A comparative study of ITS2 metabarcoding and traditional microscopic palynology as methods of identifying taxonomic origins of bee collected pollen

Identification of the taxonomic provenance of pollen collected by honey bees provides information on the diversity of plants from which pollen is collected, effectiveness of pollination services provided, and the floral sources used by bees in the production of honey. To date, microscopic palynology has been used to address questions of pollen origin. This microscopic approach can be arduous, requiring large amounts of time and experienced research personnel. Furthermore, taxonomic classification is often limited to the generic or family level given the morphological ambiguities associated with certain plant clades. We employed DNA metabarcoding to determine the taxonomic origins of four samples of multi-floral pollen collected by honey bees in early spring in central Ohio. The ribosomal Internally Transcribed Spacer 2 (ITS2) was amplified using universal primers and sequenced using next-generation Illumina Miseq technology. Paired-end sequences were then joined using PEAR and the resulting sequences were aligned to a customized ITS2 reference library, derived from Genbank, using the BLASTN algorithm. Lastly, the BLAST output was used to make taxonomic assignments with MEGAN5. In addition, we performed microscopic palynological analysis on the samples in order to compare the two approaches. The molecular technique showed promise as a fast and effective means of pollen identification with potential to provide greater taxonomic resolution, though it currently lacks the capacity for quantitative measurement provided by traditional microscopic palynology.