Effects of supplemental methionine sources in finishing pig diets on growth performance, carcass characteristics, cutting yields, and meat quality.

Supplemental methionine (Met) is widely used within the swine industry; however, data are limited regarding the effect of Met sources on carcass cutability and meat quality. The objective was to determine the effects of L-Met (LM, 99%), DL-Met (DLM, 99%), or calcium salt of DL-Met hydroxyl analog (MHA, 84%) in finishing pig diets on carcass characteristics and meat quality. At 9 weeks of age, pigs (N = 240) were allocated to 60 single-sex pens for a four-phase finishing trial that lasted 104 d. Pigs were fed a common grower diet until day 56 where pens were randomly allotted to one of the three experimental diets. For the remaining 7 wk of the finisher phase, pigs (BW = 79.9 ±â€…0.80 kg) were fed diets containing LM, DLM, or MHA, with the supplemental Met source providing 25% of standardized ileal digestible (SID) Met + cysteine (Cys) requirement based on 65% bioefficacy for MHA in comparison with LM or DLM. One pig per pen was slaughtered at the study conclusion (on day 104), and the left sides of carcasses were fabricated into subprimal cuts to determine carcass-cutting yields. Loin quality including proximate composition and shear force were measured. Hot carcass weight was not different (P = 0.34) between treatments (LM 104.5 kg; DLM 103.0 kg; MHA 101.5 kg), moreover, loin eye area was not different (P = 0.98) between treatments (LM 52.65 cm²; DLM 52.49 cm²; MHA 52.81 cm²). Boneless carcass-cutting yield was not different (P = 0.56) between treatments (LM 54.97 kg; DLM 54.82 kg; MHA 54.52 kg). Loin pH was not different (P = 0.24) between treatments (LM 5.45; DLM 5.48; MHA 5.45). However, drip loss tended to be reduced (P = 0.11) by the DLM treatment (5.58%) compared with LM (7.03%) and MHA (6.68%) treatments. Shear force was not different (P = 0.85) between treatments (LM 3.03 kg; DLM 3.06 kg; MHA 3.10 kg). However, cook loss tended to be reduced (P = 0.06) by the DLM treatment (16.20%) compared with LM (18.18%) and MHA (18.50%) treatments. These data suggest that only minimal differences in carcass cutability and meat quality can be attributed to Met source in finishing pig diets when using 65% bioefficacy for MHA relative to L-Met or DL-Met.

Dietary monoglyceride supplementation to support intestinal integrity and host defenses in health-challenged weanling pigs.

Frequent incidence of postweaning enterotoxigenic Escherichia coli (ETEC) diarrhea in the swine industry contributes to high mortality rates and associated economic losses. In this study, a combination of butyric, caprylic, and capric fatty acid monoglycerides was investigated to promote intestinal integrity and host defenses in weanling pigs infected with ETEC. A total of 160 pigs were allotted to treatment groups based on weight and sex. Throughout the 17-d study, three treatment groups were maintained: sham-inoculated pigs fed a control diet (uninfected control [UC], n = 40), ETEC-inoculated pigs fed the same control diet (infected control [IC], n = 60), and ETEC-inoculated pigs fed the control diet supplemented with monoglycerides included at 0.3% of the diet (infected supplemented [MG], n = 60). After a 7-d acclimation period, pigs were orally inoculated on each of three consecutive days with either 3 mL of a sham-control (saline) or live ETEC culture (3 × 109 colony-forming units/mL). The first day of inoculations was designated as 0 d postinoculation (DPI), and all study outcomes reference this time point. Fecal, tissue, and blood samples were collected from 48 individual pigs (UC, n = 12; IC, n = 18; MG, n = 18) on 5 and 10 DPI for analysis of dry matter (DM), bacterial enumeration, inflammatory markers, and intestinal permeability. ETEC-inoculated pigs in both the IC and MG groups exhibited clear signs of infection including lower (P < 0.05) gain:feed and fecal DM, indicative of excess water in the feces, and elevated (P < 0.05) rectal temperatures, total bacteria, total E. coli, and total F18 ETEC during the peak-infection period (5 DPI). Reduced (P < 0.05) expression of the occludin, tumor necrosis factor α, and vascular endothelial growth factor A genes was observed in both ETEC-inoculated groups at the 5 DPI time point. There were no meaningful differences between treatments for any of the outcomes measured at 10 DPI. Overall, all significant changes were the result of the ETEC infection, not monoglyceride supplementation.

Infection caused by the bacterium known as enterotoxigenic Escherichia coli (ETEC) is a common disruptor of weaned pigs' health, leading to economic losses for the producers. To determine if nutritional supplementation could help protect against these losses, weaned pigs were assigned to one of three treatments: 1) uninfected and fed a standard nursery pig diet, 2) infected with ETEC and fed the same standard diet, or 3) infected with ETEC and fed the standard diet supplemented with a combination of butyric, caprylic, and capric fatty acid monoglycerides. Growth performance was tracked throughout the 17-d study and health outcomes were measured at the peak and resolution of ETEC infection. At the peak-infection time point, pigs that were infected with ETEC had lower fecal moisture content, greater fecal bacterial concentrations, and elevated body temperatures compared with uninfected pigs. Additionally, infection reduced expression of genes related to inflammation, angiogenesis, and the intestinal barrier during the peak-infection period. Overall, all significant changes were the result of the ETEC infection, and there were no meaningful differences observed between the different treatments.

