Ovine theileriosis is a tick-borne hemoparasitic disease caused by protozoan parasites of the genus Theileria, which poses a substantial threat to sheep and goats. The disease is prevalent in northwestern China, yet studies on the diversity of its pathogens in Qinghai Province remain limited. From 2014 to 2025, 1062 blood samples were collected from sheep in 11 counties of Qinghai Province, and the Theileria spp. were investigated by PCR targeting the 18S rRNA gene. Genetic diversity analysis was performed to assess sequence variations and phylogenetic relationships. The results show that 424 samples tested positive for Theileria spp., with an overall infection rate of 39.92%, including T. uilenbergi 26.74%, T. luwenshuni 22.41%, T. ovis 18.17%, T. capreoli 0.56%, and Theileria sp. OT3 0.09%. The co-infection rate was 22.69%. Six haplotypes were detected in both T. uilenbergi and T. ovis, and fifteen in T. luwenshuni. Hap 1 was the dominant haplotype for all three species, with respective proportions of 73.08%, 77.06% and 82.17%. Phylogenetic analysis showed the strains were most closely related to those from Hunan, Turkey, Shaanxi, etc. These findings reveal a high diversity of Theileria species in Qinghai sheep, which provides crucial epidemiological insights into the transmission dynamics of ovine theileriosis in Qinghai and vital support for developing effective control strategies.

Ticks are obligate hematophagous arthropods and important vectors of various pathogens. This study identified tick species parasitizing goats in Guide County, Qinghai Province, China, and screened for Theileria and Anaplasma species. Tick specimens were identified via morphological observation and molecular analysis of 16S rRNA and cox1 gene sequences. Pathogen detection was performed using PCR targeting the 18S rRNA gene of Theileria and the groEL gene of Anaplasma, followed by sequencing and phylogenetic analysis. Morphological and molecular data consistently identified all 80 engorged tick samples collected from goats on a single farm as Hyalomma marginatum, with 97.7–99.7% sequence similarity to reference sequences. Among the 80 tick samples, the DNA detection rate for Theileria ovis was 5.0% (4/80), 17.5% for Theileria luwenshuni (14/80), and 6.3% for Anaplasma ovis (5/80). In this study, H. marginatum was identified in Guide County, Qinghai Province, thereby enriching the current knowledge of tick species distribution in this region. Furthermore, the PCR detection of T. ovis, T. luwenshuni, and A. ovis DNA in H. marginatum provides fundamental molecular surveillance data for tick-borne pathogen monitoring in this region; however, because all examined ticks were engorged, the detected DNA may have originated from the residual host blood meal, and these findings do not demonstrate infection of the ticks or vector competence.