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Development of a Duplex Insulated Isothermal PCR Assay for Rapid On-Site Detection and Differentiation of Genotypes 1 and 2 of African Swine Fever Virus
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...
Autores principales: | , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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Frontiers Media S.A.
2022
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9300913/ https://www.ncbi.nlm.nih.gov/pubmed/35873169 http://dx.doi.org/10.3389/fcimb.2022.948771 |
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author | Song, Ruilong Liu, Penggang Yang, Yang Lee, Hu Suk Chen, Changhai Wu, Xiaodong Li, Xiangdong |
author_facet | Song, Ruilong Liu, Penggang Yang, Yang Lee, Hu Suk Chen, Changhai Wu, Xiaodong Li, Xiangdong |
author_sort | Song, Ruilong |
collection | PubMed |
description | 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. |
format | Online Article Text |
id | pubmed-9300913 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-93009132022-07-22 Development of a Duplex Insulated Isothermal PCR Assay for Rapid On-Site Detection and Differentiation of Genotypes 1 and 2 of African Swine Fever Virus Song, Ruilong Liu, Penggang Yang, Yang Lee, Hu Suk Chen, Changhai Wu, Xiaodong Li, Xiangdong Front Cell Infect Microbiol Cellular and Infection Microbiology 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. Frontiers Media S.A. 2022-07-07 /pmc/articles/PMC9300913/ /pubmed/35873169 http://dx.doi.org/10.3389/fcimb.2022.948771 Text en Copyright © 2022 Song, Liu, Yang, Lee, Chen, Wu and Li https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
spellingShingle | Cellular and Infection Microbiology Song, Ruilong Liu, Penggang Yang, Yang Lee, Hu Suk Chen, Changhai Wu, Xiaodong Li, Xiangdong Development of a Duplex Insulated Isothermal PCR Assay for Rapid On-Site Detection and Differentiation of Genotypes 1 and 2 of African Swine Fever Virus |
title | Development of a Duplex Insulated Isothermal PCR Assay for Rapid On-Site Detection and Differentiation of Genotypes 1 and 2 of African Swine Fever Virus |
title_full | Development of a Duplex Insulated Isothermal PCR Assay for Rapid On-Site Detection and Differentiation of Genotypes 1 and 2 of African Swine Fever Virus |
title_fullStr | Development of a Duplex Insulated Isothermal PCR Assay for Rapid On-Site Detection and Differentiation of Genotypes 1 and 2 of African Swine Fever Virus |
title_full_unstemmed | Development of a Duplex Insulated Isothermal PCR Assay for Rapid On-Site Detection and Differentiation of Genotypes 1 and 2 of African Swine Fever Virus |
title_short | Development of a Duplex Insulated Isothermal PCR Assay for Rapid On-Site Detection and Differentiation of Genotypes 1 and 2 of African Swine Fever Virus |
title_sort | development of a duplex insulated isothermal pcr assay for rapid on-site detection and differentiation of genotypes 1 and 2 of african swine fever virus |
topic | Cellular and Infection Microbiology |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9300913/ https://www.ncbi.nlm.nih.gov/pubmed/35873169 http://dx.doi.org/10.3389/fcimb.2022.948771 |
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