ObjectivesRed wheat, the class of wheat used to make yeast bread products, is associated with reductions in colon cancer biomarkers, regardless of refinement state. We hypothesized that red wheat as well as the phenol-rich aleurone and testa layers of red wheat would reduce colonic precancerous lesions and oxidative stress, and beneficially modulate the gut microbiome in rats with diet-induced obesity. MethodsRats were divided into seven groups (12/group) and fed a normal fat diet (NFD), high-fat diet (50% of total kcal as fat, HFD), whole red wheat + HFD (whR + HFD), refined red wheat + HFD (refR + HFD), refined white wheat + HFD (refW + HFD), aleurone layer + HFD (AL + HFD), or testa layer + HFD (TL + HFD). After a 14-day adaptation period, rats received two i.p. injections of the colon-specific carcinogen 1,2-dimethylhydrazine (DMH), administered one week apart. Sixty-three days after the second injection, colons were harvested and precancerous lesions (aberrant crypt foci, ACF) were enumerated. Staining intensity of 3-nitrotyrosine (3-NT) was determined immunohistochemically in distal colon tissue. Microbial DNA from cecal contents was sequenced using a 16S rRNA metagenomic approach. Differences in alpha and beta diversity, and microbial abundances were determined. ResultsCompared to the NFD, the HFD had a greater number of ACF, regardless of size (i.e., AC/ACF). The refR + HFD had significant reductions in medium ACF (3–5 AC/ACF; 2.62 vs. 4.28), large ACF (≥6 AC/ACF; 0.06 vs. 0.45), ACF multiplicity (1.58 vs. 2.01) and 3-NT (% positivity per ACF; 2.06% vs. 4.51%) compared to the HFD. All diets containing wheat reduced large ACF number. The TL + HFD and AL + HFD demonstrated trends for reducing ACF with 8 AC (0.06 vs. 0.18) and 3-NT (2.22% vs. 4.51%), respectively, compared to the HFD. Beta diversity significantly differed between diet groups (R2 = 0.27, P = 0.001), and there was greater abundance of Faecalitalea, Fusicatenibacter, and Lactobacillus in the cecal contents of rats fed wheat-containing diets. ConclusionsRed wheat reduces precancerous lesions, oxidative stress, and beneficially modulates the gut microbiome relative to a non-wheat diet. The phenol-rich testa and aleurone layers alone had little influence on these outcomes. Funding SourcesNDSU Collaborative Seed Grant Program.