Many domestic and wild animals are susceptible to malignant catarrhal fever (MCF), which is a lymphoproliferative, multisystemic, and fatal disease. The present study was conducted on 47 cattle. The clinical examination findings revealed a persistent high fever, enlargement of superficial lymph nodes, corneal opacity, respiratory symptoms, oral lesions, and ulcerative skin lesions. Buffy coat samples were collected for laboratory analysis. Semi-nested polymerase chain reaction (PCR) assay has been used. Ovine herpesvirus-2 (OvHV-2) Deoxyribonucleic acid (DNA) was detected in the buffy coat of 21 cattle. DNA sequencing and phylogenetic analysis of OvHV-2 were performed in our study. The phylogenetic analysis of the PCR product of the Egyptian strain of OvHV-2 (Assiut) showed close similarity with OvHV-2 strains of different governorates (Beni-suef, Giza and Fayoum) of Egypt, India, Turkey, Iraq, Italy, Brazil and United Kingdom. There was no significant variation (P<0.05) between the percentages of MCF infection and the sex, age, and breed (native and mixed breed) of molecularly tested cattle. According to the climatologic conditions of Assiut governorate, there is no discernible difference (P<0.05) between the MCF infection rate and the cold and hot months. To date, this is the first report of OvHV-2 in Assiut Governorate, so it is advisable to separate susceptible cattle from sheep, especially during lambing, and to euthanize any animals that are clinically infected with MCF.