Metagenomic sequencing of dung beetle intestinal contents directly detects and identifies mammalian fauna

Abstract

SummaryCost, time, and expertise constraints limit traditional observation-based comprehensive biodiversity assessment. Therefore, surrogate focal taxa representative of wider biodiversity are commonly used as an imperfect ‘proxy’. Contemporary biodiversity assessments are also increasingly benefiting from the combination of high-throughput sequencing and metagenomic methodologies that enable identification of environmental DNA samples. However, there is a need for empirical studies combining the use of surrogate taxa with metagenomic approaches, that promise rapid and efficient biodiversity assessment.We here tested for the first time the possibility of using the intestinal contents of wild-collected dung beetles (Scarabaeidae) as a source of mammalian DNA, in a metagenomics proof-of-concept approach to directly detect and identify mammals from an area of savanna-scrub in southern Africa. Dung beetles have been purveyed as an indirect proxy measure of mammalian diversity, owing to their dependence upon vertebrate dung as a food source, and the ease with which they can be comprehensively sampled using simple and repeatable trapping protocols, achievable much faster than vertebrate surveys.Following shotgun sequencing of gut content DNA extractions from ten dung beetle species, we used in silico filters to identify mammals by searching the resulting reads against known mammalian mitochondrial DNA from online sequence repositories, matching 546 paired reads to known mitogenomes held in GenBank, and 634 reads to known mammal barcode sequences held in BOLD. Identified mammalian sequences were consistent with wild and domesticated ungulates known from the sampling site, and included blue wildebeest, plains zebra, and domestic cattle and goat. Four dung beetle samples yielded sufficient sequence data to successfully assemble the near-complete mitogenome of blue wildebeest at up to 21 X mean coverage, despite low initial DNA concentrations, unambiguously corroborating identification.It is conceptually and practically possible to rapidly and economically apply metagenomic techniques in dung beetle gut sequencing to detect the presence of mammals upon whose dung the beetles have fed. Since the approach can be readily scaled up, it may prove to be of practical use as a complement to traditional biodiversity assessment methods, and should be tested in usefulness for detecting rare, endangered or cryptic mammal species.

Unexpectedly diverse forest dung beetle communities in degraded rain forest landscapes in Madagascar

“…One way to explore this would be by identifying the food remains in the guts of different dung beetle species by molecular methods. This would inform us about the taxa these species use either as carrion or dung (Gillett, Johnson, Barr, & Hulcr, 2016;Gómez & Kolokotronis, 2017). In addition to unveiling the direct trophic interactions, identifying gut contents would enable describing how wide generalists or narrow specialists different species are, which remains largely unknown for dung beetles worldwide (Raine & Slade, 2019).…”

Carrion fly‐derived DNA metabarcoding is an effective tool for mammal surveys: Evidence from a known tropical mammal community

“…So far, the most promising approach for use across a broad range of species is mass trapping and metabarcoding of invertebrates that feed on vertebrates (Bohmann, Schnell, & Gilbert, ; Calvignac‐Spencer, Leendertz, Gilbert, & Schubert, ). Such invertebrate “samplers” tested to date include leeches (Schnell et al., ), mosquitos (Logue et al., ), dung beetles (Gillett, Johnson, Barr, & Hulcr, ) and carrion flies (Calvignac‐Spencer, Merkel et al., ; Lee, Gan, Clements, & Wilson, ; Lee, Sing, & Wilson, ; Schubert et al., ). We focus on the latter.…”

Carrion fly-derived DNA metabarcoding is an effective tool for mammal surveys: evidence from a known tropical mammal community

“…So far, the most promising approach for use across a broad range of species is mass trapping and metabarcoding of invertebrates that feed on vertebrates (Bohmann et al 2013;Calvignac-Spencer et al 2013a). Such invertebrate 'samplers' tested to date include leeches (Schnell et al 2015), mosquitos , dung beetles (Gillett et al 2016) and carrion flies (Calvignac-Spencer et al 2013b;Lee et al 2015;Schubert et al 2014).…”