Aims This study evaluated the effects of Bacillus amyloliquefaciens TL106, isolated from Tibetan pigs’ faeces, on the growth performance, immune response, intestinal barrier function, morphology of jejunum, caecum and colon, and gut microbiota in the mice with enterohaemorrhagic Escherichia coli (EHEC)‐induced intestinal diseases. Methods and Results In all, 40 female C57BL/6J mice were randomly divided into four groups: mice fed a normal diet (Control), mice oral administration of TL106 daily (Ba), mice challenged with EHEC O157:H7 on day 15 (O157) and mice oral administration of TL106 daily and challenged with EHEC O157:H7 on day 15 (Ba+O157). The TL106 was administrated to mice for 14 days, and mice were infected with O157:H7 at day 15. We found that TL106 could prevent the weight loss caused by O157:H7 infection and alleviated the associated increase in pro‐inflammatory factors (TNF‐α, IL‐1β, IL‐6 and IL‐8) and decrease in anti‐inflammatory factor (IL‐10) in serum and intestinal tissues of mice caused by O157:H7 infection (P < 0·05). Additionally, TL106 could prevent disruption of gut morphology caused by O157:H7 infection, and alleviate the associated decrease in expression of tight junction proteins (ZO‐1, occludin and claudin‐1) in jejunum and colon (P < 0·05). In caecum and colon, the alpha diversity for bacterial community analysis of Chao and ACE index in Ba+O157 group were higher than O157 group. The TL106 stabilized gut microbiota disturbed by O157:H7, including increasing Lachnospiraceae, Prevotellaceae, Muribaculaceae and Akkermansiaceae, and reducing Lactobacillaceae. Conclusions We indicated the B. amyloliquefaciens TL106 can effectively protect mice against EHEC O157:H7 infection by relieving inflammation, improving intestinal barrier function, mitigating permeability disruption and stabilizing the gut microbiota. Significance and Impact of the Study Bacillus amyloliquefaciens TL106 can prevent and treat intestinal disease induced by EHEC O157:H7 in mice, which may be a promising probiotic for disease prevention in animals.