Abstract
African swine fever (ASF) is a lethal disease in swine caused by etiologic African swine fever virus (ASFV). The global spread of ASFV has resulted in huge economic losses globally. In the absence of effective vaccines or drugs, pathogen surveillance has been the most important first-line intervention to prevent ASF outbreaks. Among numerous diagnostic methods, recombinase polymerase amplification (RPA)-based detection is capable of producing sensitive and specific results without relying on the use of expensive instruments. However, currently used gene-specific, probe-based RPA for ASFV detection is expensive and time-consuming. To improve the efficiency of ASFV surveillance, a novel directly visualized SYBR Green I-staining RPA (RPAS) method was developed to detect the ASFV genome. SYBR Green I was added to the amplified RPA products for direct visualization by the naked eye. The sensitivity and specificity of this method were confirmed using standard plasmid and inactivated field samples. This method was shown to be highly specific with a detection limit of 103 copies/μl of ASFV in 15 min at 35°C without any cross-reactions with other important porcine viruses selected. In summary, this method enables direct sample visualization with reproducible results for ASFV detection and hence has the potential to be used as a robust tool for ASF prevention and control.
Highlights
Belonging to phylum Nucleocytoviricota (NCLDV), class Pokkesviricetes, order Asfuvirales, and family Asfarviridae, African swine fever virus (ASFV) is the only member of genus Asfivirus
This study aims to improve the efficiency of ASFV surveillance
Pseudorabies virus (PRV) strain (HN1201), Japanese encephalitis virus (JEV) live vaccine strain (SA14-14-2), and porcine epidemic diarrhea virus (PEDV) strain were provided by the Henan Academy of Agricultural Sciences (Zhengzhou, China), while porcine reproductive and respiratory syndrome virus (PRRSV) SD16 strain and porcine parvovirus (PPV) strain were derived from our laboratory
Summary
Belonging to phylum Nucleocytoviricota (NCLDV), class Pokkesviricetes, order Asfuvirales, and family Asfarviridae, African swine fever virus (ASFV) is the only member of genus Asfivirus. ASFV causes African swine fever (ASF), which is acute and frequently lethal in domestic pigs and wild boars, and is characterized by high fever, hemorrhage syndrome, and a mortality rate of up to 100%. Reported in Kenya in 1921, ASF has spread to Europe in the 1950s and subsequently to North America in the 1960s (Sanchez-Vizcaino et al, 2012; European Food Safety et al, 2017). Since August 2018 when ASF was firstly reported in China, which has the largest pig consumption market in the world (Ge et al, 2018), there have been numerous ASF outbreaks within a short period of time in more than 30 Chinese provinces, causing a huge economic loss (Zhao et al, 2019; Teklue et al, 2020). ASF has spread to more than 80 countries, threatening the pig industries worldwide (Dixon et al, 2019)
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