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Rapid Detection of Actinobacillus pleuropneumoniae From Clinical Samples Using Recombinase Polymerase Amplification
Actinobacillus pleuropneumoniae (APP) is the causative agent of porcine pleuropneumonia, resulting in high economic impact worldwide. There are currently 19 known serovars of APP, with different ones being predominant in specific geographic regions. Outbreaks of pleuropneumonia, characterized by sud...
Autores principales: | , , , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Frontiers Media S.A.
2022
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8990124/ https://www.ncbi.nlm.nih.gov/pubmed/35400111 http://dx.doi.org/10.3389/fvets.2022.805382 |
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author | Stringer, Oliver W. Li, Yanwen Bossé, Janine T. Forrest, Matthew S. Hernandez-Garcia, Juan Tucker, Alexander W. Nunes, Tiago Costa, Francisco Mortensen, Preben Velazquez, Eduardo Penny, Paul Rodriguez-Manzano, Jesus Georgiou, Pantelis Langford, Paul R. |
author_facet | Stringer, Oliver W. Li, Yanwen Bossé, Janine T. Forrest, Matthew S. Hernandez-Garcia, Juan Tucker, Alexander W. Nunes, Tiago Costa, Francisco Mortensen, Preben Velazquez, Eduardo Penny, Paul Rodriguez-Manzano, Jesus Georgiou, Pantelis Langford, Paul R. |
author_sort | Stringer, Oliver W. |
collection | PubMed |
description | Actinobacillus pleuropneumoniae (APP) is the causative agent of porcine pleuropneumonia, resulting in high economic impact worldwide. There are currently 19 known serovars of APP, with different ones being predominant in specific geographic regions. Outbreaks of pleuropneumonia, characterized by sudden respiratory difficulties and high mortality, can occur when infected pigs are brought into naïve herds, or by those carrying different serovars. Good biosecurity measures include regular diagnostic testing for surveillance purposes. Current gold standard diagnostic techniques lack sensitivity (bacterial culture), require expensive thermocycling machinery (PCR) and are time consuming (culture and PCR). Here we describe the development of an isothermal point-of-care diagnostic test - utilizing recombinase polymerase amplification (RPA) for the detection of APP, targeting the species-specific apxIVA gene. Our APP-RPA diagnostic test achieved a sensitivity of 10 copies/μL using a strain of APP serovar 8, which is the most prevalent serovar in the UK. Additionally, our APP-RPA assay achieved a clinical sensitivity and specificity of 84.3 and 100%, respectively, across 61 extracted clinical samples obtained from farms located in England and Portugal. Using a small subset (n = 14) of the lung tissue samples, we achieved a clinical sensitivity and specificity of 76.9 and 100%, respectively) using lung imprints made on FTA cards tested directly in the APP-RPA reaction. Our results demonstrate that our APP-RPA assay enables a suitable rapid and sensitive screening tool for this important veterinary pathogen. |
format | Online Article Text |
id | pubmed-8990124 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2022 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-89901242022-04-09 Rapid Detection of Actinobacillus pleuropneumoniae From Clinical Samples Using Recombinase Polymerase Amplification Stringer, Oliver W. Li, Yanwen Bossé, Janine T. Forrest, Matthew S. Hernandez-Garcia, Juan Tucker, Alexander W. Nunes, Tiago Costa, Francisco Mortensen, Preben Velazquez, Eduardo Penny, Paul Rodriguez-Manzano, Jesus Georgiou, Pantelis Langford, Paul R. Front Vet Sci Veterinary Science Actinobacillus pleuropneumoniae (APP) is the causative agent of porcine pleuropneumonia, resulting in high economic impact worldwide. There are currently 19 known serovars of APP, with different ones being predominant in specific geographic regions. Outbreaks of pleuropneumonia, characterized by sudden respiratory difficulties and high mortality, can occur when infected pigs are brought into naïve herds, or by those carrying different serovars. Good biosecurity measures include regular diagnostic testing for surveillance purposes. Current gold standard diagnostic techniques lack sensitivity (bacterial culture), require expensive thermocycling machinery (PCR) and are time consuming (culture and PCR). Here we describe the development of an isothermal point-of-care diagnostic test - utilizing recombinase polymerase amplification (RPA) for the detection of APP, targeting the species-specific apxIVA gene. Our APP-RPA diagnostic test achieved a sensitivity of 10 copies/μL using a strain of APP serovar 8, which is the most prevalent serovar in the UK. Additionally, our APP-RPA assay achieved a clinical sensitivity and specificity of 84.3 and 100%, respectively, across 61 extracted clinical samples obtained from farms located in England and Portugal. Using a small subset (n = 14) of the lung tissue samples, we achieved a clinical sensitivity and specificity of 76.9 and 100%, respectively) using lung imprints made on FTA cards tested directly in the APP-RPA reaction. Our results demonstrate that our APP-RPA assay enables a suitable rapid and sensitive screening tool for this important veterinary pathogen. Frontiers Media S.A. 2022-03-25 /pmc/articles/PMC8990124/ /pubmed/35400111 http://dx.doi.org/10.3389/fvets.2022.805382 Text en Copyright © 2022 Stringer, Li, Bossé, Forrest, Hernandez-Garcia, Tucker, Nunes, Costa, Mortensen, Velazquez, Penny, Rodriguez-Manzano, Georgiou and Langford. 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 Stringer, Oliver W. Li, Yanwen Bossé, Janine T. Forrest, Matthew S. Hernandez-Garcia, Juan Tucker, Alexander W. Nunes, Tiago Costa, Francisco Mortensen, Preben Velazquez, Eduardo Penny, Paul Rodriguez-Manzano, Jesus Georgiou, Pantelis Langford, Paul R. Rapid Detection of Actinobacillus pleuropneumoniae From Clinical Samples Using Recombinase Polymerase Amplification |
title | Rapid Detection of Actinobacillus pleuropneumoniae From Clinical Samples Using Recombinase Polymerase Amplification |
title_full | Rapid Detection of Actinobacillus pleuropneumoniae From Clinical Samples Using Recombinase Polymerase Amplification |
title_fullStr | Rapid Detection of Actinobacillus pleuropneumoniae From Clinical Samples Using Recombinase Polymerase Amplification |
title_full_unstemmed | Rapid Detection of Actinobacillus pleuropneumoniae From Clinical Samples Using Recombinase Polymerase Amplification |
title_short | Rapid Detection of Actinobacillus pleuropneumoniae From Clinical Samples Using Recombinase Polymerase Amplification |
title_sort | rapid detection of actinobacillus pleuropneumoniae from clinical samples using recombinase polymerase amplification |
topic | Veterinary Science |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8990124/ https://www.ncbi.nlm.nih.gov/pubmed/35400111 http://dx.doi.org/10.3389/fvets.2022.805382 |
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