Detection and visualization of the major acidic exopolysaccharide of Ralstonia solancearum and its role in tomato root infection and vascular colonization

Exopolysaccharides play an important role in the pathogenicity of Ralstonia solanacearum. We compared in vitro and in planta exopolysaccharide product ion of the pathogenic strain AW1 with that of three related mutant strains impaired in both their exopolysaccharide production and aggressiveness on t omato. The distinction between the two hexosamine-rich exopolysaccharides, namely the N-acetyl-glucosaminorhamnan and the major N-acetylgalactosamine- containing acidic polymer was emphasized. The major acidic polymer was iden tified specifically by electron microscopy using glutaraldehyde/ruthenium r ed/uranyl acetate staining, by immunofluorescence using specific monoclonal antibodies and correlated to an appropriate biochemical analysis. The two mutant strains AW1-1 and AW-19A were totally devoid of any production of th e major exopolysaccharide in vitro or in planta whatever the technique used . Infection and vascular colonization of tomato roots by the pathogenic str ain were also compared to those of the mutant strains by light microscopy. Pathogenicity on tomato was assessed by root infection without any artifici al injury. Light microscopy showed that the two mutant strains AW1-1 and AW -19A were poorly infective and unable to invade xylem vessels, while they i nduced defence mechanisms in root tissues and appeared aggregated or degene rated within cortical infection pockets. These two mutant strains were non- pathogenic or weakly aggressive, respectively. In contrast, the pathogenic strain AW1 and the hypoaggressive AW1-41 strains, which produce large amoun ts of the major acidic exopolysaccharide in planta, were both infective and invasive, and tomato root tissues exhibited only limited defence reactions . Thus, the major acidic exopolysaccharide produced by Ralstonia solanacear um is involved in root infection and vascular colonization, though its prec ise role is still unknown.