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.

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.

The dietary protein content slightly affects the body temperature of growing pigs exposed to heat stress.

Heat stress (HS) increases body temperature (BT) and reduces feed intake in pigs. Increasing the dietary protein content may correct the reduced amino acid intake provoked by HS, but it may further increase BT. The effect of dietary protein level on BT of HS pigs was analyzed with nine ileal cannulated pigs (61.7 ± 2.5 kg body weight). A thermometer set to register BT at 5-min intervals was implanted into the ileum. There were two treatments: low-protein (10.8%) wheat-free-amino acid diet (LP); high-protein (21.6%), wheat-soybean-meal diet (HP). The study was conducted in two 10-d periods; in each period, d1 to d6 was for diet adaptation, d7 to d9 was for data analysis, and d10 for ileal sample collection. Pigs were fed at 0600 h (morning), 1400 h (midday), and 2200 h (evening), same amount each time. Following, the separate contribution of ambient temperature and thermal effect of feeding on the postprandial BT increment was analyzed in fed and fasted pigs. Ambient temperature ranged from 30.1 to 35.4 °C and relative humidity from 50% to 84%. Both ambient temperature and BT followed similar patterns. The BT of HP pigs after the morning and midday meals was higher (P < 0.05) but size of the BT increments did not differ between HP and LP pigs. Midday and evening postprandial BT were higher than postprandial morning BT (P < 0.05). The BT increment was larger and longer after the midday than after the morning and evening meals (P < 0.05). The capacity of pigs to dissipate postprandial body heat depends on the accumulated thermal load received before their meals, because the thermal load before the morning meal was smaller than that before the evening meal. The estimated contribution of thermal effect of feeding (0.42 to 0.87 °C) on the total postprandial BT increment (0.69 to 1.53 °C) was larger (P < 0.05) than that of ambient temperature (0.27 to 0.66 °C). In conclusion, these data indicate that the dietary protein level has a small effect on the BT of HS pigs regardless of feeding time. Also both the thermal effect of feeding and ambient temperature impact the BT of HS pigs, although the former had a stronger effect. This information may be useful to design better feeding strategies for pigs exposed to HS conditions.

Serum concentrations of free amino acids in growing pigs exposed to diurnal heat stress fluctuations.

In areas where the ambient temperature (AT) is above the thermo neutral (TN) zone of pigs, significant changes within a 24-h period occur, differently affecting the availability of amino acids (AA) within the same day. An experiment was conducted to analyze the serum concentrations (SC) of free AA in pigs exposed to diurnal variations in AT. Six pigs (27.1 ±1.3kg body weight) implanted with a thermometer to register the body temperature (BT) at 15-min intervals were used. Blood samples were collected on the last 3 d of the 14-d study, at 0700h (lowest AT), 1200h (mild HS), and 1600h (severe HS). The pigs received 1.2kg/d of an AA-supplemented, wheat-soybean meal diet, in two equal meals (0700 and 1900h). The AT and BT, recorded at 0700, 1200, and 1600h was: 30.6, 38.6, 41.1°C, and 38.2, 39.5, 40.3°C, respectively. The BT was significantly correlated (P < 0.001) with the AT. The SC (µM/mL) of Ile, Lys, Met, Val, Ala, Asn, and Pro were higher (P ≤ 0.01); Arg, Phe, Glu, and Tyr tended to be higher (P ≤ 0.10); but Cys was lower (P < 0.05) at 1200h than at 0700h. Lys was higher, Cys and Tyr were lower (P < 0.05), and Ile and Val tended to be higher (P ≤ 0.10) at 1600h than at 0700h. Serum Arg, Ile, Phe, Ala, Asn, Gln, Pro, Ser, and Tyr were lower (P < 0.05), and Leu and Val tended to be lower at 1600h than at 1200h. These data demonstrate that AT directly alters the BT of pigs, and that diurnal variations in AT differently affect their SC and availability of AA for growth.

Effect of heat stress on performance and expression of selected amino acid and glucose transporters, HSP90, leptin and ghrelin in growing pigs.

Exposing animals to high ambient temperature provokes heat stress (HS) that may affect cellular function and reduced productive performance. The effect of chronic exposure (21d) of pigs to high ambient temperature on expression of amino acid (b(0,+)AT, CAT1) and glucose (SGLT1, GLUT4) transporters, ghrelin, leptin and HSP90 was evaluated. Eighteen pigs (32.6kg body weight) were distributed into 3 groups: (1) pigs housed under natural high ambient temperature conditions, and fed ad libitum (HS); (2) pigs housed in an air-conditioned room at 24°C (thermo-neutral) fed ad libitum (TNad); (3) pigs housed as in (2), but pair-fed with the HS pigs (TNpf). Body temperature, respiratory frequency, weight gain, feed intake, and feed conversion ratio were measured. At d-21 pigs were euthanized and samples from stomach, duodenum, jejunum, liver, longissimus and semitendinosus muscles, and white adipose tissue were collected for mRNA analysis. In the HS room ambient temperature fluctuated every day (23.6-37.6°C). Respiratory frequency and body temperature were higher in HS pigs (P<0.001). Weight gain and feed intake of TNad were higher (P<0.001) than TNpf and HS; gain: feed ratio was not affected by ambient temperature. Expression of HSP90 was higher in duodenum and longissimus (P≤0.038) of HS compared to TNpf. Expression of ghrelin, leptin and b(0,+)AT were not affected by ambient temperature (P>0.050). CAT1 expression in liver was higher (P=0.050) but in longissimus was lower (P=0.017) in HS than in TNpf pigs. Expression of SGLT1 was higher (P=0.045) in duodenum of HS than in TNpf but it was not different in jejunum (P=0.545); GLUT4 tended to be higher in liver and semitendinosus of HS pigs (P=0.063). In conclusion, feed intake remains low whereas respiratory frequency and body temperature remain higher; and expression of HSP90, CAT1, SGLT1 and GLUT4 increases in some tissues in pigs under chronic HS conditions.