It is well established that the pectinolytic bacteria Pectobacterium atrosepticum (Pca) and Dickeya spp. are causal organisms of blackleg in potato. In temperate climates, the role of Pectobacterium carotovorum subsp. carotovorum (Pcc) in potato blackleg, however, is unclear. In different western and central European countries plants are frequently found with blackleg from which only Pcc can be isolated, but not Pca or Dickeya spp. Nevertheless, tubers vacuum-infiltrated with Pcc strains have so far never yielded blackleg-diseased plants in field experiments in temperate climates. In this study, it is shown that potato tubers, vacuum-infiltrated with a subgroup of Pcc strains isolated in Europe, and planted in two different soil types, can result in up to 50% blackleg diseased plants. All three pathogens can cause tuber rot during storage, whereas infections of seed potatoes with Pca and Dickeya spp. can also result in the occurrence of various field symptoms, including reduced emergence, chlorosis, wilting, tuber and stem rot, blackleg, haulm desiccation and plant death (Pérombelon and Kelman 1980;Pérombelon 1992;Fraaije et al. 1996;Helias et al. 2000). These symptoms can be caused by Pca as well as by Dickeya spp., making identification of the causal organism by visual observation in the field unreliable. Regulatory field inspections in seed potatoes in many countries are therefore based on recognition of this 'blackleg complex' without discriminating between the various blackleg-causing organisms.

Studies were conducted to explain the relative success of 'Dickeya solani', a genetic clade of Dickeya biovar 3 and a blackleg-causing organism that, after recent introduction, has spread rapidly in seed potato production in Europe to the extent that it is now more frequently detected than D. dianthicola. In vitro experiments showed that both species were motile, had comparable siderophore production and pectinolytic activity, and that there was no antagonism between them when growing. Both 'D. solani' and biovar 1 and biovar 7 of D. dianthicola rotted tuber tissue when inoculated at a low density of 10 3 CFU mL. In an agar overlay assay, D. dianthicola was susceptible to 80% of saprophytic bacteria isolated from tuber extracts, whereas 'D. solani' was susceptible to only 31%, suggesting that 'D. solani' could be a stronger competitor in the potato ecosystem. In greenhouse experiments at high temperatures (28°C), roots were more rapidly colonized by 'D. solani' than by biovar 1 or 7 of D. dianthicola and at 30 days after inoculation higher densities of 'D. solani' were found in stolons and progeny tubers. In co-inoculated plants, fluorescent protein (GFP or DsRed)-tagged 'D. solani' outcompeted D. dianthicola in plants grown from vacuum-infiltrated tubers. In 3 years of field studies in the Netherlands with D. dianthicola and 'D. solani', disease incidence varied greatly annually and with strain. In summary, 'D. solani' possesses features which allow more efficient plant colonization than D. dianthicola at high temperatures. In temperate climates, however, tuber infections with 'D. solani' will not necessarily result in a higher disease incidence than infections with D. dianthicola, but latent seed infection could be more prevalent.

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The real-time RT-PCR protocol of Boonham and coworkers performed extremely well in a recent ring test, comparing different methods for detection of Potato spindle tuber pospiviroid (PSTVd) in several laboratories. Since, in addition, real-time PCR technology has proved suitable for highthroughput testing, this method was chosen as the starting point for the development of a protocol for the large-scale testing of potato. The initial experiments focused on the specificity of the primers and probes with regard to different isolates of PSTVd and other (pospi-) viroids. Further experiments were performed with leaf material from both primarily and secondarily infected plants. The parameters studied were sampling position, growing-on temperature and bulking rate. In addition, different grinding, nucleic-acid extraction and disinfection methods were compared. To monitor false negatives and positives, different controls were included and tested in duplex and triplex formats. The final protocol was tested using a hundred samples from the Dutch potato-monitoring programme. The results of this pilot experiment were promising. Future plans include the development of a protocol for direct tuber testing and inter-laboratory ring testing of the protocols.