Background and Aim: Theileria annulata infection is a tick-borne disease affecting ruminants in the tropical and subtropical regions causing severe economic losses. This study aimed to characterize circulating T. annulata isolates from four governorates (administrative districts) north and south of Egypt using polymerase chain reaction (PCR)-restriction fragment length polymorphism.

Materials and Methods: Fifty samples were collected from the four governorates of Egypt and were examined by a PCR assay based on the heat shock protein 70 gene. The amplified product was subsequently digested using two restriction enzymes, Taq I and Alu I, to determine which pattern of T. annulata strains was involved.

Results: The findings revealed that one distinct pattern was observed for T. annulata isolates in the northern governorates and another one in the southern governorates. The Taq I enzyme produced three fragments (100, 175, and 270 bp), and the Alu I enzyme produced four fragments (60, 90, 125, and 270 bp).

Conclusion: This study determined the presence of two distinct circulating genotypes of T. annulata among cattle in Egypt based on PCR-RFLP using the HSP 70 gene. More studies are needed in different parts of the country to investigate the virulence and strain variance of T. annulata in cattle.

Keywords: cattle, Egypt, genotype, polymerase chain reaction-restriction fragment length polymorphism, Theileria annulata.