Purpose Data on the seroprevalence of hepatitis E virus (HEV) in heamodialysis (HD) patients are conflicting, ranging from 0 to 44%. The aim of this study was to determine the HEV seroprevalence and risk factors among HD patients in Croatia. Methods A total of 394 HD patients from six medical facilities in five Croatian cities (three sites in the continental and three sites in the coastal region) were tested for HEV IgM/IgG antibodies using an enzyme-linked immunosorbent assay. Additionally, all samples were tested for HEV RNA by RT-PCR. Sociodemographic data and risk factors were collected using a questionnaire. Results HEV IgG antibodies were detected in 110 (27.9%) patients. The seroprevalence varied significantly between dialysis centres, ranging from 5.2 to 43.4% (p = 0.001). HEV IgM antibodies were found in 0.04% of IgG positive patients. All patients tested negative for HEV RNA. Factors associated with HEV IgG seropositivity were age > 60 years (OR 8.17 ; 95% CI 1.08–62.14), living in the continental parts of the country (OR 2.58 ; 95% CI 1.55– 4.30), and transfusion of blood products (OR 1.66 ; 95% CI 1.01–2.73). After adjusting for age and gender, patients from continental regions had higher odds of HEV seropositivity compared to patients from coastal regions (OR 2.88 ; 95% CI 1.71–4.85) and those who had RBC transfusions (OR 1.70, 95% CI 1.02–2.69) compared to those who did not. Conclusion The study showed a high HEV seropositivity among HD patients in Croatia, with significant variations between geographical regions. Continental area of residence and RBC transfusion were the most significant risk factors for HEV seropositivity. Due to the high seroprevalence, routine HEV screening among HD patients, especially in transplant candidates should be considered.