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A procedure often performed following fistulotomy and advancement flap is curettage of the fistula tract after fistulotomy or after closing the internal opening. Epithelialization of the fistula tract might prevent closure of the fistula tract. The aim of this study was to assess the incidence and origin of epithelialization of the fistula tract in patients with perianal fistulae undergoing fistulotomy.Only patients with low perianal fistulae that were surgically treated by fistulotomy were included. Surgical biopsies were taken from the fistula tract from three different locations; on the proximal side at the internal opening, in the middle of the fistula tract and near the distal end close to the external opening.In the study period, 18 patients with low perianal fistulae were included. In 15 of the 18 patients, squamous epithelium was found at least in one of the biopsies taken from the fistula tract. Epithelium was predominantly found near the internal opening. There was no relation between the duration of fistula complaints and the amount of epithelialization (P = 0.301). The amount of epithelium was not related to the presence of a history of fistula surgery (P = 1.000).This study demonstrated epithelialization in the fistula tract in the majority of the patients surgically treated by fistulotomy for low perianal fistulae. Curettage of perianal fistulae must therefore be considered an essential step in the surgical treatment of perianal fistula.