearly (P < 0.01) reduced with lowering dietary NPP, and this relationship was similar regardless of whether phytase was supplemented or not. The results from this study indicated that NPP levels in laying hen diets could be reduced to 1.0 g/kg (excluding the portion of NPP released by phytase) with the inclusion of phytase, without negative effects on production performance and health of the hens, thereby diminishing P excretion into environment.

Response of nursery pigs to a synbiotic preparation of starch and an anti-Escherichia coli K88 probiotic.

Postweaning diarrhea in pigs is frequently caused by enterotoxigenic Escherichia coli K88 (ETEC). The aim of this study was to test the efficacy of E. coli probiotics (PRO) in young pigs challenged with E. coli K88. We also tested the synbiotic interaction with raw potato starch (RPS), which can be used as a prebiotic. Forty 17-day-old weaned piglets were randomly assigned to four treatments: treatment 1, positive-control diet (C), no probiotics or RPS but containing in-feed antibiotics; treatment 2, probiotic (PRO), no feed antibiotics plus a 50:50 mixture of probiotic E. coli strains UM-2 and UM-7; treatment 3, 14% RPS, no antibiotics (RPS); treatment 4, 14% RPS plus a 50:50 mixture of probiotic E. coli strains UM-2 and UM-7, no antibiotics (PRO-RPS). The pigs were challenged with pathogenic E. coli K88 strains on day 7 of the experiment (24-day-old pigs) and euthanized on day 10 of the experiment (35-day-old pigs). Probiotic and pathogenic E. coli strains were enumerated by selective enrichment on antibiotics, and microbial community analysis was conducted using terminal restriction length polymorphism analysis (T-RFLP) of 16S rRNA genes. The combination of raw potato starch and the probiotic had a beneficial effect on piglet growth performance and resulted in a reduction of diarrhea and increased microbial diversity in the gut. We conclude that the use of E. coli probiotic strains against E. coli K88 in the presence of raw potato starch is effective in reducing the negative effects of ETEC in a piglet challenge model.

Effect of plant sterol-enriched diets on plasma and egg yolk cholesterol concentrations and cholesterol metabolism in laying hens.

Egg exists as a major dietary source of cholesterol in Western diets. In North America, laying hen diets are usually devoid of cholesterol when diets are formulated to exclude animal-based products. Hence, laying hens meet their physiological cholesterol requirement through de novo synthesis. Plant sterols exert a cholesterol-lowering effect in humans by interfering with intestinal sterol absorption. However, it is unknown whether plant sterol supplementation could be effective in reducing intestinal reabsorption of biliary cholesterol in laying hens, thus modulating whole body cholesterol in favor of lower plasma and yolk cholesterol content. The current study was designed to investigate the effect of diets enriched with 0, 0.5, 1, and 2% plant sterols on cholesterol absorption, synthesis, as well as plasma, liver, and egg yolk cholesterol concentrations in laying hens. After 8 wk of plant sterol intervention (first 2 wk were acclimatization), feed intake, BW, egg weight, egg yolk weight, egg production, Haugh units, liver mass, plasma, and hepatic cholesterol concentrations did not differ as a function of plant sterol supplementation. Egg cholesterol concentrations (mg/g) fluctuated during the 6-wk experimental period. At wk 6, a minor reduction in egg yolk cholesterol concentration (mg per g of yolk, P<0.05, vs. control) was observed in hens fed 1 and 2% cholesterol-enriched diets, respectively. However, such result failed to affect total egg cholesterol content. No statistical difference was observed across treatments over 6 wk. Neither cholesterol absorption rates nor synthesis differed as a function of treatment. Results suggested that overall cholesterol content in egg yolk was not affected by feeding hens plant sterol-enriched diets over 6 wk.

Characterization of folate-dependent enzymes and indices of folate status in laying hens supplemented with folic acid or 5-methyltetrahydrofolate.

