eDNA analysis of yard waste samples reveals taxonomical diversity, sequence database limitations, and consistencies across sequencing platforms

Authors: Walker Iii, William; Neven, Lisa

Submitted to: Journal of Insect Science
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/3/2026
Publication Date: 7/8/2026
Citation: Walker III, W.B., Neven, L.G. 2026. eDNA analysis of yard waste samples reveals taxonomical diversity, sequence database limitations, and consistencies across sequencing platforms. Journal of Insect Science. 26(4). Article ieag062. https://doi.org/10.1093/jisesa/ieag062.
DOI: https://doi.org/10.1093/jisesa/ieag062

Interpretive Summary: Recent breakthroughs in advanced DNA sequencing technologies have facilitated substantial increases in the usefulness of a type of technology, known as DNA-barcoding, to identify and characterize distinct biological species at the DNA level. One application of this approach, known as environmental DNA (eDNA) analysis, involves extracting DNA from environmental samples, then sequencing barcoding marker genes present in the DNA extractions, and finally, characterizing species present in the environment by comparison of the identified sequences to known sequences deposited in public gene databases. This approach enables complex biological and ecological research questions to be examined without the need to have direct access to or observation of whole organisms. USDA researchers in Wapato, WA, assessed the efficiency and effectiveness of two different advanced DNA sequencing technology platforms in the sequencing of eDNA derived from nine common yard waste samples. For this, a single DNA barcoding marker gene was analyzed, and sequenced DNA units from each sample were compared to definitive sequences in public DNA-sequence depositories. The findings of this study highlight benefits and limitations of this methodological approach. This research will be useful to the scientific community in raising greater awareness of the approach, and may spur its utilization in examining economically relevant biological research topics in the agricultural sector.

Technical Abstract: Timely identification of biological species is often needed for various purposes including economic reasons, and advances in DNA sequencing technologies have greatly augmented the ability to identify species through the application of DNA barcoding. One such method examines environmental DNA (eDNA) to sample presence of organisms in an environment without necessarily having direct access to the whole organisms. In recent years, multiple high-throughput sequencing platforms have emerged, and there are differences in the efficiency, effectiveness and economics across these platforms. In this report, we examine application of two platforms, from PacBio and Oxford Nanopore Technologies, to sequence COI amplicons from nine barcoded yard waste samples that we previously studied for a different purpose. Here, we observed consistencies across the platforms in the identification of operational taxonomical units (OTUs) from broad swaths of life, most prominently including Bacteria, Amoebozoa, Fungi, Arthropoda, Nematoda, Spiralia and Viridiplantae. Other taxonomical groupings were also tentatively identified. However, limitations in coverage of diversity of COI sequences in the public databases rendered species level identification impossible for many of the OTUs. Insect species were the best represented across all barcoded samples and both sequencing platforms regarding percentage identity to best BLAST hits in the databases. Following this, we took an in-depth look at knowledge of presence of highly matched species in the locality from where the eDNA samples were derived. Strengths and limitations of this approach in the analysis of eDNA are discussed.