Abstract
Objective Establishing a highly sensitive real-time fluorescence quantitative PCR (qPCR) method for universal testing of epidemic African swine fever virus (ASFV) strains. Methods The ASFV p72 gene was targeted to design primer probes covering 24 p72 genotypes. The optimal amount of dimethylsulphoxide (DMSO) for qPCR amplification was determined, Various sensitivity and limit of detection (LOD) tests were performed, and clinical samples from China and imported goods were tested. Results The optimal primer-probe combination could specifically detect ASFV, 1.5% DMSO was optimal for qPCR, and LOD reached 3.2 copies/μL with good reproducibility (n = 20, p = 0.369). The method was employed to test 142 clinically suspected samples, of which 30 pig blood and 37 pig tissue samples were ASFV-positive. Moreover, the positive testing rate for ASFV was higher than for the standard qPCR method recommended by the Office International Des Epizooties (OIE), and for the commercially available kit. Thus, our method is superior for testing weakly positive samples with low virus titre, and epidemic strains present in imported goods. Conclusion Our method could be employed for universal testing of epidemic ASFV strains worldwide, ensuring wider coverage of hosts and ASFV strains/endemic strains, reducing false negatives, and benefitting early diagnosis.
Highlights
African swine fever (ASF) is a febrile, acute, and subacute highly contagious infectious disease caused by ASF virus (ASFV) [1]
Our method could be employed for universal testing of epidemic African swine fever virus (ASFV) strains worldwide, ensuring wider coverage of hosts and ASFV strains/endemic strains, reducing false negatives, and benefitting early diagnosis
The results showed that the target sequence shared 93.9100% homology with the other 23 representative ASFV strains in GenBank, and the lowest homology (93.9%) was with strain MWHOG/1
Summary
African swine fever (ASF) is a febrile, acute, and subacute highly contagious infectious disease caused by ASF virus (ASFV) [1]. The disease first broke out in Kenya in 1921, spread to many countries in Europe, Central America, Africa and Asia. On August 3, 2018, it was confirmed by the China Animal Health and Epidemiology Center that ASF was first discovered in Shenyang, China [4]. It broke out in Jiangsu, Zhejiang, and other provinces in China, and it is still spreading [5]. The only way to control the spread of disease is to kill diseased pigs, treat the source of the disease, and cut off the source of infection [8] [9]
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