In vitro symbiotic seed germination of epipactis flava (Orchidaceae) promoted by endophytic fungus, tulasnella phuhinrongklaensis

Abstract

The rheophytic orchid, Epipactis flava, is a rare and endangered terrestrial orchid that grows throughout Southeast Asia. In nature, seed germination and protocrom development of this orchid species require a symbiotic association with fungi. In this study, Tulasnella phuhinrongklaensis CMU-CR41, was investigated for its compatibility to act as a symbiont during in vitro seed germination and the protocorm development of this orchid. Percentage of symbiotic seed germination and protocorm developmental rate index of the symbiotic treatment were higher than asymbiotic seed germination treatment. The level of protocorm development was the most advanced at up to stage 3 over a course of 65 days. The formation of peloton-like structures was found in the rhizoids and cortical cells of protocorms in the symbiotic treatment. The identification of the fungal association was confirmed through molecular methods. This is the first report on the in vitro seed germination and protocorm development of E. flava using through the use of a compatible fungal symbiont.