Babesiosis, caused by Babesia ovis, is a major seasonal issue in sheep, particularly in countries like Türkiye with high Rhipicephalus bursa tick populations. Previous studies employing various methods such as microscopy, serology, or molecular techniques have reported different epidemiological data concerning ovine babesiosis. Addressing this knowledge gap, our study employed a combined nested PCR (nPCR)/indirect ELISA (iELISA) approach, analyzing blood samples collected from 414 sheep between April and July 2023 using both techniques. nPCR amplified the 18S ribosomal RNA gene of B. ovis and determined a molecular prevalence of 1.9%. Conversely, serological testing using iELISA targeted the BoSA1 antigen and revealed a significantly higher positivity rate of 59.9% for anti-B. ovis antibodies. The temporary presence of Babesia after recovery reduces nPCR sensitivity, resulting in lower molecular prevalence. However, even if Babesia is not present in the host, anti-B. ovis antibodies remain in the serum for a long time and can be detected serologically. Our study underscores the necessity of concurrently employing molecular and serological methods for an accurate assessment of B. ovis prevalence. It highlights the importance of comprehensive epidemiological approaches for effective disease management in sheep populations.

Babesia ovis is a significant tick-borne parasite of sheep, capable of causing both acute disease and long-lasting, low-grade infections. Imidocarb dipropionate (IMDP) is commonly used against babesiosis, yet whether it can completely eliminate B. ovis remains uncertain. In this study, we examined whether the parasite persists after treatment and whether such residual infections can still be transmitted. Three sheep were experimentally infected, treated with IMDP once clinical signs appeared, and then monitored for 180 days by microscopy, nested PCR, and iELISA. Fever and microscopic parasitemia resolved soon after treatment, but nPCR intermittently detected parasite DNA for several weeks. By day 180, all treated sheep were negative by nPCR and microscopy, while two still showed detectable antibodies. Blood collected at this time was transfused into naïve sheep. Two of the three recipients showed nPCR positivity at scattered time points and later seroconverted while showing no clinical signs. In contrast, Rhipicephalus bursa ticks that fed on the treated donors neither acquired the parasite nor transmitted it to recipients, likely because post-treatment parasitemia remained below the acquisition threshold. Overall, these results indicate that IMDP controls clinical disease but may not fully clear B. ovis, allowing silent transmission through blood despite negative routine tests.