Fructo-oligosaccharide (FOS) and pectin are known soluble dietary fibers and can influence gut microbiota and consequently modulate gut health. To understand the differential impact patterns of pectin vs. FOS in modulating gut microbiota in the small and large intestine, an ileal-cannulated pig model was adopted to compare the temporal and spatial effects of FOS and citrus pectin (CP) on the gut microbiota. Sixteen terminal ileal-cannulated pigs were randomly divided into 2 groups and fed with a standard diet supplemented with either 3% FOS or 3% CP for 28 d. The CP group and FOS group showed different microbial composition, especially in the feces, with time and location as major factors affecting microbiota in the CP group, and with only location contribution in the FOS group. In the feces, relative to the FOS group, the CP group showed higher abundance of and and lower abundance of and (adjusted  < 0.05), a higher level of short-chain fatty acids and a lower level of lactate at both d 14 and 25 ( < 0.05), and more copy numbers of genes encoding key enzymes related to propionate () and butyrate () production and lactate utilization () ( < 0.05), indicating a greater degree of microbial carbohydrate fermentation. In the ileum, as compared with FOS, CP increased the bacteria with high capability of fermenting amino acids, including and (adjusted  < 0.05), and the expression of enzymes responsible for amino acid fermentation (i.e. lysine decarboxylase), as well as the amino acid fermentation products (cadaverine and tyramine) ( < 0.05), indicating a greater degree of amino acid fermentation. Overall, our results highlight a differential dynamic impact of dietary CP vs. FOS on microbial composition and metabolism in the gut. The dietary CP has a stronger ability to promote microbial amino acid fermentation in the ileum and carbohydrate fermentation in the feces than FOS. These findings provide a new insight into the role of different fibers in gut nutrition and guidelines for the choice of fibers in manipulating gut health.