ASHS 2024 Conference

Commercially produced floral hemp (Cannabis sativa L.) is high in cannabidiol (CBD) concentrations relative to tetrahydrocannabinol (THC); this is intentional due to regulatory pressures requiring low THC thresholds. Given the predominant role of genotype in plant development, it is crucial to also explore environmental factors that may allow for optimization of hemp growth, yield, and quality. The goal in this study is to evaluate the extent to which an extended vegetative growing period has on height, width, and yield in irrigated raised bed production of floral hemp in North Dakota. To survey this relationship between planting date, growth, and quality parameters, mother plants were germinated from seed at the beginning of the growing season. Each treatment group of cuttings, separated by approximately two weeks, were excised from their respective cultivar mother plant for four timing treatment groups. Cultivars evaluated were ‘ACDC’, ‘Bubbatonic’, ‘Sour Elektra’, and ‘Umpqua’. The main effect of cultivar did not significantly affect any measured parameters except for height and the top 1/3 portion of dry floral biomass. Planting date treatments significantly affected the wet weight of total above-ground biomass, total dry above-ground biomass, and total floral biomass. Average total above-ground biomass and dry floral biomass was statistically different for each planting date except for the last two planting dates (June 19 and July 3). Total above-ground dry biomass averaged over cultivars, were 4070 g, 2432 g, 1323 g, and 894 g, for dates May 19, June 5, June 19, and July 3, respectively. Mean yields for total dry floral biomass, averaged over cultivars, were 1779 g, 1279 g, 784 g, and 535 g, for dates May 19, June 5, June 19, and July 3, respectively. Earlier planting dates showed an increase in height for three of the four cultivars with the exception of ‘Umpqua’. A cultivar interaction with planting date treatment occurred for the top 1/3 portion of dry floral biomass and indicated that earlier planting date increased the biomass for ‘Bubbatonic’ and ‘Sour Elektra’ while planting date did not influence dry biomass for ‘ACDC’ and ‘Umpqua’. The insights gained from assessing the impact of variable vegetative growing periods on growth and quality parameters of photoperiod-dependent floral hemp could have broader implications for optimizing production practices. Understanding the intricate interplay between genotype, environmental factors, and cultivation practices is essential for advancing sustainable and efficient hemp cultivation strategies.

Louisiana’s hot and humid climate provides the perfect environmental conditions for the growth of fungal and bacterial plant pathogens. These fungal infections are an obstacle to the success of commercial production of Cannabis sativa in the state. Two of the most recent and significant fungal diseases are southern blight caused by Sclerotium rolfsii and stem canker caused by Botrytis cinerea. However, there is a lack of formal and professional knowledge regarding fungi that infect medicinal hemp plants, and practical and effective methods for managing the casual agents of these diseases. The objective of this study was to identify natural plant products from two native plants of Louisiana, that have been reported in the Native American ethnobotanical literature to have antifungal/antibacterial properties. An in-vitro bioassay experiment was conducted using the agar plug diffusion method testing the antifungal inhibition of crude ethanol extracts from the two species against each of the two pathogens Sclerotium rolfsii and Botrytis cinerea, on four plates each of Extract 1 Diospyros virginiana L. and Extract 2 Equisetum hymale L. of 1/4PDA spiked at a dose of 250ppm, 500ppm, 750ppm, and 1000ppm against a control plate of 1/4PDA for six days. Based on this initial crude extract bioassay there is a highly significant difference in the two crude extracts (p=0.000105) when tested against Botrytis cinerea. There is also a significant difference in concentration. The test against Sclerotium rolfsii did not find any significant inhibition from either of the plant extracts tested. From our findings we will continue the research study to test the antifungal potential of crude ethanol extract as well as Hexane, chloroform, ethyl acetate soluble fractions of Extract 1 Diospyros virginiana. The goal of the study is to integrate the antifungal compounds and their application for the development of best practices in Cannabis production.

Fiber hemp (Cannabis sativa L.

Micropropagation usually involves cytokinin in single-harvest batches. We report two in vitro studies with multiple harvest: (1) fed batch process with modified physical states and (2) LED light treatments. In (1), genotypes of Cannabis sativa were observed in stationary agar (A), stationary Oasis® infused with liquid (OIL) and agitated Oasis® infused with liquid (AOIL).Fifteen explants were planted in vessels with 120 mL DKW medium harvested on 3-week cycles, with 0 or 15 mL additional media. Harvested shoots, length, and dry mass from repeated cycles were recorded. Genotypes T1 and Peach failed on multiple harvest cycles and were eliminated, although single cycle had higher quality in OIL. BaOx and Cherry1 on OIL/AOIL with additions were better quality than A in five cycles. Shoots harvested increased from 15 to 30 in cycles 1-3 in OIL/AOIL, but in A were approximately 20, while length was longest in OIL/AOIL. By cycle 3, all measured responses were decreasing until cycle 5 where a minimum of 7 shoots per vessel or more were only in OIL, but shoots were too short to plant in greenhouse. In (2), blue and supplemental far-red were observed with in vitro shoots of BaoX and Cherry1. OIL treatments were placed in LED polychromatic and dichromatic light (white, high red:blue, medium red:blue, white w/5% far-red, high red:blue 5% far-red, medium red:blue w/5% far-red, low red:blue w/ 5% far-red) at similar intensities (190-240 µmols·m-2·s-116 h-photoperiod). Media additions were made with responses recorded bi-weekly. Five randomly selected microcuttings per vessel rooted ex vitro on mist bench for 16 days. Over multi-cycles, plants treated with 5% far-red increased number and length, while plants under higher blue light increased dry mass. Shoot number increased to 28 in cycles 1-3 with far-red, and 18 without before decreasing to initial 15 during cycle 5. The accumulated shoots per vessel over 5-cycles (10-weeks) was 108 with far-red, and 84 without. Shoot length in far-red-treated plants increased from 19 - 25 mm in cycle 3 before decreasing to 10 mm in cycle 5. Plants without far-red had 10 – 15 mm length the entire experiment. Dry mass was highest during cycle 1 with blue light before decreasing 50% in cycle 3, where it remained until cycle 5. Sixty-eight percent of shoots rooted regardless of prior in vitro treatment. OIL with media addition allowed shoots to be harvested five cycles, while signaling response of far-red light allowed increased productivity and length.

