BACKGROUND: Cholangiocarcinoma (CCA), a fatal bile duct cancer, has a high incidence in Western Siberia, Russian Federation. In addition, Opisthorchis felineus (O. felineus), a bile duct dwelling trematode liver fluke is highly endemic. Closely related species have been shown cancerogenic agents in Asia. We therefore examined the association between O. felineus infection and CCA in Western Siberia. METHODS: We conducted a hospital-based, individually matched case-control study between January 2017 and August 2020 in Tomsk Oblast and Khanty-Mansiysk Autonomous Okrug, Yugra, Russian Federation. Histologically confirmed CCA patients (cases) were compared to age, sex and place of residence matched hospital controls. The examination of study participants included the diagnosis of current and past O. felineus infection, abdominal ultrasonographical assessment, physical examination, and interview on exposures to potential risk factors. RESULTS: We identified 40 patients with CCA and 160 controls. Exposures to O. felineus infection was strongly associated with CCA (OR 3.9, 95% CI 1.4-10.8, p = 0.008). Also, cases reported more often that they were currently or in the past infected by O. felineus compared to controls (OR 4.03, 95% CI 1.7 - 9.5, p = 0.001). Furthermore, cases reported river fish consumption and fishing habits significantly more often than controls (OR 5.5, 95% CI 1.5 - 19.8, p = 0.009; OR 3.3, 95% CI 1.4 - 7.7, p = 0.005). CONCLUSIONS: The study results revealed a strong significantly increased risk for CCA development in O. felineus-infected individuals. Elaboration of the guidelines on screening programs for early CCA diagnosis, prevention and treatment is socially important in endemic regions.