Non‐lethal loop‐mediated isothermal amplification assay as a point‐of‐care diagnostics tool for Neoparamoeba perurans, the causative agent of amoebic gill disease
Neoparamoeba perurans is the causative agent of amoebic gill disease (AGD). Two loop‐mediated isothermal amplification (LAMP) assays targeting the parasite 18S rRNA and the Atlantic salmon EF1α, used as internal control, were designed. The N. perurans LAMP assay did not amplify close relatives N. pe...
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Online Access: | http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7383609/ http://www.ncbi.nlm.nih.gov/pubmed/32364315 https://doi.org/10.1111/jfd.13175 |
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ftpubmed:oai:pubmedcentral.nih.gov:7383609 2023-05-15T15:31:34+02:00 Non‐lethal loop‐mediated isothermal amplification assay as a point‐of‐care diagnostics tool for Neoparamoeba perurans, the causative agent of amoebic gill disease Cano, Irene McCullough, Robin Mulhearn, Brian Gunning, Susie Waine, Ava Joiner, Claire Paley, Richard 2020-05-04 http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7383609/ http://www.ncbi.nlm.nih.gov/pubmed/32364315 https://doi.org/10.1111/jfd.13175 en eng John Wiley and Sons Inc. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7383609/ http://www.ncbi.nlm.nih.gov/pubmed/32364315 http://dx.doi.org/10.1111/jfd.13175 © 2020 Crown copyright. Journal of Fish Diseases published by John Wiley & Sons Ltd. This article is published with the permission of the Controller of HMSO and the Queen’s Printer for Scotland. This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. CC-BY J Fish Dis Original Articles Text 2020 ftpubmed https://doi.org/10.1111/jfd.13175 2020-08-02T00:32:06Z Neoparamoeba perurans is the causative agent of amoebic gill disease (AGD). Two loop‐mediated isothermal amplification (LAMP) assays targeting the parasite 18S rRNA and the Atlantic salmon EF1α, used as internal control, were designed. The N. perurans LAMP assay did not amplify close relatives N. pemaquidensis and N. branchiphila, or the host DNA. This assay detected 10(6) copies of the parasite 18S rRNA gene under 13 min and 10(3) copies under 35 min. Five “fast‐and‐dirty” DNA extraction methods were compared with a reference method and further validated by TaqMan™ qPCR. Of those, the QuickExtract buffer was selected for field tests. Seventy‐one non‐lethal gill swabs were analysed from AGD‐clinically infected Atlantic salmon. The pathogen was detected under 23 min in fish of gill score >2 and under 39 min for lower gill scores. About 1.6% of the tests were invalid (no amplification of the internal control). 100% of positives were obtained from swabs taken from fish showing gill score ˃3, but only ~50% of positives for lower gill scores. The present LAMP assay could be implemented as a point‐of‐care test for the on‐site identification of N. perurans; however, further work is required to improve its performance for lower scores. Text Atlantic salmon PubMed Central (PMC) Journal of Fish Diseases 43 7 779 790 |
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Original Articles |
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Original Articles Cano, Irene McCullough, Robin Mulhearn, Brian Gunning, Susie Waine, Ava Joiner, Claire Paley, Richard Non‐lethal loop‐mediated isothermal amplification assay as a point‐of‐care diagnostics tool for Neoparamoeba perurans, the causative agent of amoebic gill disease |
topic_facet |
Original Articles |
description |
Neoparamoeba perurans is the causative agent of amoebic gill disease (AGD). Two loop‐mediated isothermal amplification (LAMP) assays targeting the parasite 18S rRNA and the Atlantic salmon EF1α, used as internal control, were designed. The N. perurans LAMP assay did not amplify close relatives N. pemaquidensis and N. branchiphila, or the host DNA. This assay detected 10(6) copies of the parasite 18S rRNA gene under 13 min and 10(3) copies under 35 min. Five “fast‐and‐dirty” DNA extraction methods were compared with a reference method and further validated by TaqMan™ qPCR. Of those, the QuickExtract buffer was selected for field tests. Seventy‐one non‐lethal gill swabs were analysed from AGD‐clinically infected Atlantic salmon. The pathogen was detected under 23 min in fish of gill score >2 and under 39 min for lower gill scores. About 1.6% of the tests were invalid (no amplification of the internal control). 100% of positives were obtained from swabs taken from fish showing gill score ˃3, but only ~50% of positives for lower gill scores. The present LAMP assay could be implemented as a point‐of‐care test for the on‐site identification of N. perurans; however, further work is required to improve its performance for lower scores. |
format |
Text |
author |
Cano, Irene McCullough, Robin Mulhearn, Brian Gunning, Susie Waine, Ava Joiner, Claire Paley, Richard |
author_facet |
Cano, Irene McCullough, Robin Mulhearn, Brian Gunning, Susie Waine, Ava Joiner, Claire Paley, Richard |
author_sort |
Cano, Irene |
title |
Non‐lethal loop‐mediated isothermal amplification assay as a point‐of‐care diagnostics tool for Neoparamoeba perurans, the causative agent of amoebic gill disease |
title_short |
Non‐lethal loop‐mediated isothermal amplification assay as a point‐of‐care diagnostics tool for Neoparamoeba perurans, the causative agent of amoebic gill disease |
title_full |
Non‐lethal loop‐mediated isothermal amplification assay as a point‐of‐care diagnostics tool for Neoparamoeba perurans, the causative agent of amoebic gill disease |
title_fullStr |
Non‐lethal loop‐mediated isothermal amplification assay as a point‐of‐care diagnostics tool for Neoparamoeba perurans, the causative agent of amoebic gill disease |
title_full_unstemmed |
Non‐lethal loop‐mediated isothermal amplification assay as a point‐of‐care diagnostics tool for Neoparamoeba perurans, the causative agent of amoebic gill disease |
title_sort |
non‐lethal loop‐mediated isothermal amplification assay as a point‐of‐care diagnostics tool for neoparamoeba perurans, the causative agent of amoebic gill disease |
publisher |
John Wiley and Sons Inc. |
publishDate |
2020 |
url |
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7383609/ http://www.ncbi.nlm.nih.gov/pubmed/32364315 https://doi.org/10.1111/jfd.13175 |
genre |
Atlantic salmon |
genre_facet |
Atlantic salmon |
op_source |
J Fish Dis |
op_relation |
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7383609/ http://www.ncbi.nlm.nih.gov/pubmed/32364315 http://dx.doi.org/10.1111/jfd.13175 |
op_rights |
© 2020 Crown copyright. Journal of Fish Diseases published by John Wiley & Sons Ltd. This article is published with the permission of the Controller of HMSO and the Queen’s Printer for Scotland. This is an open access article under the terms of the http://creativecommons.org/licenses/by/4.0/ License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. |
op_rightsnorm |
CC-BY |
op_doi |
https://doi.org/10.1111/jfd.13175 |
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Journal of Fish Diseases |
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43 |
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7 |
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779 |
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790 |
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1766362091272273920 |