How to Detect and Prevent Microbial Contamination in Cannabis: Going Beyond Compliance Testing

In the US, public health and safety regulations for cannabis are currently being implemented state-by-state, as legalization continues solely at the state-level amid federal inaction. Although they differ across state borders, all these regulations will require cannabis samples to be tested before reaching market shelves. These tests include chemical and microbiological screenings, which aim to limit the occurrence of residual pesticides, processing chemicals, and microbials representing a potential public health risk.

Getting microbial

Microbial testing is generally focused on detecting organisms that are also screened for in other industries, such as food safety or water quality. E. coli and salmonella are classic examples of bacteria commonly associated with food-borne illness and are therefore monitored by the beef and poultry industries in particular. These organisms are legitimate health and safety concerns, and although exceedingly rare on cannabis flower or processed extracts, it’s certainly prudent for precautionary monitoring in edible products infused with cannabis – other inputs to edibles (eggs, milks, etc.) represent a potential vector of contamination.

Bacteria aren’t the only category of microorganism commonly screened for in harvest and product batches of cannabis. Total yeast and mold counts, as well as species-specific aspergillus testing, are common requirements in many states. Molds¹ are complex multicellular microbial creatures known for their spore-production and general bronchopulmonary complications in humans. Aspergillus is a type of mold, which at times has been implicated in opportunistic pathogenesis of humans.

While aspergillus is a notable quality indicator for a product like cannabis, typically inhaled during consumption, fungal communities naturally present in the crop² are far more diverse than aspergillus types alone. Microbial constituents, especially fungal varieties, naturally occur in cannabis and depend on the cultivation type, ranging from soil-grown to hydroponic, indoor, outdoor, greenhouse, glasshouse, etc. Likewise, the inputs applied by a cultivator will greatly influence the numbers and types of organisms present. Since aspergillus is a known challenge to human health³, and since its detection can fail a cannabis harvest under current regulatory regimes, we pay it particular attention in the industry. However, its presence or absence gives little indication of the occurrence of other creatures of fungal biology capable of causing crop failure and economic losses, a consequence not tied to the pass-or-fail stamp of state compliance testing.

Failed it

When a cannabis batch fails a lab’s microbial compliance test it’s extremely costly to the producer, even with an economical remediation pathway, which may still hinder the quality of the final product. Implementing an environmental monitoring program allows the cultivator or processor to establish a microbial baseline, and then monitor, detect, and pinpoint problems. Individual environments can be monitored over time and with changing activities. This can be accomplished by taking swab samples⁴ of air handling systems – surfaces typically in contact with processing steps – and other typical point sources of microbial biomass. Inputs such as soil, nutrients, compost teas, employee traffic patterns, as well as human hygiene (shoes, phones) all contribute as vectors of external microbiological introduction to the cultivation, processing, or storage environment. Swabs can easily be tested for the compliance organisms that cause a failure during regulatory testing, but far more information can be obtained if the laboratory is using a platform that can screen for and detect a wider scope of organisms.

Plant pathogens that cause crop losses are currently untested from a cannabis regulatory perspective, but are economically relevant, and therefore worthy of attention. Detection strategies for these organisms certainly exist but aren’t necessarily widely available to our industry, perhaps due to a lack of demand to date (potentially derived from the avoidance of more costly testing). Cultivators often witness symptoms in their crop that aren’t traceable to nutrient deficiency and that they can’t diagnose visually, but could potentially be pinpointed to a causative organism, such as pythium or fusarium² fungi via molecular diagnostic testing.

Producing solutions

Thus far, microbial regulatory testing at the state level is focused on public health risks as opposed to crop loss risks. Cannabis producers and processors worry about both types of risks to their batches, but only have easy access to the regulatory-focused testing. One of the best preventative measures to avoid microbial contamination is to purpose-build the production or processing environment as opposed to retrofitting a subpar facility whenever possible. Ceiling type, age, and material is surprisingly important, as is an air conditioning system’s design, age, and maintenance status. Roof integrity is immensely important, and deficiencies need to be identified immediately and rectified. Often damage may only be apparent after a heavy rainstorm, yet the microbial growth continues unseen once the moisture seeps in. Personal hygiene translates to potential contamination vectors and should be a factor in standard operating procedures at the production or processing site. Environmental monitoring programs can elucidate the effectiveness of decontamination protocols and establish the necessary periodicity of cleaning regimes. It’s important to understand microbial contamination can occur just as commonly after the cultivation stage - during harvesting, processing and curing phases as well.

