Molecular Epidemiology and Phylogeny of and in Sheep and Goats from Southern Punjab, Pakistan.

spp. are responsible for ovine and caprine theileriosis, leading to significant morbidity and mortality in small ruminants. The present study aims to investigate spp. infections in small ruminants from Southern Punjab in Pakistan, and genetic characterize revealed spp. isolates. A total of 93 sheep and 107 goats were sampled between May and August 2022. Blood smears were examined microscopically, and PCR amplification targeting the 18S rRNA gene was performed to detect spp. Additionally, specific PCR assays targeting 18S rRNA and partial sequences were used to identify and , respectively.  The prevalence of spp. was significantly higher using PCR (13.5%) compared to microscopic screening (5%). Sheep showed a higher prevalence rate (19.4%) compared to goats (8.4%) ( = 0.024). Young sheep aged ≤ 1 year were more commonly infected with spp. (41%) compared to older sheep ( = 0.006). The prevalence of spp. was higher in sheep-only herds (37.3%) compared to goat-only herds (18%) or mixed-species herds (8.1%) ( = 0.015). The prevalence rates of and were 9% and 2.5%, respectively, with four animals (2 goats and 2 sheep) showing co-infection. Phylogenetic analysis revealed that our 18S rRNA sequence clustered with previously reported sequences from sheep in Turkey, China, Spain, and goats in Tanzania. The obtained partial sequence formed a distinct cluster from other isolates in Pakistan and neighboring countries.  spp. co-circulation in Pakistani small ruminants, particularly the presence of and , highlights the need for attention from animal health decision-makers due to their financial and health impacts.