Historical Museum Specimens Research Articles

Museum epigenomics: Characterizing cytosine methylation in historic museum specimens.

Museum genomics has transformed the field of collections-based research, opening up a range of new research directions for paleontological specimens as well as natural history specimens collected over the past few centuries. Recent work demonstrates that it is possible to characterize epigenetic markers such as DNA methylation in wellpreserved ancient tissues. This approach has not yet been tested in traditionallyprepared natural history specimens such as dried bones and skins, the most common specimen types in vertebrate collections. In this study, we developed and tested methods to characterize cytosine methylation in dried skulls up to 76years old. Using a combination of ddRAD and bisulphite treatment, we characterized patterns of cytosine methylation in two species of deer mouse (Peromyscus spp.) collected in the same region in Michigan in 1940, 2003, and 2013-2016. We successfully estimated methylation in specimens of all age groups, although older specimens yielded less data and showed greater interindividual variation in data yield than newer specimens. Global methylation estimates were reduced in the oldest specimens (76years old) relative to the newest specimens (1-3years old), which may reflect post-mortem hydrolytic deamination. Methylation was reduced in promoter regions relative to gene bodies and showed greater bimodality in autosomes relative to female X chromosomes, consistent with expectations for methylation in mammalian somatic cells. Our work demonstrates the utility of historic specimens for methylation analyses, as with genomic analyses; however, studies will need to accommodate the large variance in the quantity of data produced by older specimens.

Sequence capture phylogenomics of historical ethanol-preserved museum specimens: Unlocking the rest of the vault.

Natural history collections play a crucial role in biodiversity research, and museum specimens are increasingly being incorporated into modern genetics-based studies. Sequence capture methods have proven incredibly useful for phylogenomics, providing the additional ability to sequence historical museum specimens with highly degraded DNA, which until recently have been deemed less valuable for genetic work. The successful sequencing of ultraconserved elements (UCEs) from historical museum specimens has been demonstrated on multiple tissue types including dried bird skins, formalin-fixed squamates and pinned insects. However, no study has thoroughly demonstrated this approach for historical ethanol-preserved museum specimens. Alongside sequencing of "fresh" specimens preserved in >95% ethanol and stored at -80°C, we used extraction techniques specifically designed for degraded DNA coupled with sequence capture protocols to sequence UCEs from historical museum specimens preserved in 70%-80% ethanol and stored at room temperature, the standard for such ethanol-preserved museum collections. Across 35 fresh and 15 historical museum samples of the arachnid order Opiliones, an average of 345 UCE loci were included in phylogenomic matrices, with museum samples ranging from six to 495 loci. We successfully demonstrate the inclusion of historical ethanol-preserved museum specimens in modern sequence capture phylogenomic studies, show a high frequency of variant bases at the species and population levels, and from off-target reads successfully recover multiple loci traditionally sequenced in multilocus studies including mitochondrial loci and nuclear rRNA loci. The methods detailed in this study will allow researchers to potentially acquire genetic data from millions of ethanol-preserved museum specimens held in collections worldwide.

Historical museum collections clarify the evolutionary history of cryptic species radiation in the world's largest amphibians.

Inaccurate taxonomic assessment of threatened populations can hinder conservation prioritization and management, with human‐mediated population movements obscuring biogeographic patterns and confounding reconstructions of evolutionary history. Giant salamanders were formerly distributed widely across China, and are interpreted as a single species, Andrias davidianus. Previous phylogenetic studies have identified distinct Chinese giant salamander lineages but were unable to associate these consistently with different landscapes, probably because population structure has been modified by human‐mediated translocations for recent commercial farming. We investigated the evolutionary history and relationships of allopatric Chinese giant salamander populations with Next‐Generation Sequencing methods, using historical museum specimens and late 20th‐century samples, and retrieved partial or near‐complete mitogenomes for 17 individuals. Samples from populations unlikely to have been affected by translocations form three clades from separate regions of China, spatially congruent with isolation by either major river drainages or mountain ranges. Pliocene–Pleistocene divergences for these clades are consistent with topographic modification of southern China associated with uplift of the Qinghai‐Tibet Plateau. General Mixed Yule Coalescent model analysis indicates that these clades represent separate species: Andrias davidianus (Blanchard, 1871) (northern Yangtze/Sichuan), Andrias sligoi (Boulenger, 1924) (Pearl/Nanling), and an undescribed species (Huangshan). Andrias sligoi is possibly the world's largest amphibian. Inclusion of additional reportedly wild samples from areas of known giant salamander exploitation and movement leads to increasing loss of biogeographic signal. Wild Chinese giant salamander populations are now critically depleted or extirpated, and conservation actions should be updated to recognize the existence of multiple species.

On the systematic status of the genus Oriocalotes Günther, 1864(Squamata: Agamidae: Draconinae) with the description of a new species from Mizoram state, Northeast India.

