Abstract
African swine fever virus (ASFV), the aetiological agent of African swine fever (ASF), causes lethal haemorrhagic fever in domestic pigs with high mortality and morbidity and has devastating consequences on the global swine industry. On-site rapid and sensitive detection of ASFV is key to the timely implementation of control. In this study, we developed a rapid, sensitive and instrument-free ASFV detection method based on CRISPR/Cas12a technology and lateral flow detection (named CRISPR/Cas12a-LFD). The limit of detection of CRISPR/Cas12a-LFD is 20 copies of ASFV genomic DNA per reaction, and the detection process can be completed in an hour. The assay showed no cross-reactivity with other swine DNA viruses, and has 100% agreement with real-time PCR detection of ASFV in 149 clinical samples. Overall, the CRISPR/Cas12a-LFD method provides a novel alternative for the portable, simple, sensitive, and specific detection of ASFV and may contribute to the prevention and control of ASF outbreaks.
Highlights
African swine fever virus (ASFV), the aetiological agent of African swine fever (ASF), causes lethal haemorrhagic fever in domestic pigs with high mortality and morbidity and has devastating consequences on the global swine industry
The sequence alignment result of the ASFV B646L gene indicated that the regions targeted by crRNA1 and crRNA6 were highly conserved in all genotypes, while the regions targeted by the remaining eight CRISPR RNAs (crRNAs) contained single nucleotide polymorphisms (SNPs) in different genotypes (Supplementary Fig. 2b)
After being activated by specific ASFV genomic DNA, CRISPR/Cas12a cleaves the single-stranded DNA (ssDNA) reporter molecule through its nonspecific endonuclease activity, which leads to the generation of fluorescence signals or detection via the lateral flow assay
Summary
African swine fever virus (ASFV), the aetiological agent of African swine fever (ASF), causes lethal haemorrhagic fever in domestic pigs with high mortality and morbidity and has devastating consequences on the global swine industry. On-site rapid and sensitive detection of ASFV is key to the timely implementation of control. The CRISPR/Cas12a-LFD method provides a novel alternative for the portable, simple, sensitive, and specific detection of ASFV and may contribute to the prevention and control of ASF outbreaks. The disease is caused by African swine fever virus (ASFV), which belongs to the genus Asfivirus within the Asfarviridae family[2]. The recommended diagnostic methods include polymerase chain reaction (PCR) and real-time PCR for ASFV genomic DNA10,11. These methods require an expensive instrument and professional operation, which limits their application in the field. Both loopmediated isothermal amplification (LAMP) and cross-priming amplification (CPA) are available for ASFV detection[12,13]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
