Taxonomic Identification and Molecular DNA Barcoding of Collected Wild-Growing Orchids Used Traditionally for Salep Production

Abstract

Molecular DNA barcoding combined with botanical taxonomy can be used for the identification and conservation of collected Greek orchids used for salep production as well as in the regulation of fair salep trade. A modified CTAB protocol was used for DNA extraction, amplification of barcoding regions (ITS, matK, rbcL, trnH-psbA), and sequencing. Sequencing data were assembled using Bioedit software, and the BLAST algorithm was used on the NCBI database for species identification at the genus level. Molecular barcoding data based on genetic similarity identification was in full coherence with taxonomic classification based on morphological data. The combination of ITS and matK exhibited a greater capacity to identify a species among the Greek salep samples. Out of the 53 samples examined, 52.9% were classified as Dactylorhiza spp. and 33.3% as Anacamptis spp., whereas only 6 samples were identified as Orchis spp. (11.8%). Given that a superior-quality salep beverage comes from tubers of the latter, the number of samples classified as such in northwestern Greece is unexpectedly low. A database of 53 original reference sequences from wild-growing samples of Greek origin was generated, providing a valuable resource for the identification of other salep samples from different regions. The DNA barcoding results unveiled that salep samples from northwestern Greece are related to nine members of four different genera of Orchidaceae. All species are nationally protected and covered by the CITES convention, while many of these orchids are included in the EU Directive 92/43/EEC appendix as “Other Important Species”. Thus, expedited coordinated management actions are needed to ensure their survival in the future.

Introduction to the Special Issue of Plants on “Advances in Plant Reproductive Ecology and Conservation Biology”

“…Second, the hypothesized patterns of genetic diversity were not often recovered. Greater than anticipated genetic variation was found in S. floridana , I. lacustris , and E. capitatum , but only minimal differences were identified between dioecious and cosexual species of Solanum , C. gileadensis , D. foliosa , and for species used in Salep [ 2 , 3 , 5 , 6 , 7 ]. Collectively, these studies demonstrate the important role that field and laboratory studies play in ensuring a comprehensive understanding of population biology.…”

“…Each tried to gain a better understanding of rare species using a distinct approach. Tsaballa et al [ 2 ] and Mansour et al [ 3 ] investigated economically important species that have been impacted by human development. Tsaballa et al [ 2 ] employed molecular barcoding to uncover the identity of wild species of orchids in Salep, a powder used in beverages and food for medicinal purposes that is culturally important in the eastern Mediterranean, and this has resulted in the overharvesting of orchids.…”

“…Tsaballa et al [ 2 ] and Mansour et al [ 3 ] investigated economically important species that have been impacted by human development. Tsaballa et al [ 2 ] employed molecular barcoding to uncover the identity of wild species of orchids in Salep, a powder used in beverages and food for medicinal purposes that is culturally important in the eastern Mediterranean, and this has resulted in the overharvesting of orchids. The authors recognized that Salep, in Greece, is composed of species from four genera, with the greatest percentage from Dactylorhiza Neck.…”

“…First, the breeding system and human influence are important in the current population structure of the species. This ranges from the orchid species used in Salep to the role of climate change in the shifts in weather in Australia to the modifications in the fire regime in south Florida [ 2 , 5 , 6 ]. Second, the hypothesized patterns of genetic diversity were not often recovered.…”

Qualitative Metabolite Profiling of Orchis purpurea Huds. by GC and UHPLC/MS Approaches

“…This latter is a powder obtained from the dried tubers of more than 35 species of terrestrial orchids, including species from the genera Anacamptis , Dactylorhiza , and Orchis , to cite just a few [ 4 ]. Its high consumption in the eastern Mediterranean area is causing serious risks to both consumers and orchids [ 5 ]. Thus, tubers are often illegally harvested causing conservation concerns [ 6 ], while the final product may be adulterated with potential health risks [ 7 ].…”