Assessing Botanical Authenticity of Honey Through DNA-Metabarcoding-Based Pollen Analysis: A Preliminary Study.

Honey is highly vulnerable to botanical mislabeling and fraud, highlighting the need for dedicated analytical approaches capable of reliably supporting authenticity assessment. To the best of our knowledge, this study is the first to integrate traditional melissopalynology and rbcL DNA metabarcoding for the botanical characterization of honeys collected from Apulian beekeepers. Melissopalynological analysis confirmed the labeled floral source in 6/10 (60%) samples, while four samples were reclassified. One cherry honey was identified as polyfloral, and one robinia (acacia) honey as crimson clover honey. The two samples labeled as polyfloral showed a predominance of chestnut and coriander pollen. In contrast, molecular results confirmed label declarations for chestnut and cherry honeys, whereas robinia (acacia) honeys highlighted known limitations associated with under-represented pollen taxa. In terms of biodiversity, DNA metabarcoding identified 128 plant families, whereas melissopalynology detected only 42. Despite this difference in taxonomic resolution, both approaches converged on the core botanical composition of the honey matrix, sharing 39 plant families that represent the principal nectariferous and entomophilous taxa. Overall, melissopalynology proved decisive for regulatory label verification, while DNA metabarcoding provided a broader qualitative description of the botanical composition. As a preliminary pilot study, the combined application of melissopalynology and DNA metabarcoding may contribute to a more comprehensive assessment of honey botanical origin and traceability; however, molecular detection alone cannot be considered sufficient to establish compliance with legal requirements for unifloral honey labeling.