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Genome-Informed Recombinase Polymerase Amplification Assay Coupled with a Lateral Flow Device for In-Field Detection of Dickeya Species
- Source :
- Plant Disease. 104:2217-2224
- Publication Year :
- 2020
- Publisher :
- Scientific Societies, 2020.
-
Abstract
- Dickeya spp. cause blackleg and soft rot diseases of potato and several other plant species worldwide, resulting in high economic losses. Rapid detection and identification of the pathogen is essential for facilitating efficient disease management. Our aim in this research was to develop a rapid and field-deployable recombinase polymerase amplification (RPA) assay coupled with a lateral flow device (LFD) that will accurately detect Dickeya spp. in infected plant tissues without the need for DNA isolation. A unique genomic region (mglA/mglC genes) conserved among Dickeya spp. was used to design highly specific robust primers and probes for an RPA assay. Assay specificity was validated with 34 representative strains from all Dickeya spp. and 24 strains from other genera and species; no false positives or negatives were detected. An RPA assay targeting the internal transcribed spacer region of the host genome was included to enhance the reliability and accuracy of the Dickeya assay. The detection limit of 1 fg was determined by both sensitivity and spiked sensitivity assays; no inhibitory effects were observed when 1 µl of host sap, macerated in Tris-EDTA buffer, was added to each reaction in the sensitivity tests. The developed RPA assay is rapid, highly accurate, sensitive, and fully field deployable. It has numerous applications in routine diagnostics, surveillance, biosecurity, and disease management.
- Subjects :
- 0106 biological sciences
0301 basic medicine
biology
Blackleg
Recombinase Polymerase Amplification
Dickeya
Plant Science
biology.organism_classification
01 natural sciences
Genome
DNA extraction
Microbiology
03 medical and health sciences
030104 developmental biology
Internal transcribed spacer
Agronomy and Crop Science
Gene
Pathogen
010606 plant biology & botany
Subjects
Details
- ISSN :
- 19437692 and 01912917
- Volume :
- 104
- Database :
- OpenAIRE
- Journal :
- Plant Disease
- Accession number :
- edsair.doi...........364f0eaad457c80fb786ef0a6e6e3d5e