Interpretation of the EC method for the detection of latent Corynebacterium sepedonicum infections in potato¹

Abstract

The EC mcthod for the detection of latent ring-rot infections (Corynebacterium sepedonicum) consists essentially of an indirect immunofluorescence test and a pathogenicity test on eggplant (EP). When interpreting the results of this method, care should be taken that: (a) eggplants iire grown at 2 1 "C. At 28-29°C the detection level ofthe EP was increased 10'to lo3fold to lo5 to lo6 cells m1-I and latent infections (which may go unnoticed) occurred with lo4 to lo5 cells nil-'; (b) reisolations are made from symptomless eggplants grown at the optimum temperatuie (21°C) as latent infections can also occur in these plants at concentrations of lo2 to 10' cells inl-'; (c) potato tuber extracts are tested without freezing or stored freeze-dried or in a proper protective agent. Freezing at -20°C in maceration buffer of cells of C. sepedonicutii and Pseudomonas solanacearum (the latter was included for comparison) reduced their numbers by 90-98"/0 after one day. This could easily cause viable cell numbers to drop below the detection level of the pathogenicity test.

Potato brown rot in western Europe – history, present occurrence and some remarks on possible origin, epidemiology and control strategies

“…The minimum growth temperature is 10-12°C. Survival at -20°C in simple buffer solution without cryoprotectant was 100 days, where > 90% of the bacteria died in the first few days, and 180 days when protected in tissue extract (Janse & Van Vaerenbergh, 1987). Survival at -20°C in simple buffer solution without cryoprotectant was 100 days, where > 90% of the bacteria died in the first few days, and 180 days when protected in tissue extract (Janse & Van Vaerenbergh, 1987).…”

Development of a semi-selective medium and an immunofluorescence colony-staining procedure for the detection of Clavibacter michiganensis subsp. sepedonicus in cattle manure slurry

“…The general failure to isolate Cms from natural substrates like soil or heel ends of symptomless potato tubers is the result of the strong dominance of naturally occurring saprophytes on the isolation medium (Janse and Van Vaerenbergh, 1987). The compe-tition for essential nutrients between Cms and fast growing saprophytes seems the most likely explanation for the extensive inhibition of Cms, even on media specially developed for Cms such as YGM.…”

“…If these colonies are present, isolate from Cms-type colony by puncturing the colony with a fine needle or glass capillary, followed by streaking on YGM to obtain a pure culture. 4) Identify the pure culture with IF-cell staining, fatty acid profiling, and or eggplant inoculation (Janse and Van Vaerenbergh, 1987;Lelliot and Stead, 1987). If many Cms-type colonies are present and quantification is needed perform IFC on representative areas.…”

“…Confirmation of IF-results was considered necessary because crossreacting organisms are known to occur in potato tuber samples (Miller, 1984;Janse and Van Vaerenbergh, 1987). Isolation procedures in general failed for this purpose and therefore a sensitive but time consuming pathogenicity test is now used for confirmation of IF-positive samples.…”

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“…Systematic and careful control of potato seed with the aim to eliminate Cms from seed potato production has been adopted in the Czech Republic. Bacteriological analyses of potato samples have been conducted according to the above mentioned directive of EU, where the immunofluorescence (IF) test and test of pathogenicity on eggplant were used to confirm Cms in potato tubers (JANSE & VAN VAERENBERGH 1987).…”