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Analysis of environmental DNA detected in lake sediments shows promise to become a great paleoecological technique that can provide detailed information about organism communities living in past environments. However, when interpreting sedimentary environmental DNA records, it is of crucial importance to separate ecosystem responses to large-scale environmental change from “noise” caused by changes in sediment provenance or potential post-depositional DNA mobility. In this study, we show that plant and mammalian communities reconstructed from sediments are strongly affected by sediment provenance, but unaffected by vertical mobility of DNA after sediment deposition. We observe that DNA from aquatic plants was abundant in background sediment, while embedded detrital event layers (sediment deposited under erosion events) primarily contained terrestrial plants; hence, vertical mobility of aquatic plant DNA across sediment layers was negligible within our studied lakes. About 33% of the identified terrestrial plant genera were only found in detrital sediment, suggesting that sediment origin had a strong impact on the reconstructed plant community. Similarly, DNA of some mammalian taxa (Capra hircus, Ursus arctos, Lepus, and Felis) were only or preferentially found in detrital event layers. Temporal changes across the Holocene were the main drivers of change for reconstructed plant communities, but sediment type was the second most important factor of variance. Our results highlight that erosion and sediment provenance need to be considered when reconstructing past mammalian and plant communities using environmental DNA from lake sediments.

Earthworms are soft tissue organisms that rarely leave fossils that can be used to identify species. Absence of fossils makes the natural history of earthworm species in post-glacial landscapes of Fennoscandia largely unknown. Analyses of environmental DNA (eDNA) preserved in natural archives such as lake sediments and buried soil layers (paleosols) may offer an opportunity to assess the composition of past earthworm communities. In this thesis, I explore the use of metabarcoding as an analytical method to detect DNA from earthworms that lived in past European environments. I aimed at extracting DNA from various forms of paleosols in Europe and lake sediments, but earthworm DNA is rare in these deposits and amplifying DNA from this group of soil fauna was largely unsuccessful. However, during the scientific progression of my work, I discovered that metabarcoding-based studies are sensitive to ‘tag jumping’, which is a process where sample specific labels (tags) added to sequences for identification of individual samples ‘jump’, resulting in crosstalk between samples. My results suggest that tag jumping i) is mediated by the formation of heteroduplexes (DNA with two strands from different samples), ii) affects interpretations of eDNA studies by adding species to samples where they were not originally present, and iii) makes eDNA assemblages more similar. Importantly, my results also highlight that metabarcoding can generate powerful and trustworthy reconstructions of past environments if conducted with protocols that remove the influence of tag jumps. Reconstructions of terrestrial organisms from eDNA in sediments are also enhanced by erosion events that amplify DNA signals of landliving organisms. I conclude that earthworm DNA is difficult to detect in natural archives using current metabarcoding techniques and that tag jumping, a problem rarely discussed in metabarcoding studies, constitutes a concern in parity with direct sample contamination. 

Metabarcoding of environmental DNA constitutes a state-of-the-art tool for environmental studies. One fundamental principle implicit in most metabarcoding studies is that individual sample amplicons can still be identified after being pooled with others—based on their unique combinations of tags—during the so-called demultiplexing step that follows sequencing. Nevertheless, it has been recognized that tags can sometimes be changed (i.e., tag jumping), which ultimately leads to sample crosstalk. Here, using four DNA metabarcoding data sets derived from the analysis of soils and sediments, we show that tag jumping follows very specific and systematic patterns. Specifically, we find a strong correlation between the number of reads in blank samples and their topological position in the tag matrix (described by vertical and horizontal vectors). This observed spatial pattern of artefactual sequences could be explained by polymerase activity, which leads to the exchange of the 3′ tag of single stranded tagged sequences through the formation of heteroduplexes with mixed barcodes. Importantly, tag jumping substantially distorted our data sets—despite our use of methods suggested to minimize this error. We developed a topological model to estimate the noise based on the counts in our blanks, which suggested that 40%–80% of the taxa in our soil and sedimentary samples were likely false positives introduced through tag jumping. We highlight that the amount of false positive detections caused by tag jumping strongly biased our community analyses.