ARS | Publication request: Beta 1, 4-glucanase in Glassy-Winged Sharpshooter Saliva, and its Possible Role in Infection and Movement of X. fastidiosa

Submitted to: CDFA Pierce's Disease Control Program Research Symposium
Publication Type: Proceedings
Publication Acceptance Date: September 4, 2007
Publication Date: December 12, 2007
Repository URL: http://www.cdfa.ca.gov/pdcp/Research_Symposium_Details.html
Citation: Backus, E.A., Labavitch,, J.M. 2007. Beta 1, 4-glucanase in Glassy-Winged Sharpshooter Saliva, and its Possible Role in Infection and Movement of X. fastidiosa. In: Proceedings of the 2007 Pierce's Disease Research Symposium, December 12-14, 2007, San Diego, California. p. 120-122.

Interpretive Summary: The purpose of this project is to determine whether saliva of the glassy-winged sharpshooter (GWSS) aids establishment of Xylella fastidiosa (Xf) cells that are injected into a plant during GWSS feeding. If so, the bacteria would be located near or in watery, enzymatic saliva of the insect vector, suggesting that saliva is a carrier of the bacteria during inoculation. Fluorescent labeling of EGase, the major enzymatic protein in the saliva of GWSS, will be used as a marker for watery saliva, ordinarily invisible using normal microscopy methods. This year, we: 1) completed development of a method to prepare and microscopically examine grape petioles, into which GWSS had injected Xf cells that are transformed to express green fluorescent protein (GFP), 2) dissected an additional 1500 pairs of salivary glands for protein extraction, 3) extracted and purified EGase, and 4) contracted with a commercial company to produce antibodies to EGase. Work in the coming year will complete the objectives, using confocal laser scanning microscopy to produce images of the antibody fluorescence-stained EGase and GFP-expressing Xf. A finding that Xf cells are carried by saliva would allow development of several new disease management tactics, such as engineering plants to break down or prevent the action of the saliva. In this way, initial infection could be prevented.

Technical Abstract: The purpose of this project is to determine whether 1, 4-glucanase (EGase), the major enzymatic protein in watery saliva of glassy-winged sharpshooter (GWSS) co-localizes via immunocytochemistry with the few ‘pioneer’ Xylella fastidiosa (Xf) cells that are inoculated into a plant by this vector. If it does, then this suggests that watery, enzymatic saliva of the vector is a carrier of bacteria during inoculation, and that saliva might somehow aid in this process. This year, we: 1) completed development of a method to retain fluorescence of Xf cells that are transformed to express green fluorescent protein (GFP) through the paraffin-sectioning process, 2) dissected an additional 1500 pairs of salivary glands for protein extraction, 3) extracted and purified EGase, and 4) contracted with a commercial company to produce antibodies to EGase. Work in the coming year will complete the objectives by using secondary antibodies to EGase and confocal laser scanning microscopy to co-localize the EGase with GFP Xf.