Effects of dietary supplementation with L-arginine on the endogenous losses of amino acids in growing pigs exposed to heat stress.

Exposing pigs to heat stress (HS) provokes higher death of intestinal cells, resulting in elevated endogenous intestinal losses (EIL) of amino acids (AA) and damage to intestinal epithelia. Arginine (Arg) is precursor for the synthesis of polyamines, which are involved in proliferation of intestinal cells and restoration of the intestinal epithelia. Thus the effect of adding L-Arg to diets for HS pigs on the EIL of AA was analyzed. Twelve pigs (23.1 ± 1.1 kg body weight) implanted with T-type cannulas at the end of ileum were individually housed and allowed 15-days for surgery recovery under thermoneutral (TN) conditions (22 ± 2 °C). Following, the pigs were randomly assigned to one of three treatments: TN pigs fed a semi-purified, corn starch-3% casein basal diet (TN-B); HS pigs with the basal diet (HS-B); HS pigs consuming the basal diet supplemented with 0.20% L-Arg (HS-Arg). The experiment consisted of two 9-day periods; each period included 7-days of adaptation to their respective diet, followed by a 2-day ileal digesta collection period. Digesta was collected during 12 consecutive hours each day. The pigs were fed twice a-day. Ambient temperature (AT) inside the TN and HS rooms ranged from 18.6 to 27.6 °C and from 29.5 to 40.7 °C, respectively. Body temperature followed a pattern similar to that of AT. The daily EIL of indispensable AA increased (P < 0.01) in the HS-B pigs compared to both the TN-B and the HS-Arg pigs, however, there was no EIL difference between the TN-B and the HS-Arg pigs (P > 0.05). Likewise, with the exception of serine, daily losses of endogenous dispensable AA in the HS-B pigs were higher (P < 0.01) in comparison with those of TN-B and HS-Arg pigs. In summary, HS exposure compared to TN conditions increases the loss of endogenous AA, but dietary supplementation with L-Arg helped to counteract the negative HS effect.

Effects of dietary protein content and crystalline amino acid supplementation patterns in low protein diets on intestinal bacteria and their metabolites in weaned pigs raised under Different sanitary conditions.

The objective of this experiment was to investigate the effects of dietary crude protein (CP) content and crystalline amino acids (CAA) supplementation patterns in low CP (LCP) diets on intestinal bacteria and their metabolites in weaned pigs raised under clean (CSC) or unclean sanitary conditions (USC). One hundred forty-four piglets (6.35 ±â€…0.63 kg) were assigned to one of six treatments in a 3 × 2 factorial arrangement based on CP content and sanitary conditions in a randomized complete block design to give eight replicates with three pigs per pen over a 21-d period. Diets consisted of a high CP (HCP; 21%) and two LCP (18%) diets supplemented with 9 CAA (Lys, Met, Thr, Trp, Val, Ile, Leu, His, and Phe) or only six CAA (Lys, Met, Thr, Trp, Val, and Ile) to meet the requirements. The CSC room was washed weekly, whereas the USC room had sow manure spread in the pens from the beginning of the study and was not washed throughout the experiment. Jejunum and colon digesta were sampled on day 21. Both jejunum and colon digesta were analyzed for ammonia nitrogen, short-chain fatty acids, and biogenic amines but only colon digesta was analyzed for microbiome composition (16s rRNA sequencing on MiSeq). Data were analyzed using R software for 16S rRNA and the MIXED procedure of SAS for microbial metabolites. Sanitation, CP content, and CAA supplementation patterns did not affect the diversity of colonic bacterial composition in weaned pigs. Pigs raised under USC had greater (P < 0.05) jejunal ammonia nitrogen concentration than those raised under CSC. Pigs fed LCP diets had reduced (P < 0.05) jejunal ammonia nitrogen concentration compared to those fed the HCP diet. Interactions between sanitation and dietary CP content were observed (P < 0.05) for: (1) jejunal acetate and (2) colonic spermidine and spermine, whereby (1) acetate concentrations decreased from NCP to LCP in pigs raised under the CSC but those concentrations increased under the USC, and (2) spermidine and spermine concentrations increased in LCP diets compared to HCP diet under USC, unlike CSC which did not show any difference between HCP and LCP. In conclusion, reducing dietary CP lowered ammonia nitrogen content regardless of sanitation and increased microbial metabolites in weaned pigs raised under USC. However, LCP diets with different CAA supplementation patterns did not affect bacterial diversity in weaned pigs, regardless of the hygienic conditions where the animals were housed.

Lowering dietary crude protein concentration by 3% to 4% units has been used as one of the strategies to promote growth and improve the gut health of weaned pigs. Undigested and endogenous protein could be available for microbial fermentation, and protein fermentation is considered detrimental to the gut health of the host animal. The unclean sanitary condition model mimics commercial raising conditions and stimulates a low-grade inflammatory and immune response. Ammonia nitrogen is one of the harmful metabolites derived by protein fermentation and pigs fed low-protein diets had decreased ammonia nitrogen than those fed high-protein diets. Also, pigs raised under unclean sanitation had greater ammonia nitrogen than those raised under clean sanitation. However, sanitation, protein content, and crystalline amino acids supplementation patterns did not affect the diversity of colonic bacterial composition in weaned pigs. The results obtained from the present study showed that a low protein diet could be used to improve gut health in weaned pigs.

Effect of dl-methionine supplementation above requirement on performance; intestinal morphology, antioxidant activity, and gene expression; and serum concentration of amino acids in heat stressed pigs.

