HerbalEGram: Volume 14, Issue 1, January 2017

Reviewed: Raja HA, Baker TR, Little JG, Oberlies NH. DNA barcoding for identification of consumer-relevant mushrooms: A partial solution for product certification? Food Chem. 2017;214:383-392.

Attributing the correct scientific name to dietary ingredients derived from fungal materials remains a challenge, in part due to difficulties authenticating species by chemical means and the nature of fungal taxonomic names, which are undergoing numerous revisions because of the application of molecular methods. This study evaluated the viability of the internal transcribed spacer (ITS) region — the DNA “barcode” accepted by the Consortium for the Barcode of Life for fungi identification — for DNA-based identification of medicinal and culinary mushrooms. The main goal of this study, which was conducted by academic researchers from the University of North Carolina at Greensboro and industry scientists from Procter & Gamble and New Chapter, Inc., was to demonstrate that Sanger sequencing of the ITS region is appropriate to verify the identity of fungal species.

A total of 33 samples (20 powdered mycelial samples from a commercial supplier, six grocery store mushrooms, and seven dietary supplements containing powdered fungal material) were included in the study. The results indicated that 10 samples (30%) displayed accurate binomials. Another 10 samples listed the genus name accurately, but had incorrect species names. For example, five of the six products labeled to contain reishi (Ganoderma lucidum, Ganodermataceae) actually contained either G. sichuanense or G. resinaceum.

In five (15%) of all samples, the genus or species indicated on the label was incorrect. This included four of the six samples sold as “Cordyceps sinensis” (Ophiocordycipitaceae; current name: Ophiocordyceps sinensis). Three of these products were found to contain Tolypocladium inflatum* (Ophiocordycipitaceae) and one was found to contain Inonotus sanghuang (Hymenochaetaceae).

From the remaining eight samples (24%), including two products labeled to contain cordyceps and one labeled as reishi, the authors were not able to obtain DNA for ITS sequencing, since these samples were processed to the extent that no suitable DNA could be extracted.

The analysis of the grocery store mushrooms showed that three of the six samples (50%) were correctly labeled, while in one sample, the exact species could not be identified (the fungal genus, however, was accurately labeled). In another case, suitable DNA could not be obtained to determine species identity. Interestingly, the last sample labeled to contain porcini (Boletus edulis, Boletaceae) was identified as B. shiyong, which is a recently discovered species.

The authors listed four important points for consideration when using DNA barcoding to identify mushrooms:

1. DNA barcoding via Sanger sequencing of the ITS region should be used only on materials that have been minimally processed;
2. DNA barcoding cannot be performed via Sanger sequencing on samples containing multiple species;
3. DNA barcoding works efficiently and accurately if accurate reference sequence databases are available; and
4. DNA barcoding can provide only a partial solution for product certification, and other independent techniques, such as those provided by analytical chemistry, may also serve to ensure the lack of contamination and adulteration of these substances.

The researchers hope that their study will encourage International Nucleotide Sequence Databases, such as GenBank, to work on the RefSeq database. This database consists of fully annotated ITS sequences obtained by expert mycologists for medicinal as well as commercial mushrooms and is maintained by the National Center for Biotechnology Information (NCBI).

Dietary supplements containing mycelia (the mycelium is the vegetative part of a fungus that consists of threadlike tubes called hyphae) or fruiting bodies of fungi have grown in popularity and are expected to continue gaining importance in the marketplace.^2,3 Even in the presence of the fruiting bodies, which unlike mycelia have distinct morphological characteristics, correct identification of fungal material requires considerable expertise. Authentication becomes more complicated when powdered materials or fungal extracts are used. Chemical fingerprints have been used successfully for the identification of a number of fungi, but the composition is often dominated by similar polysaccharides and terpenoids, making chemical authentication challenging.⁴

Genetic methods, such as DNA barcoding, offer an alternative way to establish the identity of medicinal and commercial mushrooms, and unlike with medicinal plants, experts have been able to agree on ITS as a suitable genetic region for DNA barcoding. However, as evidenced by the data from Raja et al., the same limitations observed with DNA barcoding for plants are also seen with fungi. As such, the method should be used only for crude raw material and in combination with other independent methods (e.g., botanical, macroscopic, microscopic, and/or chemical analyses).

* Spores from Cordyceps subsessilis (syn. Elaphocordyceps subsessilis, Ophiocordycipitaceae) are known to produce colonies of Tolypocladium inflatum, which is a phenomenon known as anamorphism. Quandt et al. suggest that the three names, C. subsessilis, E. subsessilis, and T. inflatum, should be considered synonyms.¹ Tolypocladium inflatum is of medicinal interest since it is the source of the immunosuppressant ciclosporin.

—Stefan Gafner, PhD

References

1. Quandt CA, Kepler RM, Gams W, et al. Phylogenetic-based nomenclatural proposals for Ophiocordycipitaceae (Hypocreales) with new combinations in Tolypocladium. IMA Fungus. 2014;5(1):121-134.
2. Daniells S. Five key Expo West supplement trends. NutraIngredients-USA website. Available at: www.nutraingredients-usa.com/Markets/Five-key-Expo-West-supplement-trends. Published March 16, 2016. Updated March 22, 2016. Accessed December 13, 2016.
3. Smith T, Kawa K, Eckl V, Johnson J. Sales of herbal dietary supplements in US increased 7.5% in 2015. HerbalGram. 2016;(111):67-73. Available at: http://cms.herbalgram.org/herbalgram/issue111/hg111-mktrpt.html. Accessed December 13, 2016.
4. Elsayed EA, El Enshasy H, Wadaan MAM, Aziz R. Mushrooms: a potential natural source of anti-inflammatory compounds for medical applications. Mediators Inflamm. 2014;2014:805841. doi: 10.1155/2014/805841.