Acute Microbial Protease Supplementation Increases Net Postprandial Plasma Amino Acid Concentrations After Pea Protein Ingestion in Healthy Adults: A Randomized, Double-Blind, Placebo-Controlled Trial.

BACKGROUND: Digestibility is a primary factor in determining the quality of dietary protein. Microbial protease supplementation may be a strategy for improving protein digestion and subsequent postprandial plasma amino acid availability. OBJECTIVES: To assess the effect of co-ingesting a microbial protease mixture with pea protein on postprandial plasma amino acid concentrations. DESIGN: A mixture of 3 microbial protease preparations (P3) was tested for proteolytic efficacy in an in vitro static simulation of gastrointestinal digestion. Subsequently, in a randomized, double-blind, placebo-controlled crossover trial, 24 healthy adults (27 ± 4 y; 12 females, 12 males) ingested 25 g pea protein isolate (20 g protein, 2.2 g fat) with either P3 or maltodextrin placebo (PLA). Blood samples were collected at baseline and throughout a 0‒5 h postprandial period and both the early (0-2 h) iAUC and total (0-5 h) iAUC were examined. RESULTS: Plasma glucose concentrations decreased in both conditions (P < 0.001), with higher concentrations after P3 ingestion compared with PLA (P < 0.001). Plasma insulin concentrations increased for both conditions (P < 0.001) with no difference between conditions (P = 0.331). Plasma total amino acid (TAA) concentrations increased over time (P < 0.001) with higher concentrations observed for P3 compared with PLA (P = 0.010) during the 0‒5 h period. There was a trend for elevated essential amino acid (EAA) concentrations for P3 compared with PLA (P = 0.099) during the 0‒5 h postprandial period but not for leucine (P = 0.282) or branched-chain amino acids (BCAA, P = 0.410). The early net exposure (0‒2 h iAUC) to amino acids (leucine, BCAA, EAA, and TAA) was higher for P3 compared with PLA (all, P < 0.05). CONCLUSIONS: Microbial protease co-ingestion increases plasma TAA concentrations (0-5 h) and leucine, BCAA, EAA, and TAA availability in the early postprandial period (0‒2 h) compared with ingesting pea protein with placebo in healthy adults.

Organ growth and fermentation profiles of broilers differing in body growth rate.

This study sought to determine the relationship among broiler performance, organ development, and indicators of microbiota colonization. A total of 1,200 two-day-old male Ross 308 broiler chicks, divided among 3 cohorts of equal size, were housed in battery cages, and allotted based on body weight. On study d 11, birds were weighed, and birds with BW gain within the 10th and 90th percentiles were assigned to the Slow and Fast groups, respectively. Birds (n = 30 for each group) selected on d 11 were provided water and a corn-soybean meal-based diet ad libitum while maintained individually through study d 25 (i.e., a 14-d growth period). Parameters regarding growth performance, organ and intestine weights and lengths, and intestinal volatile fatty acid concentrations were measured. All data were analyzed by one-way ANOVA using the Mixed procedure of SAS. Fast birds exhibited greater (P < 0.001) BW gain and feed intake than slow birds, but feed conversion ratio (FCR) did not differ (P = 0.19). Additionally, Slow birds had higher (P < 0.05) relative weights (% of BW) for nearly all organs on d 11 and 25, most notably the gizzard, proventriculus, pancreas, and liver. Conversely, intestinal sections were longer (P < 0.05) in the Fast birds. Measurement of gut histomorphology did not show any notable differences between growth rate groups in terms of villi height, crypt depth, or their ratio for either time-point (P > 0.05). In terms of volatile fatty acid concentrations of luminal contents, acetate concentrations were 10.2% higher (P < 0.001) in the ileum of the Slow birds compared with Fast birds on d 25. Overall, the findings suggest that total BW gain is influenced by the development of metabolically active organs, as supported by lower weight gain in Slow birds with relatively larger organ weights and shorter intestinal lengths than their Fast counterparts. The general lack of differences in fermentation end-product concentrations in luminal contents does not rule out influence of the microbiota on growth rate of broilers, which warrants further investigation.

The effect of chronic, non-pathogenic maternal immune activation on offspring postnatal muscle and immune outcomes.

