The Cypriot mouflon (Ovis orientalis ophion), a once almost extirpated species of wild sheep, is under strict surveillance because it can be threatened by likely transmission of pathogenic bacteria, such as Anaplasma spp., Rickettsia spp., and Coxiella burnetii, primarily from domestic ungulates. We collected 77 blood samples from Cypriot mouflons and 663 of their ectoparasites (Rhipicephalus turanicus, Rhipicephalus sanguineus, Rhipicephalus bursa, Hyalomma anatolicum excavatum, Hyalomma marginatum, Haemaphysalis punctata, Haemaphysalis sulcata, and Ixodes gibossus) and tested them by polymerase chain reaction and sequencing. Twenty-three mouflon blood samples (30%) were positive for C. burnetii, 23 (30%) for Rickettsia spp., and 8 (10%) for Anaplasma ovis. Of 109 pools of ectoparasites, 32.1% were positive for C. burnetii, 28.4% for Rickettsia spp., and 10.9% for A. ovis; 11.9% were positive for both C. burnetii and Rickettsia spp., 6.4% for both Rickettsia spp. and A. ovis, and 2.8% for all three pathogens. This is the first survey that records the presence of tick-borne pathogens, both in the Cypriot mouflon and in ticks parasitizing it.