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Animal Nutrition Conference of Canada 2021
The following technical article is related to the event::
Animal Nutrition Conference of Canada 2021

Diet complexity and L-threonine supplementation: effects on growth performance, immune response, and intestinal integrity in nursery pigs

Published on: 8/26/2021
Author/s : Bonjin Koo, Chengbo Yang, and Charles Martin Nyachoti. / Department of Animal Science, University of Manitoba Winnipeg, MB.
Summary

Despite higher economic value of a simplified nursery pig diet compared to a conventional complex diet, a simple diet may stimulate the pig’s immune system and impair intestinal integrity. Dietary Thr supplementation is required when pigs are immunologically challenged to maintain the gut health. Thus, a study was performed to investigate the effects of diet complexity and L-Thr supplementation on growth performance, immune response and gut integrity in nursery pigs. Thirty-two pigs (body weight 7.23 ± 0.48 kg) were randomly assigned to dietary treatments in a 2 × 2 factorial arrangement based on diet complexity (complex or simple) and dietary Thr content. The complex diet contained fish meal, plasma protein and dried whey to mimic a conventional nursery diet. The simple diet was formulated with corn, wheat, and soybean meal. L-Thr was supplemented to each diet to supply either 100% (STD Thr) or 115% (SUP Thr) of the NRC (2012) requirement for Thr. Pigs were individually housed and fed the diets ad libitum for 14 d. All data were analyzed using the MIXED procedure of SAS. Diet complexity, dietary Thr content, and their interactions were considered main effects. Dietary treatment did not affect growth performance. However, pigs fed the simple diet had greater (P < 0.05) plasma interleukin (IL)-10 and IL-6 concentrations compared to those fed the complex diet on d 7 and d 14, respectively. Simple diet-fed pigs tended to show greater (P < 0.10) expression of genes encoding for tumor necrosis factor-α, claudin-1, and zonula occludens-1 in the jejunum compared to complex diet-fed pigs. The SUP Thr increased (P < 0.05) villus height and goblet cell density in villi and crypts in the jejunum and deepened (P < 0.05) crypts in the proximal colon. The SUP Thr resulted in the upregulation (P < 0.05) of occludin gene expression and tended to downregulate (P = 0.10) IL-6 gene expression in the jejunum. Interactions (P < 0.05) between diet complexity and L-Thr supplementation level were observed in goblet cell density in the crypt in the jejunum. In conclusion, SUP Thr in a simple diet would be a nutritional strategy to fortify the gut integrity in nursery pig production, but it does not prevent the induction of inflammation caused by simple diet in nursery pigs.

