A Rapid Diagnostic for Detection of Aphelenchoides besseyi and A. fujianensis Based on Real-Time PCR
Aphelenchoides besseyi and A. fujianensis have been frequently found in mixed populations associated with forage grass seed in Brazil. The morphological similarity between both species has previously led A. fujianensis to be erroneously identified as A. besseyi. A. besseyi is a quarantine pest in ma...
| Veröffentlicht in: | Plant disease. - 1997. - 102(2018), 3 vom: 01. März, Seite 519-526 |
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| 1. Verfasser: | |
| Weitere Verfasser: | , , , , |
| Format: | Online-Aufsatz |
| Sprache: | English |
| Veröffentlicht: |
2018
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| Zugriff auf das übergeordnete Werk: | Plant disease |
| Schlagworte: | Journal Article Research Support, Non-U.S. Gov't Validation Study DNA Primers DNA, Helminth DNA, Ribosomal |
| Zusammenfassung: | Aphelenchoides besseyi and A. fujianensis have been frequently found in mixed populations associated with forage grass seed in Brazil. The morphological similarity between both species has previously led A. fujianensis to be erroneously identified as A. besseyi. A. besseyi is a quarantine pest in many countries that import Brazilian forage seed; however, there is no current evidence suggesting that A. fujianensis is a plant-parasitic species. Two real-time polymerase chain reaction (qPCR) diagnostics were developed to detect each species and an operational envelope was established. A set of primers and hydrolysis probes for each species was designed targeting the large subunit (LSU) region. To assess their specificity, primers and probes sets were tested with samples of nontarget Aphelenchoides and Paraphelenchus sp. also frequently associated with forage seed. Experiments using dilutions of purified plasmid standards underpinned the sensitivity of the qPCR assays, which detected as few as 10 copies of target nematode ribosomal DNA. Thus, the developed diagnostics were sufficiently sensitive to detect DNA extracted from a fragment of a single target nematode. There was a positive correlation between copy number of the target species and nematode abundance, suggesting the potential of this method for quantification. Evidence of intra-individual variability among cloned sequences of the LSU region in a single A. besseyi population is also reported |
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| Beschreibung: | Date Completed 14.02.2019 Date Revised 10.12.2019 published: Print-Electronic Citation Status MEDLINE |
| ISSN: | 0191-2917 |
| DOI: | 10.1094/PDIS-08-17-1160-RE |