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Background: Theileriosis is one of the most prevalent infectious diseases of livestock in the Arabian Peninsula,\ud and causes high rates of mortality and morbidity in sheep and cattle. However, there is a paucity of information\ud on the distribution of Theileria spp. over the whole region and their impact on different hosts. The present study\ud carried out a country-wide molecular survey for Theileria spp. of livestock in Oman across four governorates. The\ud aim of the survey was to define the prevalence of Theileria spp. in cattle, sheep and goats, highlight risk factors\ud for infection and identify the main tick species involved in parasite transmission.\ud Material and methods: A total of 2020 animals were examined in the survey consisting of sheep [n = 592], goats\ud [n = 981] and cattle [n = 447]. All three species were raised and co-grazed on the same farms. Theileria\ud parasites were detected using PCR-RFLP and RLB of the 18S rRNA gene. Cloning and sequencing of the 18S rRNA\ud was carried out on 11 T. lestoquardi isolates from Ash-Sharqiyah, and Ad-Dhahira governorates, and phylogenetic\ud relationships were inferred using additional sequences of T. lestoquardi, T. annulata and T. ovis available in\ud GenBank.\ud Results: Theileria spp. prevalence was 72.3%, 36.7% and 2.7% among cattle, sheep and goats, respectively.\ud Strong similarity in results was obtained using RLB and PCR-RFLP for detection of Theileria spp. however, RLB\ud detected a higher rate of mixed infection than PCR-RFPL (P < 0.001). Theileria annulata was the only parasite\ud detected in cattle, while sheep and goats carried T. ovis, T. lestoquardi and T. annulata as well as Theileria spp.\ud OT1. Of the four Theileria spp. detected in small ruminants, overall T. ovis was most prevalent (sheep [33.4%],\ud goats [2.0%]), whereas T. lestoquardi was less prevalent (sheep [22.0%], goats [0.5%]). A large proportion of\ud infected sheep (19%) carried mixed infection of T. ovis and T. lestoquardi. However, single T. lestoquardi\ud infections (3.0%) were less prevalent than T. ovis infections (14.5%). Risk of Theileria spp. infection was\ud significantly higher for exotic breeds, relative to native breeds, of cattle (p = 0.00002) and sheep (p = 0.005).\ud Phylogenetic analysis placed T. lestoquardi in Oman in the same clade as other T. lestoquardi strains isolated from\ud the same regional area (Iraq and Iran). The main tick species, identified on the examined animals, Hyalomma\ud anatolicum, was widely distributed and was found in all of the surveyed governorates.\ud Conclusion: Theileria spp. are widespread in Oman with variable prevalence detected in different regions. Two\ud economically important hosts, cattle and sheep are at high risk from virulent T. annulata and T. lestoquardi,\ud respectively. The survey indicates extensive exposure to ticks and transmission of infection that has a significant\ud economic impact. The higher prevalence of T. lestoquardi as mixed rather than single infection requires further\ud investigation.