Recently, a T-to-C polymorphism at position -295 in the promoter region of the human interleukin-16 (IL-16) gene was reported. The expression of IL-16 is increased in inflammatory bowel disease, in particular in Crohn's disease. However, data concerning the IL-16 promoter polymorphism in inflammatory bowel disease are lacking. Thus, the current study aimed at the assessment of this polymorphism in Crohn's disease and ulcerative colitis. One hundred three patients with Crohn's disease, 100 patients with ulcerative colitis, and 120 healthy unrelated controls were genotyped for the promoter polymorphism. Furthermore, patients with Crohn's disease were stratified according to disease localization and the respective clinical phenotype (fistulizing, fibrostenotic, or inflammatory). The frequencies of the T allele (P < 0.01) and the TT genotype (P < 0.01) were significantly increased in patients with Crohn's disease compared to the controls, regardless of the disease phenotype or the site of intestinal involvement. An association with ulcerative colitis was not observed. Herein a new association between a promoter polymorphism of the IL-16 gene and Crohn's disease was observed and correlates with the previously described increased mucosal expression of IL-16 in inflammatory bowel disease.