Short-chain fructo-oligosaccharides enhances intestinal barrier function by attenuating mucosa inflammation and altering colonic microbiota composition of weaning piglets

Figures & data

Table 1. Composition and nutrient level of basal diet (air dry-basis).

Ingredients	Content (%)	Nutrient level^c	Content
Maize	30.83	DE / (MJ/kg)	15.09
Extruded maize	24.00	CP (%)	20.88
Extruded soybean	6.00	Ca (%)	0.80
Dehulled soybean meal (44% CP)	10.00	AP (%)	0.41
Soybean protein concentrate	10.00	SID Lys (%)	1.35
Whey powder	5.00	SID Met (%)	0.46
Fish meal	5.00	SID Thr (%)	0.80
Soybean oil	1.80	SID Trp (%)	0.22
Sucrose	3.00	–	–
Glucose	2.00	–	–
Salt	0.30	–	–
Limestone	0.72	–	–
CaHPO4	0.52	–	–
Chloride choline	0.10	–	–
Vitamin premix^a	0.05	–	–
Mineral premix^b	0.30	–	–
DL-Met (98.5%)	0.13	–	–
L-Lys·HCl (78%)	0.18	–	–
L-Thr (98.5%)	0.07	–	–
Total	100.00	–	–

^aThe vitamin premix provided the following per kg of the diet: retinyl acetate, 0.95 mg; cholecalciferol, 0.02 mg; DL-α-tocopheryl acetate, 28 mg; menadione, 2 mg; riboflavin, 1 mg; pyridoxine, 3 mg; vitamin B12, 48 μg; D-pantothenic acid, 30 mg; folic acid, 2.0 mg; biotin, 1 mg.

^bThe mineral premix provided the following per kg of the diet: Fe (ferrous sulphate), 100 mg; Cu (copper sulphate), 6 mg; Mn (manganese oxide), 4 mg; Zn (zinc sulphate), 100 mg; I (potassium iodate), 0.14 mg; Se (odium selenite), 0.35 mg.

^cNutrient levels were calculated values.

DE: Digestible energy; CP: Crude protein; Ca: Calcium; AP: Available phosphorus; SID Lys: Standard ileal digestible lysine; SID Met: Standard ileal digestible methionine; SID Thr: Standard ileal digestible threonine; SID Trp: Standard ileal digestible tryptophan.

Table 2. Nucleotide sequences of primers used to measure targeted genes.

Gene symbols	Nucleotide sequence of primers (5′-3′)	Accession No.	Annealing temperature
ACTB	F: TCTGGCACCACACCTTCT R: TGATCTGGGTCATCTTCTCAC	XM_003124280.3	60
TOP2B	F: AACTGGATGATGCTAATGATGCT R: TGGAAAAACTCCGTATCTGTCTC	NM_001258386.1	60
TBP	F: GATGGACGTTCGGTTTAGG R: AGCAGCACAGTACGAGCAA	DQ178129	59
YWHAZ	F: ATGCAACCAACACATCCTATC R: GCATTATTAGCGTGCTGTCTT	DQ178130	60
IL-1β	F: ACGTGCAATGATGACTTTGTCTG R: AGAGCCTTCAGCATGTGTGG	NM_214055.1	58
IL-6	F: TTCACCTCTCCGGACAAAAC R: TCTGCCAGTACCTCCTTGCT	NM_214399.1	60
TNF-α	F: CATGATCCGAGACGTGG R: CTCGAAGTGCAGTAGGC	NM_214022.1	62
IL-10	F: TAATGCCGAAGGCAGAGAGT R: GGCCTTGCTCTTGTTTTCAC	NM_214041.1	60
ZO1	F: ATCTCGGAAAAGTGCCAGGA R: CCCCTCAGAAACCCATACCA	XM_003480423.3	61
ZO2	F: CCAGGAAGCACAGAATGCAA R: AAGTCTGGCGGGACCTCTCT	XM_005660148.2	61
OCLN	F: CATGGCTGCCTTCTGCTTCATTGC R: ACCATCACACCCAGGATAGCACTCA	NM_001163647.2	65
CLDN1	F: TATGACCCCATGACCCCAGT R: GCAGCAAAGTAGGGCACCTC	NM_001244539.1	59
CLDN2	F: TTCCTCCCTGTTCTCCCTGA R: CACTCTTGGCTTTGGGTGGT	NM_001161638.1	62

All the primers were designed according to the sequence which can be reached by corresponding accession number and can only amplify the isoform listed above.