The intestinal morphology and function can be compromised in pigs exposed to heat stress (HS), partly due to increased production of reactive-oxygen species. Because methionine (Met) functions as intracellular antioxidant, the requirement of Met may be increased in HS-pigs. The effect of dietary supplementation with dl-Met above requirement on performance, small intestine morphology, antioxidant enzymes activity, amino acid transporters expression, and serum concentration (SC) of free AA in HS-pigs was evaluated. A basal wheat-soybean meal diet was formulated to meet 100% Met requirement with the other indispensable AA exceeding at least 20% their requirement. Sixty individually housed pigs (23.0 ± 2.4 kg BW, 12 pigs per treatment) were randomly assigned to five treatments: TN100, thermal-neutral (22.7 °C) housed pigs fed the basal diet; HS100, HS120, HS140, HS160; HS-pigs (29.6 °C to 39.4 °C) fed the basal diet supplemented with dl-Met to contain 0%, 20%, 40%, and 60% dl-Met above the requirement, respectively. Pigs had free access to feed and water during the 21-d trial. Blood samples were collected on day 18 to analyze the absorptive AA-SC. The effect of ambient temperature (HS100 vs. TN100), as well as the linear and quadratic effects of increasing Met levels in the diets for HS-pigs were analyzed. The HS100 pigs gained less weight than TN100 and HS120 pigs (P < 0.01); gain:feed was also higher in HS120 pigs than in HS100 pigs (P ≤ 0.05). Feed intake of TN100 pigs was higher than that of HS-pigs fed the dl-Met supplemented diets (P < 0.05). Villi height reduced in pigs HS, but Met supplementation quadratically increased it (P < 0.05). Superoxide dismutase and catalase activities, reduced glutathione concentration, and relative expression of B0AT2 in ileum decreased (P < 0.05), but glutathione peroxidase activity increased in HS-pigs. dl-Met supplementation linearly affected catalase and glutathione peroxidase activities, as well as the relative expression of b0,+AT in jejunum (P < 0.05) of HS-pigs. The SC of Ile, Leu, Lys, Phe, and Val were higher in HS100 pigs than in TN100 pigs (P < 0.05). Graded levels of supplemental dl-Met in diets for HS-pigs linearly decreased SC of Ile, Leu, and Val (P < 0.05), tended to decrease His, Lys, and Thr (P < 0.10), and increased Met (P < 0.01). In conclusion, HS had negative effect on weight gain and intestinal morpho-physiology; however, it was ameliorated by adding 20% Met above the requirement in diets for growing pigs.

The exposure of pigs to ambient temperature above their comfort zone affects several functions of the small intestine, especially those related with digestion of feed and absorption of nutrients, which in turn reduces the availability of nutrients for growth. Amino acids such as methionine are involved in multiple functions of intestinal cells. Thus, methionine supplementation may help pigs to overcome the negative impact of their exposure to high ambient temperature. Indeed, methionine supplementation to the diet increased growth rate and feed efficiency of pigs housed under heat stress, which was presumably associated with an improvement in the utilization of the absorbed amino acids.

Apparent and standardised ileal amino acid digestibilities in heat-stressed pigs fed wheat-soybean meal diets supplemented with l-arginine and dl-methionine.

Heat stress (HS) exposure may damage the small intestine epithelia of pigs affecting the digestibility and absorption of amino acids (AA). Arg and Met can enhance antioxidant and intestinal cell proliferation activity, thus supplementing them in diets might alleviate epithelial damage and correct the reduced AA digestibility. The effect of adding extra l-Arg and dl-Met to diets on the apparent (AID) and standardised ileal digestibility (SID) of AA was analysed in a 10-day experiment conducted with 10 ileal-cannulated HS pigs (25.3 ± 2.4 kg body weight). The pigs were divided into two treatments: Control, wheat-soybean meal diet supplemented with l-Lys, l-Thr, dl-Met and l-Trp; and control diet plus 0.20% l-Arg and 0.20% dl-Met (Arg + Met). Following an 8-day diet adaptation period, ileal digesta was continuously collected for 12 h, starting at 0700, on Days 9 and 10. All pigs were daily exposed to 29.6-36.1°C; ileal temperature was continuously monitored at 15-min intervals. Feed was provided twice a day. The ileal temperature ranged from 40.3 to 41.5°C. Daily ileal outflow of His, Ile, Leu, Phe, Thr, Ser and Tyr decreased (p < 0.05), and that of Arg, Val and Pro tended to decrease (p < 0.10) in the Arg + Met pigs. The AID of Arg, His, Met, Thr and Tyr, and the SID of His, Met, and Thr increased in pigs fed the Arg + Met diet (p < 0.05). Thr and Val had the lowest AID values whereas Arg, Met, and Lys had the highest values. Arg (r = 0.64) and Met (r = 0.84) intake were highly correlated with their AID values; Met intake was highly correlated with its SID value (r = 0.72). Valine and Arg had the lowest whereas Arg had the highest SID values. In conclusion, supplementing l-Arg and dl-Met above the requirement decreases the ileal outflow of several AA, and increases the AID and SID of some essential AA in HS pigs.

Ileal alkaline phosphatase is upregulated following functional amino acid supplementation in Salmonella Typhimurium-challenged pigs.

We recently showed that functional amino acid (FAA) supplementation improves growth performance and immune status of Salmonella Typhimurium (ST)-challenged pigs, which was further improved by a longer adaptation period. It is expected that the effects are associated with increased activity of intestinal alkaline phosphatase (IAP). The objective of this study was to evaluate the effects of FAA supplementation and adaptation period on the ileal, cecal, and colonic activity of IAP in weaned pigs challenged with ST. In experiment 1, a total of 32 mixed-sex weanling pigs were randomly assigned to dietary treatments in a 2 × 2 factorial arrangement with low (LP) or high protein (HP) content and basal (FAA-) or FAA profile (FAA+; Thr, Met, and Trp at 120% of requirements) as factors. In experiment 2, a total of 32 mixed-sex weanling pigs were randomly assigned to one of four dietary treatments, being FAA- fed throughout the experimental period (FAA-) or an FAA profile fed only in the post-inoculation (FAA + 0), for 1 wk pre- and post-inoculation (FAA + 1), or throughout the experimental period (FAA + 2). In experiments 1 and 2, after a 7- and 14-d adaptation period, respectively, pigs were inoculated with saline solution containing ST (3.3 and 2.2 × 109 CFU/mL, respectively). Plasma alkaline phosphatase was measured on days 0 and 7 post-inoculation in experiment 1, and IAP (ileum, cecum, and colon) was measured in experiments 1 and 2. Correlations among ileal IAP and serum albumin and haptoglobin, plasma superoxide dismutase (SOD), malondialdehyde (MDA), and reduced:oxidized glutathione, ileal myeloperoxidase, ST shedding and ileal colonization, and post-inoculation average daily gain, feed intake (ADFI), and gain:feed were also analyzed. In experiment 1, plasma alkaline phosphatase was decreased with ST inoculation and the overall content was increased in LP-FAA+ compared with LP-FAA- (P < 0.05). Moreover, ileal IAP was increased in FAA+ compared with FAA- pigs in both studies (P < 0.05) regardless of adaptation time (P > 0.05). IAP was positively correlated with MDA and ADFI and negatively correlated with SOD and ST shedding in experiment 1 (P < 0.05). These results demonstrate a positive effect of FAA supplementation, but not adaptation period, on ileal alkaline phosphatase activity in Salmonella-challenged pigs, which may be associated with improvements in antioxidant balance.

