PARATRANSGENESIS FOR CONTROL OF PIERCE'S DISEASE: MANIPULATION OF ENDOPHYTIC BACTERIA FOR PARATRANSGENIC CONTROL OF PIERCE'S DISEASE

• Author(s): Lampe, David; Miller, Thomas; Bextine, Blake; Cooksey, Donald; Lauzon, Carol;
• Abstract: The Glassy-Winged Sharpshooter (GWSS) is the principal vector of Xylella fastidiosa, the causative agent of Pierce's disease, in the California wine country. One strategy to control the spread of Xylella by the GWSS is to make the insect refractory to transmission of the bacterium. One can imagine at least two ways this can be done. The first is to genetically engineer the population of sharpshooters directly to become refractory. This transgenic method is being seriously discussed by biologists wishing to halt the spread of malaria in Africa, via the direct genetic manipulation of Anopheles gambiae populations (Kiszewski and Spielman 1998). This method is fraught with potential difficulties of many sorts and, in any case, has not been tried to date. The second method is to manipulate the insect vector indirectly by manipulating its gut flora. This technique is termed paratransgenesis and has many potential advantages over the direct transgenic approach. First, bacteria are far easier to manipulate genetically. Secondly, bacteria can be made to secrete or carry very specific agents of control, like single chain antibodies. Paratransgenesis has been attempted to control Chagas' disease in S. America (Beard et al. 2001) and a form of it is being developed to deliver therapeutic agents in mouse models of human disease, for eventual applications for humans (Beninati et al. 2000). In attempting to create transgenic gut symbionts of the GWSS several problems present themselves immediately. The final transgenic strain will need to be stable (i.e., the exogenous DNA not contained in a virus or bacterial transposon), the exogenous DNA should be incorporated into the chromosome and not borne on a plasmid, no drug markers should be left in the strain, and as little exogenous DNA should be transferred as possible. We have developed a genetic modification system that meets those requirements based on the mariner family of eukaryotic transposable elements. These elements are active in all domains of life when appropriately manipulated, but do not occur naturally in prokaryotes. Thus stable strains of GWSS gut symbionts can be created that should be suitable for release into the wild for the control of X. fastidiosa.
• Publication Date: Dec 2002
• Journal: 2002 Pierce's Disease Research Symposium