Authentication of honey based on a DNA method to differentiate Apis mellifera subspecies: Application to Sicilian honey bee (A. m. siciliana) and Iberian honey bee (A. m. iberiensis) honeys

Honey contains intrinsic markers that can be used to identify its origin. In this study, we used the honey as source of honey bee DNA and developed a test to detect the entomological origin of the honey by identifying the Apis mellifera subspecies using an informative mitochondrial DNA region. We then applied this method for the authentication of A. m. siciliana and A. m. iberiensis honeys. DNA was extracted from 60 honey samples produced in several Italian regions (including Sicily and close minor islands), Portugal, Spain and other countries. PCR primers were designed to amplify a fragment of 85 bp (A. mellifera C lineage; highly frequent in A. m. ligustica, the Italian honey bee that is frequently implicated in human introductions worldwide), or 138 bp (M lineage; characteristic of A. m. mellifera) or 152 bp (A lineage; of the honey bee subspecies of African origin). All sampled Sicilian honeys (that were from A. m. siciliana) showed only the fragment of 152 bp, confirming its expected origin. All honeys from A. m. iberiensis showed only the fragment of 152 bp or only the fragment of 138 bp or both, in agreement with the hybrid origin of Iberian honey bee populations. All other analyzed honeys showed the fragments of 85 bp or 85 + 138 bp, suggesting that they were produced from other subspecies. This authentication system could be a useful tool to support conservation genetic programs that rely on marketing links between honey bee genetic resources and the honey they produce.