Direct PCR meets high-throughput sequencing - metabarcoding of aquatic insect communities without DNA purification

Abstract

Aquatic emergent insect communities form an important link between aquatic and terrestrial ecosystems, yet studying them is costly and time-consuming as they are usually diverse and superabundant. Metabarcoding is a valuable tool to investigate arthropod community compositions, however high-throughput applications, such as for biomonitoring, require cost-effective and user-friendly procedures. To investigate if the time-consuming and labour-intensive DNA purification step can be omitted in metabarcoding, we studied the difference in detection rates and individual read abundance using standard DNA purification versus direct PCR protocols. Metabarcoding with and without DNA purification was performed with artificially created communities of known composition as well as on natural communities both of the dipteran family Chironomidae to compare detection rates, individual read abundances and presence-absence community composition. We found that the here presented novel approach of direct PCR metabarcoding did not alter detection rates and had a minor effect on individual read abundances in artificially created communities. Furthermore, presence-absence community compositions of natural chironomid communities were highly comparable using both approaches. In conclusion, we showed that direct PCR protocols can be applied in chironomid metabarcoding approaches, with possible application for a wider range of arthropod taxa, enabling us to study communities more efficiently in the future.

Direct PCR meets high-throughput sequencing – metabarcoding of chironomid communities without DNA extraction

“…Raw sequences were deposited in GenBank SRA archive and are available with the Bio-Project accession number PRJNA989176. Data and R scripts are available from Zenodo https://doi.org/10.5281/zenodo.8074454 (Röder and Schwenk 2023).…”