Description: Paratuberculosis is caused by Mycobacterium avium subsp.paratuberculosis (Map) and affects many animal species, with ruminants as usual hosts. Due to the lack of successful treatment, infection leads to chronic wasting and death of the affected animals, causing serious economic losses in addition to the spread of public fear connected to the possible role of Map in Crohn`s disease. It is also a common disease of ruminants in Slovenia. Because of the lack of prevalence data since 2001, the purpose of this work was to estimate the seroprevalence of paratuberculosis in cattle herds in Slovenia. Animals older than two years in 20 percentage of herds, originating from all different areas of Slovenia were tested in 2008 for the presence of antibodies against Map. A total of 38,374 sera from 6,779 cattle herds were initially examined by the in-house ELISA screening method, and positive or suspect sera were subjected to second screening and final confirmation by applying the Pourquier ELISA Paratuberculosis kit (Institut Pourquier, France). A positive result was obtained in 228 (0.59 percentage) animals from 188 (2.77 percentage) herds, resulting in true-prevalence (TP) estimates of 3.96 percentage at animal and 18.49 percentage at herd level. Currently, TP of paratuberculosis in cattle in Slovenia is lower than at the time of monitoring in 1999, when a comparable number of animals were tested but has remained similar at the herd level. Compared to many European countries, both the animal and the herd prevalences in Slovenia are fairly low, which can be partly attributed to the existence of numerous ` family ` farms, with a small number of animals per herd, since Map more easily spreads through an infected herd than among different herds. If the breeding strategy changes and animal trade with other countries increases, the present favorable situation in Slovenia will probably also change