A nine-week feeding trial was conducted to investigate changes in the intestinal microbiota of turbot in response to alternate feeding between terrestrially sourced oil (TSO)- and fish oil (FO)-based diets. The following three feeding strategies were designed: (1) continuous feeding with the FO-based diet (FO group); (2) weekly alternate feeding between soybean oil (SO)- and FO-based diets (SO/FO group); and (3) weekly alternate feeding between beef tallow (BT)- and FO-based diets (BT/FO group). An intestinal bacterial community analysis showed that alternate feeding reshaped the intestinal microbial composition. Higher species richness and diversity of the intestinal microbiota were observed in the alternate-feeding groups. A PCoA analysis showed that the samples clustered separately according to the feeding strategy, and among the three groups, the SO/FO group clustered relatively closer to the BT/FO group. The alternate feeding significantly decreased the abundance of Mycoplasma and selectively enriched specific microorganisms, including short-chain fatty acid (SCFA)-producing bacteria, digestive bacteria (Corynebacterium and Sphingomonas), and several potential pathogens (Desulfovibrio and Mycobacterium). Alternate feeding may maintain the intestinal microbiota balance by improving the connectivity of the ecological network and increasing the competitive interactions within the ecological network. The alternate feeding significantly upregulated the KEGG pathways of fatty acid and lipid metabolism, glycan biosynthesis, and amino acid metabolism in the intestinal microbiota. Meanwhile, the upregulation of the KEGG pathway of lipopolysaccharide biosynthesis indicates a potential risk for intestinal health. In conclusion, short-term alternate feeding between dietary lipid sources reshapes the intestinal microecology of the juvenile turbot, possibly resulting in both positive and negative effects.