Next-Generation Sequencing Reveals the Total Yeast and Mold Species Present in Greenhouse-Grown Cannabis Inflorescences—Implications for Quality Control

Next-generation sequencing (NGS) was conducted to determine the composition of the fungal and yeast species (mycobiome) within inflorescences of seven THC-containing genotypes of cannabis (Cannabis sativa L.). They were grown under conventional hydroponic cultivation or under organic production in commercial greenhouses. Up to 26 fungal and yeast species were identified from DNA sequence homologies to representative species in databanks. The mycobiome composition varied according to cultivation method, cannabis genotype, and post-harvest drying. Highest numbers of total fungal reads were present in organically grown inflorescences. Stem tissues supporting the inflorescences contained up to 50% of total fungal reads. Using scanning electron microscopy, fungal spore-like structures and mycelium were observed within the stem tissues. The genera represented by >1% of total fungal reads within inflorescences were Alternaria, Aspergillus, Botrytis, Candida, Fusarium, Penicillium, Trichoderma and Rhizophagus. Post-harvest drying of the inflorescences reduced total fungal reads by 50–65%, depending on cannabis genotype and fungal species. Detached trichome glands subjected to NGS analysis revealed a range of air-disseminated fungi, including Penicillium, Fusarium, Beauveria, Trichoderma, Aspergillus, Botrytis, Alternaria and Aureobasidium. Spore-like structures were observed attached to the external surfaces of glandular trichomes using scanning electron microscopy. The occurrence of mycotoxin-producing species of Aspergillus was rare, while species of Alternaria, Fusarium and Penicillium, recognized as potential mycotoxin producers, were more frequent. The most prevalent fungal species reads among 12 dried cannabis samples were Penicillium chrysogenum, P. citrinum and Alternaria spp.; Botrytis and Fusarium spp. were also present in six samples. The use of biological control products containing Trichoderma and Beauveria during flower development increased fungal reads of these species in fresh inflorescences, with some carry-over into dried samples. The mycorrhizal fungus Rhizophagus irregularis was detected in inflorescence tissues. Reads of a number of previously unreported fungal species were present within inflorescences at very low frequencies. Their origins may be from the surrounding environment or the growing substrate used during cannabis cultivation. Human DNA was found in a number of dried samples, possibly from physical handling of post-harvest inflorescences. These results indicate the diversity and prevalence of certain genera of fungi/yeasts in greenhouse-cultivated cannabis inflorescences.