A mismatch between early developmental diet and adulthood may increase obesity risk. Our objective was to determine the effects of re-matching rats to their weaning diets high in protein or fiber after transient high-fat/high-sucrose challenge in adulthood. We hypothesize that a long-term high fiber diet will be associated with a gut microbiota and hepatic gene expression reflective of reduced adiposity. Wistar rat pups were fed a control (C), high prebiotic fiber (HF), or high protein (HP) diet from 3-15 weeks of age; a high-fat/high-sucrose diet from 15-21 weeks; their respective C, HF, or HP diets from 21-25 weeks. Gut microbiota of cecal contents and hepatic gene expression were measured when rats were terminated at 25 weeks of age. HF rats had higher total bacteria, bifidobacteria and Bacteroides/Prevotella spp than C and HP at 25 weeks (P < 0.05). Firmicutes, especially Clostridium leptum, decreased in HF compared to C and HP (P < .05). The ratio of Firmicutes:Bacteroidetes was markedly lower in HF versus C and HP at 25 weeks (P < .05). HF decreased hepatic cholesterol content compared to HP and C at 25 weeks. HF and HP increased 3-hydroxy-3-methylglutaryl-CoA reductase mRNA and decreased lecithin-cholesterol acyltransferase mRNA compared to C (P < .05). In conclusion, re-matching rats to a HF but not HP diet attenuated the typical increase in Firmicutes:Bacteroidetes ratio associated with consumption of a high fat diet. Lower hepatic cholesterol with long-term HF diet intake may be related to alterations in gut microbiota and hepatic lipid metabolism.