The effect of culture media on asymbiotic seed germination in Epipactis flava, an endangered rheophytic orchid in Thailand

Epipactis flava is a rheophytic orchid, which is very rare lifestyle within Orchidaceae. It has been considered an endangered orchid species and included in the Thailand red data. Naturally, germination of orchid seeds, including E. flava, is due to the lack of endosperm. Hence, in vitro seed germination has been effectively employed for the propagation of endangered orchids. Nonetheless, in vitro seed germination of E. flava has never been reported. Thus, the germination of immature E. flava seeds (6 weeks after pollination) was examined and investigated in five semi-solid culture media (modified VW, MS, BM, MM, and KC). The result showed that BM medium could promote germination faster than the other media, approximately 20.0% of seeds were germinated within 2 weeks after culture, reaching 63.4% after 4 weeks of culture. However, at 8 weeks after culture the germination rates of BM (70.2%), modified VW (69.2%), and MM (65.8%) media were not significantly different. In terms of protocorm development, BM medium also stimulated a higher stage of development (stage 5) than the other media. Up to 54.0% of seeds cultured in BM medium reached stage 5 of development by 8 weeks after culture. Conversely, KC medium had the lowest germination rate (8.2%) and no protocorm development in stage 5. In summary, BM is the most effective medium for in vitro germination of E. flava.