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Establishment of a Dual Real-Time PCR Assay for the Identification of African Swine Fever Virus Genotypes I and II in China

Since the first outbreak of ASFV genotype II in China in 2018, ASF has posed a significant threat to the swine industry. After the emergence of genotype I in China in 2020, the epidemic prevention and control have become more difficult. No effective commercial vaccine is currently available, and the...

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Autores principales: Gao, Qi, Feng, Yongzhi, Yang, Yunlong, Luo, Yizhuo, Gong, Ting, Wang, Heng, Gong, Lang, Zhang, Guihong, Zheng, Zezhong
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9198542/
https://www.ncbi.nlm.nih.gov/pubmed/35720851
http://dx.doi.org/10.3389/fvets.2022.882824
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author Gao, Qi
Feng, Yongzhi
Yang, Yunlong
Luo, Yizhuo
Gong, Ting
Wang, Heng
Gong, Lang
Zhang, Guihong
Zheng, Zezhong
author_facet Gao, Qi
Feng, Yongzhi
Yang, Yunlong
Luo, Yizhuo
Gong, Ting
Wang, Heng
Gong, Lang
Zhang, Guihong
Zheng, Zezhong
author_sort Gao, Qi
collection PubMed
description Since the first outbreak of ASFV genotype II in China in 2018, ASF has posed a significant threat to the swine industry. After the emergence of genotype I in China in 2020, the epidemic prevention and control have become more difficult. No effective commercial vaccine is currently available, and the disease is difficult to eradicate; therefore, the identification of the ASFV genotype is critical to establish biosafety control measures. In this study, a dual real-time PCR detection method based on B646L and E183L genes was developed to distinguish between ASFV genotypes I and II by specifically amplifying the genotype I E183L gene. The method is strongly specific, detects B646L and E183L genes simultaneously, and does not cross-react with PEDV, PCV, PRRSV, PRV, and CSFV. The double real-time PCR detection of ASFV genotypes I and II showed a B646L amplification curve, and only genotype I showed an E183L amplification curve, consistent with our expectations. The method has high sensitivity and the lowest copy numbers detected for recombinant plasmids B646L and E183L were 1.07 × 10(2) and 3.13 × 10(4) copies/μL, respectively. The method is reproducible, and the coefficient of variation for detecting the coefficient of variation (CV) values of the two recombinant plasmids was <2%. Seven samples were positive and 277 were negative, and the results of the two methods were consistent. The dual real-time PCR presented in this study provides a rapid detection method for the identification of ASFV genotypes I and II, which may lead to improving efficient prevention and control measures for ASF in China.
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spelling pubmed-91985422022-06-16 Establishment of a Dual Real-Time PCR Assay for the Identification of African Swine Fever Virus Genotypes I and II in China Gao, Qi Feng, Yongzhi Yang, Yunlong Luo, Yizhuo Gong, Ting Wang, Heng Gong, Lang Zhang, Guihong Zheng, Zezhong Front Vet Sci Veterinary Science Since the first outbreak of ASFV genotype II in China in 2018, ASF has posed a significant threat to the swine industry. After the emergence of genotype I in China in 2020, the epidemic prevention and control have become more difficult. No effective commercial vaccine is currently available, and the disease is difficult to eradicate; therefore, the identification of the ASFV genotype is critical to establish biosafety control measures. In this study, a dual real-time PCR detection method based on B646L and E183L genes was developed to distinguish between ASFV genotypes I and II by specifically amplifying the genotype I E183L gene. The method is strongly specific, detects B646L and E183L genes simultaneously, and does not cross-react with PEDV, PCV, PRRSV, PRV, and CSFV. The double real-time PCR detection of ASFV genotypes I and II showed a B646L amplification curve, and only genotype I showed an E183L amplification curve, consistent with our expectations. The method has high sensitivity and the lowest copy numbers detected for recombinant plasmids B646L and E183L were 1.07 × 10(2) and 3.13 × 10(4) copies/μL, respectively. The method is reproducible, and the coefficient of variation for detecting the coefficient of variation (CV) values of the two recombinant plasmids was <2%. Seven samples were positive and 277 were negative, and the results of the two methods were consistent. The dual real-time PCR presented in this study provides a rapid detection method for the identification of ASFV genotypes I and II, which may lead to improving efficient prevention and control measures for ASF in China. Frontiers Media S.A. 2022-06-01 /pmc/articles/PMC9198542/ /pubmed/35720851 http://dx.doi.org/10.3389/fvets.2022.882824 Text en Copyright © 2022 Gao, Feng, Yang, Luo, Gong, Wang, Gong, Zhang and Zheng. 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 Veterinary Science
Gao, Qi
Feng, Yongzhi
Yang, Yunlong
Luo, Yizhuo
Gong, Ting
Wang, Heng
Gong, Lang
Zhang, Guihong
Zheng, Zezhong
Establishment of a Dual Real-Time PCR Assay for the Identification of African Swine Fever Virus Genotypes I and II in China
title Establishment of a Dual Real-Time PCR Assay for the Identification of African Swine Fever Virus Genotypes I and II in China
title_full Establishment of a Dual Real-Time PCR Assay for the Identification of African Swine Fever Virus Genotypes I and II in China
title_fullStr Establishment of a Dual Real-Time PCR Assay for the Identification of African Swine Fever Virus Genotypes I and II in China
title_full_unstemmed Establishment of a Dual Real-Time PCR Assay for the Identification of African Swine Fever Virus Genotypes I and II in China
title_short Establishment of a Dual Real-Time PCR Assay for the Identification of African Swine Fever Virus Genotypes I and II in China
title_sort establishment of a dual real-time pcr assay for the identification of african swine fever virus genotypes i and ii in china
topic Veterinary Science
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9198542/
https://www.ncbi.nlm.nih.gov/pubmed/35720851
http://dx.doi.org/10.3389/fvets.2022.882824
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