Functional amino acid (FAA) supplementation has been shown to improve gut health and antioxidant defense in weaned piglets challenged with Salmonella Typhimurium (ST), regardless of dietary protein content. The beneficial effects were further improved when pigs were adapted to FAA for 2 wk prior to the ST challenge. Recent evidence has shown that intestinal alkaline phosphatase (IAP), which may be influenced by nutritional factors, attenuates intestinal inflammation, possibly due to gut microbiota modulation. This study is the first to identify that ileal IAP activity is increased following FAA supplementation in ST-challenged pigs, regardless of adaptation period. Moreover, ileal IAP activity correlated with systemic markers of antioxidant defense, which highlights the enzyme's role in attenuating systemic infection. Overall, the development of feeding strategies with positive effects on IAP activity is of interest, due to the enzyme's central role on the gut and whole-body homeostasis and health.

Effect of arginine supplementation on the morphology and function of intestinal epithelia and serum concentrations of amino acids in pigs exposed to heat stress.

The exposure of pigs to heat stress (HS) appears to damage their intestinal epithelia, affecting the absorption of amino acids (AAs). Arg is involved in the restoration of intestinal epithelial cells but HS reduces Arg intake. The effect of dietary supplementation with Arg on the morphology of intestinal epithelia, AA transporter gene expression, and serum concentration (SC) of free AAs in HS pigs was analyzed. Twenty pigs (25.3 ± 2.4 kg body weight) were randomly assigned to two dietary treatments: Control (0.81% Arg), wheat-soybean meal diet supplemented with l-Lys, l-Thr, dl-Met, and l-Trp, and the experimental diet where 0.16% free l-Arg was supplemented to a similar Control diet (+Arg). All pigs were individually housed and exposed to HS, fed ad libitum with full access to water. The ambient temperature (AT), recorded at 15-min intervals during the 21-d trial, ranged on average from 29.6 to 39.4 °C within the same day. Blood samples were collected on day 18 at 1600 hours (AT peak); serum was separated by centrifugation. At the end of the trial, five pigs per treatment were sacrificed to collect samples of mucosa scratched from each small intestine segment. The expression of AA transporters in intestinal mucosa and the SC of AAs were analyzed. Villi height (VH) was higher (P < 0.01) in the duodenum, jejunum, and ileum but the crypt depth did not differ between the Control and the +Arg pigs. Supplementation of l-Arg increased the mRNA coding for the synthesis of the cationic AA transporter b0,+ (P < 0.01) and the neutral AA transporter B0 (P < 0.05) in the duodenum by approximately 5-fold and 3-fold, respectively, but no effect on mRNA abundance was observed in the jejunum and ileum. The supplementation of l-Arg increased serum Arg, His, Met, Thr, Trp, and urea (P < 0.05) and also tended to increase Val (P < 0.10) but did not affect Ile, Lys, Leu, and Phe. These results indicate that supplementing 0.16% l-Arg to the Control diet may help to improve the function of the small intestine epithelium, by increasing the VH, the abundance of AA transporters, and the SC of most indispensable AAs in pigs exposed to HS conditions. However, the lack of effect of supplemental Arg on both Lys SC and weight gain of pigs suggests that increasing the Lys content in the +Arg diet might be needed to improve the performance of HS pigs.

A longer adaptation period to a functional amino acid-supplemented diet improves growth performance and immune status of Salmonella Typhimurium-challenged pigs.

We recently showed that dietary supplementation with key functional amino acids (FAA) improves growth performance and immune status of Salmonella Typhimurium (ST)-challenged pigs. It is not known if ST-challenged pigs will benefit from a longer adaptation period to FAA. The objective of this study was to evaluate the effects of different adaptation periods to diets containing FAA above requirements for growth on performance and immune response of weaned pigs subsequently challenged with ST. A total of 32 mixed-sex weanling pigs (11.6 ± 0.3 kg) were randomly assigned to 1 of 4 dietary treatments, being a basal amino acid (AA) profile fed throughout the experimental period (FAA-) or a functional AA profile (FAA+; Thr, Met, and Trp at 120% of requirements) fed only in the postinoculation (FAA+0), for 1 wk pre- and postinoculation (FAA+1), or throughout the experimental period (FAA+2). After a 14-d adaptation period, pigs were inoculated with ST (2.15 × 109 CFU/mL). Growth performance, body temperature, fecal score, acute-phase proteins, oxidant/antioxidant balance, score for ST shedding in feces and intestinal colonization, and fecal and digesta myeloperoxidase (MPO) were measured pre- and postinoculation. Postinoculation body temperature and fecal score, serum haptoglobin, plasma superoxide dismutase (SOD), malondialdehyde (MDA), and fecal MPO were increased while serum albumin and plasma reduced glutathione (GSH):oxidized glutathione (GSSG) were reduced compared to pre-inoculation (P < 0.05). Average daily gain and G:F were greater in FAA+2 pigs compared to FAA- pigs (P < 0.05). Serum albumin was higher in FAA+2 and FAA+1 compared to FAA+0 and FAA- pigs (P < 0.05) while FAA+2 pigs had lower haptoglobin compared to FAA- (P < 0.05). Plasma SOD was increased and GSH:GSSG was decreased in FAA- pigs compared to the other treatments (P < 0.05). Score for ST shedding in feces was progressively lower from d 1 to 6 regardless of treatment (P < 0.05) and was lower in FAA+2 pigs compared to FAA- and FAA+0 (P < 0.05). Counts of ST in colon digesta were higher in FAA- and FAA+0 pigs compared to FAA+2 (P < 0.05). Fecal and colonic digesta MPO were lower in FAA+2 and FAA+1 pigs compared to FAA- (P < 0.05). These results demonstrate a positive effect of a longer adaptation period to FAA-supplemented diets on performance and immune status of weaned pigs challenged with Salmonella.

