Johne`s disease is economically and zoonotically an important disease in dairy animals and required continuous surveillance. The study was carried out to investigate the prevalence of Paratuberculosis (Johne's disease) in one of dairy cattle farm in Egypt. A total of 964 serum samples were collected from 964 dairy cattle and were examined by ELISA. Out of them 155 (16.1%) samples reacted positively. The results revealed that the age and lactation period have significant effect of prevalence of Johne's disease in dairy cattle. The disease was mostly prevalent in animals over 6 years of age and in early stage of lactation. Furthermore, the disease is more prevalent in heavy lactating animals in comparison with other examined cattle. In contrast, the pregnancy stage showed no significant effect on occurrence of the disease in dairy cattle. The result of the current study demonstrate that Mycobacterium avium subsp. paratuberculsosis (MAP) is circulating in dairy farm in Egypt and further studies require to reveal the epidemiology of the disease