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The impact of extraction method and pollen concentration on community composition for pollen metabarcoding - PubMed

  • ️Mon Jan 01 2024

. 2024 Aug 6;12(5):e11601.

doi: 10.1002/aps3.11601. eCollection 2024 Sep-Oct.

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The impact of extraction method and pollen concentration on community composition for pollen metabarcoding

Arne Devriese et al. Appl Plant Sci. 2024.

Abstract

Premise: Plants and pollinators closely interact with each other to form complex networks of species interactions. Metabarcoding of pollen collections has recently been proposed as an advantageous method for the construction of such networks, but the extent to which diversity and community analyses depend on the extraction method and pollen concentration used remains unclear.

Methods: In this study, we used a dilution series of two pollen mixtures (a mock community and pooled natural pollen loads from bumblebees) to assess the effect of mechanical homogenization and two DNA extraction kits (spin column DNA extraction kit and magnetic bead DNA extraction kit) on the detected pollen richness and community composition.

Results: All species were successfully detected using the three methods, even in the most dilute samples. However, the extraction method had a significant effect on the detected pollen richness and community composition, with simple mechanical homogenization introducing an extraction bias.

Discussion: Our findings suggest that all three methods are effective for detecting plant species in the pollen loads on insects, even in cases of very low pollen loads. However, our results also indicate that extraction methods can have a profound impact on the ability to correctly assess the community composition of the pollen loads on insects. The choice of extraction methodology should therefore be carefully considered to ensure reliable and unbiased results in pollen diversity and community analyses.

Keywords: Illumina sequencing; bumblebees; metabarcoding; plant–pollinator interactions; pollen.

© 2024 The Author(s). Applications in Plant Sciences published by Wiley Periodicals LLC on behalf of Botanical Society of America.

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Figures

Figure 1
Figure 1

Relative read abundance (proportions of reads detected for each species) in the mock community samples (A) and bumblebee samples (B) for each dilution step, shown for three different extraction methods. Species are ordered based on their expected proportion per sample in the mock community.

Figure 2
Figure 2

Non‐metric multidimensional scaling (NMDS) of the mock community pollen assemblage (A) and bumblebee pollen assemblage (B). Ellipses indicate 95% confidence level.

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