Microbial composition and in vitro fermentation patterns of human milk oligosaccharides and prebiotics differ between formula-fed and sow-reared piglets

Abstract

The microbial composition and in vitro fermentation characteristics of human milk oligosaccharides (HMO), lacto-N-neotetraose (LNnT), a 2:1 mixture of polydextrose (PDX) and galactooligosaccharides (GOS), and short-chain fructooligosaccharides (scFOS) by pooled ascending colonic microbiota from 9- and 17-d-old formula-fed (FF) and sow-reared (SR) piglets were assessed. pH change and gas, SCFA, and lactate production were determined after 0, 2, 4, 8, and 12 h of incubation. In most donor groups, the pH change was greater for scFOS fermentation and lower for PDX/GOS than for other substrates. LNnT fermentation produced larger amounts of gas, total SCFA, acetate, and butyrate than did the other substrates, whereas HMO and scFOS produced higher amounts of propionate and lactate, respectively. In general, pH change, total SCFA, acetate, and propionate production were greater in pooled inoculum from FF and 9-d-old piglets, whereas SR-derived inoculum produced higher amounts of butyrate and lactate after 4 h fermentation. Gut microbiota were assessed by 16S ribosomal RNA V3 gene denaturing gradient gel electrophoresis analysis and real-time qPCR. Microbial structures differed among the 4 groups before fermentation, with higher counts of Bifidobacterium in SR piglets and higher counts of Clostridium cluster IV, XIVa, and Bacteroides vulgatus in FF piglets. Lactobacillus counts were higher in 9-d-old piglets than in 17-d-old piglets, regardless of diet. Bifidobacterium, Bacteroides, and clostridial species increased after 8 and 12 h fermentation on most substrates. In summary, piglet diet and age affect gut microbiota, leading to different fermentation patterns. HMO have potential prebiotic effects due to their effects on SCFA production and microbial modulation.

FIGURE 1

Changes in pH after 2, 4, 8, and 12 h in vitro fermentation of oligosaccharides with pooled ascending colonic contents from FF9 (A), FF17 (B), SR9 (C), and SR17 (D) donor groups. The effects of substrate, donor, and time of fermentation and their interactions (substrate × time, substrate × donor, donor × time, and substrate × time × donor) on fermentation were significant (P < 0.0001) for pH change. Values are least-squares means (pooled SEM = 0.012); n = 3. For each donor group, labeled means at a time without a common letter differ, P < 0.05. FF9, 9-d-old formula-fed piglets; FF17, 17-d-old formula-fed piglets; GOS, galactooligosaccharides; HMO, human milk oligosaccharides; LNnT, lacto-N-neotetraose; PDX, polydextrose; scFOS, short-chain fructooligosaccharides; SR9, 9-d-old sow-reared piglets; SR17, 17-d-old sow-reared piglets.

FIGURE 2

16S rRNA V3 gene DGGE profile (A) and PCA (B) of ascending colonic microbiota from 9- and 17-d-old formula-fed and sow-reared piglets before in vitro fermentation. The first principal component (PC1) explained 35.3% of the variation and shows the difference between FF17 and SR17 groups; the third principal component (PC3) explained 9.62% of the variation and shows the difference between FF9 and SR9. *Indicates piglets used for in vitro fermentation. DGGE, denaturing gradient gel electrophoresis; FF9, 9-d-old formula-fed piglets; FF17, 17-d-old formula-fed piglets; Mr, DGGE marker that was made by the mixture of randomly selected bands; PCA, principal components analysis; SR9, 9-d-old sow-reared piglets; SR17, 17-d-old sow-reared piglets.