The montane agamid lizard genus Oriocalotes is currently considered monotypic, represented by the species, O. paulus. The systematic status of this taxon has remained questionable since its initial descriptions in the mid-1800s. A detailed molecular and morphological study was carried out to assess the validity of this genus, and its systematic position within the Asian agamid subfamily, Draconinae. Freshly collected and historical museum specimens from the type locality of O. paulus were examined morphologically, along with additional samples collected from localities in Mizoram state, Northeast India. Utilising newly generated molecular sequences (two mitochondrial and three nuclear genes), combined with those previously published for representative genera from the subfamilies Draconinae and Agaminae, Maximum Likelihood and Bayesian phylogenetic trees were constructed. Phylogenetic results suggest that Oriocalotes is part of the widespread South and Southeast Asian radiation of Calotes. Comparative morphological studies (including external morphology, hemipenis and osteology) between Oriocalotes and related genera further support this systematic placement. Oriocalotes is herein regarded as a junior subjective synonym of Calotes. Calotes paulus comb. nov. is also assigned a lectotype and given a detailed redescription based on the lectotype, paralectotypes and additional topotypic material. Furthermore, the specimens collected from Mizoram populations are found to be morphologically and genetically distinct from Calotes paulus comb. nov., and are described herein as a new species, Calotes zolaiking sp. nov.

Refined age and geological context of two of Australia’s most important Jurassic vertebrate taxa (Rhoetosaurus brownei and Siderops kehli), Queensland

Australia’s Jurassic vertebrate fossil record remains extremely sparse with only two dinosaur taxa and two temnospondyl amphibians identified to date. Of these, the spectacular and extremely well-preserved giant amphibian, Siderops kehli, and the only known pre-Cretaceous sauropod in Australia, Rhoetosaurus brownei, are perhaps the most important. The age of both specimens, and the stratigraphic context of Rhoetosaurus brownei, are weakly constrained and imprecisely defined, limiting our understanding of their evolutionary relationships within a broader Gondwanan context. To clarify and contextualise the evolutionary relationships and ages of these two iconic Jurassic taxa, we used UPb detrital zircon geochronology to date the sandstone matrix from around the bones of the historic museum specimens. The robust maximum depositional age for Siderops was calculated at 176.6 Ma ± 2 Ma, indicating that it is no older than late Toarcian, which refines existing biostratigraphic estimates. The Rhoetosaurus maximum depositional age was determined to be 162.6 ± 1.1 Ma, no older than early Oxfordian, demonstrating that the fossils are younger than expected, and definitely recovered from the Walloon Coal Measures.

Chiseling Away at the Dogma of Dietary Specialization in Dipodomys Microps

Dipodomys microps, the chisel-toothed kangaroo rat, is heralded as one of few mammalian herbivores capable of dietary specialization. Throughout its range, the diet of D. microps is thought to consist primarily of Atriplex confertifolia (saltbush), a C4 plant, and sparing amounts of C3 plants. Using stable isotopes of carbon and nitrogen as natural diet tracers, we asked whether D. microps is an obligate specialist on saltbush. We analyzed hair samples of D. microps for isotopes from historic and recent museum specimens (N = 66). A subset of samples (N = 17) from 2017 that were associated with field notes on plant abundances were further evaluated to test how local saltbush abundance affects its inclusion in the diet of D. microps. Overall, we found that the chisel-toothed kangaroo rat facultatively specializes on saltbush and that the degree of specialization has varied over time and space. Moreover, saltbush abundance dictates its inclusion in the diet. Furthermore, roughly a quarter of the diet is comprised of insects, and over the past century, insects have become more prevalent and saltbush less prevalent in the diet. We suggest that environmental factors such as climate change and rangeland expansion have caused D. microps to include more C3 plants and insects.

A new caruncle-bearing fanged frog (Limnonectes, Dicroglossidae) from Laos and Thailand.

A new species of the dicroglossid frog genus Limnonectes is described from recent and historical museum specimens collected in central and southern Laos and northeastern Thailand. Limnonectessavansp. nov. has males that bear a caruncle on top of the head, and most closely resembles L.dabanus from adjacent southern Vietnam and eastern Cambodia. However, the new species is readily distinguished from L.dabanus, and all other caruncle-bearing species of Limnonectes in mainland Southeast Asia, by its adult and larval morphology, mitochondrial DNA, and advertisement call. Its description brings the total number of caruncle-bearing species of Limnonectes to six.

Cryptic diversity and ranavirus infection of a critically endangered Neotropical frog before and after population collapse

AbstractMesoamerican amphibian declines in apparently pristine and protected habitats have been severe, especially at elevations above 500 m sea level and have been linked to emerging diseases and a changing climate. The Craugastor punctariolus species series of direct developing frogs is endemic to the region and used to be comprised of 33 species, seven of which have known populations at present. One of these, C. ranoides, endemic to southern Nicaragua and Costa Rica, was historically found in cloud forest sites of Área de Conservación Guanacaste (ACG) in north‐west Costa Rica and extended into dry forest sites 20 km distant. Here, C. ranoides declined and disappeared from high elevation sites between the mid‐1980s and early 1990s, but populations persisted in the lowland dry forest. We compared the genetic richness and ranavirus infection status of C. ranoides from extant dry forest populations to historic museum specimens of now extinct ACG cloud forest populations using DNA sequence diversity at two mitochondrial loci and molecular screening for ranavirus. Extant dry forest populations of C. ranoides formed a monophyletic group which included historic specimens sampled at cloud forest sites. However, the extirpated ACG cloud forest population contained additional diversity: samples formed a divergent clade with unknown spatial distribution. Ranavirus was detected in both current and museum samples of C. ranoides and sequences from a 267‐nucleotide region of the major capsid protein gene shared 100% sequence identity with one another and with Frog virus 3. Our findings document cryptic diversity within an endangered species that has demonstrated no recovery in cloud forests and raises questions about Ranavirus and its potential link to the amphibian declines in this system. The presence of the same C. ranoides clade within present day and historical samples suggests a potential for effective translocation and repopulation of extirpated cloud forest populations.

