Establishment and Application of a Quantitative PCR Method for E248R Gene of African Swine Fever Virus

SIMPLE SUMMARY: African swine fever (ASF) is the most serious animal disease that endangers the pig industry in China, causing the heaviest economic loss so far. Effective prevention and control of ASF is necessary for China’s pig industry, and also the focus and hotspot of global swine industry disease prevention and control research. In view of this, rapid, specific and sensitive diagnosis of ASF is of great significance. In this study, ASF virus (ASFV) E248R gene was selected to be the target for establishing a real-time PCR method which did not cross-react with other porcine viruses that could cause similar symptoms. The detection methods can be used for the efficient detection of ASFV infection and the recombinant live-vectored virus-expressing antigen protein of ASFV. ABSTRACT: ASF has caused huge economic losses to China’s swine industry. As clinical symptoms of ASF were difficult to distinguish from classical swine fever and porcine reproductive and respiratory syndrome (PRRS), rapid and effective differential diagnosis of ASFV seems very important to control the spread of the disease. In this study, the ASFV E248R gene was selected to be the target for establishing a real-time PCR method. TaqMan real-time PCR for the detection of ASFV E248R gene did not cross-react with other porcine viruses that could cause similar symptoms. The results of the repeatability test showed that the coefficients of variation between and within groups were lower than 1.977%. This method can be used for the rapid detection and early diagnosis of ASF. Meanwhile, the recombinant PRRS virus (PRRSV)-expressing E248R gene of ASFV was constructed and rescued by using the reverse genetic platform of live-attenuated PRRSV vaccine. The ASFV E248R gene can be detected by using this real-time PCR detection method, confirming that the ASFV E248R gene could be stably amplified in PRRSV genome at least 20 cell passages. The detection methods can be used for the efficient detection of the ASFV infection and recombinant PRRSV live vector virus-expressing ASFV antigen protein.