Tulasnella calospora (UAMH 9824) retains its effectiveness at facilitating orchid symbiotic germination in vitro after two decades of subculturing.

Abstract

Background: The technique of symbiotic germination-using mycorrhizal fungi to propagate orchids from seed in vitro-has been used as one method to cultivate orchids in North America and abroad for > 30 years. A long-held assumption is that mycorrhizal fungi used for this purpose lose their effectiveness at germinating seeds over time with repeated subculturing.

Results: We provide evidence for the lingering efficacy of one particular strain of Tulasnella calospora (266; UAMH 9824) to stimulate seed germination exemplified by the North American terrestrial orchid, Spiranthes cernua, as a case study. This fungus was originally acquired from roots from Spiranthes brevilabris in 1999 and sub-cultured during the two decades since. Seeds inoculated with the fungus in vitro developed to an advanced protocorm stage after 16 days, and leaf elongation was pronounced after 42 days. In a pilot study, seedlings co-cultured with Tulasnella calospora 266 were deflasked after 331 days and later transferred to soil under greenhouse conditions where they eventually initiated anthesis. During the course of two decades, seeds of 39 orchid species, cultivars and hybrids spanning 21 genera, germinated in vitro co-cultured with Tulasnella calospora 266. These orchids included temperate terrestrials and tropical epiphytes alike.

Conclusions: The sustained effectiveness of this fungus is noteworthy because it argues against the concept of mycorrhizal fungi losing their symbiotic capability through prolonged subculturing. This study serves as an example of why in situ habitat preservation is essential for the conservation of orchids as a source of potentially useful mycorrhizal fungi.