Prebiotic Xylo-Oligosaccharides Modulate the Gut Microbiome to Improve Innate Immunity and Gut Barrier Function and Enhance Performance in Piglets Experiencing Post-Weaning Diarrhoea

Abstract

During commercial pig production, weaning is a major stressor that disrupts the gut microbiome, compromises intestinal barrier integrity, and increases the susceptibility of piglets to pathogens. This often results in post-weaning diarrhoea (PWD), leading to growth retardation, morbidity, and economic loss. This study investigated the effects of dietary xylo-oligosaccharide (XOS) supplementation on the growth performance and gut health of 216 piglets with naturally occurring PWD. Piglets received either 0 (CON), 50 (XOS-50), or 500 (XOS-500) mg XOS/kg feed from weaning at 28 days of age (d1) for 54 days. XOS-500 significantly improved body weight at d22 and d54, but had no effect on average daily gain, daily feed intake (DFI), or feed conversion ratio. The intestinal microbiota alpha-diversity was unaffected by XOS, though jejunal beta diversity differed between CON and XOS-500 groups at d22. Jejunal Chao richness correlated positively with d54 body weight, while ileal Chao richness correlated negatively with DFI. Salmonella was present in all diet groups but did not differ in abundance; however, the levels were negatively correlated with alpha diversity. XOSs increased Lactobacillus (d22, d54) and Clostridium_XI (d22), while reducing Veillonellaceae spp. (d22). XOSs reduced jejunal goblet cell (GC) density at d22 but increased duodenal and jejunal GCs and reduced duodenal crypt depth at d54. XOSs upregulated the genes for the tight junction proteins CLDN2, CLDN3, ALPI, and ZO-1, while downregulating the cytokine IL-8. These findings highlight XOSs' potential to improve growth and gut health in weaning piglets with naturally occurring PWD, to maintain productivity and enhance welfare.

Keywords: dietary carbohydrates; growth; histology; immunity; intestinal health; microbiota; oligosaccharides; pigs; prebiotics.

Figure 1

Body weights of control and XOS-fed piglets at d1, d7, d14, d22, and d54 post-weaning. Bars represent the average body weight across the replicate pens for each diet. Error bars represent standard error of the mean. n = 12 pens per diet group. Asterisks indicate statistically significant pairwise differences between diet groups as determined by Tukey’s HSD test: *, p < 0.05.

Figure 2

ADG, DFI, and FCR of piglets on control or XOS diets from d1 to d54. Boxplots display the median, first quartile, and third quartile of ADG, DFI, and FCR. Box whiskers extend to the smallest and largest values within 1.5 times the interquartile range. n = 12 pens per diet group. p-values from Kruskal–Wallis tests.

Figure 3

Correlations between d54 alpha diversity and performance metrics. (A) Jejunal Chao richness and d54 body weight for CON (n = 6), XOS-50 (n = 9), and XOS-500 (n = 7) pigs. (B) Ileal Chao richness and DFI from d1 to d54 for CON (n = 11), XOS-50 (n = 12), and XOS-500 (n = 11) pigs. Green points, CON; orange points; XOS-50; purple points, XOS-500. r_s, Spearman’s rank correlation coefficient; p, Spearman’s rank test p-value.

Figure 4

The relative abundance of bacterial taxa annotated to OTUs at the phylum and genus level identified from the GIT digesta of piglets fed a control or XOS-supplemented diet. UC = unclassified at the phylum or genus level. CD = CON duodenum; X50D = XOS-50 duodenum; X500D = XOS-500 duodenum; CJ = CON jejunum; X50J = XOS-50 jejunum; X500J = XOS-500 jejunum; CI = CON ileum; X50I = XOS-50 ileum; X500I = XOS-500 ileum; CCa = CON caecum; X50Ca = XOS-50 caecum; X500Ca = XOS-500 caecum; CCo = CON colon; X50Co = XOS-50 colon; X500Co = XOS-500 colon; CR = CON rectum; X50R = XOS-50 rectum; X500R = XOS-500 rectum. Sample sizes for each day, GIT location, and diet group are as follows: d22—duodenum (CON: n = 10, XOS-50: n = 9, XOS-500: n = 9), jejunum (CON: n = 10, XOS-50: n = 10, XOS-500: n = 9), ileum (CON: n = 12, XOS-50: n = 12, XOS-500: n = 12), caecum (CON: n = 12, XOS-50: n = 12, XOS-500: n = 12), colon (CON: n = 12, XOS-50: n = 12, XOS-500: n = 12), and rectum (CON: n = 9, XOS-50: n = 12, XOS-500: n = 12); d54—duodenum (CON: n = 6, XOS-50: n = 9, XOS-500: n = 7), jejunum (CON: n = 6, XOS-50: n = 9, XOS-500: n = 7), ileum (CON: n = 11, XOS-50: n = 12, XOS-500: n = 11), caecum (CON: n = 12, XOS-50: n = 12, XOS-500: n = 12), colon (CON: n = 12, XOS-50: n = 12, XOS-500: n = 11), and rectum (CON: n = 12, XOS-50: n = 12, XOS-500: n = 11).