ACTB: β-actin; TOP2B: topoisomerase (DNA) IIb; TBP: TATA box binding protein; YWHAZ: tyrosine 3-monooxygenase/tryptophan 5-monooxygenase activation protein; IL-1β: interleukin 1 beta; IL-6: interleukin 6; TNF-α: tumour necrosis factor alpha; IL-10: interleukin 10; ZO1: zona occludens 1; ZO2: zona occludens 2; OCLN: occludin; CLDN1: claudin 1; CLDN2: claudin 2.

Table 3. Primer sequences used to analyse abundances of selected microbes.

Microbes	Nucleotide sequence of primers (5′-3′)
Universal	F: ACTCCTACGGGAGGCAGCAGT
	R: TATTACCGCGGCTGCTGGCAC
Firmicutes	F: TGAAACTCAAAGGAATTGACG
	R: ACCATGCACCACCTGTC
Bacteroidetes	F: GGAACATGTGGTTTAATTCGATGAT
	R: AGCTGACGACAACCATGCAG
Lactobacillus spp.	F: AGCAGTAGGGAATCTTCCA
	R: CACCGCTACACATGGAG
Prevotella spp.	F: CACCAAGGCGACGATCA
	R: GGATAACGCCTGGACCT
Bacteroides spp.	F: CGATGGATAGGGGTTCTGAGAGGA
	R: GCTGGCACGGAGTTAGCCGA
Ruminococcus spp.	F: GAGTGAAGTAGAGGTAAGCGGAATTC
	R: GCCGTACTCCCCAGGTGG
Roseburia spp.	F: TACTGCATTGGAAACTGTCG
	R: CGGCACCGAAGAGCAAT
Bifidobacterium spp.	F: GCGTGCTTAACACATGCAAGTC
	R: CACCCGTTTCCAGGAGCTATT

Table 4. Effects of short-chain fructo-oligosaccharides on growth performance and diarrhoea index of weaning piglets.

Items	Control group	scFOS group	p Value
Initial body weight, kg	7.02 ± 0.16	7.03 ± 0.13	.84
Final body weight, kg	14.99 ± 0.41	15.52 ± 0.49	<.05
Average daily feed intake, g	477.50 ± 16.66	486.11 ± 22.50	.40
Average daily weight gain, g	284.82 ± 9.81	302.98 ± 13.08	<.05
Feed to gain ratio	1.68 ± 0.03	1.60 ± 0.02	<.05
Diarrhoea index	0.24 ± 0.04	0.19 ± 0.02	<.05

scFOS: short-chain fructo-oligosaccharides.

Table 5. Effects of short-chain fructo-oligosaccharides on serum intestinal permeability biomarkers and cytokines concentrations of weaning piglets.

Items	Control group	scFOS group	p Value
Diamine oxidase, U/L	8.71 ± 1.54	7.38 ± 0.94	.06
D-lactate, μg/mL	23.14 ± 1.50	20.47 ± 1.69	<.05
Endotoxin, EU/mL	195.55 ± 13.56	201.38 ± 12.11	.38
IL-1β, pg/mL	179.02 ± 12.27	162.10 ± 14.18	<.05
IL-6, pg/mL	45.36 ± 4.81	31.90 ± 5.06	<.05
IL-10, pg/mL	113.59 ± 7.09	114.87 ± 11.24	.79
TNF-α, pg/mL	218.77 ± 21.49	186.04 ± 23.44	<.05

scFOS: short-chain fructo-oligosaccharides; EU: endotoxin units; IL-1β: interleukin 1 beta; IL-6: interleukin 6; IL-10: interleukin 10; TNF-α: tumour necrosis factor alpha.

