Journal of Economic Entomology (2004) 97, 757-763

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Blake Bextine, Shu-Jen Tuan, Harris Shaikh, Matthew Blua and T.A. Miller (2004)
Evaluation of methods for extracting Xylella fastidiosa DNA from the glassy-winged sharpshooter
Journal of Economic Entomology 97 (3), 757-763
Abstract: The recent spread of the plant pathogenic bacterium Xylella fastidiosa Wells et al. by an invasive vector species, Homalodisca coagulata Say, in southern California has resulted in new epidemics of Pierce's disease of grapevine. Our goal is to develop an efficient method to detect low titers of X. fastidiosa in H. coagulata that is amenable to large sample sizes for epidemiological studies. Detection of the plant pathogenic bacterium X. fastidiosa in its insect vector is complicated by low titers of bacteria, difficulty in releasing it from the insect mouthparts and foregut, and the presence of substances in the insect that inhibit polymerase chain reaction (PCR). To select the optimal protocol for DNA extraction to be used with PCR, we compared three standard methods and 11 commercially available kits for relative efficiency of X. fastidiosa DNA extraction in the presence of insect tissue. All of the protocols tested were proficient at extracting DNA from pure bacterial culture (1 × 105 cells), and all but one protocol successfully extracted sufficient bacterial DNA in the presence of insect tissue. Three DNA extraction techniques, immunomagnetic separation, the DNeasy Tissue kit (Qiagen, Hercules, CA), and Genomic DNA Purification kit (Fermentus, Hanover, MD), were compared more closely using a dilution series of X. fastidiosa (5,000-0 cells) with and without insect tissue present. The DNeasy Tissue kit was the best kit tested, allowing detection of 5 × 103 X. fastidiosa cells with an insect head background.
(The abstract is excluded from the Creative Commons licence and has been copied with permission by the publisher.)
Link to article at publishers website
Database assignments for author(s): Blake Bextine, Matthew J. Blua, Thomas A. Miller

Research topic(s) for pests/diseases/weeds:
transmission/dispersal of plant diseases
surveys/sampling/distribution


Pest and/or beneficial records:

Beneficial Pest/Disease/Weed Crop/Product Country Quarant.


Xylella fastidiosa Grapevine (Vitis) U.S.A. (SW)
Homalodisca vitripennis U.S.A. (SW)