Histomonas meleagridis is a flagellated protozoan parasite and the aetiological agent of histomonosis (histomoniasis or blackhead disease), a severe disease in poultry with very limited options for treatment. In the present investigation an inactivated vaccine, based on destroyed parasites and administered by intramuscular injection, failed to induce an efficient protection against a severe challenge. By cloacal infection of 14-day-old turkeys with cloned protozoa passaged in vitro for 95, 215 or 295 times, respectively, severe attenuation could be demonstrated as none of the infected birds died. Following infection with one of the higher passages, the birds resisted the challenge with virulent parasites. Using this approach no mortality due to histomonosis could be recorded in all of the 42 birds subjected to vaccination, either through direct infection or kept as in-contact birds, whereas all of the control animals died. Pathological lesions in the liver and caeca were only noticed in a few birds. The absence of parasitic DNA in the liver of vaccinated birds confirmed the effect of vaccination. For the first time vaccination of turkeys is reported to induce a solid protection against a severe challenge, both based on a clonal culture of Histomonas meleagridis.