Dietary trace minerals can impact gut flora, which can further affect intestinal health. However, dietary balance pattern of trace minerals for intestinal health of broilers needs to be explored. The present study was conducted to investigate the effect of dietary pattern of Cu, Fe, Mn, Zn and Se on the intestinal morphology, microbiota, short chain fatty acid concentrations, antioxidant status and the expression of tight junction proteins in broilers. A total of 240 1-d-old Arbor Acres male broilers were randomly assigned to 1 of 5 treatments with 6 replicate cages of 8 birds per cage for each treatment. The birds were fed the corn-soybean meal basal diet supplemented with 5 combination patterns of trace minerals for 42 d. The dietary treatments were as follows: the inorganic sources were added to the diet based on the recommendations of the current National Research Council (NRC, T1) and Ministry of Agriculture of P. R. China (MAP) (T2) for broiler chicks, respectively; the inorganic sources were added to the diet at the levels based on our previous results of inorganic trace mineral requirements for broilers (T3); the organic sources were added to the diet at the levels considering the bioavailabilities of organic trace minerals for broilers described in our previous studies (T4); and the organic sources were added to the diet based on the recommendations of the current MAP for broiler chicks (T5). The results showed that broilers from T1 had lower (P < 0.05) crypt depth (CD) and higher (P < 0.05) villus height : CD in duodenum on d 21 and lower CD (P < 0.05) in jejunum on d 42 than those from T3 and T4. Broilers from T1, T3 and T5 had higher (P < 0.05) Shannon index in cecum on d 21 than those from T4. Broilers from T1 had higher (P < 0.05) abundance of Lactobacillus in ileum on d 21 than those from T2 and T3. Broilers from T1, T2 and T5 had higher (P < 0.05) valeric acid concentrations in cecum on d 42 than those from T3 and T4. In addition, Birds from T2 had higher (P < 0.05) Claudin-1 mRNA level in jejunum on d 42 than those from T3 and T4. And birds from T3, T4 and T5 had higher (P < 0.05) Occludin protein expression levels in duodenum on d 42 than those from T2. These results indicate that dietary pattern of Cu, Fe, Mn, Zn and Se influenced gut flora and intestinal health of broilers, and the appropriate pattern of Cu, Fe, Mn, Zn and Se in the diet for intestinal health of broilers would be Cu 12 mg, Fe 229 mg, Mn 81 mg, Zn 78 mg and Se 0.24 mg/kg (1 to 21 d of age), and Cu 11 mg, Fe 193 mg, Mn 80 mg, Zn 73 mg and Se 0.22 mg/kg (22 to 42 d of age), when the trace minerals as inorganic sources were added to diets according to the recommendations of the current NRC.