The medical potential of cannabis (Cannabis sativa L.) is based on the complex chemical profile, comprising hundreds of secondary metabolites including cannabinoids, terpenoids and flavonoids. Cultivation conditions were demonstrated to affect plant development, function and production of secondary metabolites in cannabis. Understanding regulation of plant response to environmental conditions is key for development of optimal chemical profile for modern medicine. We have recently demonstrated sensitivity of the secondary metabolite profile in medical cannabis to mineral nutrition, with considerable responses to N, P, and K nutrition. Therefore, knowledge on the cannabis plant response to fertigation schemes is essential for the optimization of cultivation for production of high quality standardized material for the medical market, as well as for development of plant products containing specific desirable phytochemical profiles. In the talk, we will discuss our recent results concerning the potential of additional macronutrients and micronutrients to regulate plant development and the profile of active secondary metabolites in ‘drug-type’ medical cannabis. In pot experiments under controlled conditions, we demonstrated differential induction of changes in the cannabinoid and terpene profiles in ‘drug-type’ medical cannabis also by Ca, Mg, Zn and Mn. Furthermore, rate of uptake and deposition in the plants of individual macronutrients and micronutrients changes between the vegetative and the reproductive developmental stages, and along the reproductive phase.

The emergence of a thriving hemp industry in the U.S. will depend on the breeding of high-yielding regionally adapted cultivars. Despite the latest research efforts, little is known regarding the genetic basis of important agronomic traits in hemp. The objective of this research was to identify and characterize genomic regions associated with seed morphology and quality traits. F 2 mapping populations were developed by crossing hemp germplasm bred or cultivated for cannabinoids (‘FL 58’ × ‘TJ’s CBD’), grain (GVA- H-20-1179 × ‘Picolo’), or fiber (‘Si-1’ × GVA-H-21-1003) market classes. These populations were investigated due to their variation in seed size and seed crude protein. The cannabinoid, grain, and fiber populations were grown and seed was harvested in 2021, 2022, and 2023, respectively. Harvested seeds were phenotyped for thousand seed weight (TSW) and crude protein content predicted by near-infrared (NIR) spectroscopy. The high-cannabinoid population was genotyped using an Illumina array, while the fiber and grain populations were genotyped using an Agilent SureSelect Custom Target Enrichment Probe Set. Marker-associated sequences were aligned to the CBDRx v.2.0 reference genome to align the physical and genetic maps. The TSW and protein content in the cannabinoid population ranged from 9.62 to 23.93 g and 19.25 to 31.89 %, respectively. In contrast, the TSW of the fiber and grain populations ranged from 7.34 to 45.17 g and 8.73 to 31.42 g, respectively. Numerous quantitative trait loci (QTL) of varying effect sizes were identified genome-wide. Notably, in the high- cannabinoid population, major and minor effect QTL for TSW were detected on Chr01 corresponding to 642 kb and 5.56 Mb genetic regions, respectively. Our results in the cannabinoid population highlight the importance of developing more than one F 2 mapping population in a given cross to capture the effect of more alleles due to high heterozygosity in hemp and evaluating distinct pedigrees to sample additional alleles in diverse genetic backgrounds. Narrowing the region around or identifying candidate genes will allow the development of high-throughput molecular markers for beneficial alleles across mapping pedigrees. These findings will accelerate hemp breeding programs through the implementation of marker-assisted selection for high-yielding and high-quality hemp cultivars for grain production.

Cannabis (Cannabis sativa L.) is cultivated for its cannabinoids, which have applications for therapeutic and recreational use. This phenomic evaluation explores accumulation of 16 cannabinoids of interest and associated morphological changes in Cannabis flowers. Eight cultivars of interest were grown in containers within an environmentally controlled greenhouse for 150 days (72 days reproductive). Light intensity, light duration, temperature, and relative humidity were regulated. Monitoring floral development, we observed a consistent increase in cannabinoid concentration as flowers matured, peaking in advanced stages of development. This accumulation pattern was consistent across diverse cultivars, which indicates this accumulation pattern to be the result of a fundamental biological mechanism. Concurrent with cannabinoid accumulation, we noted morphological changes in trichomes, which are widely utilized as markers of maturation within industry. Trichomes transitioned from sparse and translucent to abundant, enlarged, and displaying orange/amber hues as flowers matured, signifying floral maturation and trichome senescence. Importantly, a significant linear correlation emerged between cannabinoid accumulation and trichome morphological changes across all cultivars. This underscores a tight relationship between cannabinoid biosynthesis and trichome development, shaped by genetic factors. In summary, our findings demonstrate the intricate relationship between cannabinoid accumulation and floral morphology in Cannabis. Insights gained are invaluable for cultivar selection, breeding, and cultivation practices aimed at optimizing cannabinoid quantity and time to harvest. Understanding the underlying molecular mechanisms of cannabinoids promises tailored approaches for the optimization of cannabinoid production and the fostering of therapeutic and industrial advancements in Cannabis.