The objective was to determine the effects of maternal inflammation on offspring muscle development and postnatal innate immune response. Sixteen first-parity gilts were randomly allotted to repeated intravenous injections with lipopolysaccharide (LPS; n = 8, treatment code INFLAM) or comparable volume of phosphate buffered saline (CON, n = 8). Injections took place every other day from gestational day (GD) 70 to GD 84 with an initial dose of 10 µg LPS/kg body weight (BW) increasing by 12% each time to prevent endotoxin tolerance. On GD 70, 76, and 84, blood was collected at 0 and 4 h postinjection via jugular or ear venipuncture to determine tumor necrosis factor (TNF)-α, interleukin (IL)-6, and IL-1ß concentrations. After farrowing, litter mortality was recorded, and the pig closest to litter BW average was used for dissection and muscle fiber characterization. On weaning (postnatal day [PND] 21), pigs were weighed individually and 2 barrows closest to litter BW average were selected for another study. The third barrow closest to litter BW average was selected for the postnatal LPS challenge. On PND 52, pigs were given 5 µg LPS/kg BW via intraperitoneal injection, and blood was collected at 0, 4, and 8 h postinjection to determine TNF-α concentration. INFLAM gilt TNF-α concentration increased (P < 0.01) 4 h postinjection compared to 0 h postinjection, while CON gilt TNF-α concentration did not differ between time points. INFLAM gilt IL-6 and IL-1ß concentrations increased (P = 0.03) 4 h postinjection compared to 0 h postinjection on GD 70, but did not differ between time points on GD 76 and 84. There were no differences between INFLAM and CON gilts litter mortality outcomes (P ≥ 0.13), but INFLAM pigs were smaller (P = 0.04) at birth and tended (P = 0.09) to be smaller at weaning. Muscle and organ weights did not differ (P ≥ 0.17) between treatments, with the exception of semitendinosus, which was smaller (P < 0.01) in INFLAM pigs. INFLAM pigs tended (P = 0.06) to have larger type I fibers. INFLAM pig TNF-α concentration did not differ across time, while CON pig TNF-α concentration peaked (P = 0.01) 4 h postinjection. TNF-α concentration did not differ between treatments at 0 and 8 h postinjection, but CON pigs had increased (P = 0.01) TNF-α compared to INFLAM pigs 4 h postinjection. Overall, maternal immune activation did not alter pig muscle development, but resulted in suppressed innate immune activation.

Maternal inflammation or immune activation impacts fetal development and subsequently the offspring's postnatal performance. In particular, maternal immune activation may be detrimental to fetal muscle development and alter postnatal immune responses, both of which are vital in determining livestock efficiency. However, understanding the relationship between maternal immune activation and offspring development is difficult as many models use a live pathogen. This introduces many confounding factors, including increased mortality, persistent postnatal infection, and potential copathogens. Therefore, the objective of this study was to determine the effect of maternal inflammation on offspring muscle development and postnatal inflammatory response using repeated injections of a nonpathogenic immune stimulant. Each injection successfully induced an inflammatory response as indicated by increased rectal temperature and circulating inflammatory markers. The gestational challenge did not result in increased litter mortality. Further, muscle development was not altered in piglets exposed to gestational inflammation. However, when challenged with the same immune stimulant given to the dams, pigs exposed to maternal inflammation had a remarkably suppressed immune response compared to controls. Overall, maternal inflammation independent of infection affected offspring immune function, but not muscle development.

The probiotic Lacticaseibacillus rhamnosus HN001 influences the architecture and gene expression of small intestine tissue in a piglet model.

This study investigated the effects of Lacticaseibacillus rhamnosus HN001 supplementation on the architecture and gene expression in small intestinal tissues of piglets used as an animal model for infant humans. Twenty-four 10-d-old entire male piglets (4·3 (sd 0·59) kg body weight) were fed an infant formula (IF) (control) or IF supplemented with 1·3 × 105 (low dose) or 7·9 × 106 (high dose) colony-forming units HN001 per ml of reconstituted formula (n 8 piglets/treatment). After 24 d, piglets were euthanised. Samples were collected to analyse the histology and gene expression (RNAseq and qPCR) in the jejunal and ileal tissues, blood cytokine concentrations, and blood and faecal calprotectin concentrations. HN001 consumption altered (false discovery rate < 0·05) gene expression (RNAseq) in jejunal tissues but not in ileal tissues. The number of ileal goblet cells and crypt surface area increased quadratically (P < 0·05) as dietary HN001 levels increased, but no increase was observed in the jejunal tissues. Similarly, blood plasma concentrations of IL-10 and calprotectin increased linearly (P < 0·05) as dietary HN001 levels increased. In conclusion, supplementation of IF with HN001 affected the architecture and gene expression of small intestine tissue, blood cytokine concentration and frequencies, and blood calprotectin concentrations, indicating that HN001 modulated small intestinal tissue maturation and immunity in the piglet model.

The dual sugar test (lactulose plus mannitol) is an unreliable indicator of feed withdrawal-associated changes in intestinal permeability in broiler chickens.

