Background. There is a paucity of evidence documenting the pathogenicity of Dientamoeba fragilis, an intestinal protozoan common in children. As case-reports on successful treatment are numerous, many authors advocate treatment, despite no placebo-controlled trials being available. Metronidazole is often used for treatment, though eradication rates are relatively low (60-80%). In the present study we determined the clinical and microbiological efficacy of metronidazole in Danish children. Methods. In this parallel placebo-controlled double-blinded trial, children aged 3-12 years with >4 weeks of gastrointestinal symptoms were allocated using block randomization in a 1:1 ratio to a 10-day course of oral metronidazole or placebo. Primary outcome was change in level of gastrointestinal symptoms, measured on a visual-analog-scale (VAS), and secondary outcome was eradication of D. fragilis infection. Participants, care givers, investigators and sponsor were all blinded to group assignment. The trial was registered with clinicaltrials.gov (NCT01314976) prior to start. Results. Of 96 participants, 48 were allocated to the metronidazole and placebo group each. Mean VAS change from pre- to post-treatment did not differ significantly (p=0.8) between the metronidazole (-1.8 CI (-2.5, -1.1) and the placebo group (-1.6 CI (-2.3, -0.9). Eradication of D. fragilis was significantly greater in the metronidazole group, although it declined rapidly from 62.5% two weeks after end of treatment to 24.9% eight weeks after end of treatment. Conclusions. These findings do not provide evidence to support routine metronidazole treatment of D. fragilis positive children with chronic gastrointestinal symptoms. Study funded by Statens Serum Institut.