Figure 5

Discriminative taxa identified by ALDEx2 and ANCOM-BC2 between control and XOS-fed pigs. (A) Differentially abundant OTUs in the duodenum, jejunum, and ileum between control and XOS-fed pigs. (B) Differentially abundant OTUs in the caecum, colon, and rectum between control and XOS-fed pigs. Shapes without a black outline represent differentially abundant OTUs between CON and XOS-50 pigs; shapes with a black outline represent differentially abundant OTUs between CON and XOS-500 pigs. Sample sizes for each day, GIT location, and diet group are as follows: d22—duodenum (CON: n = 10, XOS-50: n = 9, XOS-500: n = 9), jejunum (CON: n = 10, XOS-50: n = 10, XOS-500: n = 9), ileum (CON: n = 12, XOS-50: n = 12, XOS-500: n = 12), caecum (CON: n = 12, XOS-50: n = 12, XOS-500: n = 12), colon (CON: n = 12, XOS-50: n = 12, XOS-500: n = 12), and rectum (CON: n = 9, XOS-50: n = 12, XOS-500: n = 12); d54—duodenum (CON: n = 6, XOS-50: n = 9, XOS-500: n = 7), jejunum (CON: n = 6, XOS-50: n = 9, XOS-500: n = 7), ileum (CON: n = 11, XOS-50: n = 12, XOS-500: n = 11), caecum (CON: n = 12, XOS-50: n = 12, XOS-500: n = 12), colon (CON: n = 12, XOS-50: n = 12, XOS-500: n = 11), and rectum (CON: n = 12, XOS-50: n = 12, XOS-500: n = 11). A list of bacterial taxa annotated to the OTUs presented in this figure is shown in Table 3.

Figure 6

Correlations between alpha diversity and HSP60/invA ratios. Green points, CON (n = 49); orange points, XOS-50 (n = 73); purple points, XOS-500 (n = 68). r_s, Spearman’s rank correlation coefficient; p, Spearman’s rank test p-value.

Figure 7

Differences in upper intestinal gut architecture and GC expression between control and XOS-fed pigs. Gut architecture and GC density changes on d22 in the jejunum (A–C) and ileum (D,E), and on d54 in the duodenum (F,G), jejunum (H), and ileum (I). *, p < 0.05; **, p < 0.01; ***, p < 0.001; Kruskal–Wallis and Dunn’s tests.

Figure 8

Changes in expression of CLDN2, CLDN3, ALPI, ZO-1, and IL-8 in piglet GIT tissues. Gene expression was quantified by qRT-PCR, and fold change was calculated using the 2^−ΔΔct method. Fold changes were then log₂-transformed, where all CON values are zero by definition and, therefore, are omitted from the plot, allowing upregulation and downregulation to be visualised as deviations from the x-axis. Expression of each gene of interest was normalised to the average of housekeeping genes GAPDH and RPL4 for each sample. Orange bars represent XOS-50 log₂ (fold changes), and purple bars represent XOS-500 log₂ (fold changes). An asterisk directly above or below a bar indicates a significant difference compared to the CON group, while an asterisk with a connecting line between bars denotes a significant difference between XOS-50 and XOS-500 pigs. For each sample and gene of interest, ct values >35 were excluded from analysis, and outliers were removed using the 1.5 × IQR method. Sample sizes for each gene and diet group presented are as follows: CLDN2 (CON: n = 10, XOS-50: n = 11, XOS-500: n = 10), CLDN3 (CON: n = 11, XOS-50: n = 12, XOS-500: n = 11), ALPI (CON: n = 9, XOS-50: n = 11, XOS-500: n = 9), ZO-1 (CON: n = 9, XOS-50: n = 10, XOS-500: n = 11), and IL-8 (CON: n = 11, XOS-50: n = 11, XOS-500: n = 11). *, p < 0.05.