Sensitive, Accurate and Rapid Detection of the Northern Root-Knot Nematode, <i>Meloidogyne hapla</i>, Using Recombinase Polymerase Amplification Assays
Rapid and reliable diagnostics of root-knot nematodes are critical for selections of effective control against these agricultural pests. In this study, recombinase polymerase amplification (RPA) assays were developed targeting the IGS rRNA gene of the northern root-knot nematode, <i>Meloidogyn...
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| Format: | Article |
| Language: | English |
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MDPI AG
2021-02-01
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| Series: | Plants |
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| Online Access: | https://www.mdpi.com/2223-7747/10/2/336 |
| _version_ | 1797411985014390784 |
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| author | Sergei A. Subbotin Julie Burbridge |
| author_facet | Sergei A. Subbotin Julie Burbridge |
| author_sort | Sergei A. Subbotin |
| collection | DOAJ |
| description | Rapid and reliable diagnostics of root-knot nematodes are critical for selections of effective control against these agricultural pests. In this study, recombinase polymerase amplification (RPA) assays were developed targeting the IGS rRNA gene of the northern root-knot nematode, <i>Meloidogyne hapla.</i> The RPA assays using TwistAmp<sup>®</sup> Basic, TwistAmp<sup>®</sup> exo and TwistAmp<sup>®</sup> nfo kits (TwistDx, Cambridge, UK) allowed for the detection of <i>M. hapla</i> from crude extracts of females, eggs and juveniles without a DNA extraction step. The results of the RPA assays using real-time fluorescence detection (real-time RPA) in series of crude nematode extracts showed reliable detection after 13 min with a sensitivity of 1/100 of a second-stage juvenile and up to 1/1000 of a female in reaction tubes. The results of the RPA assays using lateral flow dipsticks (LF-RPA) showed reliable detection within 30 min with a sensitivity of 1/10 of a second-stage juvenile and 1/1000 of a female in reaction tubes. The RPA assay developed here is a successful tool for quick, accurate and sensitive diagnostics of <i>M. hapla</i>. The application of the LF-RPA assay has great potential for diagnosing infestation of this species in the lab, field or in areas with a minimal laboratory infrastructure. |
| first_indexed | 2024-03-09T04:54:27Z |
| format | Article |
| id | doaj.art-c2eed1c8e3c1445987ba7ee185d5ce91 |
| institution | Directory Open Access Journal |
| issn | 2223-7747 |
| language | English |
| last_indexed | 2024-03-09T04:54:27Z |
| publishDate | 2021-02-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Plants |
| spelling | doaj.art-c2eed1c8e3c1445987ba7ee185d5ce912023-12-03T13:07:54ZengMDPI AGPlants2223-77472021-02-0110233610.3390/plants10020336Sensitive, Accurate and Rapid Detection of the Northern Root-Knot Nematode, <i>Meloidogyne hapla</i>, Using Recombinase Polymerase Amplification AssaysSergei A. Subbotin0Julie Burbridge1Plant Pest Diagnostic Center, California Department of Food and Agriculture, 3294 Meadowview Road, Sacramento, CA 95832, USAPlant Pest Diagnostic Center, California Department of Food and Agriculture, 3294 Meadowview Road, Sacramento, CA 95832, USARapid and reliable diagnostics of root-knot nematodes are critical for selections of effective control against these agricultural pests. In this study, recombinase polymerase amplification (RPA) assays were developed targeting the IGS rRNA gene of the northern root-knot nematode, <i>Meloidogyne hapla.</i> The RPA assays using TwistAmp<sup>®</sup> Basic, TwistAmp<sup>®</sup> exo and TwistAmp<sup>®</sup> nfo kits (TwistDx, Cambridge, UK) allowed for the detection of <i>M. hapla</i> from crude extracts of females, eggs and juveniles without a DNA extraction step. The results of the RPA assays using real-time fluorescence detection (real-time RPA) in series of crude nematode extracts showed reliable detection after 13 min with a sensitivity of 1/100 of a second-stage juvenile and up to 1/1000 of a female in reaction tubes. The results of the RPA assays using lateral flow dipsticks (LF-RPA) showed reliable detection within 30 min with a sensitivity of 1/10 of a second-stage juvenile and 1/1000 of a female in reaction tubes. The RPA assay developed here is a successful tool for quick, accurate and sensitive diagnostics of <i>M. hapla</i>. The application of the LF-RPA assay has great potential for diagnosing infestation of this species in the lab, field or in areas with a minimal laboratory infrastructure.https://www.mdpi.com/2223-7747/10/2/336diagnosticsroot-knot nematoderecombinase polymerase amplification |
| spellingShingle | Sergei A. Subbotin Julie Burbridge Sensitive, Accurate and Rapid Detection of the Northern Root-Knot Nematode, <i>Meloidogyne hapla</i>, Using Recombinase Polymerase Amplification Assays Plants diagnostics root-knot nematode recombinase polymerase amplification |
| title | Sensitive, Accurate and Rapid Detection of the Northern Root-Knot Nematode, <i>Meloidogyne hapla</i>, Using Recombinase Polymerase Amplification Assays |
| title_full | Sensitive, Accurate and Rapid Detection of the Northern Root-Knot Nematode, <i>Meloidogyne hapla</i>, Using Recombinase Polymerase Amplification Assays |
| title_fullStr | Sensitive, Accurate and Rapid Detection of the Northern Root-Knot Nematode, <i>Meloidogyne hapla</i>, Using Recombinase Polymerase Amplification Assays |
| title_full_unstemmed | Sensitive, Accurate and Rapid Detection of the Northern Root-Knot Nematode, <i>Meloidogyne hapla</i>, Using Recombinase Polymerase Amplification Assays |
| title_short | Sensitive, Accurate and Rapid Detection of the Northern Root-Knot Nematode, <i>Meloidogyne hapla</i>, Using Recombinase Polymerase Amplification Assays |
| title_sort | sensitive accurate and rapid detection of the northern root knot nematode i meloidogyne hapla i using recombinase polymerase amplification assays |
| topic | diagnostics root-knot nematode recombinase polymerase amplification |
| url | https://www.mdpi.com/2223-7747/10/2/336 |
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