Although cannabis is a resilient plant (hence the moniker “weed”), and it forms trichomes that either defend via their structure, or chemicals like terpenes that have antimicrobial, antibacterial, and antiviral properties, there are still threats by microorganisms that must be addressed. Most recently, discussions on hop latent viroid, for example, have revealed the massive loss in cannabis crops devastated by this disease.
Losing plants is obviously tragic, and not just for financial reasons, although the lost revenue from not being able to sell your crops and lost operational costs such as wasted resources (e.g., humans, energy, water) certainly stack up. It’s evident that testing crops entering your facility and those you’re actively cultivating is critical to the success of your grow. I recently spoke with Pathogen Dx CEO, Milan Patel, to learn more.
Jason S. Lupoi, Ph.D.: What are the most common sources for bacteria/powdery mildew (PM)? What tests exist for determining the presence of PM?
Milan Patel: Powdery mildew is similar to a hybrid organism in that it’s a mutation of both bacterial and fungal organisms. As such, it can’t be cultured so Petri dish testing doesn’t work. Only molecular PCR [polymerase chain reaction] tests like our Envirox test can be used to test for it.
Its presence is a function of the environmental conditions. Powdery mildew thrives in environments with high humidity, compromised or poor air circulation, insufficient lighting, and in over-fertilized crops. It spreads easily when plants are touching each other so closely that it transmits simply by physical contact.
What are some of the most common threats within a grow house, and what are some that may not necessarily be common, but can be very detrimental?
We’ve identified roughly 50 plant pathogen threats that impact plant health. The most common threats within a grow house include E. Coli, Salmonella and the four Aspergillus species. Less common but more detrimental threats are organisms such as Mucor, Botrytis, Fusarium and Pythium. These bugs are soil-bearing so they affect the quality of the plant by attacking the roots, causing rot and degrading soil health. Then there are the viruses – we have seen Hop Latent Viroid (HLVd) significantly impact growers’ crop quality and yield. This particular virus is devastating, so quicker and earlier identification will enable the grower to isolate the virus and take necessary precautions before it spreads throughout the whole grow. The only method to detect HLVd is by real time (RT)-PCR or real-time quantitative reverse transcription (qRT)-PCR and there are several companies that produce single organism tests. PathogenDx is in the process of developing a multiplexed assay for the detection of several plant viruses from a single sample to deliver a better, more efficient test for cannabis grows/cultivations.
What kinds of tests does PathogenDx offer for these threats? How do the tests work?
We offer AOAC-certified QuantX Gen 2, the only molecular total yeast and mold that quantifies the amount of yeast and mold in the sample. Our other regulated test includes Detectx, which identifies pathogenic species (E. coli, Salmonella, Aspergillus, Listeria for edibles, Staphylococcus, Pseudomonas). Our non-regulated Envirox test monitors environmental conditions and is able to test for up to 50 pathogens in a single surface swab or air collection from a grow.
How does PathogenDx’s PCR differ from traditional qPCR?
PathogenDx PCR technology is unique in that we use endpoint PCR, which offers much better sensitivity compared to traditional qPCR technology. The amplification reaction is taken to its “endpoint” where the amplification primer is fully consumed, while in traditional qPCR, the reaction stops working after hitting a certain threshold. The reaction stops on its own because, in standard qPCR, the primer concentration is high enough that it is no longer limiting and the reaction stops.
Generally, the endpoint seen in the PCR is due to classical end-product inhibition. This is mostly due to end product competition with the byproducts of nucleotide triphosphate incorporation into the DNA amplicon product and also via end-product competition with the amplicon product itself, which at high amplicon product concentration begins to bind Taq polymerase non-specifically.
The second difference is that qPCR is limited to either sequential testing of the sample and, in some cases, limited multiplex testing (where the number of species that can be evaluated simultaneously is no more than four). Meanwhile, we have 50 different probes in triplicate printed at the bottom of our array wells for a total of 150 probes which allows us to identify the exact sequence of a specific pathogen species and even a sub-strain of that pathogen. The benefit here is a much greater degree of specificity in the test.
What improvements have been made to your cannabis testing system? How has the move to the 96-well plate changed processing time?
We’ve learned a lot from our agile development of COVID-19 diagnostics and applied those learnings to our cannabis testing products. These new improvements bring the cannabis industry up to the same level of standards and performance as diagnostics platforms used in the food, agriculture, and human diagnostics sectors. We have singularly focused on delivering a series of improvements that bring significant benefits to our customers in this sector:
– Our expanded “foil covered” 96-well microarray allows for increased throughput, lower cost per sample processing, and the flexibility to accommodate varying sample volumes by removing the costs of unused wells. This is an industry first as previously, labs had to either wait for a full complement of samples to complete a 96-well plate, or they would lose out on the economics of unused wells which can be very costly. For example, if a lab is only running 36 samples, they’d have to either wait for more samples, or take a loss on the other 60 wells in the plate. With the foil-covered wells, lab technicians are able to section off the foil for only the number of wells that the lab needs to run for that day’s sample volume, whether it’s 12 or 60 samples, without any loss of unused wells as they can be run on another day or shift. This does not impact the turn-around time of the lab by having to wait for the full complement of samples. It’s the only such offering in the market.
– We’ve introduced our AOAC-certified Quantx fungal assay, the only molecular test in the market to qualify CFUs [colony forming units] for yeast and mold in a single shift, helping labs and growers obtain results in hours versus days.
– The new custom PathogenDx Reporting Portal increases consumer ease and efficiency by replacing DropBox and reducing the number of steps necessary to obtain lab results and certificates of analyses.
– Lastly, we’ve developed our Detectx-Alive test, which isolates live DNA from deceased cells in any bacterial or fungal safety test and delivers results in six hours or less.