Identification of botanical origin of bee-collected mixed pollen samples: a comparison between palynological and DNA metabarcoding methods
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Identification of botanical origin of mixed pollen samples has several applications, including unraveling plant-pollinator interactions, determining botanical origin of honey, monitoring allergy-related airborne pollen sources, or even monitoring pesticide use in crops. These applications have typically been addressed using light microscopy, a costly approach that often provides low taxonomic resolution. However, with high-throughput sequencing (HTS) becoming increasingly affordable, DNA metabarcoding is emerging as a promising alternative to classical palynology. In addition to be time- and cost-effective for large sample sizes, metabarcoding has the potential to allow identification of pollen mixtures at the species level. However, before it can be widely employed in pollen analysis, the reliability of this molecular tool must be appraised. Herein, we compared the two approaches on 61 bee-collected pollen samples from eight European
countries. The samples were homogenized and split into two sub-samples. One set of 61 sub-samples was analyzed by palynology experts from the “Institut für Bienenkunde”, Germany, and the other one was subjected to HTS, using ITS2 as the barcode, in the labs of CIMO and CIBIO. Comparisons of the relative abundances at the family level show no significant differences (P ≥ 0.1057, Wilcoxon signed-rank test) and high correlation values (0.2736 ≤ r ≤ 1.000, Pearson’s correlation) between the two approaches. The highest correlation values were observed for Italian samples (0.7245 ≤ r ≤ 0.9842; global r = 0.8958) and the lowest for Greek samples (0.0266 ≤ r ≤ 0.9703; global r = 0.5149). These results suggest that ITS2 metabarcoding offers a reliable alternative to classical palynology and this approach is now being employed in the European project INSIGNIA (https://www.insignia-bee.eu/), which is developing a standard protocol for using the honey bee as a tool for environmental monitoring.
