This study was conducted to investigate the seroprevalence of non-structure protein for foot and mouth disease virus (FMDV) and identify FMDV serotypes. A total of 3600 serum samples (1080 buffaloes and 2520 cattle) were collected randomly from different breeds, age, and sex at El Beheira, El Dakahlia, and El Giza. Nonstructure protein of FMDV was detected in 41 of 1080 (3.8%) Buffalo and 185 of 2520 (7.3%) cattle. The case fatality was 14.6 and 17.8% in the vaccinated buffalo and cattle, respectively. The prevalence of nonstructural protein FMDV serotyping O was 8.3, 7.5 and 2.9% in El Beheira, El Dakahlia, and El Giza, respectively. Moreover, the case fatality was 8% (El Beheira), 26.3% (El Dakahlia), and 20% (El Giza). The circulation of FMDV was prevalent during the winter season of the year. The frequency of positive case was significantly different between species, sex, and age, while it was non-significant with different breeds. The mean values of antibodies of a non-structural protein of FMDV were the highest in male cattle at 5 months to 1 year of age. Additionally, seven epithelial tissues were collected from tongue; buccal mucosa and teat of recently sudden dead animals. The obtained sequences by reverse transcription polymerase chain reaction (RT-PCR) were registered in GenBank under accession numbers MF980930.1, MF991123.1, MF991124.1, and MF991125.1. Phylogenetic analysis revealed that the obtained sequences belonged to deposited FMDV type O (KP121442.1) with similarity ratios of 98, 100, 99, and 99%, respectively. Also, the deduced amino acids of the obtained sequences are related to capsid protein of VP1 gene of FMDV.