Key result
The duplex insulated isothermal PCR assay rapidly detected and differentiated ASFV genotypes I and II with a detection limit of 20 copies per reaction and successfully identified all 58 clinical samples as genotype II.
Why the study?
With genotype I African swine fever virus newly reported alongside genotype II and no commercial vaccine available, rapid onsite differentiation of both genotypes was needed.
Does the duplex iiPCR assay accurately and rapidly detect and differentiate ASFV genotypes I and II in clinical samples?
Does the duplex iiPCR assay accurately and rapidly detect and differentiate ASFV genotypes I and II in clinical samples?
The developed duplex iiPCR assay provides a rapid, sensitive, and specific on-site tool for detecting and differentiating ASFV genotypes I and II within 40 minutes, allowing for direct use of non-hemolytic serum samples.
Answers are grounded in this paper and cite their sources.
May enable rapid onsite ASFV I/II differentiation; leaves open prospective field validation before adoption.
Genotype II African swine fever virus (ASFV) has been plaguing Asian pig industry since 2018. Recently, genotype I ASFV was reported for the first time in China. Since there is no commercial vaccine available against ASFV, early onsite detection and quick culling procedures are commonly used by many countries all over the world. It is important that the above two genotypes of ASFV could be quickly differentiated during onsite detection at the same time. In this study, we established a sensitive and simple Fluorescent Probe Hydrolysis-Insulated isothermal PCR (iiPCR) that can detect and differentiate two genotypes of ASFV within 40 minutes. The positive or negative results of tested samples were displayed on the screen of the device automatically after PCR amplification was complete. The detection limit of the iiPCR was tested to be 20 copies for both genotype I and genotype II ASFVs. There was no cross-reactivity with other swine viruses by using the established iiPCR. Fifty-eight ASFV positive samples confirmed by National ASF Reference Laboratory were subjected to the established duplex iiPCR for genotype differentiation. The results showed that all these ASFV-positive samples belong to genotype II. At last, we found serum samples could be directly used as the templates for iiPCR without comprising sensitivity and specificity. Therefore, the duplex iiPCR established in study provide a useful tool for ASFV onsite detection and genotype differentiation.
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Song et al. (2022) studied African Swine Fever Virus (ASFV) (n=68). Duplex insulated isothermal PCR (iiPCR) vs. Real-time PCR was evaluated on Detection limit and genotype differentiation accuracy. The duplex insulated isothermal PCR assay rapidly detected and differentiated ASFV genotypes I and II with a detection limit of 20 copies per reaction and successfully identified all 58 clinical samples as genotype II.
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