Despite its importance in poultry research, there is lack of standardized and practical techniques to measure intestinal permeability in a noninvasive manner. Therefore, this research sought to standardize a procedure using lactulose (Lac) and mannitol (Man) to measure intestinal barrier function in broilers. Twenty-one-day-old male and female Ross 308 birds were orally gavaged (either 2 mL/kg BW or fixed 3 mL per bird) with a solution containing 5 to 25 g Lac and 1 to 5 g Man dissolved in pure water to reach 100 mL of final solution. Feed withdrawal (FW; 2-24-h duration) prior to dosing with Lac and Man (LacMan) was mainly used to induce graded intestinal permeability. Blood samples were collected at 60-, 90-, or 120-min after LacMan dosing using serum or plasma (K2EDTA and/or Na-Heparin) blood tubes. Lac and Man concentrations were quantified by HPLC. Plasma samples collected 90-min after LacMan dosing elicited the least variable response (22.4% vs. 22.8% or 23.4% CV when compared with 60- and 120-min sampling time-points, respectively), and both markers were detectable after administering a solution containing the lowest concentration of Lac and Man. However, analytical problems arose when using Na-Heparin anticoagulant as high glucose levels interfered with Lac quantification. Upon improving the chromatographic technique, it became evident that a 24-h FW increased (P < 0.01) Lac concentrations. In the last trial, a more severe glucose interference was observed, resulting in no Lac detection within an entire treatment group. Twelve hours of FW increased (P < 0.01) Man concentration in the plasma of birds receiving the solution containing 3 g Man, but had no effect on the birds receiving the solution containing 5 g Man. A 24-h FW did not affect the Man concentrations in birds receiving the solution containing either 3 or 5 g Man. With inconsistency of Lac detection throughout our trials, it was concluded that the combination of Lac and Man is an unreliable marker to predict intestinal barrier function in broilers.

Determining the safety and efficacy of dietary supplementation with 3'-sialyllactose or 6'-sialyllactose on growth, tolerance, and brain sialic acid concentrations.

Sialylated oligosaccharides, including 3'-sialyllactose (3'-SL) and 6'-sialyllactose (6'-SL), comprise a large portion of human milk and have been known to support development over the first year of life. While research has investigated the impact of early-life supplementation, longer-term supplementation remains relatively unexplored. Consequently, the following study assesses the impact of supplementation of either 3'-SL or 6'-SL on growth performance, tolerance, and brain sialic acid concentrations. Two-day-old piglets (n = 75) were randomly assigned to a commercial milk replacer ad libitum without or with 3'-SL or 6'-SL (added at 0.2673% on an as-is basis). Daily body weight and feed disappearance were recorded to assess growth performance and tolerance. Pigs were euthanized for sample collection on postnatal day 33 (n = 30) or 61 (n = 33), respectively. Across growth performance, clinical chemistry and hematology, histomorphology, and sialic acid quantification, dietary differences were largely unremarkable at either time-point. Overall, SA was well-tolerated both short-term and long-term.

Metabolite profiling of peripheral blood plasma in pigs in early postnatal life fed whole bovine, caprine or ovine milk.

Ruminants' milk is commonly used for supplying nutrients to infants when breast milk is unavailable or limited. Previous studies have highlighted the differences between ruminants' milk composition, digestion, absorption, and fermentation. However, whether consuming different ruminants' milk impact the appearance of the circulatory blood metabolites in the early postnatal life is not well understood. The analysis conducted here aimed to determine the effect of feeding exclusively whole milk from bovine, caprine or ovine species to pigs, approximately 7 days-old for 15 days, on circulatory blood plasma metabolites. Relative intensities of plasma metabolites were detected using a liquid chromatography-mass spectrometry based metabolomic approach. Seven polar and 83 non-polar (lipids) metabolites in plasma were significantly different (false discovery rate < 0.05) between milk treatments. These included polar metabolites involved in amino acid metabolism and lipids belonging to phosphatidylcholine, lysophosphatidylcholine, sphingomyelin, and triglycerides. Compared to the caprine or bovine milk group, the relative intensities of polar metabolites and unsaturated triglycerides were higher in the peripheral circulation of the ovine milk group. In contrast, relative intensities of saturated triglycerides and phosphatidylcholine were higher in the bovine milk group compared to the ovine or caprine milk group. In addition, correlations were identified between amino acid and lipid intake and their appearance in peripheral blood circulation. The results highlighted that consuming different ruminants' milk influences the plasma appearance of metabolites, especially lipids, that may contribute to early postnatal life development in pigs.

Assessing the urinary concentration of nitrofurantoin and its antibacterial activity against Escherichia coli, Staphylococcus pseudintermedius, and Enterococcus faecium isolated from dogs with urinary tract infections.

Nitrofurantoin, a broad-spectrum nitrofuran class antibiotic, is applied as a first-line antibiotic in treating human urinary tract infections (UTIs) due to its great efficacy and high achievable concentration. The interest in using this antibiotic in companion animals has increased due to the growing demand for effective antibiotics to treat UTIs caused by multidrug-resistant bacteria. Currently, the susceptibility interpretations for nitrofurantoin are based on the breakpoints set for humans, while the canine-specific breakpoints are still unavailable. In this study, we assessed the concentration of nitrofurantoin reaching the dog's urine using the recommended oral dosing regimen. In addition, we examined the efficacy of this breakpoint concentration against the common canine UTI pathogens, Escherichia coli, Staphylococcus pseudintermedius, and Enterococcus faecium. Eight experimental beagle dogs were treated with ~5 mg/kg of nitrofurantoin macrocrystal PO 8qh for 7 days. The urine samples were collected via cystocentesis at 2, 4, and 6 h after administration on day 2 and day 7 and used to quantify nitrofurantoin concentrations by ultra-high performance liquid chromatography. The results showed that 26.13-315.87 µg/mL nitrofurantoin was detected in the dogs' urine with a mean and median concentration of 104.82 and 92.75 µg/mL, respectively. Additionally, individual dogs presented with urinary nitrofurantoin concentrations greater than 64 µg/mL for at least 50% of the dosing intervals. This concentration efficiently killed E. coli, and S. pseudintermedius, but not E. faecium strains carrying an MIC90 value equal to 16, 16, and 128 µg/mL, respectively. Taken together, these results suggest that the value of 64 µg/mL may be set as a breakpoint against UTI pathogens, and nitrofurantoin could be an effective therapeutic drug against E. coli and S. pseudintermedius for canine UTIs.

