The objective of this study was to investigate the effects of Lactobacillus reuteri LR1, a new strain isolated from the feces of weaned pigs, on the growth performance, intestinal morphology, immune responses, and intestinal barrier function in weaned pigs. A total of 144 weaned pigs (Duroc × Landrace × Yorkshire, 21 d of age) with an initial BW of 6.49 ± 0.02 kg were randomly assigned to 3 dietary treatments with 8 replicate pens, each of per treatment and 6 pigs. Pigs were fed a basal diet (CON, controls), the basal diet supplemented with 100 mg/kg olaquindox and 75 mg/kg aureomycin (OA) or the basal diet supplemented with 5 × 1010 cfu/kg L. reuteri LR1 for a 14-d period. At the end of study, the ADG, ADFI, and G:F were calculated, and 1 randomly selected pig from each pen was euthanized for sample collection. The LR1 increased ADG (22.73%, P < 0.05) compared with CON. The villus height of the ileum was increased (P < 0.05) and crypt depth in duodenum was reduced (P < 0.05), along with increased (P < 0.05) villus height to crypt depth ratio of the jejunum and ileum by LR1 compared with CON and OA. LR1 increased (P < 0.05) ileal mucosal content of IL-22 and transforming growth factor-β compared with OA. Compared with CON, LR1 increased (P < 0.05) and OA decreased (P < 0.05) the ileal content of secretory immunoglobulin A (sIgA), and the abundance of transcripts of porcine β-defensin 2 and protegrin 1-5. Compared with CON, LR1 increased (P < 0.05) tight junction protein zonula occludens-1 and occludin transcripts in the mucosa of the jejunum and ileum, and those of mucin-2 in ileal mucosa. The relative expression of toll-like receptor 2 (TLR2) and TLR4 were increased (P < 0.05) in ileal mucosa in pigs fed LR1 compared with CON. In conclusion, these data indicated that dietary LR1 supplementation at 5 × 1010 cfu/kg improved growth performance, intestinal morphology, and intestinal barrier function in weaned pigs.