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Besnoitiosis is an emerging parasitic disease of equids. Italy is one of the few European countries where the circulation of Besnoitia spp. antibodies was demonstrated. In this study, a case of clinical besnoitiosis in two donkeys in northern Italy is reported. The two animals were clinically examined. Serum and blood samples were analyzed for the detection of Besnoitia spp. antibodies and for hematology, biochemistry, and enzyme activity, respectively. ITS-1 PCR and sequencing were carried out on DNA extracted from skin biopsies. Clinical