Purification methods to reduce interference by dextran sodium sulfate with quantification of gene expression in intestinal tissue samples from a piglet model of colitis.

Dextran sodium sulfate (DSS) is commonly used to induce intestinal (i.e., colonic) inflammation in a variety of animal models. However, DSS is known to cause interference when using quantitative-real time polymerase chain reaction (qRT-PCR) methods, thereby invalidating accurate and precise measurement of tissue gene expression. Therefore, the goal of this study was to determine whether different mRNA purification methods would reduce DSS-interference. Colonic tissue samples were collected at postnatal days (PND) 27 or 28 from pigs that had not been administered DSS (Control), and two independent groups of pigs that received 1.25 g of DSS/kg of BW/d (DSS-1 and DSS-2) from PND 14 to 18. Tissue samples collected were subsequently stratified into three purification methods (i.e., 9 total treatment × method combinations), including: 1) no purification, 2) purification with lithium chloride (LiCl), or 3) purification using spin column filtration. All data were analyzed using a one-way ANOVA in the Mixed procedure of SAS. The average RNA concentrations across all treatments were between 1,300 and 1,800 µg/µL for all three in vivo groups. Although there were statistical differences among purification methods, the 260/280 and 260/230 ratios fell between acceptable limits of 2.0 to 2.1 and 2.0 to 2.2, respectively, for all treatment groups. This confirms the RNA quality was adequate and not influenced by purification method in addition to suggesting the absence of phenol, salts, and carbohydrate contamination. For pigs in the Control group that did not receive DSS, qRT-PCR Ct values of four cytokines were achieved, though these values were not altered by purification method. For pigs that had undergone DSS dosing, those tissues subjected to either no purification or purification using LiCl did not generate applicable Ct values. However, when tissues derive from DSS-treated pigs underwent spin column purification, half of the samples from DSS-1 and DSS-2 groups generated appropriate Ct estimates. Therefore, spin column purification appeared to be more effective than LiCl purification, but no method was 100% effective, so caution should be exercised when interpreting gene expression results from studies where animals are exposed to DSS-induced colitis.

Dextran sodium sulfate (DSS) is a chemical used to experimentally induce colonic inflammation in animal models. However, DSS causes chemical inhibition of processes involved with quantitative real-time polymerization chain reaction, thereby inhibiting the measurement of gene expression in tissues. In this study, differing methods of RNA purification were applied to remove DSS inhibition. Because no purification methods were 100% effective in alleviating this interference, caution should be exercised when interpreting gene expression results from studies where animals are exposed to DSS-induced colitis.

Underpinning the Food Matrix Regulation of Postexercise Myofibrillar Protein Synthesis by Comparing Salmon Ingestion With the Sum of Its Isolated Nutrients in Healthy Young Adults.

BACKGROUND: Protein is most commonly consumed as whole foods as opposed to single nutrients. However, the food matrix regulation of the postprandial muscle protein synthetic response has received little attention. OBJECTIVES: The purpose of this study was to assess the effects of eating salmon (SAL) and of ingesting the same nutrients as an isolated mixture of crystalline amino acids and fish oil (ISO) on the stimulation of postexercise myofibrillar protein synthesis (MPS) and whole-body leucine oxidation rates in healthy young adults. METHODS: Ten recreationally active adults (24 ± 4 y; 5 men, 5 women) performed an acute bout of resistance exercise, followed by the ingestion of SAL or ISO in a crossover fashion. Blood, breath, and muscle biopsies were collected at rest and after exercise during primed continuous infusions of L-[ring-2H5]phenylalanine and L-[1-13C]leucine. All data are presented as means ± SD and/or mean differences (95% CIs). RESULTS: Postprandial essential amino acid (EAA) concentrations peaked earlier (P = 0.024) in the ISO group than those in the SAL group. Postprandial leucine oxidation rates increased over time (P < 0.001) and peaked earlier in the ISO group (1.239 ± 0.321 nmol/kg/min; 63 ± 25 min) than those in the SAL group (1.230 ± 0.561 nmol/kg/min; 105 ± 20 min; P = 0.003). MPS rates for SAL (0.056 ± 0.022 %/h; P = 0.001) and ISO (0.046 ± 0.025 %/h; P = 0.025) were greater than the basal rates (0.020 ± 0.011 %/h) during the 0- to 5-h recovery period, with no differences between conditions (P = 0.308). CONCLUSION: We showed that the postexercise ingestion of SAL or ISO stimulate postexercise MPS rates with no differences between the conditions. Thus, our results indicate that ingesting protein from SAL as a whole-food matrix is similarly anabolic to ISO in healthy young adults. This trial was registered at www. CLINICALTRIALS: gov as NCT03870165.

