A retrospective serological survey was carried out by the indirect immunofluorescence assay (IFA) and complement fixation (CF) test in a sample of 425 healthy residents (240 females and 185 males) to investigate whether Lymphocytic choriomeningitis virus (LCMV) circulates in the rural area of the northern Croatian island of Vir, which is known to be endemic for murine typhus. The overall prevalence of LCMV antibodies detected by IFA was found to be 36% (155 out of 425) and 13% (54 out of 425) by CF. No significant difference in the LCMV seroprevalence was observed with respect to age and gender by any of the methods used. The results show that inhabitants in the area studied are clearly being exposed to LCMV and the infection of the population occurs in the first 10 years of life. Equal prevalence of IgG and especially of short-lived CF antibodies in the elderly in comparison to other age groups and children can be explained by reinfections and boosting of antibodies through permanent contact with the virus. To the best of our knowledge this study reports the first evidence of endemic LCMV in Croatia and one of the highest LCMV human prevalence reported worldwide to date. The epidemiological association between LCMV and other zoonoses which can be expected in this community as well as the etiology of summer influenza-like illness along the coastal area of Croatia has to be explored further.