Somatic cell selection with thaxtomin A as a positive selection agent was used to isolate variants of potato cv. Russet Burbank with strong to extreme resistance to common scab. Glasshouse and field trials identified 51 variants with significantly reduced disease incidence (frequency of infected tubers) and severity (tuber lesion coverage) compared with the parent cultivar. The most promising variants exhibited extreme disease resistance, rarely showing lesions, which were invariably superficial and shallower than those on the parent. Resistance traits were consistently expressed both in 10 glasshouse and two field trials at different locations, with varied inoculum and disease pressure. Disease-resistant variants differed in their response to thaxtomin A in tuber slice bioassays. Of 23 variants tested, 10 showed reduced thaxtomin A susceptibility, with the remaining 13 responding similar to that of the parent. Thus, toxin tolerance was not the only factor responsible for observed disease resistance; however, four of the five most disease-resistant variants had enhanced thaxtomin A tolerance, suggesting that this factor is important in the expression of strong disease resistance. Pathogenicity and toxin tolerance remained stable over a 6-year period, demonstrating that selected phenotypes were robust and genetic changes stable. The majority of disease-resistant variants had tuber yields equivalent to the parent cultivar in glasshouse trials. This suggests that selection for disease resistance was not associated with negative tuber attributes and that certain variants may have commercial merit, worthy of further agronomic testing.

Production of the phytotoxin thaxtomin A by pathogenic Streptomyces spp. is essential for induction of common scab disease in potato. Prior studies have shown that foliar application of sublethal concentrations of 2,4-dichlorophenoxyacetic acid (2,4-D) and other auxin or auxin-like compounds significantly reduced severity and occurrence of common scab in subsequently produced tubers. However, the means of disease suppression by these compounds was not known. We confirm the disease suppressive activity of 2,4-D. Detailed tuber physiological examination showed that lenticel numbers, lenticel external dimensions, and periderm thickness and structure, physiological features believed to be critical to Streptomyces scabiei infection, were not substantially changed by 2,4-D treatments, negating a possible mechanism for disease suppression through alteration of these structures. In contrast, our studies show accumulation of 2,4-D in tubers of treated plants occurs and is associated with an enhanced tolerance to thaxtomin A. Applying 2,4-D to cultures of S. scabiei did not significantly alter in vitro growth of the pathogen. Thaxtomin A production by the pathogen was inhibited by 2,4-D, but only at the highest rate tested (1.0 mM), which is at least 200-fold more than is found in 2,4-D treated tubers. These data suggest 2,4-D has no direct effect on the pathogen or its virulence. Confirmatory evidence from studies with Arabidopsis thaliana seedlings demonstrated that the auxins 2,4-D and IAA ameliorate thaxtomin A toxicity. The evidence presented whereby auxin treatment inhibits toxicity of thaxtomin A secreted by the pathogen suggests a novel indirect means of disease suppression.

Thaxtomin A, a key phytotoxin produced by plant pathogenic Streptomyces sp., is implicit in common scab disease expression in potato. Primary targets and modes of action of thaxtomin A toxicity in plant cells are not well understood. In this work, early signalling events associated with thaxtomin A toxicity were studied using the ion-selective microelectrode ion flux estimation (MIFE) technique. Thaxtomin A-induced changes in net ion fluxes were measured across the plasma membrane (PM) of root and pollen tube tissue in Arabidopsis thaliana and tomato. Within a minute after toxin application, a rapid and short-lived Ca2+ influx was observed. Well ahead of the marked inhibition of root growth, a significant shift towards net H+ efflux across the PM occurred in all tissues. Similar to root tissues, thaxtomin A significantly modified ion flux profiles from growing pollen tubes. Thaxtomin A was more effective in young, physiologically active tissues (root elongation zone or pollen tube apex), suggesting a higher density of thaxtomin A-binding sites in these regions. Overall, our data provide the first evidence that thaxtomin A triggers an early signalling cascade, which may be crucial in plant-pathogen interactions. It also suggests a possible interaction between thaxtomin A and PM auxin receptors, as revealed from experiments on the auxin-sensitive ucu2-2/gi2 A. thaliana mutant.

A somatic cell selection approach, developed using thaxtomin A as a positive selection agent, recovered 113 potato cell colonies of cv. Iwa from which 39 regenerated plants within a single subculture period. In glasshouse pathogenicity trials, 13 of these regenerated variants exhibited significantly greater resistance to common scab disease than the unselected parent cultivar, both in mean tuber surface coverage with lesions and the proportion of disease free tubers. The best variant had on average an 85-86% lower disease score (or 91-92% less estimated tuber surface cover) than the unselected parent cultivar. The disease-resistant variants varied in their response to thaxtomin A in detached leaflet and tuber slice bioassays. This suggested that the cell selection procedure and thaxtomin A screening had limited success in selecting for toxin tolerance, and that thaxtomin A tolerance was not necessarily the driving factor for induction of disease resistance observed. Variation in relative necrosis scores between leaflet and tuber assays for individual variants possibly suggests differential expression of thaxtomin A tolerance in the different tissues. Yield estimates from the glasshouse grown potatoes showed the majority of disease-resistant variants had tuber yields equivalent to the unselected parent cultivar. This suggests that selection for disease resistance was not associated with negative tuber yield attributes and that these variants may have commercial merit and are worthy of further agronomic testing.

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