SYMBIOTIC CONTROL OF PIERCES DISEASE: CONSTRUCTION OF TRANSGENIC STRAINS OF ALCALIGENES XYLOSOXIDANS DENITRIFICANS EXPRESSING SURFACE ANTI-XYLELLA FACTORS AS MICROBIAL PESTICIDES FOR PIERCES DISEASE CONTROL

• Author(s): Lampe, David; Miller, Thomas; Bextine, Blake; Cooksey, Donald; Lauzon, Carol; Lindow, Steven; Richards, Frank;
• Abstract: The glassy-winged sharpshooter (GWSS) is the principle vector of the xylem-limited bacterium Xylella fastidiosa (Xf), which causes Pierces disease (PD) in grapes. Limiting the spread of this pathogen by rendering GWSS incapable of pathogen transmission (paratransgenesis) is a promising method of pathogen control. Paratransgenesis seeks to modify the phenotype of an organism indirectly by modifying its symbiotic bacteria to confer vector-incompetence Paratransgenic approaches to disrupt pathogen infection of humans are being developed by several groups. These include interference with the ability of triatomid bugs to transmit pathogens causing Chagas disease 1, interference with HIV attachment to its target cells in the reproductive tracts of humans 2, and the elimination of persistent Candida infections from biofilms in chronically infected patients 3. Paratransgenesis has also been applied to deliver cytokines mammalian guts to relieve colitis 4; 5. Thus, the method has wide applicability. Alcaligenes xylosoxidans denitrificans (Axd) is Gram negative, beta proteobacterial species that can colonize the GWSS foregut and cibarium, as well as various plant tissues, including xylem. It is non-pathogenic in insects, plants and healthy humans. Given these characteristics, Axd has become the focus of our paratransgenesis efforts to control PD in grapes. Over the past two years we developed the technology to stably modify Axd by inserting genes into its chromosome and also isolated as single chain antibody that recognized an epitope on the surface of the Pierces Disease strain of Xf. 6. We report here the construction of strains of Axd that express an anti-Xylella single chain antibody (scFv) on the outer surface of Axd as fusions to three different heterologous outer membrane proteins. In each case, strains of varying fitness were recovered as measured by growth rate as compared to wild-type strains.
• Publication Date: Dec 2004
• Journal: 2004 Pierce's Disease Research Symposium