Four culture media were evaluated for their ability to induce Phytophthora palmivora growth and produce culture filtrate (CF), and to determine a CF concentration and culture period effective for in vitro screening of black rot resistance in Dendrobium cv. ‘Earsakul’. Mycelial fresh weights of P. palmivora cultured in potato dextrose broth (PDB; the most commonly used medium for fungi), pea sucrose broth (PSB; a medium frequently used for Phytophthora spp.), and Murashige and Skoog broth (MSB; the most popular plant tissue culture medium) were found to be significantly higher than that in the newly developed modified oat meal broth (MOMB). When the total proteins of CFs were analysed with SDS-PAGE, a protein band of 10.5 kDa MW was found in CFs from all media with the highest level in PSB. LC-MS/MS analysis identified this protein as α-elicitin that had an identical amino acid sequence to the α-elicitin hibernalin of P. hibernalis and syringicin from P. syringae. The optimum conditions for in vitro selection of Dendrobium for black rot resistance using α-elicitin-containing CFs were also determined by evaluating the CF toxicity on Dendrobium protocorm-like bodies (PLBs) when cultured in all media supplemented with 0, 30, 50 and 100% CFs for seven, 14 and 21 d. The levels of PLB necrosis varied according to medium types, CF concentrations and culture periods. The maximum percentage of PLB necrosis (100%) was obtained in PSB supplemented with 50 and 100% CFs, and the severity of PLB necrosis was highest when treated with 100% CF for 14 and 21 d.

The effects of culture media on growth and proliferation of ‘Earsakul’ dendrobium (Dendrobium) protocorm-like bodies (PLBs) were evaluated in a two-step culture. After culturing on each of the four first step media for 4 months and on each of the four second step media for 4 months, the greatest total PLB fresh weight, increase in number of PLBs and growth rates were obtained when using Vacin and Went medium 1 (VW1) in both culture steps compared with those in 15 other medium combinations. Starting from 0.5 g of PLBs, culturing on VW1 for 8 months achieved a total of 415.25 g of PLBs, a multiplication rate of 830-fold. The supplementation of tomato (Solanum lycopersicum) and substitution of ‘Hom Thong’ banana [Musa acuminata (AAA group)] with ‘Khai’ banana [M. acuminata (AA group)] in this new medium promoted growth and proliferation of dendrobium PLBs 2.4-fold over the control medium, suggesting its usefulness in commercial micropropagation.

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