Inactivation of Multiple Virulence Genes Reduces the Ability of Pseudomonas solanacearum to Cause Wilt Symptoms. Timothy P. Denny. Departments of Plant Pathology(1) and Microbiology(2), University of Georgia, Athens 30602 U.S.A. Brian F. Carney(1), and Mark A. Schell(1,2). Departments of Plant Pathology(1) and Microbiology(2), University of Georgia, Athens 30602 U.S.A. MPMI 3:293-300. Accepted 17 April 1990. Copyright 1990 The American Phytopathological Society.

Previous research on Pseudomonas solanacearum had shown that endoglucanase (encoded by egl), endopolygalacturonase (encoded by pglA), and extracellular polysaccharide (encoded by multiple eps genes) are each virulence factors involved in wilt disease of tomato. We have now constructed egl pglA and egl eps double mutants of P. solanacearum and compared them to the single mutants for production of extracellular enzymes in culture and for virulence by both stem and root inoculation procedures. The egl and egl eps mutants produced no endoglucanase activity, but unexpectedly overproduced polygalacturonase (PG) activity by 30 to 140%. Inactivation of pglA in either a wild-type or egl9::Tn5 background reduced total PG activity approximately 25 to 50%. Viscometry data indicated that the pglA mutants were deficient in endo-PG activity, but retained exo-PG activity. Virulence of the pglA and egl pglA mutants was not affected when 10⁶ bacteria were injected into stems of tomato plants or when 10⁹ bacteria were poured onto the soil; when stems were inoculated with 10⁴ bacteria of the mutants, the plants wilted 1 to 2 days slower than those inoculated with the parent strains. In contrast, inactivation of egl consistently reduced the ability of P. solanacearum to wilt tomato plants regardless of the strain, the type of assay, or the inoculum concentration tested. The egl eps mutant, which did not wilt plants in two-thirds of the stem inoculation experiments and appeared not to infect plants when added to the soil, was the least virulent of the defined mutants. The root inoculation assay was also used to show the close relationship between the appearance of wilt symptoms and reduced water use by the plant, strengthening the hypothesis that wilt is the result of vascular dysfunction.