Lumpy skin disease (LSD) is a devastating viral disease in cattle that has recently spread from Africa to Middle Eastern countries. This study aimed to investigate the relationship between Lumpy Skin Disease Virus (LSDV) isolates from different regions of Iran and the origin and spread of these viruses. In this study, 184 out of 228 suspected outbreaks of Lumpy Skin Disease were confirmed based on clinical signs in 22 provinces. Samples were collected from 9 provinces, totaling 34 samples, and were tested using Real-Time PCR at the Veterinary Diagnostic Laboratory of Khorasan Razavi Province. Out of 34 samples, 26 samples from 7 provinces were positive in the Real-Time PCR assay. Fifteen Lumpy Skin Disease virus samples were subjected to Sanger-dideoxy direct sequencing. NCBI-Blast search of the examined sequences revealed that the most homologous sequences that most of these sequences aligned with were Lumpy Skin Disease Virus sequences (GenBank MN642592.1). The phylogenetic tree constructed from the obtained sequences showed the highest similarity to African isolates, such as Kenya (GenBank MN072619.1), and Asian isolates, such as India (GenBank MIN995838.1). It was concluded that the outbreak of Lumpy Skin Disease in Iran was likely caused by the introduction of Lumpy Skin Disease viruses from neighboring countries. The obtained P32 nucleotide sequence data is a valuable resource for understanding the genetic nature and molecular epidemiology of local Lumpy Skin Disease virus isolates.

Keywords