Use of FTA Technology to Extract Wheat streak mosaic virus and Candidatus Liberibacter Solanacearum from Single Vectors
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Detection and quantification of plant pathogens is a fundamental component of plant disease epidemiology. For vector-borne diseases, pathogen detection and quantification within the vector is often necessary to understand development and spread of disease. However, extraction and testing of pathogens from a single vector can be expensive and tedious, especially with large numbers of samples. Therefore, to simplify sample collection and reduce cost of analysis, we used a new nucleic acid elution approach using existing Flinders Technology Associates (FTA) technology from Whatman GE Healthcare. We applied this method in two very distinct vector-borne pathogen systems for Wheat streak mosaic virus (Tritimovirus) (McKinney 1937) and Candidatus Liberibacter solanacearum (Liefting et al. 2009). Total DNA and RNA were extracted from adult and nymphal psyllids (family Psyllidae) and wheat curl mite, Aceria tosichella Keifer, respectively using FTA cards (Whatman GE Health care, Pittsburgh, PA), and results were compared to extraction with DNeasy/RNeasy® kits (Qiagen Inc., Valencia, CA). Additionally, composite samples of multiple mite and psyllid vectors were used to compare the efficiencies of both methods. FTA elution and kit extractions were significantly different (P < 0.001). Slightly lower cycle threshold (Ct) values of 27.8 ± 0.19 and 25.5 ± 0.17 and 29.8 ± 0.09 and 27.45 ± 0.05 were determined for the composite adult and nymphal psyllids for the FTA and kit extractions, respectively. Wheat curl mite Ct values averaged 32.5 ± 0.75 and 27.6 ± 1.5 for the FTA elution and kit extractions, respectively. Coefficient of variation values were found to be lower for the FTA elution method when compared to the kit extractions for both the adult and nymphal psyllid (0.7 and 0.9%) and wheat curl mite (2.3 and 5.5%). Despite lower Ct values the FTA elution method is more cost efficient, protects nucleic acid integrity, and proved to be a valuable and reliable tool for detection and comparative studies of both pathogens within a single-vector sample.
author list (cited authors)
Price, J. A., Simmons, A., Bass, J., & Rush, C. M.