Effect of specific elicitors of Cladosporium fulvum on tomato suspension cells: evidence for the involvement of active oxygen species

Intercellular fluid (IF) obtained from tomato (Lycopersicon esculentum L.) leaflets colonized by Cladosporium fulvum Cooke contains specific elicitors that induce necrosis in tomato cultivars resistant to the race of C. fulvum used to produce the IF. The responses of cell-suspension cultures produced from tomato lines near-isogenic for resistance genes Cf 4 and Cf 5 to IF produced from leaves infected by races 4 (virulent on Cf 4 but not Cf 5 plants), 2.4.5, and 2.4.5.9 (both virulent on Cf 4 and Cf 5 plants) were used to investigate the possibility that active oxygen (AO) species were involved in the initial host reaction to these elicitors. Concurrently, the same assays were used to determine if the cell lines retained the elicitor specificity of the original plants. An IF/cell combination that gives an incompatible reaction in leaves (race 4 IF and Cf 5 cells) showed reduced oxygen uptake and increases in malonaldehyde (a product of lipid peroxidation); cytochrome c reducing activity, which was inhibited by superoxide dismutase (SOD) (an assay for superoxide); luminol-dependent chemiluminescence (an assay for several AO species); activity of extracellular peroxidases; and extracellular phenolic compounds. In contrast, compatible combinations (IF from races 2.4.5 or 2.4.5.9 and Cf 4 or Cf 5 cells; race 4 IF and Cf 4 cells) did not exhibit any of these changes. The addition of catalase, SOD, ascorbate (a scavenger of superoxide), mannitol (a scavenger of the hydroxyl radical), KCN, or salicyl hydroxamic acid (both inhibitors of peroxidases) prior to IF treatment reduced the IF-induced increases in malonaldehyde and extracellular phenolics. Catalase was an effective inhibitor of the IF-induced changes in oxygen uptake and cytochrome c reducing activity. These results demonstrate the specificity of the IF-induced cell responses and confirm that AO species are involved in the initial cell response