Functional amino acid supplementation, regardless of dietary protein content, improves growth performance and immune status of weaned pigs challenged with Salmonella Typhimurium.

High dietary protein may increase susceptibility of weaned pigs to enteric pathogens. Dietary supplementation with functional amino acids (FAA) may improve growth performance of pigs during disease challenge. The objective of this study was to evaluate the interactive effects of dietary protein content and FAA supplementation above requirements for growth on performance and immune response of weaned pigs challenged with Salmonella. Sixty-four mixed-sex weanling pigs (13.9 ± 0.82 kg) were randomly assigned to dietary treatments in a 2 × 2 factorial arrangement with low (LP) or high protein (HP) content and basal (AA-) or FAA profile (AA+; Thr, Met, and Trp at 120% of requirements) as factors. After a 7-d adaptation period, pigs were inoculated with either a sterile saline solution (CT) or saline solution containing Salmonella Typhimurium (ST; 3.3 × 109 CFU/mL). Growth performance, body temperature, fecal score, acute-phase proteins, oxidant/antioxidant balance, ST shedding score in feces and intestinal colonization, fecal and digesta myeloperoxidase (MPO), and plasma urea nitrogen (PUN) were measured pre- and postinoculation. There were no dietary effects on any measures pre-inoculation or post-CT inoculation (P > 0.05). Inoculation with ST increased body temperature and fecal score (P < 0.05), serum haptoglobin, plasma superoxide dismutase (SOD), malondialdehyde (MDA), PUN, and fecal MPO, and decreased serum albumin and plasma reduced glutathione (GSH):oxidized glutathione (GSSG) compared with CT pigs (P < 0.05). ST-inoculation reduced average daily gain (ADG) and feed intake (ADFI) vs. CT pigs (P < 0.05) but was increased by AA+ vs. AA- in ST pigs (P < 0.05). Serum albumin and GSH:GSSG were increased while haptoglobin and SOD were decreased in ST-inoculated pigs fed AA+ vs. AA- (P < 0.05). PUN was higher in HP vs. LP-fed pigs postinoculation (P < 0.05). Fecal ST score was increased in ST-inoculated pigs on days 1 and 2 postinoculation and declined by day 6 (P < 0.05) in all pigs while the overall score was reduced in AA+ vs. AA- pigs (P < 0.05). Cecal digesta ST score was higher in HP vs. LP-fed pigs and were lower in AA+ compared with AA- fed pigs in the colon (P < 0.05). Fecal and digesta MPO were reduced in ST pigs fed AA+ vs. AA- (P < 0.05). These results demonstrate a positive effect of FAA supplementation, with minimal effects of dietary protein, on performance and immune status in weaned pigs challenged with Salmonella.

Bioavailability of the calcium salt of dl-methionine hydroxy analog compared with dl-methionine for nitrogen retention and the preference of nursery pigs for diets based on the 2 forms of methionine.

Experiments were conducted to determine the relative bioavailability (RBV) of the calcium salt of the hydroxy analog of dl-methionine (MHA-Ca, 84%) to dl-methionine (dl-Met, 99%) as Met sources fed to pigs. In experiment 1, 42 crossbred barrows (initial BW of 15.0 ± 0.7 kg) were allotted to 7 treatments in an N-balance study. The basal diet (BD) was formulated to contain 15.4% CP and 0.22% Met (70% of requirement). Diets included (1) BD, (2) BD + 0.025% dl-Met, (3) BD + 0.050% dl-Met, (4) BD + 0.075% dl-Met, (5) BD + 0.038% MHA-Ca, (6) BD + 0.077% MHA-Ca, and (7) BD + 0.115% MHA-Ca. An increase in dietary inclusion rates of both Met sources linearly increased (P < 0.01) N retained (g/d) and N retention (% of intake). Using linear slope-ratio regression, the RBV value of MHA-Ca to dl-Met for N retained (g/d) was 63.0% on a product-to-product basis (75.0% on an equimolar basis). In experiment 2, 40 crossbred barrows (initial BW of 15.5 ± 1.5 kg) were allotted to 5 treatments in another N-balance study. The BD was formulated to contain 17.0% CP and 0.22% Met (70% of requirement). Diets included (1) BD, (2) BD + 0.030% dl-Met, (3) BD + 0.060% dl-Met, (4) BD + 0.046% MHA-Ca, and (5) BD + 0.092% MHA-Ca. Increasing levels of dl-Met or MHA-Ca increased N retained (g/d) and N retention (% of intake) linearly (P < 0.001) and quadratically (P < 0.05). Using linear slope-ratio regression, a product-to-product RBV value of MHA-Ca to dl-Met was 68.4% (81.4% on an equimolar basis) for N retained (g/d). In experiment 3, 276 pigs (12 barrow and 11 gilt replicates; initial BW of 7.09 ± 1.1 kg) were used in 3 diet preference studies. Pigs were randomly allotted to 1 of 3 treatment comparisons of feed choice: (1) BD (0.23% Met) or BD + 0.07% dl-Met; (2) BD or BD + 0.0825% MHA-Ca, and (3) BD + 0.07% dl-Met or BD + 0.0825% MHA-Ca. Pigs consumed a higher percentage (55 vs. 45%; P = 0.008) of their total feed intake from the diet supplemented with 0.07% dl-Met in Comparison 1, but a lower percentage (45 vs. 55%; P = 0.003) of their total feed intake from the diet supplemented with 0.0825% MHA-Ca in Comparison 2. There was no diet preference for dl-Met or MHA-Ca in Comparison 3. The observed Met source preference differences occurred in the barrow replicates but not in the gilt replicates. These results demonstrated the mean RBV of MHA-Ca to dl-Met of 65.7% on a product-to-product (wt/wt) basis or 78.2% on an equimolar basis and that a preference for Met sources was observed in barrows but not in gilts.

Dietary Supplementation of dl-Methionine Potently Induces Sodium-Dependent l-Methionine Absorption in Porcine Jejunum Ex Vivo.

