Background: The etiology of Crohn's disease has so far remained unknown, but the current research has concentrated upon autoimmunity and/or mycobacterial infection. With the appearance of the highly sensitive polymerase chain reaction (PCR), the possibility of detecting nucleic acids from even a few present microorganisms has emerged. Methods: We have used a sensitive variant of the polymerase chain reaction (nested primer PCR) for detection of a multi-copy insertional element (IS900) specific for Mycobacterium paratuberculosis. This technique was applied to DNA extracted from fresh as well as paraffin-embedded intestinal tissue obtained from patients undergoing resection of inflamed intestinal tissue due to Crohn's disease. Results: In fresh intestinal tissue from 11 of 24 patients with Crohn's disease, 2 of 10 patients with ulcerative colitis and 3 of 28 patients with other colonic disorders specific M. paratuberculosis DNA was found. In paraffin embedded tissue from 4 of 58 patients with Crohn's disease and from none of 72 control patients (55 patients with ulcerative colitis and 17 patients with colonic cancer), specific M. paratuberculosis DNA was found. Conclusions: Whether these microorganisms were connected to the inflammatory bowel disease, or their presence were mere coincidence, cannot be stated. We find this presence interesting and encouraging for further investigations.