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A probe-based real-time PCR assay for the detection of Neospora caninum in clinical samples from cattle
- Barry, Rhiannon, Nissly, Ruth H., Feria, Willard, Thirumalapura, Nagaraja, Tewari, Deepanker, Jayarao, Bhushan M., Kuchipudi, Suresh V.
- Veterinary parasitology 2019 v.269 pp. 2-6
- Chlamydia, DNA, Leptospira, Neospora caninum, Protozoa, cattle, detection limit, fetus, parasites, pathogens, quantitative polymerase chain reaction, rapid methods, synthetic genes, tissues
- Neospora caninum is an apicomplexan protozoan parasite that is a leading cause of abortion in cattle. Detection of parasite-specific DNA by PCR is a highly sensitive method for identifying the presence of N. caninum in a variety of tissues. We developed and validated a probe-based real-time PCR assay targeting the conserved Nc5 gene of N. caninum. Using N. caninum strain Nc-1 genomic DNA and a synthetic gene fragment as amplification standards, we determined the PCR amplification efficiency and the limit of detection to be 95.60% and 3 copies, respectively. Five pathogens frequently associated with bovine abortions, namely bovine viral diarrhea virus types I and II, bovine alphaherpesvirus-1, Chlamydia, and Leptospira, were tested to ensure analytical exclusivity. A total of 103 clinical samples from aborted fetuses were tested concurrently with a standard conventional PCR and the new probe-based real-time PCR assay. All tested samples showed 100% agreement between these two assays. In conclusion, the probe-based real-time PCR assay facilitates accurate and rapid detection of N. caninum from abortions in cattle.