The present study concerned with phylogentic analysis of three genes (gapA, p 40 pseudogene and uvrC) related to adhesion and virulence of Mycoplasma bovis (M. bovis) in cows and buffaloes. In this study, 625 milk samples were collected from clinical and subclinical mastitis in cows and buffaloes in two Egyptian Governorates Fayoum (south of Cairo Governorate) and Dakahlia (south of Cairo Governorate) for the detection of M. bovis infection by isolation and PCR. Mycoplasma infection was higher in cows (31.6%) than buffaloes (14.3%) suffered from clinical mastitis at Fayoum. The incidence of clinical mastitis in cows (20%) is higher than that in buffaloes (10%) at Dakahlia. Concerning subclinical cases, the incidence was higher at Dakahlia (30.19% and 38.3%) than Fayoum (20.6% and 12%) in cows and buffaloes respectively. Phylogentic analysis of nucleotides and amino acids of gapA gene showed 100% identity of the two isolates. The study proved that p40 gene is present in bovine as pseudogene and its protein did not expressed. The amino acids of uvrC gene of our field isolates showed 50 a.a substitutions. In conclusion M. bovis isolate isolated in the current study had identical gapA gene in both cow and buffalo in DNA and amino acid sequences (aa), whereas uvrC gene showed 50 aa substitutions in which could affect the antigenicity and p40 gene showed no protein expression and present in bovine as pseudogene.