BACKGROUND: Methionine is an essential amino acid (AA) with many fundamental roles. Humans often supplement l-Met, whereas dl-Met and dl-2-hydroxy-4-(methylthio)butanoic acid (dl-HMTBA) are more frequently used to supplement livestock. OBJECTIVES: The study aimed to investigate whether dietary Met source alters the absorptive capacity for Met isomers in the small intestine of piglets. METHODS: A total of 27 male 10-wk-old piglets in 3 feeding groups received a diet supplemented with 0.21% dl-Met, 0.21% l-Met, or 0.31% dl-HMTBA to meet the Met + cystine requirement. After ≥10 d, absorptive fluxes of d-Met or l-Met were measured at a physiological concentration of 50 µM and a high concentration of 5 mM in duodenum, middle jejunum, and ileum ex vivo. Data were compared by 2-factor ANOVA. RESULTS: Across diets, fluxes of both Met isomers at both tested concentrations increased from duodenum to ileum by a factor of ∼2-5.5 (P < 0.05). Pigs supplemented with dl-Met had greater (P < 0.085) absorptive fluxes at 50 µM l-Met (0.50, 2.07, and 3.86 nmol · cm-2 · h-1) and d-Met (0.62, 1.41, and 1.19 nmol · cm-2 · h-1) than did pigs supplemented with dl-HMTBA (l-Met: 0.28, 0.76, and 1.08 nmol · cm-2 · h-1; d-Met: 0.34, 0.58, and 0.64 nmol · cm-2 · h-1) in duodenum, jejunum, and ileum, respectively. Only in jejunum of dl-Met-fed pigs, fluxes at 50 µM l-Met were reduced by the omission of luminal Na+ (from 3.27 to 0.86 nmol · cm-2 · h-1; P < 0.05) and by a cocktail of 22 luminal AAs (to 1.05 nmol · cm-2 · h-1; P < 0.05). CONCLUSIONS: Dietary supplementation of dl-Met increases the efficiency of l-Met and d-Met absorption at physiologically relevant luminal Met concentrations along the small intestine of pigs, including a very prominent induction of an Na+-dependent transport system with preference for l-Met in the mid-jejunum. Dietary supplementation with dl-Met could be a promising tool to improve the absorption of Met and other AAs.

The effect of reduced dietary glycine and serine and supplemental threonine on growth performance, protein deposition in carcass and viscera, and skin collagen abundance of nursery pigs fed low crude protein diets.

Thirty five barrows (initial body weight [BW]: 15.1 ± 1.0 kg) were used to determine the effect of partially replacing Gly + Ser with Thr in reduced crude protein (CP) diets on growth performance, protein deposition in carcass and viscera, and skin collagen abundance during the late nursery phase to 25 kg BW. Pigs were individually fed one of five iso-nitrogenous diets (n = 7) for 21 d. The basal diet met estimated essential amino acids (AA) requirements by using all essential AA plus Gly and Ser in free form (CON; 12.1% CP; as-fed, analyzed contents). The remaining four diets were formulated by reducing total Gly and Ser concentrations to 60% or 20% of the CON diet. The N removed with Gly and Ser was replaced with either crystalline Thr or Glu. Total analyzed Thr made up either 1.59% (T1; 12.5% CP) or 2.34% (T2; 12.2% CP) of the Thr-supplemented diets, and total analyzed Glu made up either 3.47% (G1; 12.7% CP) or 4.64% (G2; 12.9% CP) of the Glu-supplemented diets. Pigs were slaughtered on day 21 to determine body composition and skin collagen abundance via bright field microscopy. Overall, average daily gain (ADG) and G:F and final carcass weights were greater for pigs fed diets supplemented with Glu (G1 + G2) vs. those fed diets supplemented with Thr (T1 + T2; P < 0.05, P = 0.060, and P = 0.050 for ADG, G:F, and final carcass weight, respectively); intermediate values were observed for CON. Nitrogen retention in carcass plus viscera and the AA profile of deposited protein in the carcass were not influenced by dietary treatment. Pigs fed the T2 and G2 diets had greater retention of Thr (vs. CON and G2) and Glu (vs. CON and T2) in the viscera protein, respectively (P < 0.05). The apparent utilization efficiency of standardized ileal digestible Thr for protein deposition in carcass plus viscera was less for pigs fed T2 (15.1%) vs. those fed CON (56.7%) or G2 (58.6% ± 2.9%) diets (P < 0.001). Only pigs fed T1 had skin collagen abundance not different from CON; pigs fed G1, G2, and T2 had reduced skin collagen abundance compared with CON and T1 (P < 0.01). Using Glu as an N source when Gly and Ser were reduced to 60% and 20% of CON in reduced CP diets maintained ADG for pigs between 15 and 25 kg BW, whereas supplying Thr as a N source reduced ADG and carcass weight. When dietary Gly and Ser were supplied at 60% of CON, only Thr supplementation rescued skin collagen abundance. Therefore, supplemental Thr at excess levels is not sufficient to replace N from Gly and Ser in reduced CP diets fed to late nursery pigs, despite supporting skin collagen abundance as a secondary indicator of Gly status.

Dietary protein-bound or free amino acids differently affect intestinal morphology, gene expression of amino acid transporters, and serum amino acids of pigs exposed to heat stress.

