Paratuberculosis (Johne’s disease) is an OIE listed notifi able economically important contagious mycobacterial disease of ruminants caused by Mycobacterium avium subspecies paratuberculosis. The present study reports the seroprevalence of paratuberculosis in bovines of the rural area from fi ve different agro-climatic zone of Punjab. A total of 736 serum samples from both cattle and buffalo herds were evaluated for the presence of antibodies to MAP using a commercially available paratuberculosis screening enzyme-linked immunosorbent assay (ELISA) kit. Twenty three animals were positive and the seroprevalence was found as 3.125%. Highest prevalence was recorded in western plain zone (6.66%) followed by western zone (3.07%), central zone (2.68%), sub mountain undulating zone (2%) and nil in undulating plain zone