1. Poster Print Size: This poster template is 24” high by 48” wide. It can be used to print any poster with a 1:2 aspect ratio including 30x60, 36x72, 42x84, and 48x96. Placeholders: The various elements included in this poster are ones we often see in medical, research, and scientific posters. Feel free to edit, move, add, and delete items, or change the layout to suit your needs. Always check with your conference organizer for specific requirements. Image Quality: You can place digital photos or logo art in your poster file by selecting the Insert, Picture command, or by using standard copy & paste. For best results, all graphic elements should be at least 150-200 pixels per inch in their final printed size. For instance, a 1600 x 1200 pixel photo will usually look fine up to 8“-10” wide on your printed poster. To preview the print quality of images, select a magnification of 100% when previewing your poster. This will give you a good idea of what it will look like in print. If you are laying out a large poster and using half-scale dimensions, be sure to preview your graphics at 200% to see them at their final printed size. Please note that graphics from websites (such as the logo on your hospital's or university's home page) will only be 72dpi and not suitable for printing. [This sidebar area does not print.] Change Color Theme: This template is designed to use the built-in color themes in the newer versions of PowerPoint. To change the color theme, select the Design tab, then select the Colors drop-down list. The default color theme for this template is “Office”, so you can always return to that after trying some of the alternatives. Printing Your Poster: Once your poster file is ready, visit www.genigraphics.com to order a high-quality, affordable poster print. Every order receives a free design review and we can deliver as fast as next business day within the US and Canada. Genigraphics® has been producing output from PowerPoint® longer than anyone in the industry; dating back to when we helped Microsoft® design the PowerPoint® software. US and Canada: 1-800-790- 4001 Email: info@genigraphics.com [This sidebar area does not print.] Cannabis Microbiome Sequencing Reveals Penicillium Paxilli and the Potential for Paxilline Drug Interactions with Cannabidiol Kevin McKernan 1, Jessica Spangler 1, Yvonne Helbert 1, Ryan Lynch 1, Adrian Devitt-Lee 1, Lei Zhang 1, Wendell Orphe 1, Ted Foss 1, Chris Hudalla 2, Matthew Silva 2, Cindy Orser 3,Douglas Smith 1 Affiliations: 1 Medicinal Genomics Corporation, Woburn, MA; 2 ProVerde Laboratories, Milford, MA, 3 DigiPath Labs Jessica Spangler Medicinal Genomics Corporation Email: Jessica.Spangler@medicinalgenomics.com Research and Development Contact 1. McKernan, K.J., Spangler, J., Helbert, Y., Zhang, L. & Tadigotla, V. DREAMing of a patent-free human genome for clinical sequencing. Nat Biotechnology 31, 884-887 (2013). 2.McKernan, K.J. et al. Expanded genetic codes in next generation sequencing enable decontamination and mitochondrial enrichment. PLoS One 9, e96492 (2014). 3.Stewart, E.J. Growing Unculturable Bacteria. Journal of Bacteriology 194, 4151-4160 (August 2012). 4.Kusari, P., Kusari, S., Spiteller, M., & Kayser, O. Endophytic fungi harbored in Cannabis sativa L.: diversity and potential as biocontrol agents against host plant-specific phytopathogens. Fungal Diversity 60, 137–151 (2013). 5. Langouet, S. t al. Inhibition of CYP1A2 and CYP3A4 by oltipraz results in reduction of aflatoxin B1 metabolism in human hepatocytes in primary culture. Cancer research 55, 5574-5579 (1995). 6. Langouet, S. et al. Metabolism of aflatoxin B1 by human hepatocytes in primary culture. Advances in experimental medicine and biology 387, 439-442 (1996). 