Rapid and sensitive RPA-Cas12a-fluorescence assay for point-of-care detection of African swine fever virus.

African swine fever (ASF) is a serious contagious disease that causes fatal haemorrhagic fever in domestic and wild pigs, with high morbidity. It has caused devastating damage to the swine industry worldwide, necessitating the focus of attention on detection of the ASF pathogen, the African swine fe...

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Autores principales:	Jinyu Fu, Yueping Zhang, Guang Cai, Geng Meng, Shuobo Shi
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2021
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/ef4c2251d8524aa2988a6e1a3288f3a7
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spelling	oai:doaj.org-article:ef4c2251d8524aa2988a6e1a3288f3a72021-12-02T20:06:49ZRapid and sensitive RPA-Cas12a-fluorescence assay for point-of-care detection of African swine fever virus.1932-620310.1371/journal.pone.0254815https://doaj.org/article/ef4c2251d8524aa2988a6e1a3288f3a72021-01-01T00:00:00Zhttps://doi.org/10.1371/journal.pone.0254815https://doaj.org/toc/1932-6203African swine fever (ASF) is a serious contagious disease that causes fatal haemorrhagic fever in domestic and wild pigs, with high morbidity. It has caused devastating damage to the swine industry worldwide, necessitating the focus of attention on detection of the ASF pathogen, the African swine fever virus (ASFV). In order to overcome the disadvantages of conventional diagnostic methods (e.g. time-consuming, demanding and unintuitive), quick detection tools with higher sensitivity need to be explored. In this study, based on the conserved p72 gene sequence of ASFV, we combined the Cas12a-based assay with recombinase polymerase amplification (RPA) and a fluorophore-quencher (FQ)-labeled reporter assay for rapid and visible detection. Five crRNAs designed for Cas12a-based assay showed specificity with remarkable fluorescence intensity under visual inspection. Within 20 minutes, with an initial concentration of two copies of DNA, the assay can produce significant differences between experimental and negative groups, indicating the high sensitivity and rapidity of the method. Overall, the developed RPA-Cas12a-fluorescence assay provides a fast and visible tool for point-of-care ASFV detection with high sensitivity and specificity, which can be rapidly performed on-site under isothermal conditions, promising better control and prevention of ASF.Jinyu FuYueping ZhangGuang CaiGeng MengShuobo ShiPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 16, Iss 7, p e0254815 (2021)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Jinyu Fu Yueping Zhang Guang Cai Geng Meng Shuobo Shi Rapid and sensitive RPA-Cas12a-fluorescence assay for point-of-care detection of African swine fever virus.
description	African swine fever (ASF) is a serious contagious disease that causes fatal haemorrhagic fever in domestic and wild pigs, with high morbidity. It has caused devastating damage to the swine industry worldwide, necessitating the focus of attention on detection of the ASF pathogen, the African swine fever virus (ASFV). In order to overcome the disadvantages of conventional diagnostic methods (e.g. time-consuming, demanding and unintuitive), quick detection tools with higher sensitivity need to be explored. In this study, based on the conserved p72 gene sequence of ASFV, we combined the Cas12a-based assay with recombinase polymerase amplification (RPA) and a fluorophore-quencher (FQ)-labeled reporter assay for rapid and visible detection. Five crRNAs designed for Cas12a-based assay showed specificity with remarkable fluorescence intensity under visual inspection. Within 20 minutes, with an initial concentration of two copies of DNA, the assay can produce significant differences between experimental and negative groups, indicating the high sensitivity and rapidity of the method. Overall, the developed RPA-Cas12a-fluorescence assay provides a fast and visible tool for point-of-care ASFV detection with high sensitivity and specificity, which can be rapidly performed on-site under isothermal conditions, promising better control and prevention of ASF.
format	article
author	Jinyu Fu Yueping Zhang Guang Cai Geng Meng Shuobo Shi
author_facet	Jinyu Fu Yueping Zhang Guang Cai Geng Meng Shuobo Shi
author_sort	Jinyu Fu
title	Rapid and sensitive RPA-Cas12a-fluorescence assay for point-of-care detection of African swine fever virus.
title_short	Rapid and sensitive RPA-Cas12a-fluorescence assay for point-of-care detection of African swine fever virus.
title_full	Rapid and sensitive RPA-Cas12a-fluorescence assay for point-of-care detection of African swine fever virus.
title_fullStr	Rapid and sensitive RPA-Cas12a-fluorescence assay for point-of-care detection of African swine fever virus.
title_full_unstemmed	Rapid and sensitive RPA-Cas12a-fluorescence assay for point-of-care detection of African swine fever virus.
title_sort	rapid and sensitive rpa-cas12a-fluorescence assay for point-of-care detection of african swine fever virus.
publisher	Public Library of Science (PLoS)
publishDate	2021
url	https://doaj.org/article/ef4c2251d8524aa2988a6e1a3288f3a7
work_keys_str_mv	AT jinyufu rapidandsensitiverpacas12afluorescenceassayforpointofcaredetectionofafricanswinefevervirus AT yuepingzhang rapidandsensitiverpacas12afluorescenceassayforpointofcaredetectionofafricanswinefevervirus AT guangcai rapidandsensitiverpacas12afluorescenceassayforpointofcaredetectionofafricanswinefevervirus AT gengmeng rapidandsensitiverpacas12afluorescenceassayforpointofcaredetectionofafricanswinefevervirus AT shuoboshi rapidandsensitiverpacas12afluorescenceassayforpointofcaredetectionofafricanswinefevervirus
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Rapid and sensitive RPA-Cas12a-fluorescence assay for point-of-care detection of African swine fever virus.

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