Cannabis Chemovar Nomenclature Misrepresents Chemical and Genetic Diversity; Survey of Variations in Chemical Profiles and Genetic Markers in Nevada Medical Cannabis Samples
Ulrich Reimann-Philipp, Mark Speck, Cindy Orser, Steve Johnson, Aaron Hilyard, Helen Turner, Alexander J. Stokes, and Andrea L. Small-Howard
Cannabis and Cannabinoid Research, Volume X, Number X, 2019
Mary Ann Liebert, Inc.
Introduction : Medical cannabis patients receive clinical benefits from the secondary metabolites of the plant, which contain a variety of cannabinoids and terpenoids in combinations that can be used to classify the chemovars. State-regulated medical cannabis programs rely on breeder-reported ‘‘strain’’ names both within diversion control systems and to describe the medical cannabis products that are sold to patients in medical cannabis dispensaries. In state-regulated medical cannabis programs, there is no conventional nomenclature system that correlates the breeder-reported names with their profiles of active ingredients, and these ‘‘strain’’ names are invalid as they refer to chemical differences properly referred to as to chemovars.
Materials and Methods : To determine the actual levels of chemical diversity represented in 2662 samples of Cannabis flower collected between January 2016 and June of 2017 in Nevada, chemical profile data were measured from these samples by a state-qualified third-party testing laboratory. Principal component analysis (PCA) was used to define clusters in data sets representing both cannabinoids and terpenoids, cannabinoids only, or terpenoids only.
Results : The PCA of the terpenoid only data set revealed three well-defined clusters. All three terpenoids only data clusters had high tetrahydrocannabinolic acid synthase, but the terpene profiles listed in reverse-order of abundance best defined these chemovars. The three chemovars in Nevada were labeled with 396 breederreported sample names, which overestimate the diversity and do not inform patients regarding chemical properties Representative DNA samples were taken from each chemovar to determine whether the genetic diversity was greater than the chemical diversity. The limited genotyping experiment was based on DNA sequence polymorphisms. The genetic analysis revealed twelve distinct genetic clades, which still does not account for the entirety of the 396 reported sample names. The finite genotypes did not correlate with the chemotypes determined for the samples. This suggests that either the DNA-markers used were too narrowly restricted for factual separation or that environmental factors contributed more significantly to the chemical profiles of cannabis than genetics.
Conclusion : The three chemovars and twelve genotypes reflect low medical diversity on the market in Nevada during its ‘‘medical use only’’ phase. Furthermore, the 396 breeder-reported sample names within this set imply a false sense of diversity of products in Nevada dispensaries.
Keywords : cannabis; chemical profile; chemovars; genetic diversity; medical cannabis; nomenclature
Legal medical marijuana usage is increasing in the United States due to the expansion of state-regulated medical cannabis programs and the broadening scope of existing state medical marijuana programs to include ‘‘adult use.1’’ Medical cannabis patients benefit from the secondary metabolites of the plant, which include a variety of cannabinoids and terpenoids in combinations that can be used to classify the varieties of Cannabis.2,3 The major stakeholders in these state-run programs (i.e., cultivators, extractors, processors, retailers, testing labs, and state-regulating agencies) characterize cannabis products and regulate cannabis materials based on the breeder-reported name. Despite this, none of the stakeholders in medical cannabis programs is required to prove the chemovar-identity of the products.
The genus Cannabis contains a single species, Cannabis sativa.4 Most of the cannabis available on the U.S. market are hybrid varieties defined by properties such as psychoactive and medicinal effects, appearance, yield, taste, and odor.5 These varieties are called ‘‘strains’’ in the common vernacular, but should properly be referred to as chemovars as they do not meet the scientific definition of a strain such as that used for bacteria or viruses.6,7 The vast majority of commercially cultivated Cannabis plants are produced through cloning.5 Clonal propagation ensures that plants are genetically identical to the mother plant.8 In contrast to varieties propagated through seeds, requiring a lengthy process of backcrossing and inbreeding to achieve consistency, new chemovars can be created much faster by clonal propagation. New chemovars are constantly generated and enter the market, resulting in thousands of different breeder-reported names without any scientific naming convention.
Most state-regulated medical marijuana programs rely on the breeder-reported names for tracking cannabis products, and these are also the primary cannabis product designation. Naming conventions for cannabis chemovars are poorly or not at all defined, and there are currently thousands of chemovars available in states that have legalized marijuana usage. This creates a confusing situation for patients, who depend on the identifications and potency data on the packaging. If the commercial names bear little consistent relationship to either genotype or chemical phenotype, then they should not be the primary basis provided to patients for decision-making.3,9
Some cannabis researchers have suggested that chemovars should be identified based on their chemical profiles, as informed by principal component analysis (PCA),2,3,10 while others advocate for a genetically based labeling system.11–14 Currently, genetically identical chemovars may be labeled in dispensaries with different names, or different genotypes might be sold under the same name in state licensed dispensaries. Furthermore, genetically identical samples that are grown under different environmental conditions may produce different profiles of active ingredients.15
Medical marijuana patients are entitled to consistent composition and potency of cannabinoids and terpenoids. 16 Some states require disclosure of complex chemical profile information; whereas others only require disclosure of tetrahydrocannabinol levels and cannabidiol levels.1 The potential for mislabeling of chemovars, inconsistent chemical profiles of marijuana products, and often limited testing data make it difficult or impossible for many patients to obtain a consistent chemical profile of the product. The relationships between the breeder-reported names, genotype, and chemotype have not been rigorously evaluated.
In this study, we investigate variation among chemovars based on the variability of chemical profiles and the relationship of these chemical profiles to three genetic markers. We focused on the state of Nevada, which started to grant licenses for commercial medical marijuana in November 2014. Medical marijuana products sold in Nevada are tested by a third-party laboratory for cannabinoid and terpene content and for microbial and pesticide safety, thus, testing data are available for analysis. Chemical profiles of Cannabis flower samples collected between January 2016 and June 2017 were generated by a major Nevada testing laboratory. During the period of data collection, Nevada was amedical-only cannabis state, and so, all tested samples were destined for medically oriented use. Data cleaning consisted of correcting for misspellings and eliminating different names that were apparently used for the same samples and not counting samples only identified by numbers. PCA data for all samples were produced using algorithms to reveal probable clusters. We then compared the average content of individual cannabinoids and terpenes within the resultant clusters. Finally, representative samples were selected for DNA sequence analysis to reveal the genetic diversity within the cannabis samples. DNA sequence data from fragments of three genes involved in the biosynthesis of cannabinoids and terpenes were the basis of the genotypic analysis. The variation in these sequences was then used to establish the genetic relationship of the samples and the relationship of the genetic data to the chemical profiles.can.2018.0063