Ancient mitochondrial genomes recovered from small vertebrate bones through minimally destructive DNA extraction: Phylogeography of the New Zealand gecko genus Hoplodactylus

Abstract

Methodological and technological improvements are continually revolutionizing the field of ancient DNA. Most ancient DNA extraction methods require the partial (or complete) destruction of finite museum specimens, which disproportionately impacts small or fragmentary subfossil remains, and future analyses. We present a minimally destructive ancient DNA extraction method optimized for small vertebrate remains. We applied this method to detect lost mainland genetic diversity in the large New Zealand diplodactylid gecko genus Hoplodactylus, which is presently restricted to predator‐free island and mainland sanctuaries. We present the first mitochondrial genomes for New Zealand diplodactylid geckos, recovered from 19 modern, six historical/archival (1898–2011) and 16 Holocene Hoplodactylus duvaucelii sensu latu specimens, and one modern Woodworthia sp. specimen. No obvious damage was observed in post‐extraction micro‐computed tomography reconstructions. All “large gecko” specimens examined from extinct populations were found to be conspecific with extant Hoplodactylus species, suggesting their large relative size evolved only once in the New Zealand diplodactylid radiation. Phylogenetic analyses of Hoplodactylus samples recovered two genetically (and morphologically) distinct North and South Island clades, probably corresponding to distinct species. Finer phylogeographical structuring within Hoplodactylus spp. highlighted the impacts of Late Cenozoic biogeographical barriers, including the opening and closure of Pliocene marine straits, fluctuations in the size and suitability of glacial refugia, and eustatic sea‐level change. Recent mainland extinction obscured these signals from the modern tissue‐derived data. These results highlight the utility of minimally destructive DNA extraction in genomic analyses of less well studied small vertebrate taxa, and the conservation of natural history collections.

Application of palaeogenetic techniques to historic mollusc shells reveals phylogeographic structure in a New Zealand abalone

“…Preliminary shotgun sequencing data (not reported; available at: Walton et al, 2022 , https://doi.org/10.5061/dryad.3ffbg79mq ) generated from double‐stranded Illumina sequencing libraries created following the single‐tube protocol of Carøe et al ( 2018 ) yielded a very low proportion (<0.1%, Table S4 ) of target (haliotid mitogenomic) reads. Accordingly, we employed a single‐stranded Illumina sequencing library approach (Gansauge et al, 2017 ; described in full in Scarsbrook et al, 2022 , supporting information methods “C”) to improve recovery of highly fragmented DNA. Quantitative and indexing PCRs were carried out as above.…”

“…Bait generation and hybridization capture methods (described in full in: Scarsbrook et al, 2022 ) were adapted from Maricic et al ( 2010 ), Horn ( 2012 ) and González‐Fortes and Paijmans ( 2019 ) to selectively enrich haliotid mitogenomic DNA and to reduce the number of off‐target (i.e., exogenous) sequences recovered, such as from shell‐boring organisms. Biotinylated DNA baits were generated from long‐range PCR product from an ethanol‐preserved mainland specimen of H. virginea (M.333472, Table 1 ).…”

“…Biotinylated DNA baits were generated from long‐range PCR product from an ethanol‐preserved mainland specimen of H. virginea (M.333472, Table 1 ). Hybridization reactions contained: 1× hybridization buffer and blocking agent (both: Agilent), 2 μM each of BO1.P5.F and BO1.P7.F custom blocking oligos (Scarsbrook et al, 2022 ) and a 5:1 ratio of library to bait mix, and were incubated for 48 h at 55°C.…”

It is time for ancient DNA to sweat the small stuff

“…There are three possible solutions to sampling fossils for biomolecules: (i) accept that damage is made to the fossil; (ii) use fragmented bone that is of little use morphologically—[methods such as bulk‐bone metabarcoding (Murray et al, 2013) and ZOOMs (Buckley et al, 2009) take this approach to study biomolecules, and identify fragmentary fossil targets]; or (iii) develop new methods that are minimally destructive. Scarsbrook et al (2023) explore the latter of these approaches.…”

“…But here's the rub -extracting DNA from the bones of this 'small stuff' is not only difficult, it often destroys the sample. In this issue, Scarsbrook et al (2023) describe a new way to study the ancient (or historical) DNA of small vertebrates that is minimally destructive. The authors use the method to reconstruct the dynamic evolutionary history of New Zealand geckos and make new insights into how remnant populations should be managed.…”