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Objective: To assess hepatitis E virus (HEV) seroprevalence in HIV patients and blood donors from one region in Poland. Methods: A group of 490 persons (244 HIV patients and 246 blood donors) aged 18–55 years were examined using the anti-HEV IgG assay (Wantai Biological Pharmacy Enterprise, Beijing, China). An analysis of the association between certain factors and the presence of this HEV exposure marker was conducted in both groups. Results: An HEV seropositivity rate of 50.2% was found. There was no difference in HEV seroprevalence between blood donors (49.6%, 122/246) and HIV patients (50.8%, 124/244) (p = 0.569). The anti-HEV IgG positivity rate increased with age as follows: 36.2% (59/163) in persons aged 18–30 years, 52.0% (92/177) in individuals aged 31–40 years and 63.3% (95/150) in those aged 41–55 years. HEV infection occurred in 56.4% (31/55) of people who had never travelled abroad. Conclusions: Wielkopolska Region in west-central Poland is an area hyperendemic for HEV infection. In this part of Poland, the exposure of HIV-positive persons to this virus is not greater than that of healthy blood donors.