ARS | Publication request: Competitive binding influences Xf vector load: Confocal and SEM images of GFP-expressing Xf in GWSS foreguts

Submitted to: Symposium Proceedings
Publication Type: Proceedings
Publication Acceptance Date: June 22, 2007
Publication Date: July 18, 2007
Repository URL: http://ucce.ucdavis.edu/files/filelibrary/6274/40698.pdf
Citation: Backus, E.A. 2007. Competitive binding influences Xf vector load: Confocal and SEM images of GFP-expressing Xf in GWSS foreguts. In: Proceedings of the National Viticulture Research Conference, July 9-11, 2007, Davis, CA. p. 11-12.

Interpretive Summary: Glassy-winged sharpshooters, Homalodisca vitripennis, like all insects that transmit plant pathogens, acquire Xylella fastidiosa bacteria (causative agent of Pierce’s Disease of grape) by swallowing grape xylem fluids containing bacteria. X. fastidiosa then grows in regions of the vector’s foregut near the mouth parts. Past research has shown that these bacteria are dislodged from the foregut during feeding, and are expelled (inoculated) directly into the plant, but the exact process is not understood. We have now developed a confocal microscopy technique to view X. fastidiosa cells tranformed to express green fluorescent protein (GFP), inside the foregut. Using this technique, a preliminary comparison was made of the location and size of bacterial colonies in the foreguts of lab-reared vs. field-collected GWSS, before acquisition of GFP-X. fastidiosa, after acquisition, and after inoculation. Large colonies of GFP-X. fastidiosa were seen in foreguts of lab-reared GWSS compared to those from field-collected GWSS. Lab-reared GWSS also inoculated many more GPF-X. fastidiosa than did field-collected GWSS. Thus, sharpshooters in the field have competing microbes which take up critical space in the foregut before the GFP-X. fastidiosa can be acquired, leading to reduced acquisition and inoculation of X. fastidiosa. Biological control of X. fastidiosa transmission might be possible in the future using competing microbes.

Technical Abstract: Like all hemipteran vectors of plant pathogens, glassy-winged sharpshooters, Homalodisca vitripennis acquire their load of Xylella fastidiosa (Xf ) via ingestion of fluids containing bacteria. X. fastidiosa then colonizes the anterior regions of the vector’s foregut, the cibarium (or sucking pump) and precibarium. Properties of Xf inoculation support that bacteria are dislodged from the foregut during feeding, and are expelled directly into the plant during inoculation, but the exact process is not understood. We have now developed a confocal microscopy technique to view Xf cells tranformed to express green fluorescent protein (GFP). Using this technique, a preliminary comparison (n = 6) of foregut colonization of lab-reared vs. field-collected GWSS, before acquisition of GFP-X. fastidiosa, after acquisition, and after inoculation. For acquistion, insects were allowed to feed for 7 – 8 days on symptomatic, GFP-X. fastidiosa infected grape. For inoculation, acquired insects were allowed to make a few probes on either healthy grapevines or artificial diet; discharge of Xf was later confirmed by confocal microscopy and/or PCR. Microbial colonization of foreguts of ‘clean’, lab-reared GWSS differed greatly from ‘dirty’ field-collected GWSS. Clean GWSS acquired many more GPF-Xf than did dirty GWSS. Preliminary, post-acquisition inoculation assays with these insects suggests that originally clean, ‘maximally loaded’ GWSS inoculate far more Xf than do originally dirty, ‘topped off’ GWSS. Thus, both acquisition and inoculation of X. fastidiosa appear to be influenced by availability of microbial binding sites for Xf within the precibarium.