Impaired neurogenesis with reactive astrocytosis in the hippocampus in a porcine model of acquired hydrocephalus.

BACKGROUND: Hydrocephalus is a neurological disease with an incidence of 0.3-0.7 per 1000 live births in the United States. Ventriculomegaly, periventricular white matter alterations, inflammation, and gliosis are among the neuropathologies associated with this disease. We hypothesized that hippocampus structure and subgranular zone neurogenesis are altered in untreated hydrocephalus and correlate with recognition memory deficits. METHODS: Hydrocephalus was induced by intracisternal kaolin injections in domestic juvenile pigs (43.6 ± 9.8 days). Age-matched sham controls received similar saline injections. MRI was performed to measure ventricular volume, and/or hippocampal and perirhinal sizes at 14 ± 4 days and 36 ± 8 days post-induction. Recognition memory was assessed one week before and after kaolin induction. Histology and immunohistochemistry in the hippocampus were performed at sacrifice. RESULTS: The hippocampal width and the perirhinal cortex thickness were decreased (p < 0.05) in hydrocephalic pigs 14 ± 4 days post-induction. At sacrifice (36 ± 8 days post-induction), significant expansion of the cerebral ventricles was detected (p = 0.005) in hydrocephalic pigs compared with sham controls. The area of the dorsal hippocampus exhibited a reduction (p = 0.035) of 23.4% in the hydrocephalic pigs at sacrifice. Likewise, in hydrocephalic pigs, the percentages of neuronal precursor cells (doublecortin+ cells) and neurons decreased (p < 0.01) by 32.35%, and 19.74%, respectively, in the subgranular zone of the dorsal hippocampus. The percentage of reactive astrocytes (vimentin+) was increased (p = 0.041) by 48.7%. In contrast, microglial cells were found to decrease (p = 0.014) by 55.74% in the dorsal hippocampus in hydrocephalic pigs. There was no difference in the recognition index, a summative measure of learning and memory, one week before and after the induction of hydrocephalus. CONCLUSION: In untreated juvenile pigs, acquired hydrocephalus caused morphological alterations, reduced neurogenesis, and increased reactive astrocytosis in the hippocampus and perirhinal cortex.

Formula supplementation with human and bovine milk oligosaccharides modulates blood IgG and T-helper cell populations, and ex vivo LPS-stimulated cytokine production in a neonatal preclinical model.

Introduction: Human milk contains structurally diverse oligosaccharides (HMO), which are multifunctional modulators of neonatal immune development. Our objective was to investigate formula supplemented with fucosylated (2'FL) + neutral (lacto-N-neotetraose, LNnt) oligosaccharides and/or sialylated bovine milk oligosaccharides (BMOS) on immunological outcomes. Methods: Pigs (n=46) were randomized at 48h of age to four diets: sow milk replacer formula (CON), BMOS (CON + 6.5 g/L BMOS), HMO (CON + 1.0 g/L 2'FL + 0.5 g/L LNnT), or BMOS+HMO (CON + 6.5 g/L BMOS + 1.0 g/L 2'FL + 0.5 g/L LNnT). Blood and tissues were collected on postnatal day 33 for measurement of cytokines and IgG, phenotypic identification of immune cells, and ex vivo lipopolysaccharide (LPS)-stimulation of immune cells. Results: Serum IgG was significantly lower in the HMO group than BMOS+HMO but did not differ from CON or BMOS. The percentage of PBMC T-helper cells was lower in BMOS+HMO than the other groups. Splenocytes from the BMOS group secreted more IL-1ß when stimulated ex vivo with LPS compared to CON or HMO groups. For PBMCs, a statistical interaction of BMOS*HMO was observed for IL-10 secretion (p=0.037), with BMOS+HMO and HMO groups differing at p=0.1. Discussion: The addition of a mix of fucosylated and sialylated oligosaccharides to infant formula provides specific activities in the immune system that differ from formulations supplemented with one oligosaccharide structure.

Dietary supplementation of 3'-sialyllactose or 6'-sialyllactose elicits minimal influence on cognitive and brain development in growing pigs.

Sialylated human milk oligosaccharides (HMO), such as 3'-sialyllactose (3'-SL) and 6'-sialyllactose (6'-SL), are abundant throughout lactation and at much higher concentrations than are present in bovine milk or infant formulas. Previous studies have suggested that sialylated HMO may have neurocognitive benefits in early life. Recent research has focused on infant formula supplementation with key nutrients and bioactives to narrow the developmental gap between formula-fed and breastfed infants. Herein, we investigated the impact of supplemental 3'-SL or 6'-SL on cognitive and brain development at two time-points [postnatal days (PND) 33 and 61]. Two-day-old piglets (N = 75) were randomly assigned to commercial milk replacer ad libitum without or with 3'-SL or 6'-SL (added in a powdered form at a rate of 0.2673% on an as-is weight basis). Cognitive development was assessed via novel object recognition and results were not significant at both time-points (p > 0.05). Magnetic resonance imaging was used to assess structural brain development. Results varied between scan type, diet, and time-point. A main effect of diet was observed for absolute volume of white matter and 9 other regions of interest (ROI), as well as for relative volume of the pons on PND 30 (p < 0.05). Similar effects were observed on PND 58. Diffusion tensor imaging indicated minimal differences on PND 30 (p > 0.05). However, several dietary differences across the diffusion outcomes were observed on PND 58 (p < 0.05) indicating dietary impacts on brain microstructure. Minimal dietary differences were observed from myelin water fraction imaging at either time-point. Overall, sialyllactose supplementation had no effects on learning and memory as assessed by novel object recognition, but may influence temporally-dependent aspects of brain development.

