Comparative aspects of laboratory testing for the detection of Toxoplasma gondii and its differentiation from Neospora caninum as the etiologic agent of ovine abortion.

Histologic examination of aborted material is an essential component in the diagnosis of ovine toxoplasmosis. However, the detection of Toxoplasma gondii in histologic sections, and its differentiation from the closely related protozoan Neospora caninum, is challenging. We developed a chromogenic in situ hybridization (ISH) assay for the identification of T. gondii in paraffin-embedded tissue samples. We examined retrospectively the archived placental tissue of 200 sheep abortion submissions for the presence of T. gondii by immunohistochemistry (IHC), ISH, and real-time PCR (rtPCR). All placental samples that tested positive for T. gondii by rtPCR (9 of 200) were also positive by IHC, with inconclusive IHC staining in an additional 7 rtPCR-negative cases. Further testing for N. caninum of all 200 placentas by rtPCR revealed 7 Neospora-positive cases. T. gondii ISH was positive in 4 of 9 IHC-positive samples and 1 of the 7 N. caninum rtPCR-positive samples. Real-time PCR was used as the reference standard for specificity and sensitivity calculations regarding placenta samples. Specificity of ISH and IHC was 99% and 96-100%, respectively. The sensitivity of ISH (44%) was quite low compared to IHC (100%). The exclusive use of ISH for the detection of T. gondii, and thus for the diagnosis of ovine toxoplasmosis, was not acceptable. However, combined with rtPCR, both ISH and IHC can be useful detection methods to improve histologic evaluation by visualizing the parasite within tissue sections.