Pigs exposed to heat stress (HS) increase body temperature in which can damage the intestinal epithelia and affect the absorption and availability of amino acids (AA). Protein digestion and metabolism further increase body temperature. An experiment was conducted with six pairs of pigs (of 47.3 ± 1.3 kg initial body weight) exposed to natural HS to assess the effect of substituting dietary protein-bound AA by free AA on morphology and gene expression of intestinal epithelial and serum concentration (SC) of free AA. Treatments were: high protein, 21.9% crude protein (CP) diet (HShp) and low protein, 13.5% CP diet supplemented with crystalline Lys, Thr, Met, Trp, His, Ile, Leu, Phe, and Val (HSaa). The HShp diet met or exceeded all AA requirements. The HSaa diet was formulated on the basis of ideal protein. Pigs were fed the same amount at 0700 and 1900 hours during the 21-d study. Blood samples were collected at 1700 hours (2.0 h before the evening meal), 2030 hours, and 2130 hours (1.5 and 2.5 h after the evening meal). At the end, all pigs were sacrificed to collect intestinal mucosa and a 5-cm section from each segment of the small intestine from each pig. Villi measures, expression of AA transporters (y+L and B0) in mucosa, and SC of AA were analyzed. Ambient temperature fluctuated daily from 24.5 to 42.6 °C. Weight gain and G.F were not affected by dietary treatment. Villi height tended to be larger (P ≤ 0.10) and the villi height:crypt depth ratio was higher in duodenum and jejunum of pigs fed the HSaa diet (P < 0.05). Gene expression of transporter y+L in jejunum tended to be lower (P < 0.10) and transporter B0 in the ileum was lower (P < 0.05) in HSaa pigs. Preprandial (1700 hours) SC of Arg, His, Ile, Leu, Thr, Trp, and Val was higher (P < 0.05), and Phe tended to be higher (P < 0.10) in HShp pigs. At 2030 hours (1.5 h postprandial), serum Lys, Met, and Thr were higher in the HSaa pigs (P < 0.05). At 2130 hours (2.5 h), Arg, His, Ile, Phe, and Trp were lower (P < 0.05); Met was higher (P < 0.05); and Lys tended to be higher (P < 0.10) in HSaa pigs. In conclusion, feeding HS pigs with low protein diets supplemented with free AA reduces the damage of the intestinal epithelia and seems to improve its absorption capacity, in comparison with HS pigs fed diets containing solely protein-bound AA. This information is useful to formulate diets that correct the reduced AA consumption associated with the decreased voluntary feed intake of pigs under HS.

The effect of supplementing glycine and serine to a low crude protein diet on growth and skin collagen abundance of nursery pigs1.

A total of 96 newly weaned barrows (initial body weight [BW]: 6.3 ± 0.5 kg) were used to determine the effect of a low crude protein (CP) diet supplemented with Gly and Ser on growth and skin collagen abundance. Barrows were assigned to one of three experimental diets in a three-phase feeding program fed for 35 days (n = 8; pen was the experimental unit): 1) corn-soybean meal diet (CON; 20.3% to 23.1% CP; as-fed, analyzed contents); 2) low CP diet (14.8% to 21.4% CP) supplemented with Gly and Ser (G + S) to the same concentrations as CON; 3) low CP diet supplemented with Glu to maintain the same CP concentration as the G + S diet (GLU; 15.0% to 22.1% CP). On days 21 and 35, eight pigs per treatment were euthanized for the determination of physical and chemical body composition and skin collagen abundance. Pigs fed the CON diet had greater overall ADG and final BW compared to pigs fed GLU and G + S (P < 0.01). Over the entire 35-day experimental period, ADFI was not influenced by dietary treatment but G:F tended to be greater for pigs fed CON than G + S (P = 0.084), while intermediate values were observed for GLU. Carcass weights on days 21 and 35 were greater for pigs fed CON than G + S or GLU (P < 0.01). Viscera weights on day 21 were greater for CON than G + S and GLU (P < 0.05) and on day 35 were greater for CON than G + S (P < 0.05) with intermediate values observed for GLU. The N intake (g/d) between days 0 and 35 was greater for CON than G + S or GLU (P < 0.05) and N retention in combined carcass and viscera was greater for CON than G + S (P < 0.01) with intermediate values observed for GLU. No treatment effects were observed for efficiency of N utilization. Between days 0 and 21 however, the efficiency of using dietary N for N retention in carcass and viscera tended to be less for pigs fed CON vs. GLU (73.8% vs. 91.6%), while intermediate values were observed for G + S (84.3%; P = 0.095). Pigs fed CON and G + S diets had greater skin collagen abundance than pigs fed GLU on days 21 and 35 (P < 0.01). Supplementing low CP diets with Glu or with Gly and Ser at the levels used in the current study did not maintain ADG or combined carcass and viscera N retention and only the G + S diet supported skin collagen abundance not different from pigs fed CON. The importance of meeting essential AA requirements for growth are well accepted, but supplementing specific NEAA may be needed when feeding reduced CP diets to newly weaned pigs to support secondary indicators of AA status, such as skin collagen abundance.

Bioavailability of valine in spray-dried L-valine biomass is not different from that in crystalline L-valine when fed to weanling pigs1.

An experiment was conducted to test the hypothesis that Val from a spray dried L-Val fermentation biomass (Val-FB; 64.4% L-Val) has a bioavailability of 100% relative to Val from L-Val (98% L-Val) when fed to weanling pigs. A Val-deficient basal diet containing 0.63% standardized ileal digestible (SID) Val was formulated. Six additional diets were prepared by supplementing the basal diet with 0.08%, 0.17%, or 0.25% L-Val or 0.12%, 0.25%, or 0.37% Val-FB to create experimental diets from both Val sources that contained 0.71%, 0.79%, or 0.87% SID Val. Two hundred twenty-four weaned pigs (6.87 ± 0.64 kg initial BW) were allotted to a randomized complete block design with 7 diets, 4 pigs per pen, and 8 replicate pens per diet. Diets were fed for 20 d. At the conclusion of the experiment, a blood sample from 1 pig per pen was analyzed for blood urea nitrogen (BUN) and plasma free AA. A linear regression model was used to estimate the relative bioavailability (RBV) of Val in Val-FB relative to Val from L-Val. Results indicated that the final BW and ADG were greater (P < 0.01) for pigs fed diets supplemented with Val-FB than pigs fed diets supplemented with L-Val. The ADFI decreased (linear, P ≤ 0.01), whereas G:F increased (linear, P < 0.01) by increasing inclusion of both Val sources in the diets. Regardless of source of dietary Val, BUN values were reduced (linear and quadratic, P < 0.01) as the concentration of Val in the diet increased. Pigs fed diets supplemented with L-Val had increased (linear and quadratic, P ≤ 0.05) concentrations of Val and Arg in plasma, and plasma concentrations of Ile, Leu, Lys, Ala, Cys, and Pro linearly increased (P < 0.05). There was also an increase (linear, P < 0.05) in plasma concentrations of Ile, Leu, Met, Ala, Asp, Cys, and Pro as Val-FB was added to the diets, and the concentration of Val in plasma increased (linear and quadratic, P < 0.05). Using L-Val as the standard, the RBV of Val in Val-FB as determined by ADG, G:F, and final BW was 146%, 135%, and 143%, respectively, with 95% confidence intervals of 99% to 191%, 83% to 187%, and 70% to 217%, respectively. In conclusion, the linear regression estimated a RBV of at least 100% for Val in Val-FB relative to Val from L-Val, and pigs fed diet supplemented with Val-FB had greater final BW, ADG, and G:F than pigs fed diets supplemented with the same amount of Val from L-Val.