Table 6. Effects of short-chain fructo-oligosaccharides on mRNA expression of tight junction proteins in jejunum and ileum of weaning piglets.

Genes	Control group	scFOS group	p Value
Jejunum
ZO1	1.00 ± 0.20	1.51 ± 0.16	<.05
ZO2	1.00 ± 0.40	1.09 ± 0.29	.62
OCLN	1.00 ± 0.30	1.37 ± 0.35	<.05
CLDN1	1.00 ± 0.35	1.81 ± 0.30	<.05
CLDN 2	1.00 ± 0.26	1.10 ± 0.18	.39
Ileum
ZO1	1.00 ± 0.12	1.39 ± 0.08	<.05
ZO2	1.00 ± 0.46	1.36 ± 0.42	.12
OCLN	1.00 ± 0.26	1.40 ± 0.17	<.05
CLDN1	1.00 ± 0.25	1.44 ± 0.19	<.05
CLDN2	1.00 ± 0.17	1.20 ± 0.31	.13

scFOS: short-chain fructo-oligosaccharides; ZO1: zona occludens 1; ZO2: zona occludens 2; OCLN: occludin; CLDN1: claudin 1; CLDN2: claudin 2.

Table 7. Effects of short-chain fructo-oligosaccharides on mRNA expression of cytokines in jejunum and ileum of weaning piglets.

Genes	Control group	scFOS group	p Value
Jejunum
IL-1β	1.00 ± 0.17	0.66 ± 0.13	<.05
IL-6	1.00 ± 0.33	0.74 ± 0.25	.10
TNF-α	1.00 ± 0.27	0.87 ± 0.28	.37
IL-10	1.00 ± 0.11	1.15 ± 0.20	.09
Ileum
IL-1β	1.00 ± 0.23	0.87 ± 0.25	.32
IL-6	1.00 ± 0.18	0.74 ± 0.08	<.05
TNF-α	1.00 ± 0.28	1.10 ± 0.23	.45
IL-10	1.00 ± 0.14	1.12 ± 0.14	.10

scFOS: short-chain fructo-oligosaccharides; IL-1β: interleukin 1 beta; IL-6: interleukin 6; TNF-α: tumour necrosis factor alpha; IL-10: interleukin 10.

Table 8. Effects of short-chain fructo-oligosaccharides on short-chain fatty acids profile in colonic chyme of weaning piglets.

Items	Control group	scFOS group	p Value
Acetic acid, μmol/g content	53.58 ± 5.81	65.15 ± 6.91	<.05
Propionic acid, μmol/g content	32.18 ± 3.43	34.60 ± 2.63	.13
Butyric acid, μmol/g content	16.31 ± 4.11	20.16 ± 3.21	.06

scFOS: short-chain fructo-oligosaccharides.

Table 9. Effects of short-chain fructo-oligosaccharides on relative abundance of selected bacteria in colonic chyme of weaning piglets.

Microbes (%)	Control group	scFOS group	p Value
Firmicutes	40.72 ± 8.08	40.48 ± 5.29	.95
Bacteroidetes	41.77 ± 4.98	49.16 ± 4.33	<.05
Lactobacillus	1.80 ± 0.62	3.96 ± 1.07	<.05
Prevotella	20.36 ± 4.94	27.02 ± 6.44	<.05
Bacteroides	0.37 ± 0.26	0.46 ± 0.10	.40
Ruminococcus	0.46 ± 0.31	0.26 ± 0.13	.11
Roseburia	0.21 ± 0.09	0.26 ± 0.13	.86
Bifidobacterium	0.12 ± 0.06	0.90 ± 0.44	<.05

scFOS: short-chain fructo-oligosaccharides.