Pollen DNA metabarcoding identifies regional provenance and high plant diversity in Australian honey

Abstract

Accurate identification of the botanical components of honey can be used to establish its geographical provenance, while also providing insights into honeybee ( Apis mellifera L.) diet and foraging preferences. DNA metabarcoding has been demonstrated as a robust method to identify plant species from pollen and pollen‐based products, including honey. We investigated the use of pollen metabarcoding to identify the floral sources and local foraging preferences of honeybees using 15 honey samples from six bioregions from eastern and western Australia. We used two plant metabarcoding markers, ITS2 and the trnL P6 loop. Both markers combined identified a total of 55 plant families, 67 genera, and 43 species. The trnL P6 loop marker provided significantly higher detection of taxa, detecting an average of 15.6 taxa per sample, compared to 4.6 with ITS2. Most honeys were dominated by Eucalyptus and other Myrtaceae species, with a few honeys dominated by Macadamia (Proteaceae) and Fabaceae. Metabarcoding detected the nominal primary source provided by beekeepers among the top five most abundant taxa for 85% of samples. We found that eastern and western honeys could be clearly differentiated by their floral composition, and clustered into bioregions with the trnL marker. Comparison with previous results obtained from melissopalynology shows that metabarcoding can detect similar numbers of plant families and genera, but provides significantly higher resolution at species level. Our results show that pollen DNA metabarcoding is a powerful and robust method for detecting honey provenance and examining the diet of honeybees. This is particularly relevant for hives foraging on the unique and diverse flora of the Australian continent, with the potential to be used as a novel monitoring tool for honeybee floral resources.

DNA metabarcoding using nrITS2 provides highly qualitative and quantitative results for airborne pollen monitoring

“…However, these taxa were only present in <5% maximum relative abundance in the selected samples. In a recent study, Milla et al (2021) found instead that trn L performed better than nrITS2 in Australian honey samples. However, as the authors indicate, DNA in these honey samples was degraded by long storage, causing the more stable and much shorter trn L P6 loop to be better preserved than nrITS2.…”

Preservation methods of honey bee-collected pollen are not a source of bias in ITS2 metabarcoding

“…Due to high taxonomic resolution, ITS2 has been one of the markers of choice in pollen metabarcoding (e.g. Bell et al, 2017;Cornman et al, 2015;Danner et al, 2017;Jones et al, 2021;Keller et al, 2015;Milla et al, 2021;Nürnberger et al, 2019;Richardson et al, 2015b;Sickel et al, 2015). The problem is that PCR using ITS2 barcodes may suffer from non-specificity, resulting in co-amplification of fungal contaminants (Cheng et al, 2016).…”

The Honey Bee Apis mellifera: An Insect at the Interface between Human and Ecosystem Health

“…Of course, microscopic analysis and DNA metabarcoding applied to bee pollen and honey provide information on plant taxa on which honey bees forage, but, given that pollen composition in bee matrices is largely influenced by floristic local biodiversity and flowering phenology [ 18 , 19 , 20 , 21 , 22 , 23 ], these data may improve our understanding of the local biodiversity of flowering plants.…”