State-mandated compliance testing for cannabis products has certainly resulted in industry-wide protection of public health, but also has presented challenges and opportunities with respect to what is not monitored. Cannabis testing is being implemented at the state level as medical or adult-use markets become legitimized, but the lack of standardization across the industry presents unique challenges, and also currently misses testing offerings that could significantly benefit the industry. Producers of edibles and infused beverages certainly face additional and unique situations. Microbial growth potential is different depending on the degree of dryness for flower (aka the ‘cured’ status of cannabis buds), or the exact type of candy and other infused product. Testing for public-health relevant organisms beyond salmonella and E. coli are being considered in many states in order to monitor edible and infused products. Still, the industry needs to reiterate important questions, such as what are the prevalence patterns of microbial communities in cannabis? And which organisms represent smart targets for indicator organism testing? Can we effectively limit microbial growth on biomass and edibles by limiting moisture content and water activity, two orthogonal laboratory proxies indicating microbial growth potential? Are these metrics useful as on-site, in-process monitoring tools during the drying (curing) process, analogous to similar in-process QA/QC testing in the food industry?

It turns out we are well-equipped in this day and age to empower our knowledge of microbial communities in cannabis. The use of molecular diagnostic tools bypasses the need to culture⁵ an organism in order to detect it, by targeting the organism’s genetic material instead. While culture-based detection of microbes has been the gold standard for microbiologists since the early 1900s, these techniques require specific environmental conditions and nutrient cocktails which allow a particular microorganism (such as E. coli) to reproduce and become detectable as a colony forming unit on a petri dish. This perspective on the microbial world has been limiting for us humans. But with the advent of affordable DNA sequencing capabilities⁶, analysts can now collect all the genomes present in a sample and amplify DNA segments, thereby identifying the microbial constituents by reading their genetic barcode. Such molecular methods are leveraged in compliance testing to screen for and detect specific organisms, like E. coli, but they can also provide more general information by detecting and identifying all microbial genomes in a sample. The ability to fully characterize a natural microbial community genetically is like a superpower to microbiologists.

Prevention goes hand-in-hand with knowledge. Due to a general lack of microbial screening during cultivation, often a crop infection can be undetected until it’s too late - or go misdiagnosed as a nutrient deficiency that presents similar symptoms of yellowed or wilted leaves, for example. Perhaps with more routine testing at the grow, we could soon elucidate strain-specific vulnerability to plant pathogens. Perhaps there are particular microbes which augment the health and vitality of the plant⁷, much like probiotics confer health and immunity benefits to humans. We have some indications that the microbiome associated with cannabis can influence terpene content in the flowers. The ability to characterize the microbial populations present in a sick crop could inform on potential areas of concern that aren’t addressed by current regulations, documenting for the industry the microbial challenges faced by cultivators. Such approaches would even detect the DNA of the notorious russet mite⁸ and its relatives, microscopic arachnids with extremely destructive potential to cannabis crops. Leveraging these powerful techniques to create a deeper knowledge base beyond the current compliance testing regimes could thereby pay great economic dividends in terms of protecting plant health and crop yields.

References 

1. Mold | General Information: Facts about Stachybotrys chartarum and other Molds | CDC https://www.cdc.gov/mold/stachy.htm (accessed Apr 14, 2019).
2. Punja, Z. K.; Collyer, D.; Scott, C.; Lung, S.; Holmes, J.; Sutton, D. Pathogens and Molds Affecting Production and Quality of Cannabis Sativa L. Front. Plant Sci. 2019, 10. https://doi.org/10.3389/fpls.2019.01120.
3. Aspergillosis Statistics | Types of Fungal Diseases | CDC https://www.cdc.gov/fungal/diseases/aspergillosis/statistics.html (accessed Mar 23, 2020).
4. Marsh, A. S. FAQ: Microbiology of Built Environments: Report on an American Academy of Microbiology Colloquium Held in Washington, DC, in September 2015; American Society for Microbiology: Washington (DC), 2016.
5. Fredricks, D. N.; Relman, D. A. Sequence-Based Identification of Microbial Pathogens: A Reconsideration of Koch’s Postulates. CLIN MICROBIOL REV 1996, 9, 16.
6. DNA Sequencing Costs: Data https://www.genome.gov/27541954/dna-sequencing-costs-data/ (accessed Aug 2, 2018).
7. Winston, M. E.; Hampton-Marcell, J.; Zarraonaindia, I.; Owens, S. M.; Moreau, C. S.; Gilbert, J. A.; Hartsel, J.; Kennedy, S. J.; Gibbons, S. M. Understanding Cultivar-Specificity and Soil Determinants of the Cannabis Microbiome. PLoS ONE 2014, 9 (6), e99641. https://doi.org/10.1371/journal.pone.0099641.
8. McPartland, J. M.; Hillig, K. W. The Hemp Russet Mite. J. Ind. Hemp 2003, 8 (2), 107–112. https://doi.org/10.1300/J237v08n02_10.