7. Yamaori, S., Ebisawa, J., Okushima, Y., Yamamoto, I. & Watanabe, K. Potent inhibition of human cytochrome P450 3A isoforms by cannabidiol: role of phenolic hydroxyl groups in the resorcinol moiety. Life sciences 88, 730-736 (2011). References Introduction MGC’s current PathogINDICAtor® assays for use with SenSATIVAx® DNA Extraction are listed below: Presence/Absence Tests E.coli E.coli STEC only Salmonella Aspergillus (A.niger, A.fumigatus, & A.flavus species) Background MGC Microbial Assays We explored ITS PCR both before and after culturing on 3M Total Yeast and Mold (3M-TYM) and Biomerieux Tempo cartridges (BMX- TYM) to monitor the microbial diversity before and after culturing. To our surprise many bacterial species are identified growing on the Yeast and Mold cultures generating False Positive Yeast and Mold results. Many species grow specifically on one culture platform or the other. Traditional Cannabis microbial safety testing relies on counting colony forming units (CFU/g) that grow on a petri dish, 3M film or in a culture based system (Marcu, 2013). These systems take 3-5 days to culture fungi or microbes to detectable limits and cannot discern harmful microbes from beneficial microbes and often fail to detect fungi that synthesize compounds contra-indicated in the medicinal use of cannabinoids (McKernan et al.). As a result of this lack of specificity, overuse of fungicides is common in Colorado and California in 2016 (Wurzer, 2016).Marcu, 2013Wurzer, 2016 An ideal microbial detection system would discern pathogenic from beneficial microbes. This can be challenging to do with selective medias and culturing conditions but can easily be accomplished with DNA based methods. We propose a 2 step method that first depletes or captures beneficial microbial DNA (Bacillus, Trichoderma etc.) onto a magnetic particle. The supernatant of this particle thus contains the non- beneficial microbes and can be tested by qPCR using ITS primers, as described in McKernan et al. This qPCR system can be coupled with a next generation sequencing assay known as the PathoSEEK™ microbial identification test for Cannabis and Hemp that sequences the ITS PCR products to generate Operational Taxonomic Unit (OTUs) estimates for each species. We utilize these methods to quantitate the microbial diversity present on Cannabis samples both before and after culturing of microbes using both 3M and Biomerieux products. This data reveal substantial bacterial growth in commercial Yeast and Mold detection platforms and itemization of beneficial and harmful microbes in current testing regimes. Figure 1: Sample containing plant and microbe DNA. (Plant DNA = Green, Microbial DNA = blue). X-axis is cycles (every 1.5 minutes). Y-axis is relative fluorescence units in a log 10 scale. Figure 2: Sample with no microbe DNA present. (Plant DNA = Green, Microbial DNA detection = Blue) Threshold Tests Total Yeast & Mold (18S) Total Aerobic Bacteria (16S) Total Enterobacteriaceae (16S) Total Coliform Examples of PathogINDICAtor® Assay Data Internal Control Microbial DNA John Geanacopoulos Medicinal Genomics Corporation Email: info@medicinalgenomics.com Sales Contact www.medicinalgenomics.com Experimental Design Figure 3: Sequencing Before and After culturing reveals differential growth of bacteria on Total Yeast and Mold BMX assays. Of note is the rapid growth of Bacillus often used as beneficial microbes. The rapid change in 16S contributions during culturing implies a “Heisenberg uncertainty” in microbial testing platforms where the act of measuring the risks, alters the accurate quantitation of risks. Figure 4: Sequencing of 16S amplicons (bacteria) from 3M Yeast and Mold (TYM) plates demonstrates media selective growth in ROI1 and ROI3. ROI1 demonstrate Bacteria that selectively grow on 3M TYM petri films. R0I3 demonstrate bacteria that grow BMX TYM Tempo cartridges but do not grow on 3M TYM petri films. Of note, BMX Tempo cartridges are known to contain Chloramphenicol. ROI2 demonstrates bacteria that grow in both platforms. Note the scales on the charts are dynamic. Future Directions - Beneficial Microbial DNA Depletion Figure 5: Sequencing of 18S amplicons (fungi) from 3M and BMX TYM plate indicates some consistency between TYM tests for growing yeast and molds, but highlights the importance of filtering out beneficial microbes like Trichoderma. Conclusions Accurate microbial testing should be capable of speciating risks and discounting beneficial microbes. Failure to do so encourages fungicidal use that is likely more dangerous than the microbial risks.