Hemodialysis (HD) patients are vulnerable to transfusion-transmitted infections such as hepatitis B virus (HBV). HBV infection with undetectable hepatitis B surface antigens (HBsAg) is described as occult HBV and can lead to serious complications. The aim of this study was to evaluate the prevalence of occult HBV and concomitant factors in HD patients. Using a cross-sectional design, clinical and epidemiological data were obtained from May to September 2009 in 11 different HD units in Guilan province in northern Iran. After serological testing for HBV surface antigens in 514 HD patients using a third-generation enzyme-linked immunosorbent assay kit (Diapro, Milano, Italy), HBsAg-negative patients were tested for HBV DNA using a Qiagen PCR kit (Artus Qiagen GmbH, Hilden, Germany). After omission of seven HBsAg-positive patients, 507 patients were included in the study, 280 (55.2%) of whom were male and 227 (44.8%) were female. Patients ranged in age from 16 to 66 years (mean 53.2 years). No HBV DNA was detected in HBsAg-negative patients. Some 59 patients (11.6%) were anti-hepatitis C virus positive and 32 (6.3%) were hepatitis C virus positive according to polymerase chain reaction. The study results indicate that occult HBV infection is not a significant health problem in HD patients in Guilan province.