Effect of supplemental threonine above requirement on growth performance of Salmonella typhimurium challenged pigs fed high-fiber diets1.

It was shown previously that high dietary fiber (DF) and immune system stimulation (ISS) with systemic Escherichia coli lipopolysaccharide independently increased the threonine (Thr) requirement to maximize growth performance and protein deposition (PD). However, no additive effects on the Thr requirement were observed when both DF and ISS were present. The objective of the present study was to investigate whether supplementing Thr to meet previously estimated requirements for high DF and systemic immune challenge would maintain performance of pigs exposed to an enteric immune challenge when fed high DF. A total of 128 pigs (22.6 ± SD = 1.6 kg initial BW) were assigned to 1 of 4 dietary treatments in a 2 × 2 factorial arrangement in a randomized complete block design (n = 8 pens/treatment and 4 pigs/pen) for 28 d. Treatments were a low-fiber (LF; 13% total DF) or high-fiber (HF; 20% total DF) diet with either a standard (STD; 0.65% SID) or supplemental (SUP; 0.78% SID) Thr level. After a 7-d adaptation, pigs were orally inoculated with 2 mL (2.3 × 109 CFU/mL) of Salmonella typhimurium (ST). Blood samples and rectal swabs were obtained and rectal temperature recorded to determine clinical responses and ST shedding. On day 7 postinoculation, 1 pig/pen was euthanized and mesenteric lymph nodes, spleen, and digesta (ileum, cecum, and colon) were sampled to assess ST colonization and translocation. Body weight and feed intake were recorded on day 0, 7, and 21 postinoculation to calculate ADG, ADFI, and G:F. Rectal temperature increased (P < 0.05) 24 h postinoculation and remained elevated at day 6. Serum albumin concentration decreased (P < 0.05), whereas haptoglobin concentration increased (P < 0.05) postinoculation. There was no fiber or Thr effect (P > 0.05) on ST counts in the ileum and cecum, but a fiber × Thr interaction (P < 0.05) was observed in the colon. Supplemental Thr improved (P < 0.05) growth performance in LF- and HF-fed challenged pigs. However, performance of supplemented HF challenged pigs was less than (P < 0.05) supplemented LF challenged pigs. These results suggest that Thr supplemented to meet requirements for high DF and systemic immune challenge was not sufficient to maintain growth performance of pigs fed HF diets and challenged with an enteric pathogen.

Effects of dietary methionine supplementation on growth performance, intestinal morphology, antioxidant capacity and immune function in intra-uterine growth-retarded suckling piglets.

This study investigated the effects of dietary supplementation with L -methionine (L -Met), DL -methionine (DL -Met) and calcium salt of the methionine hydroxyl analog (MHA-Ca) on growth performance, intestinal morphology, antioxidant capacity and immune function in intra-uterine growth-retarded (IUGR) suckling piglets. Six normal birthweight (NBW) female piglets and 24 same-sex IUGR piglets were selected at birth. Piglets were fed nutrient adequate basal diet supplemented with 0.08% L -alanine (NBW-CON), 0.08% L -alanine (IUGR-CON), 0.12% L -Met (IUGR-LM), 0.12% DL -Met (IUGR-DLM) and 0.16% MHA-Ca (IUGR-MHA-Ca) from 7 to 21 days of age respectively (n = 6). The results indicated that IUGR decreased average daily milk (dry matter) intake and average daily gain and increased feed conversion ratio of suckling piglets (p < 0.05). Compared with the NBW-CON piglets, IUGR also impaired villus morphology and reduced antioxidant capacity and immune homeostasis in the intestine of IUGR-CON piglets (p < 0.05). Supplementation with L -Met enhanced jejunal villus height (VH) and villus area and ileal VH of IUGR piglets compared with IUGR-CON piglets (p < 0.05). Similarly, DL -Met supplementation increased VH and the ratio of VH to crypt depth in the jejunum compared with IUGR-CON pigs (p < 0.05). Supplementation with L -Met and DL -Met (0.12%) tended to increase reduced glutathione content and reduced glutathione: oxidized glutathione ratio and decrease protein carbonyl concentration in the jejunum of piglets when compared with the IUGR-CON group (p < 0.10). However, supplementation with MHA-Ca had no effect on the intestinal redox status of IUGR piglets (p > 0.10). In conclusion, supplementation with either L -Met or DL -Met has a beneficial effect on the intestinal morphology and antioxidant capacity of IUGR suckling piglets.

Substitution of Dietary Sulfur Amino Acids by DL-2-hydroxy-4-Methylthiobutyric Acid Increases Remethylation and Decreases Transsulfuration in Weaned Piglets.