The conversion of folic acid (FA) to the biologically active 5-methyltetrahydrofolate (5-MTHF) is necessary for the deposition of folate in the egg. A study was conducted to compare egg folate concentrations, indices of folate status, and activities of folate-dependent enzymes in response to equimolar intake of either FA or 5-MTHF in laying hens. Forty-eight laying hens, 24 wk of age, from 2 different strains (Shaver White and Shaver Brown) were randomly assigned to receive 1 of 3 (n = 8 per strain) dietary treatments: 1) basal diet with no supplemental folate, 2) basal diet + 10 mg/kg of FA, or 3) basal diet + 11.3 mg/kg of 5-MTHF for 3 wk. A completely randomized design with 3 dietary treatments and 2 laying hen strains in a 3 x 2 factorial arrangement was used. Data were subjected to ANOVA, using the PROC GLM procedure of SAS. Plasma homocysteine, serum, and egg folate concentrations; hepatic serine hydroxymethyltransferase; and methionine synthase activity were affected by dietary folate supplementation but not by its form (FA and 5-MTHF). Relative to control hens, plasma homocysteine was decreased (P < 0.05) by 14.2%, whereas serum and egg folate were increased (P < 0.05) by 78.3 and 61.8%, respectively, in hens consuming either folate compound. Hepatic serine hydroxymethyltransferase and methionine synthase activity were increased and decreased (P < 0.05), respectively, in folate-fed birds compared with control-fed birds. Hepatic dihydrofolate reductase was influenced by both the addition and form of dietary folate, being higher (P < 0.05) in FA-fed birds than in 5-MTHF and control-fed birds. Feed efficiency was improved (P < 0.05) in 5-MTHF-fed birds relative to FA-fed birds. Strain of hen influenced serum folate and plasma homocysteine concentrations but not other indices of folate metabolism. Overall, FA and 5-MTHF have equivalent effects in enhancing egg folate concentrations and improving folate status in laying hens. Also, supplementation and form of folate may modulate the activity of folate-dependent enzymes.

Proton-coupled folate transporter (PCFT): molecular cloning, tissue expression patterns and the effects of dietary folate supplementation on mRNA expression in laying hens.

1. The aim was to investigate the molecular characterisation and effects of dietary folate supplementation on mRNA concentrations of the proton-coupled folate transporter (PCFT) in chicken. 2. Twenty-four (n = 8/treatment) laying hens were allocated at random to one of three dietary treatments: a basal diet without supplemental folate, or the same diet with either 10·00 mg/kg folic acid or 11·30 (equimolar) mg/kg 5-methyltetrahydrofolate (5-MTHF) for 21 d. The PCFT cDNA was cloned using the TA cloning system, and mRNA concentrations were determined by real-time polymerase chain reaction (PCR). 3. The PCFT cDNA (1643 bp) was cloned from the duodenum and its transcripts were widely distributed in chicken tissues. Duodenal and caecal PCFT mRNA was not affected by dietary treatments; however, jejunal PCFT mRNA was significantly reduced in hens fed the 5-MTHF diet versus the basal diet. 4. A broad tissue distribution of PCFT transcripts is indicative of a vital role of PCFT in the folate transport in chicken. The expression of jejunal PCFT mRNA was down-regulated by dietary 5-MTHF supplementation. These findings contribute to an understanding of the development of folate transport and the PCFT system in avian systems involving laying hens.

Lipidomic characterization of omega-3 polyunsaturated fatty acids in phosphatidylcholine and phosphatidylethanolamine species of egg yolk lipid derived from hens fed flaxseed oil and marine algal biomass.

Membrane phospholipids, including phosphatidylcholine (PC) and phosphatidylethanolamine (PE), consist of distinct fatty acids occupying the sn-1 and sn-2 positions, reflecting the highly regulated nature of lipid biosynthesis. However, little is known about the influence of dietary lipids on the positional nature of fatty acids in tissues, including the enrichment of omega-3 polyunsaturated fatty acid (PUFA) in chicken egg yolk phospholipids. This study was undertaken to characterize the PC and PE species in egg lipids derived from Lohmann hens (n=10/treatment) randomly allocated to either a control (no supplementation), a flaxseed oil (FO) or a marine algal oil (MA) diet. Each of the FO or MA diets supplied three levels of total omega-3 PUFA (0.20, 0.40 and 0.60% of diet) that were provided for 6 weeks. A combination of multiplexed mass spectrometry (MS) experiments are used to determine total, isobaric, and position molecules for PC and PE in egg yolk. The distribution of phospholipids in the yolk was predominantly PC over PE (~72 vs. 23%, respectively) across treatments. The longer chain PUFA existed in the sn-2 position in the PC and PE. Although docosahexaenoic acid (22:6) formed isomers with fatty acids 16:0, 18:0 and 18:1; it was preferentially enriched in the egg in combination with 16:0 with both the FO and MA-fed groups in both lipid pools. All 22:6-containing isomers were enriched by ~2-fold more (P < 0.0001) with MA than FO, however, all isomers exhibited a plateau with the FO-fed group. In addition, the MS analyses of PCs revealed several isobaric species containing eicosapentaenoic acid (EPA, 20:5), however, in the PE, EPA formed only one isomer (i.e. in combination with 16:0). These results may assist to elucidate potential aspects regulating the limited enrichment of omega-3 PUFA, particularly EPA and docosahexaenoic acid (22:6) in chicken eggs.

