Abstract: This study was carried out to understanding the seroprevalence of H9N2 avian influenza virus among hospital staff in Namazi hospital in Shiraz which is situated at the Southwest of Iran. A total of 300 serum samples from hospital staff and 300 control serum samples from people who came for physical check-up (control group) were collected. Hemagglutination-Inhibition (HI) tests were conducted to determine the individual serological status. The seropositive rate (SPR, defined as the proportion with HI titer = 1:40) of antibodies to H9N2 influenza virus and its Geometric Mean Titer (GMT) were calculated and compared among these groups.