Introduction
Nursery pig diets have been conventionally formulated with animal protein sources and dairy products (e.g., fish meal, plasma meal, and whey protein). However, this has resulted in a complex diet composition and high feed costs. Many attempts have therefore been made to simplify the conventional complex diet by increasing the proportion of soybean meal as a way to save on feed costs in nursery pig production. Previous studies (Skinner et al. 2014; Koo et al. 2017) confirmed that simple diets do not compromise the growth performance at the end of the nursery phase. 
However, concerns remain that the large amounts of dietary fiber, fermentable protein, and antigenic compounds in soybean meal may stimulate the pig’s immune system and impair intestinal integrity (Koo et al. 2017). This may result in poor growth performance in pigs raised in a commercial swine barn environment (Pastorelli et al. 2012). Threonine is a major component of mucins and γ-globulins (Wang et al. 2009). Previous studies have reported that dietary L-Thr supplementation over the NRC (2012) recommended levels improved intestinal morphology and immune status by regulating immunoglobulins and cytokines in pigs. We hypothesized that pigs fed a simple diet would elicit systemic and gut inflammation and show impairment in their gut integrity and barrier function when compared with pigs fed a complex diet and that L-Thr supplementation would ameliorate the inflammation and the gut impairment. The objective of this study was to investigate the effects of diet complexity and L-Thr supplementation levels on the growth performance, immune response, intestinal integrity, and microbial metabolites in nursery pigs.
Methods
Thirty-two male piglets (TN 70 × TN Tempo; Topigs Norsvin, Winnipeg, MB) with an initial body weight of 7.23 ± 0.48 kg (28-d-old) were randomly assigned to a 2 × 2 factorial arrangement based on diet complexity and the levels of dietary L-Thr supplementation. Pigs were individually housed for 14 d and had ad libitum access to feed and water. The complex diet contained animal protein sources (e.g., fish meal, spray-dried animal plasma) and a dairy product (e.g., dried whey) to mimic a conventional nursery diet. These animal protein sources and dairy product were replaced with soybean meal (30.3%) to make the simple diet. The two respective diets were supplemented with L-Thr to supply the standard NRC (2012) level of Thr (STD Thr) or 15% over the standardized ileal digestible (SID) Thr requirement (SUP Thr) for pigs weighing 9 kg. On day 7 and 14, blood samples from the jugular vein of all the pigs were collected for determination of inflammatory cytokine concentrations. On day 14, all pigs were euthanized to collect jejunum and colon samples and their contents for morphology, mRNA gene expression, and volatile fatty acids (VFA) and ammonia-nitrogen (NH3-N). All data were analyzed using the MIXED procedure of SAS (version 9.4; SAS Inst. Inc., Cary, NC, USA) with each animal used as the experimental unit. The model included diet complexity, the level of L-Thr supplementation, and their interaction. 
Results 
Pigs fed the simple diet had greater (P < 0.05) plasma interleukin (IL)-10 and IL-6 concentrations compared to those fed the complex diet on day 7 and day 14, respectively (Table 1). Simple diet-fed pigs tended to show greater (P < 0.10) expression of genes encoding for tumor necrosis factor-α, claudin-1, and zonula occludens-1 in the jejunum compared to complex diet-fed pigs (Figure 1). The simple diet–fed pigs had greater (P < 0.05) concentrations of NH3-N in the jejunum digesta than did complex diet–fed pigs (Table 1).
Table 1. Effect of diet complexity and Thr supplementation on plasma cytokine concentrations, jejunal morphology, and microbial metabolites in jejunum and colon digesta.
Effect of diet complexity and Thr supplementation on plasma cytokine concentrations, jejunal morphology, and microbial metabolites in jejunum and colon digesta.
Figure 1. Effect of diet complexity and Thr supplementation on relative mRNA gene expression (2-ΔΔCt) in jejunum. The values were expressed. The gene expression values are expressed in relation to the simple-STD Thr diet group. IL = interleukin; TNF-α = tumor necrosis factor alpha; CLDN1 = claudin 1; OCLN = occludin; ZO-1 = zonula 1; MUC2 = mucin 2.
Figure 1. Effect of diet complexity and Thr supplementation on relative mRNA gene expression (2-ΔΔCt) in jejunum. The values were expressed. The gene expression values are expressed in relation to the simple-STD Thr diet group. IL = interleukin; TNF-α = tumor necrosis factor alpha; CLDN1 = claudin 1; OCLN = occludin; ZO-1 = zonula 1; MUC2 = mucin 2.
SUP Thr increased (P < 0.05) the density of goblet cells, where mucin is secreted, in villi as well as in crypts in the jejunum, regardless of diet complexity. Furthermore, SUP Thr–group showed higher villi and upregulated (P = 0.04) occludin (a tight junction protein) in the jejunum, regardless of diet complexity. Evidence is growing that dietary Thr supplementation beneficially modifies the gut microbiota composition because the stimulated mucins (glycoconjugates) can serve as substrates for microorganisms (Chen et al. 2017). Interestingly, SUP Thr in the simple diet decreased (P < 0.05) NH3 content in the jejunum to the level observed for the complex diet. Conversely, SUP Thr in the simple diet increased (P < 0.05) total VFA production in the colon compared to that in the complex diet. The SUP Thr tended to suppress (P = 0.07) the expression of IL-6 in the jejunum, but the suppressing effect was larger with a simple diet (Interaction P = 0.10). However, there was no difference in plasma IL-6 and IL-10 concentrations between the SUP Thr and the STD Thr groups. Despite various changes in gut health indicators, no differences in the growth performance of pigs were observed among dietary treatments.
Conclusion
Feeding a simple diet to nursery pigs resulted in systemic and intestinal inflammation. SUP Thr decreased NH3 production in the jejunum and resulted in downregulation of IL-6 gene expression in the jejunum. SUP Thr also improved gut integrity and architecture. The benefits of SUP Thr seems to be greater with a simple diet than with a complex diet. Therefore, SUP Thr in combination with a simple diet could be a beneficial cost-saving strategy while maintaining gut health.
Published in the proceedings of the Animal Nutrition Conference of Canada 2020. For information on the event, past and future editions, check out https://animalnutritionconference.ca/.

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