Effects of additional dosage of vitamin D3, vitamin D2, and 25-hydroxyvitamin D3 on calcium and phosphorus utilization, egg quality and bone mineralization in laying hens.

Vitamin D is essential for the metabolism of calcium (Ca) and phosphorus (P) in birds. The objective of the study was to evaluate the effect of different isoforms of dietary vitamin D on Ca and P utilization, egg quality, and bone mineralization of laying hens. A total of 42 Lohmann white laying hens at 57 wk of age were randomly assigned to 7 dietary treatments for 6 wk. Dietary treatments were: 3,000 IU/kg Vit D3 as control, and control with additional 3,000 IU/kg 25-hydroxyvitamin D3 (T1), 9,000 IU/kg 25-hydroxyvitamin D3 (T2), 3,000 IU/kg vitamin D3 (T3), 9,000 IU/kg vitamin D3 (T4), 3,000 IU/kg of vitamin D2 (T5), or 9,000 IU/kg of vitamin D2 (T6). Egg production and egg quality were measured weekly. Fecal samples were collected at weeks 2 and 6 to measure Ca and P utilization. After 6 wk, the left tibia and femurs were collected to measure bone mineral density (BMD) and bone mineral content (BMC). A 1-way ANOVA with Tukey HSD means separation test was used for statistical analysis. There were no significant differences in egg production, egg quality, BMD, or BMC of tibia and femurs among the treatments (P > 0.05). T6 significantly reduced feed intake (P < 0.05). The apparent total tract digestibility (ATTD) of Ca was higher (P < 0.012) in treatments supplemented with additional vitamin D, irrespective of forms. The ATTD of P was higher (P < 0.0001) in T5 compared to the other treatments at both time points. The utilization of Ca and P by laying hens can be improved through the addition of different isoforms of vitamin D in diets. However, additional vitamin D supplementation to laying hens, regardless of forms, had no effect on either bone mineralization or measures of egg quality.

Effect of dietary n-3 polyunsaturated fatty acids on the composition of cecal microbiome of Lohmann hens.

Supplementation of n-3 fatty acids to poultry diets is widely acknowledged for its role in enhancing poultry products, however, little is known about the compositional responses of gut microbial communities to type and dosage of these supplements.  Here, we compared the effects of n-3 polyunsaturated fatty acids (PUFA), supplied as alpha-linolenic acid (ALA) or docosahexaenoic acid (DHA), on the composition of bacterial communities in ceca of laying hens.  Corn-soybean basal diets were supplemented with either flaxseed oil (FO, ALA-rich) or marine algal biomass (MA, DHA-rich), and each supplied 0.20 and 0.60% of total n-3 PUFA in the diet.  Lohmann LSL-Classic laying hens (n = 10/treatment) were randomly allocated to one of the 4 diets.  After 8 weeks of feeding, blood, liver and cecal digesta samples were obtained for plasma glucose, fatty acids, and short chain fatty acids analyses, respectively.  The gut bacterial communities were characterized using genomic DNA extracted from cecal contents, whereby the V3-V4 hypervariable region of the 16S rRNA gene was sequenced using the Illumina Miseq® platform.  Firmicutes and Bacteroidetes were the predominant phyla in both the FO- and MA-fed groups.  The relative abundance of Tenericutes, often associated with immunomodulation, was relatively higher (P<0.0001) in the FO than MA group.  Although the relative abundance of Bacteroides was greater for the FO- than the MA-fed group, this genus was negatively correlated (P<0.05) with total n-3 PUFA in the liver at higher dosages of both FO- and MA-fed hens.  Higher dose of FO (0.60%) and both dosages of MA (0.20 and 0.60%) substantially enriched several members of Firmicutes (e.g., Faecalibacterium, Clostridium and Ruminococcus) which are known to produce butyrate.  Moreover, co-occurrence network analysis revealed that, in the FO 0.60- and MA 0.20-fed hens, Ruminococcaceae was the most influential taxon accounting for about 31% of the network complexity.  These findings demonstrate that supplementation of different type and level of n-3 PUFA in hens' diets could enrich microbial communities with potential role in lipid metabolism and health.