Wildlife can act as reservoir of different tick-borne pathogens of veterinary and zoonotic importance. To investigate the role of wild ruminants as reservoir of piroplasm infection, 28 red deer, 69 roe deer and 38 chamois from Northern Spain were examined by reverse line blot (RLB) hybridization. The survey detected a prevalence of 85·7% in red deer, 62·3% in roe deer and 28·9% in chamois. Four different piroplasms were identified: Theileria sp. OT3 (previously described in sheep) as the most prevalent (85·7% in red deer, 46·4% in roe deer and 26·3% in chamois); Theileria sp. 3185/02 (previously described in a red deer in Central Spain) more abundant in red deer (53·6%) than in roe deer (10·1%) but absent from chamois; Babesia divergens detected in 6 roe deer; Theileria ovis present in 1 chamois. Mixed infections (Theileria sp. OT3 and Theileria sp. 3185/02) were only found in red and roe deer. Sequencing analysis of the 18S rRNA gene confirmed the RLB results and showed 99·7% identity between Theileria sp. 3185/02 and T. capreoli, suggesting that they are the same species. Tick distribution and contact of wild ruminants with domestic animals are discussed in terms of piroplasm infection. The results suggest that a considerable number of wildlife ruminants are asymptomatic carriers that may serve as reservoirs of the infection posing a serious concern in terms of piroplasmosis control.