Effects of early postnatal life nutritional interventions on immune-microbiome interactions in the gastrointestinal tract and implications for brain development and function.

The gastrointestinal (GI) microbiota has co-evolved with the host in an intricate relationship for mutual benefit, however, inappropriate development of this relationship can have detrimental effects. The developing GI microbiota plays a vital role during the first 1,000 days of postnatal life, during which occurs parallel development and maturation of the GI tract, immune system, and brain. Several factors such as mode of delivery, gestational age at birth, exposure to antibiotics, host genetics, and nutrition affect the establishment and resultant composition of the GI microbiota, and therefore play a role in shaping host development. Nutrition during the first 1,000 days is considered to have the most potential in shaping microbiota structure and function, influencing its interactions with the immune system in the GI tract and consequent impact on brain development. The importance of the microbiota-GI-brain (MGB) axis is also increasingly recognized for its importance in these developmental changes. This narrative review focuses on the importance of the GI microbiota and the impact of nutrition on MGB axis during the immune system and brain developmental period in early postnatal life of infants.

Lacticaseibacillus rhamnosus HN001 alters the microbiota composition in the cecum but not the feces in a piglet model.

The probiotic Lacticaseibacillus rhamnosus strain HN001 has been shown to have several beneficial health effects for both pediatric and maternal groups, including reduced risk of eczema in infants and gestational diabetes and postnatal depression in mothers. While L. rhamnosus HN001 appears to modify immune and gut barrier biomarkers, its mode of action remains to be fully elucidated. To gain insights into the role of HN001 on the infant microbiome, the impacts of L. rhamnosus HN001 supplementation was studied in 10-day old male piglets that were fed either infant formula, or infant formula with L. rhamnosus HN001 at a low (1.3 × 105 CFU/ml) or high dose (7.9 × 106 CFU/ml) daily for 24 days. The cecal and fecal microbial communities were assessed by shotgun metagenome sequencing and host gene expression in the cecum and colon tissue was assessed by RNA-seq. Piglet fecal samples showed only modest differences between controls and those receiving dietary L. rhamnosus HN001. However, striking differences between the three groups were observed for cecal samples. While total lactobacilli were significantly increased only in the high dose L. rhamnosus HN001 group, both high and low dose groups showed an up to twofold reduction across the Firmicutes phylum and up to fourfold increase in Prevotella compared to controls. Methanobrevibacter was also decreased in HN001 fed piglets. Microbial genes involved in carbohydrate and vitamin metabolism were among those that differed in relative abundance between those with and without L. rhamnosus HN001. Changes in the cecal microbiome were accompanied by increased expression of tight junction pathway genes and decreased autophagy pathway genes in the cecal tissue of piglets fed the higher dose of L. rhamnosus HN001. Our findings showed supplementation with L. rhamnosus HN001 caused substantial changes in the cecal microbiome with likely consequences for key microbial metabolic pathways. Host gene expression changes in the cecum support previous research showing L. rhamnosus HN001 beneficially impacts intestinal barrier function. We show that fecal samples may not adequately reflect microbiome composition higher in the gastrointestinal tract, with the implication that effects of probiotic consumption may be missed by examining only the fecal microbiome.

Xylanase supplementation of pelleted wheat-based diets increases growth efficiency and apparent metabolizable energy and decreases viscosity of intestinal contents in broilers.

This study was designed to test graded supplementation of a thermostable xylanase in pelleted, wheat-based diets fed to broiler chickens over a 28-d period. A total of 600 Ross 708 male broilers were allotted to 1 of 5 dietary treatments: positive control (PC), negative control (NC; 125 kcal of AME/kg diet reduction relative to PC), and NC supplemented with 10, 15, or 30 g/ton of xylanase. Wheat-soybean meal-based diets were pelleted and fed in 2 feeding phases (14-d each). Study outcomes included growth performance, AME, and ileal digesta viscosity with 20 battery cages of 6 birds per treatment. Data were analyzed by 1-way ANOVA along with estimation of Pearson correlation coefficients. Whereas no difference between NC and PC was observed for BW gain, NC birds exhibited increased (P < 0.05) feed intake during each feeding phase and overall, which caused improvements (P < 0.05) in feed conversion ratio (FCR) for PC vs. NC birds. The analyzed AME of PC birds was 112 kcal/kg of diet greater (P < 0.05) than for NC birds, though no differences in digesta viscosity were observed. Xylanase supplementation of the NC diet at 15 or 30 g/ton elicited overall improvements (P < 0.05) in BW gain beyond the PC, while the 30 g/ton level equalized feed intake with the PC. Regardless of level, xylanase supplementation improved (P < 0.05) the FCR relative to the NC, thereby equalizing the response with the PC. Similarly, supplementation with any xylanase level increased (P < 0.05) AME over the NC, making all treatments synonymous with the PC. Digesta viscosity of all xylanase-supplemented treatments was decreased relative to both the NC and PC treatments. Overall, this study provided clear evidence that addition of a thermostable xylanase to pelleted wheat-based diets elicited improvements in growth performance of broilers concomitant with a reduction in digesta viscosity and elevation of analyzed dietary AME content.