BACKGROUND: DL-2-hydroxy-4-methylthiobutyric acid (DL-HMTBA), an L-methionine (L-Met) hydroxyl analogue, has been suggested to be a dietary L-Met source. How dietary DL-HMTBA compared with L-Met affects whole-body L-Met kinetics in growing individuals is unknown. OBJECTIVES: We determined to what extent DL-HMTBA supplementation of an L-Met-deficient diet affects whole-body L-Met and L-cysteine (L-Cys) kinetics, protein synthesis (PS), and the L-Met incorporation rate in liver protein (L-MetInc) compared with L-Met and DL-Met supplementation in a piglet model. METHODS: Forty-five, 28-d-old weaned piglets (male, German Landrace) were allocated to 4 dietary groups: L-Met-deficient diet [Control: 69% of recommended L-Met plus L-Cys supply; 0.22% standardized ileal digestible (SID) L-Met; 0.27% SID L-Cys; n = 12] and Control diet supplemented equimolarly to 100% of recommended intake with either L-Met (n = 12; LMET), DL-Met (n = 11; DLMET), or DL-HMTBA (n = 10; DLHMTBA). At 47 d of age, the piglets were infused with L-[1-13C; methyl-2H3]-Met and [3,3-2H2]-Cys to determine the kinetics and PS rates. Plasma amino acid (AA) concentrations, hepatic mRNA abundances of L-Met cycle and transsulfuration (TS) enzymes, and L-MetInc were measured. RESULTS: During feed deprivation, L-Met kinetics did not differ between groups, and were ≤3 times higher in the fed state (P < 0.01). Remethylation (RM) was 31% and 45% higher in DLHMTBA than in DLMET and Control pigs, respectively, and the RM:transmethylation (TM) ratio was 50% higher in DLHMTBA than in LMET (P < 0.05). Furthermore, TS and the TS:TM ratio were 32% lower in DLHMTBA than in LMET (P < 0.05). L-MetInc was 42% lower in DLMET and DLHMTBA than in L-Met-deficient Control pigs, whereas plasma AA and hepatic mRNA abundances were similar among DL-HMTBA-, L-Met-, and DL-Met-supplemented pigs. CONCLUSIONS: In piglets, DL-HMTBA compared with L-Met and DL-Met supplementation increases RM and reduces the TS rate to conserve L-Met, but all 3 Met isomers support growth at a comparable rate.

Extra dietary protein-bound or free amino acids differently affect the serum concentrations of free amino acids in heat-stressed pigs1.

Pigs exposed to heat stress (HS) reduce feed intake and consequently the consumption of AA. Adding extra protein-bound or free AA to the diet may correct the reduced AA intake of HS pigs. However, extra protein-bound AA may further increase the body heat load, whereas extra free AA does not affect the heat load of HS pigs. Two experiments were conducted. In Exp. 1, the performance depression because of HS, compared with thermal neutrality, was determined with 30 pigs (31.1 ± 1.2 kg BW) fed diets with AA only as protein or as a mix of protein and free AA. Heat stress pigs consumed 18 to 25% less Lys and Thr than thermal neutral. In Exp. 2, the effect of extra dietary protein-bound or free AA on performance and serum concentration of AA in 25 HS pigs (33.6 ± 0.65 kg BW) was evaluated. Treatments were as follows: CON, wheat-soybean meal-free Lys-Thr-Met diet; xP diet, 26% more protein than the CON diet; xAA diet, 24% or more of each AA than the recommended level. Pigs were fed ad libitum. Blood samples were collected between 1600 and 1700 h, when pigs were exposed to the highest ambient temperature (around 41.3 °C). Body temperature ranged daily from 39.9 to 41.1 °C. The performance data were reported already. Pigs fed the xP diet consumed more of all indispensable AA and dispensable AA than the CON pigs (P < 0.05), and more Arg, Ile, Asp, Glu, Gly, and Ser (P < 0.05) than the xAA pigs. Except for Arg, xAA pigs consumed more indispensable AA than the CON pigs (P < 0.05). Serum Arg, His, Lys, Phe, Thr, Trp, and Val, was higher (P < 0.05) in xP than in CON pigs. Except for Ile serum, indispensable AA were higher in xAA than in CON pigs (P < 0.05). Serum Ile, Leu, Thr, and Val were higher (P < 0.05), and Met tended to be higher (P < 0.10) in xAA than in xP pigs. The difference of Ile, Leu, Met, Thr, and Val between the CON and the xAA pigs was larger than that between the CON and the xP pigs (P < 0.05). Serum Asn and Tyr were higher, and Cys and Glu were lower (P < 0.05) in xP than in CON pigs. Serum Cys tended to be lower in xAA than in CON pigs (P < 0.10). Asp and Glu were higher (P < 0.05) in xAA pigs than in xP pigs. In conclusion, these serum AA results combined with the reported performance data indicate that extra free AA in diets for HS pigs may help to correct the reduced AA availability and performance of HS pigs, although higher levels of specific AA such as Ile and Met might be needed.

Effects of dietary supplementation of l-methionine vs. dl-methionine on performance, plasma concentrations of free amino acids and other metabolites, and myogenesis gene expression in young growing pigs.

Methionine (Met), the second or third limiting amino acid (AA) in typical swine diets, plays important roles in promoting swine health and growth, especially, muscle growth. Whereas dl-Met products have been used in swine industry for many years, l-Met products have been developed recently. This research was conducted to study the effects of supplemental l-Met or dl-Met on nutrient metabolism, muscle gene expression, and growth performance of pigs. Twenty crossbred young barrows (initial body weight [BW] 21.2 ± 2.7 kg) were randomly assigned to 20 individual pens and two dietary treatments according to a completely randomized design with pigs serving as the experiment unit (n = 10). Two corn and soybean meal-based diets (diets 1 and 2) were formulated to meet or exceed the recommended requirements for energy, AA, and other nutrients (NRC. 2012. Nutrient requirements of swine, 11th ed. Washington, DC: The National Academies Press; AMINODat 5.0). Crystalline l-Met and dl-Met were supplemented to diets 1 and 2 (both at 0.13%, as-fed basis), respectively. After 4 wk of an ad libitum feeding trial, BW and feed intake were measured to calculate average daily gain (ADG), average daily feed intake (ADFI), and gain-to-feed ratio (G:F). Blood samples were collected from the jugular vein for analyses of plasma AA and metabolite concentrations. The longissimus dorsi muscle samples were collected for analysis of myogenesis gene expression. Data were analyzed using Student's t-test. There were no differences (P = 0.56 to 0.94) in ADG, ADFI, or G:F between pigs fed the two experimental diets and no differences between diets were observed in plasma free AA concentrations. No differences were observed between pigs fed the two diets in expression of mRNA for eight myogenesis-related genes, which were myogenic differentiation 1, myogenin, myogenic factors 5, muscle regulatory factor 4 (a.k.a. myogenic factors 6), and myocyte enhancer factors 2A, 2B, 2C, and 2D. In conclusion, results of this experiment indicate that the bioefficacy of l-Met is not different from that of dl-Met, which is likely because of an efficient conversion of d-Met to l-Met by pigs.