Creating genetic reference datasets: Indirect sampling of target species using terrestrial leeches as sample “collectors”

Abstract

Sampling genetic material from rare and often secretive mammals can be difficult in challenging environments such as tropical rainforests. Large-scale sampling is important however for resolving species' taxonomic uncertainties, as well as to help provide genetic material for reference databases that can be used with DNA forensics to combat the illegal wildlife trade. Environmental DNA (eDNA) and invertebrate-derived DNA (iDNA) offer a promising way to overcome this sampling bottleneck, though to date e/iDNA studies have been primarily used to obtain information about species presence using barcoding or metabarcoding approaches. In this study, we collected 4,710 terrestrial leeches across six protected areas in the Annamite Mountains of Viet Nam and Laos and sequenced fragments (223-694 bp) of mt-Cytb and/or mt-Dcr of the mitogenomes from six species that are frequently found in the illegal wildlife trade. Although amplification success was generally low (<50%) because DNA was highly degraded, we showed that e/iDNA samples can be an important source of genetic material for difficult to sample species and show promise as a way to fill existing sampling gaps. Because e/iDNA sampling has the potential to generate much-needed genetic data to improve taxonomic assessments, this approach can be used to support conservation management by contributing to genetic databases that are necessary to determine the origin of wildlife products or to inform the reintroduction of confiscated animals.

Assessing environmental DNA metabarcoding and camera trap surveys as complementary tools for biomonitoring of remote desert water bodies

“…However, to rely on cameras implies the risk of losing data (over 40 days, 1/14 cameras were lost) and demands repeated visits which may be complicated in remote areas (at least installation and recovery of cameras), where eDNA sampling offers valuable study opportunities (McInnes et al, 2021). Also, CT does not offer the same options as eDNA in terms of possible population genetic studies (Bohmann et al, 2018; Nguyen et al, 2021; Sigsgaard et al, 2016, 2020; Tilker et al, 2020; Wilting et al, 2021) and the extension to other phyla (as, e.g., invertebrates, plants, and bacteria). eDNA sampling is appealing due to its sensitivity, standardization, and non‐invasiveness, as well as the independence from taxonomic experts for taxa identification.…”

“…Environmental DNA from terrestrial animals has been mostly assessed by analyzing scats (De Barba et al, 2014; Kartzinel et al, 2015; Swift et al, 2018), soil (Leempoel et al, 2020; Yoccoz et al, 2012; Zinger et al, 2019), stomach content samples (Kennedy et al, 2019; Masonick et al, 2019; Soininen et al, 2013), leeches blood meals (Abrams et al, 2019; Nguyen et al, 2021; Tilker et al, 2020; Weiskopf et al, 2018; Wilting et al, 2021), or carrion flies (Calvignac‐Spencer et al, 2013; Gogarten et al, 2020; Rodgers et al, 2017; Schubert et al, 2015). Bulk tissue samples (mixtures of, e.g., insects or other macroinvertebrate specimens) are also increasingly used not only to assess invertebrate diversity but also as an indirect way to sample vertebrate DNA (Lynggaard et al, 2019).…”

Comparison of mosquito and fly derived DNA as a tool for sampling vertebrate biodiversity in suburban forests in Berlin, Germany

“…These arthropods have been limited to hematophagous insects such as mosquitoes, ticks, blackflies, triatomine bugs, and sandflies. In recent years, the extension from vectors to other invertebrates such as leeches (Alfano et al, 2021; Schnell et al, 2012; Wilting et al, 2021) and flies (Calvignac‐Spencer et al, 2013) and the development of the metabarcoding protocol have enabled the sampling of vertebrates through invertebrate samples beyond the scope of disease transmission. This resulted in the development of invertebrate‐derived DNA (iDNA) within the umbrella of environmental DNA and its use as monitoring tool, also for rare and elusive species (Nguyen et al, 2021).…”

Analysing the effects of distance, taxon and biomass on vertebrate detections using bulk-collected carrion fly iDNA

“…While it is possible to obtain DNA from reptile faeces to conduct monitoring and analysis [65][66][67], this is usually dependent on the collection of fresh samples [65]. Additionally, iDNA degrades quickly within the digestive tract of invertebrates [27,68]. It is worth noting that despite attempts to detect various different classes of animals, overall detection was poor throughout the study, and no significant differences in detection were observed between taxonomic classes.…”