The effects of different capsaicin doses on gastrointestinal inflammation and the gut microbiota structure in Escherichia coli O157:H7-infected mice were investigated. A mouse model of E. coli O157:H7 infection was established, and successfully infected mice were randomly divided into control, infected, and capsaicin-treated groups, receiving 15, 7.5, and 3.75 mg/kg capsaicin treatment, respectively, for seven days. The food intake and body weight were recorded. Levels of inflammatory factors, including tumor necrosis factor-α and interleukin-6 in the serum, and the inflammatory marker myeloperoxidase in the intestinal tissue, were determined. Histopathological changes in the colon and gastric tissues were observed. Additionally, changes in the mouse gut microbiota were identified through 16S rDNA sequencing. The results revealed that administration of 15 mg/kg capsaicin substantially reduced the levels of inflammatory factors in the serum and intestinal tissue of E. coli O157:H7-infected mice, improved gastrointestinal barrier damage and inflammation, and increased the abundance of Akkermansia, Anaeroplasma, and Cetobacterium. The relative abundance of Akkermansia increased by 48% compared to that in the control group. In contrast, capsaicin intake substantially reduced the abundance of Alistipes, Enterococcus, and Yokenella. In conclusion, dietary intake of 15 mg/kg capsaicin can improve gastrointestinal inflammation and the gut microbiota structure in mice infected with E. coli O157:H7. These findings provide insights for treatment interventions and dietary strategies for mice with E. coli O157:H7 infection.