Ixodes ricinus is a potential vector for some of the tick-borne microorganisms that can cause significant diseases in animals and humans. This study aims to determine the prevalence of Anaplasma, Rickettsia, Bartonella, and Francisella species in host-seeking ticks collected from the forest areas in the Sinop region located in the northernmost part of Turkey. Between May and July 2017, a total of 135 tick pools formed from 2571 of the 2734 ticks collected out of the vegetation. Samples of each pool were homogenized and analyzed by PCR. Infection prevalence was statistically analyzed in view of the maximum likelihood estimation (MLE) with a 95% confidence interval (CI). DNA of the infectious agents was determined only in the adult and nymph pools of I. ricinus. MLE values of Anaplasma spp. and Bartonella spp. in 58 pools formed from 517 of I. ricinus adults were 1.20% (95% CI: 0.50-2.49) and 0.80% (95% CI: 0.26-1.91), respectively. In 42 pools generated from 1222 of I. ricinus nymph, MLE values of infection prevalence for Anaplasma spp. and Bartonella spp. were calculated to be 0.17% (95% CI: 0.03-0.54) and 0.34% (95% CI: 0.11-0.82) in respective order. MLE values for Rickettsia spp. were 7.55% (95% CI: 5.21-10.69) and 0.52% (95% CI: 0.22-1.083) for the adult and nymph I. ricinus, respectively. The DNA of Francisella tularensis was not detected in any tick pool. The outcomes of this research are the first molecular evidence of Bartonella spp. and Bartonella henselae in questing I. ricinus in Turkey. The results also suggested that I. ricinus plays considerable roles in enzootic transmission cycles of Anaplasma phagocytophilum, B. henselae, and Rickettsia monacensis in the northernmost region of Turkey.