Comparison of extraction methods to detect porcine circovirus-2 and porcine reproductive and respiratory syndrome virus in semen samples

Since, Porcine Circa Virus-2 (PCV2) and Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) are shed in semen for a long period of time after infection and artificial insemination is a common practice in pig farms, semen is an important source of spreading these viruses to naive populations. Therefore, rapid and accurate detection of PCV2 or PRRSV in semen could be critical at the standpoint of disease prevention and control. In this study, the performance of three different extraction methods on semen samples was compared to determine an optimal extraction method for semen samples to detect PCV2 or PRRSV. Seven or 115 semen samples were collected from the boars experimentally challenged with PRRSV or PCV2, respectively. These two sets of the semen samples were processed by three different extraction methods: High-Throughput Total Nucleic Acids Isolation kit (HT-TNA), High-Throughput Viral RNA Isolation kit with modified procedure (mHT-VR) and DNeasy kit or RNeasy kit for PCV2 or PRRSV, respectively. Then, the efficiency of the extraction methods were compared by conducting the same real-time PCR for each virus. HT-INA and mHT-VR kits were faster and more convenient to process semen samples and HT-TNA kit showed higher or compatible sensitivity as compared to the other two methods while all three methods demonstrated 100% specificity. In conclusion, the HT-TNA Extraction Method could significantly reduce testing time and effort to process semen samples and showed better sensitivity for detection of PCV2 and PRRSV in semen.