Development and Evaluation of a Semi-Nested PCR Method Based on the Gene for the Detection of Infections in Goats.

We developed and evaluated a semi-nested PCR assay for the detection of infection in goats based on the sequence of the gene. Following in silico screening, the specificity of the primers was assessed using reference DNA samples, including , , , , , , , and . To determine the sensitivity of the method, blood infected with 2% parasitemia of was diluted to 10-fold serial dilutions. The method specifically amplified a 438 bp fragment of DNA, but did not demonstrate cross-amplification with the other hemoparasites tested. The sensitivity assay indicated that this PCR method was able to detect infection at a dilution of 10 of 2% parasitemia (0.074 parasites/200 µL). Ninety-seven blood samples collected from goats were used to analyze for , and the infection was detected in 18.5% of the goats. Additionally, the method was also applied to 44 field DNA samples that were detected to be positive for by reverse line blotting (RLB), and showed 84.1% agreement. The findings revealed that newly developed semi-nested PCR can detect infections in goats with high sensitivity and specificity.