Culture Plating vs DNA Testing for Microorganisms in Cannabis w/ Kyle Boyar (From BTS #29)
I'll start with the ancient technology which is plaguing It's been around for some time, and basically this is where you take your sample. You add some sort of. Growth medium right DPW TRIPTYCH soy broth. Yeah Classic Classic growth mediums basically. You put that in with your sample. Typically a one gram sample. It really just depends on what state you're in other industries use lot larger sample sizes. Something like you know anywhere from five twenty five grams, or even upwards of fifty grams really just depends on what you're testing right, but since candidates is a very precious commodity. People don't want to give up their. You Know Ultra high quality stuff more than a gram, so that's typically what we see out there in the marketplace. But, basically you're, you're homogenising your sample. Usually the stomach or By hand massaging and then you're taking that and you're taking a small sub sample of that, and then pleading it as a basically placing that inaugurating the plate, and then you know streaking it or basically spreading it uniformly across the plate surface. And then you're incubating bat depending on what you're testing for, you may incubate it for you know. From Twenty five to twenty seven sees what's typically done for Aspergillus five seven days seven days type thing. But it's going to differ for every different microorganism that you're testing for and then you're basically taking whatever you dilute it at, so you're counting. The colony is on the plate, and then we'll play it by that dilution factor, and now you have a colony forming unit gram readout. That is typically your standard readout that you see in just about everywhere. I mean while QBC are is. Very commonly accepted now in the cannabis industry when it comes to things like total count tests, you do have to convert that seek value or cycle of quantitative will get into a bit more later, but you have to convert that value into something that is acceptable for regulators, which is that a few Graham Now plaguing. Good because it is only enumerated, what is actually viable within the sample, right? And so a lot of people aren't concerned about what's dead in a sample, or what was left over and so that is an advantage of planning a disadvantage of planning is that some of these things don't grow very well and culture It depends again. What type of mediums here using what types of Brasier using There's a number of factors that contribute to how well something cultures on a plate so I would say those are kind of like the high, A. The highpoints of pleading and the potential drawbacks of it but then there's molecular methods like Qpr. QVC are right, so this is quantitative polymerase chain reaction so for those of your listeners that are familiar with polymerase chain reaction. Now this is just monitoring that reaction in real time using different flora fours. And so you're basically typically QC are run is forty cycles You are looking at you know. How quickly does amplify so and then at the point at which crosses the threshold, so you set that threshold to remove any background, the point at which your QBC our signal crosses that threshold is what we call a cycle of quantitative, so that is the point at which. which it is crossing that threshold line and the actual cycle number so basically the earlier of a C Q, You have that means that the earlier or sorry. The the greater presence of your target there is because it's a amplifying quicker, whereas if it's amplifying later than that means you had less of your target, so that would be a higher cq. Value, right? And so I guess the disadvantage with QBC are is that you aren't just selectively? Amplifying is alive. You are getting everything DNA DNA. So you're going to amplify everything and some people push back and say well. Hey, isn't that to our detriment? You know. If you have a bunch of dead stuff on there well, why am I failing for dead stuff? That just doesn't make sense well a couple of things there right so in the context of things like Aspergillus or micro toxin producing. Well. If there's a bunch of that leftover well, that could be indicative that you have mycotoxin your sample now. Of course, many states have opted for mycotoxin testing things like apple. Toxins and ochre toxin. And so that that checks that box right, but still it's good to know and a lot of people have this misconception that because you're only. Oh, sorry, 'cause! You're amplifying all of these things, right? That you're gonNA fail more as a result, but actually we find that. At least in my experience, it was the opposite. And I think the reasoning for that is. and. I guess this is another drawback of planning that I forgot to mention in. This is a perfect tie-in. Well, there's a lack of selectivity. Typically when you're planning something. You're using right selective growth media. That are supposed to foster the growth of a particular class of microbial contaminants now. There's grows mediums. There's also antibiotics that are used for selection in this case, and as we all know, antibiotics are being A. Microorganism, becoming more and more resistance, these things and just because you select for it, based on an antibiotic or some sort of growth medium doesn't necessarily mean that off target organism growth isn't going to happen so what? The the publications that medicinal! Of have done of showed is that there is a significant amount of Tara growth happening especially in the context of police mold plates. And depending on what state you're in typically limits around ten thousand CFU's program. But if you're getting off growth, you're counting dots that are actually not e- mold. You're counting oftentimes bacteria, and so this of course leads inflated accounts, more failures, and so people have this aversion to moving toward QVC are because they think that's going to happen. More frequently, but really it's the opposite because you're using targeted primers, you are selectively amplifying only for the class of organisms that you're looking for. You don't have the potential for off target growth in that case.