期刊
VIROLOGICA SINICA
卷 36, 期 2, 页码 220-230出版社
SPRINGER
DOI: 10.1007/s12250-020-00323-1
关键词
African swine fever (ASFV); Recombinase polymerase amplification (RPA); CRISPR-lwCas13a; Sensitive diagnosis
类别
资金
- National Natural Science Foundation of China [31522057, 31872451, 31720103917, 31872452]
A nucleic acid detection method using RPA-CRISPR was developed for diagnosing African swine fever, demonstrating high sensitivity and the ability to distinguish ASFV genomic DNA from other porcine viral DNA/RNA without cross-reactivity. The method showed 100% coincidence rate for diagnosing ASFV clinical DNA samples, highlighting its potential for clinical quarantine of ASFV in the swine industry and food security.
African swine fever (ASF) is an infectious disease caused by African swine fever virus (ASFV) with clinical symptoms of high fever, hemorrhages and high mortality rate, posing a threat to the global swine industry and food security. Quarantine and control of ASFV is crucial for preventing swine industry from ASFV infection. In this study, a recombinase polymerase amplification (RPA)-CRISPR-based nucleic acid detection method was developed for diagnosing ASF. As a highly sensitive method, RPA-CRISPR can detect even a single copy of ASFV plasmid and genomic DNA by determining fluorescence signal induced by collateral cleavage of CRISPR-lwCas13a (previously known as C2c2) through quantitative real-time PCR (qPCR) and has the same or even higher sensitivity than the traditional qPCR method. A lateral flow strip was developed and used in combination with RPA-CRISPR for ASFV detection with the same level of sensitivity of TaqMan qPCR. Likewise, RPA-CRISPR is capable of distinguishing ASFV genomic DNA from viral DNA/RNA of other porcine viruses without any cross-reactivity. This diagnostic method is also available for diagnosing ASFV clinical DNA samples with coincidence rate of 100% for both ASFV positive and negative samples. RPA-CRISPR has great potential for clinical quarantine of ASFV in swine industry and food security.
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