From museum drawer to tree: Historical DNA phylogenomics clarifies the systematics of rare dung beetles (Coleoptera: Scarabaeinae) from museum collections

Abstract

Although several methods exist for extracting and sequencing historical DNA originating from dry-preserved insect specimens deposited in natural history museums, no consensus exists as to what is the optimal approach. We demonstrate that a customized, low-cost archival DNA extraction protocol (∼€10 per sample), in combination with Ultraconserved Elements (UCEs), is an effective tool for insect phylogenomic studies. We successfully tested our approach by sequencing DNA from scarab dung beetles preserved in both wet and dry collections, including unique primary type and rare historical specimens from internationally important natural history museums in London, Paris and Helsinki. The focal specimens comprised of enigmatic dung beetle genera (Nesosisyphus, Onychothecus and Helictopleurus) and varied in age and preservation. The oldest specimen, the holotype of the now possibly extinct Mauritian endemic Nesosisyphus rotundatus, was collected in 1944. We obtained high-quality DNA from all studied specimens to enable the generation of a UCE-based dataset that revealed an insightful and well-supported phylogenetic tree of dung beetles. The resulting phylogeny propounded the reclassification of Onychothecus (previously incertae sedis) within the tribe Coprini. Our approach demonstrates the feasibility and effectiveness of combining DNA data from historic and recent museum specimens to provide novel insights. The proposed archival DNA protocol is available at DOI 10.17504/protocols.io.81wgbybqyvpk/v3.

Fig 1. Dung beetle tree.

(A) Species tree inferred using 1,497 UCEs and the 50% complete dataset; dry-preserved historical museum specimens are indicated in bold. Collapsed branches are proportional to the number of samples in each lineage. Hollow dots indicate fully supported nodes and nodes with numbers indicate bootstrap values < 100. (Oni) Onitini; (Ont) Onthophagini + Oniticellini; (Onc) Oniticellini; (M2, M1) Madagascan endemic lineages; (Sc) Scarabaeini; (Ph) Phanaeini; (Aus) Australasian endemic genera. For details, see S1 Table. (B) Dorsal view of the holotype of Neosisyphus rotundatus before and after DNA extraction using the archival protocol.

Fig 2. Morphology of Onychothecus tridentigeris.

Dorsal habitus of male (A) and female (B); ventral view of female (C); right wing in dorsal view, with radial posterior vein 1 (RP1) indicated (D); left elytron in lateral view, indicating the numbered elytral striae and the lateral carina (E); hind tarsus, with the modified terminal tarsomere concealing the claws (F); right protibia of male, in dorsal view (G); aedeagus in lateral (H) and dorsal (I) views; endophallites (J) (abbreviations follow Tarasov & Génier (2015) [33]).

Fig 3. Summary of UCE data resulting from two DNA extraction methods (archival extraction protocol in red and standard extraction in blue) and beetle genomes from GenBank (in green, only in B).

Violin plots illustrate the kernel density and boxplots display the median and variation. (A) Distribution of read length generated per sample, demonstrating that the density of shorter reads was generally higher from archival extractions. (B) Distribution of the number of UCE loci per sample, demonstrating that the greatest density of samples that generated large numbers of captured loci resulted from archival extractions.