In Vitro Propagation of Rheophytic Orchid, Epipactis flava Seidenf.-A Comparison of Semi-Solid, Continuous Immersion and Temporary Immersion Systems.

Boworn Kunakhonnuruk,Anupan Kongbangkerd,Phithak Inthima

doi:10.3390/biology8040072

Abstract

Epipactis flava Seidenf. is an endangered Thai rheophytic orchid that has recently shown a rapid decrease in its natural habitat, prompting an urgent need for conservation using ex situ reintroduction methods. Temporary immersion system (TIS) has been successfully applied for large-scale propagation in various plants species. Propagation efficiency of E. flava using TIS was investigated and compared with conventional semi-solid system (SSS) and liquid continuous immersion system (CIS). The highest percentage of new shoot and shoot bud formation was obtained from TIS, followed by CIS and SSS, respectively. Growth parameters as indicated by number of new shoots, shoot buds, shoot height and leaves per explant were significantly higher using TIS than with SSS and CIS. Moreover, the maximum number of new shoots and shoot buds per replication were reliably obtained from TIS higher than SSS and CIS. After acclimatization, the highest survival percentage of plantlets was observed in TIS (76.7%), with 60% surviving after eight weeks of transplantation in artificial stream. TIS was determined as the most suitable culture system for in vitro mass propagation of E. flava compared to CIS and SSS.

Highlights

A rheophytic lifestyle appears to be very rare in Orchidaceae, and the only rheophytic orchid found in Thailand is Epipactis flava [1]
Only 10 natural sites of E. flava have been found in Nan, Kanchanaburi and Tak Provinces [1] and its current conservation status is classified as an endangered species [4,5]
Explants of E. flava with two to three buds differentiated from rhizomes that were cultured in semi-solid system (SSS), continuous immersion system (CIS) and temporary immersion system (TIS) using the same medium

Summary

Introduction

A rheophytic lifestyle appears to be very rare in Orchidaceae, and the only rheophytic orchid found in Thailand is Epipactis flava [1]. Plant tissue culture is recognized as a high-performance tool for ex situ conservation of endemic and endangered orchid species [6,7,8]. For in vitro mass propagation, conventional techniques using semi-solid or shake-flask cultures are labor intensive during the subculturing period. To overcome these problems, bioreactor systems have been developed and improved. Several novel bioreactor systems have shown promise, including temporary immersion system (TIS), which are recognized as a key process for mass propagation and commercial exploitation of plant tissue cultures [9]. TIS and other novel plant bioreactors based on micropropagation have been used to increase plant culture multiplication rates and successfully applied commercially [10,11,12] for mass production of medicinal

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Conclusion