We investigated the effect of a 12-d exposure to 34°C plus dietary inclusion of the probiotic Bacillus licheniformis on the egg production, gut morphology, and intestinal mucosal immunity of laying hens. Ninety-six commercial hens (Hy-Line Brown) at the age of 60 wk were randomly allocated to 4 groups. After a period of laying rate adjustment (14 d), all the hens were subjected to 2 temperature treatments (12 d). Birds in 1 group were raised at 21°C and fed a basal diet, and birds in the other 3 groups were raised at 34°C and fed a basal diet supplemented with 0, 10(6), or 10(7) cfu of B. licheniformis per gram of feed, respectively. Rearing at 34°C depressed egg production and feed intake (P < 0.05). Compared with birds kept at 21°C, birds kept at 34°C had elevated serum levels of tumor necrosis factor-α (d 6), IL-1 (d 6 and 12), and corticosterone (d 6); decreased villus height (ileum: d 6; cecum: d 6 and 12) and ratio of villus height to crypt depth (ileum: d 6; cecum: d 6 and 12); fewer intraepithelial lymphocytes (ileum: d 6; cecum: d 6) and IgA-secreting cells (ileum: d 6; cecum: d 6 and 12); and more mast cells (ileum: d 6; cecum: d 6 and 12; P < 0.05). The number of goblet cells in the cecum increased at d 6 in heat-treated birds, and then deceased at d 12 (P < 0.05). Moreover, morphological examination showed injury to the villi of birds kept at 34°C. In general, inclusion of 10(7) cfu/g of B. licheniformis in the diet of heat-stressed hens was effective in overcoming the observed decline in egg production and feed intake, restoring the impaired villus structure, and sustaining a balanced mucosal immune response. Therefore, the probiotic B. licheniformis may be useful for ameliorating the adverse influence of heat on the egg production and gut health of laying hens.