BACKGROUND: The emergence of tick-borne disease is increasing because of the effects of the temperature rise driven by global warming. In Turkey, 19 pathogens transmitted by ticks to humans and animals have been reported. Based on this, this study aimed to investigate tick-borne pathogens including Hepatozoon spp., Theileria spp., Babesia spp., Anaplasma spp., Borrelia spp., and Bartonella spp. in tick samples (n = 110) collected from different hosts (dogs, cats, cattle, goats, sheep, and turtles) by molecular methods. METHODS: To meet this objective, ticks were identified morphologically at the genus level by microscopy; after DNA isolation, each tick sample was identified at the species level using the molecular method. Involved pathogens were then investigated by PCR method. RESULTS: Seven different tick species were identified including Rhipicephalus sanguineus, R. turanicus, R. bursa, Hyalomma marginatum, H. anatolicum, H. aegyptium, and Haemaphysalis erinacei. Among the analyzed ticks, Hepatozoon spp., Theileria spp., Babesia spp., and Anaplasma spp. were detected at rates of 6.36%, 16.3%, 1.81%, and 6.36%, respectively while Borrelia spp. and Bartonella spp. were not detected. Hepatozoon spp. was detected in R. sanguineus ticks while Theileria spp., Babesia spp., and Anaplasma spp. were detected in R. turanicus and H. marginatum. According to the results of sequence analyses applied for pathogen positive samples, Hepatozoon canis, Theileria ovis, Babesia caballi, and Anaplasma ovis were identified. CONCLUSION: Theileria ovis and Anaplasma ovis were detected for the first time to our knowledge in H. marginatum and R. turanicus collected from Turkey, respectively. Also, B. caballi was detected for the first time to our knowledge in ticks in Turkey.