In Egypt, limited studies were performed to investigate Jhone’s disease (JD) caused by Mycobacterium avium subspecies paratuberculosis (MAP). This study was concerned with diagnosis of MAP in cattle dairy farms using indirect-ELISA technique and amplification of IS900 using PCR technique for confirmation of infection and investigated adequate diagnostic markers for MAP through determining hemato-biochemical profile in cattle dairy farms. Screening of MAP was done by indirect ELISA between the periods from January 2017 to June 2018. A total of 200 cows: suffered from persistent diarrhea (n=17) and apparently healthy cows in contact with them (n=183) were used in this study. Positive JD was reported in 62 cases (31%) and negative JD was reported in 138 cases (69%). Out of the positive JD, 17 cases were clinically affected by persistent diarrhea and weight loss (clinical JD), where 45 cases were apparently healthy (sub-clinical JD). Typical colonies of MAP were isolated from the fecal samples of clinically affected cows with positive staining of MAP bacilli. Amplification of IS900 using PCR technique confirmed the presence of MAP infection. Phylogenetic analysis revealed identical similarities with MAP strains in India and Egypt. Hemato-biochemical investigations revealed significant decrease in RBCs, PCV, total protein, globulin and glucose with non-significant change in oxidative stress markers in clinically affected cows. This study is considered one of the limited researches that highlight the role of MAP infection in dairy farms in Egypt.