Oral gamma-cyclodextrin-encapsulated tributyrin supplementation in young pigs with experimentally induced colitis.

Disruption of intestinal integrity and barrier function due to tissue inflammation has negative implications on overall growth and well-being in young pigs. In this study, we investigated the effects of oral gamma-cyclodextrin-encapsulated tributyrin (TBCD) in young pigs experiencing dextran sodium sulfate (DSS)-induced colitis. Pigs (n = 32 boars) were weaned from the sow at postnatal day (PND) 2, allotted to treatment based on the litter of origin and body weight (BW), and reared artificially over a 26-d feeding period. Treatment groups included: 1) nutritionally adequate (control) milk replacer, no DSS (Control n = 8), 2) control milk replacer plus oral DSS (DSS, n = 7), and 3) control diet supplemented with 8.3 g of TBCD per kg of reconstituted milk replacer plus oral DSS (TBCD + DSS, n = 8). Colitis was induced by administering DSS at 1.25 g of DSS/kg BW daily in a reconstituted milk replacer from PND 14-18. Milk replacer and water were provided ad libitum throughout the 26-d study. All the data were analyzed using a one-way ANOVA using the MIXED procedure of SAS. Control and DSS pigs had similar BW throughout the study, while TBCD + DSS pigs exhibited decreased (P < 0.05) BW starting at approximately PND 15. Additionally, average daily gain (ADG) before and after initiation of DSS dosing, along with over the total study duration, was decreased (P < 0.05) in pigs receiving TBCD + DSS compared with the Control. Milk disappearance was decreased (P < 0.05) in TBCD + DSS pigs when compared with Control and DSS groups. Both the concentration and molar ratio of cecal butyrate concentrations were increased (P < 0.05) in TBCD + DSS pigs compared with the Control group. The DSS and TBCD + DSS treatments also increased (P < 0.05) butyrate concentrations in the luminal contents with the proximal colon compared with Control. TBCD + DSS and DSS pigs had increased (P < 0.05) mucosal width in the distal colon compared with Control, thereby indicating heightened intestinal inflammation. Overall, oral supplementation of encapsulated tributyrin increased the concentration of butyrate in the colon, but was unable to mitigate the negative effects of DSS-induced colitis.

There are negative implications in young pigs when the integrity and function of the intestine are disrupted due to colonic inflammation. Volatile compounds have been used as dietary supplements to alleviate intestinal inflammation, but little work has been completed on the use of encapsulated tributyrin in newly weaned pigs. In this study, pigs received 1 of 3 treatments: 1) a standard milk replacer without the induction of intestinal inflammation, 2) the same standard milk replacer with the induction of intestinal inflammation, or 3) milk replacer supplemented with encapsulated tributyrin with the induction of intestinal inflammation. Throughout the study period, growth performance was decreased in pigs receiving supplemental tributyrin compared with other treatments. Additionally, experimentally induced colitis increased butyrate concentrations in the cecum, while tributyrin supplementation increased butyrate concentrations in the proximal colon. Pigs undergoing intestinal inflammation had increased thickness of the mucosal layer in the distal colon compared with sham-challenged pigs. Overall, the supplementation of encapsulated tributyrin increased colonic butyrate concentrations, but did not mitigate the negative effects of inflammation in the large intestine.

The effects of ruminant milk treatments on hippocampal, striatal, and prefrontal cortex gene expression in pigs as a model for the human infant.

While infant formula is usually bovine milk-based, interest in other ruminant milk-based formulas is growing. However, whether different ruminant milk treatments with varying nutrient compositions influence the infant's brain development remains unknown. The aim was to determine the effects of consuming bovine, caprine, or ovine milk on brain gene expression in the early postnatal period using a pig model of the human infant. Starting at postnatal day 7 or 8, pigs were exclusively fed bovine, ovine, or caprine milk for 15 days. The mRNA abundance of 77 genes in the prefrontal cortex, hippocampus, and striatum regions was measured at postnatal day 21 or 22 using NanoString. The expression level of two hippocampal and nine striatal genes was most affected by milk treatments, particularly ovine milk. These modulatory genes are involved in glutamate, gamma-aminobutyric acid, serotonin, adrenaline and neurotrophin signaling and the synaptic vesicle cycle. The expression level of genes involved in gamma-aminobutyric acid signaling was associated with pigs' lactose intake. In contrast, milk treatments did not affect the mRNA abundance of the genes in the prefrontal cortex. This study provides the first evidence of the association of different ruminant milk treatments with brain gene expression related to cognitive function in the first 3 months of postnatal life.