"The problem is that you cannot tell from the product which species it is," says researcher Margret Veltman at the Natural History Museum.
A mild fragrance of flowers with a small sprinkle of cinnamon. In Greek and Turkish bars, salep is a well-known and popular drink on cool days, and an exotic feature for western tourists in search of local specialties. However, the drink is made from dried and ground tubers from wild orchids harvested in the countryside of Turkey and Iran. These orchids are all on the CITES list of endangered or potentially endangered species, and trading in such tubers is only allowed under strict conditions.
"These tubers are sold in markets and bazaars. The entire value chain, from the tubers being harvested to ending up with the end-users, is not very traceable and therefore difficult to control. It is also not possible to tell from the dried roots which species they come from. Moreover, it is often difficult to extract enough DNA to perform good genetic analyses," says Veltman.
CITES (Convention on International Trade in Endangered Species of Wild Fauna and Flora) is an international agreement intended to ensure that international trade in species of wild plants and animals does not threaten the species' survival. More than 40,900 species, including 6,610 animal species and 34,310 plant species, are currently covered by CITES trade restrictions.
Uncovering more details
She recently defended her Ph.D., where she developed a set of genetic "baits" that can be used to fish for DNA evidence of the origin of salep powder. Which orchid species does the powder come from, and where is it harvested? These could become useful tools in the fight against the illegal trade of this popular raw material.
"So far, we have used genetic methods where one sequences a specific part of the genome, what we call DNA barcoding," Veltman explains.
Such barcoding is relatively affordable, and databases for species determination are already readily available. But the method falls short when more nuanced information is needed to differentiate individuals at the population level. Regular DNA barcoding usually only looks at a few genetic markers, while targeted DNA capture can retrieve hundreds or thousands of markers from various locations on the genome. Thus, targeted DNA capture can be used to extract more information about the sample being examined, such as geographical origin and which population it comes from.
"The principle of this new method is that you use a set of genetic baits to fish out exactly the DNA fragments we are interested in from the sample," explains Veltman.
Using cells' own genetic mechanisms
The specially designed bait consists of RNA, a molecule that plays a key role when genes are functioning inside our cells. When a protein is to be formed, an RNA copy of the relevant DNA section that codes for the protein is made. This RNA copy then serves as a recipe for protein synthesis. It is this copying function that the researchers exploit to fish for specific DNA sequences in the sample.
"Our 'baits' consist of such RNA strands, which bind to the corresponding DNA strand in the sample. They hybridize. Then we only retain these hybrids and can disregard all the others," explains Veltman.
The catch – the carefully selected DNA strands – can then be compared to a standard set of gene sequences for the different species one is looking for. However, the method presents challenges, not least because of the structure of the tubers, Veltman says.
"In roots, there is a lot of starch, which makes it difficult to extract DNA. All the starch causes a gel-like substance to form that encases the DNA”.
The starch in the roots also inhibits the enzyme activity necessary to replicate the DNA before analysis. In developing the method, Veltman used specimens of orchid roots from various museum collections.
"Sometimes the DNA in the samples wasn't even measurable. Not because there's little DNA in the material, but because it's locked inside all the starch," explains Veltman.
Orchid species under pressure
Challenges notwithstanding. The results from Veltman's analyses suggest that the orchid populations in Turkey and Iran are under pressure.
"We are seeing changes in how harvesting is done. People are starting to harvest new species, in new areas. The tubers are becoming increasingly smaller in weight and size, indicating that younger individuals are being harvested more frequently."
Professor Hugo De Boer at the Natural History Museum says the new DNA tools will make it possible to do much more precise identification of plants that are traded.
"Identifying plants in trade is important for both biodiversity and health. We need to be able to identify plants that can cause harm, or plants that are important to us, plants that have significant economic value. Many plants can be identified just by looking at them, but it is not easy in cases like with timber, allergenic pollen, or herbal medicines. Then we need tools that can identify them and find the source," says de Boer.
Veltman believes that more attention is needed around the illegal sale of endangered species.
"Human activity has a tremendous impact on the nature around us. A multitude of species are disappearing. Salep is often described as an exciting and exotic drink that must be tried, but the truth is that it promotes a product that is absolutely not sustainable," she says.
The research has been financed by the EU's Horizon 2020 program.
