Honey bulk DNA metagenomic analysis to identify honey biological composition and monitor honey bee pathogens

Honey's DNA mixture originates from various organisms like plants, arthropods, fungi, bacteria, and viruses. Conventional methods like melissopalynological analysis and targeted honey DNA metabarcoding offer a limited view of honey's biological composition. We conducted a honey bulk DNA metagenomic analysis to characterize the honey's taxonomic composition and identify honey bee-related pathogens and parasites based on 266 Estonian and 103 foreign honey samples. 70.4% of the DNA in Estonian honey was derived from green plant families like Brassicaceae, Rosaceae, Fabaceae, and Pinaceae. Geographical distribution analysis revealed distinct botanical compositions between Estonian mainland and island samples. The bacterial family Lactobacillaceae was prevalent overall, reflecting the honey bee microbiota in honey. We detected 12 honey bee pathogens and parasites, including Paenibacillus larvae, Nosema ceranae, Varroa destructor, and Aethina tumida. In conclusion, the study underscores the potential of bulk DNA-based non-targeted metagenomic approaches for monitoring honey bee health, environment, and honey composition, origin, and authenticity.