Museum genomics reveals the speciation history of Dendrortyx wood-partridges in the Mesoamerican highlands

Natural history collections are increasingly valued as genomic resources. Their specimens reflect the combined efforts of collectors and curators over hundreds of years. For many rare or endangered species, specimens are the only readily available source of DNA. We leveraged specimens from a historical collection to study the evolutionary history of wood-partridges in the genus Dendrortyx. The three Dendrortyx species are found in the highlands of central Mexico and Central America south to Costa Rica. One of these species is endangered, and in general, Dendrortyx are secretive and poorly represented in tissue collections. We extracted DNA from historical museum specimens and sequenced ultraconserved elements (UCEs) and mitochondrial DNA (mtDNA) to assess their phylogeny and divergence times. Phylogenies built from hundreds to thousands of nuclear markers were well resolved and largely congruent with an mtDNA phylogeny. The divergence times revealed an unusually old avian divergence across the Isthmus of Tehuantepec in the Pliocene around 3.6 million years ago. Combined with other recent studies, our results challenge the general pattern that highland bird divergences in Mesoamerica are relatively young and influenced by the Pleistocene glacial cycles compared to the older divergences of reptiles and plants, which are thought to overlap more with periods of mountain formation. We also found evidence for monophyletic genetic lineages in mountain ranges within the widespread D. macroura, which should be investigated further with integrative taxonomic methods. Our study demonstrates the power of museum genomics to provide insight into the evolutionary histories of groups where modern samples are lacking.

A century of decline: Loss of genetic diversity in a southern African lion‐conservation stronghold

AbstractAimThere is a dearth of evidence that determines the genetic diversity of populations contained within present‐day protected areas compared with their historical state prior to large‐scale species declines, making inferences about a species’ conservation genetic status difficult to assess. The aim of this paper was to demonstrate the use of historical specimens to assess the change in genetic diversity over a defined spatial area.LocationLike many other species, African lion populations (Panthera leo) are undergoing dramatic contractions in range and declines in numbers, motivating the identification of a number of lion‐conservation strongholds across East and southern Africa. We focus on one such stronghold, the Kavango–Zambezi transfrontier conservation area (KAZA) of Botswana, Namibia, Zambia and Zimbabwe.MethodsWe compare genetic diversity between historical museum specimens, collected during the late 19th and early 20th century, with samples from the modern extant population. We use 16 microsatellite markers and sequence 337 base pairs of the hypervariable control region (HVR1) of the mitochondrial genome. We use bootstrap resampling to allow for comparisons between the historical and modern data.ResultsWe show that the genetic diversity of the modern population was reduced by 12%–17%, with a reduction in allelic diversity of approximately 15%, compared to historical populations, in addition to having lost a number of mitochondrial haplotypes. We also identify a number of “ghost alleles” in the historical samples which are no longer present in the extant population.Main ConclusionsWe argue a rapid decline in allelic richness after 1895 suggests the erosion of genetic diversity coincides with the rise of a European colonial presence and the outbreak of rinderpest in the region. Our results support the need to improved connectivity between protected areas in order to prevent further loss of genetic diversity in the region.

Historical Genomes Reveal the Genomic Consequences of Recent Population Decline in Eastern Gorillas

Many endangered species have experienced severe population declines within the last centuries [1, 2]. However, despite concerns about negative fitness effects resulting from increased genetic drift and inbreeding, there is a lack of empirical data on genomic changes in conjunction with such declines [3-7]. Here, we use whole genomes recovered from century-old historical museum specimens to quantify the genomic consequences of small population size in the critically endangered Grauer's and endangered mountain gorillas. We find a reduction of genetic diversity and increase in inbreeding and genetic load in the Grauer's gorilla, which experienced severe population declines in recent decades. In contrast, the small but relatively stable mountain gorilla population has experienced little genomic change during the last century. These results suggest that species histories as well as the rate of demographic change may influence how population declines affect genome diversity.

Next-generation museum genomics: Phylogenetic relationships among palpimanoid spiders using sequence capture techniques (Araneae: Palpimanoidea)

Historical museum specimens are invaluable for morphological and taxonomic research, but typically the DNA is degraded making traditional sequencing techniques difficult to impossible for many specimens. Recent advances in Next-Generation Sequencing, specifically target capture, makes use of short fragment sizes typical of degraded DNA, opening up the possibilities for gathering genomic data from museum specimens. This study uses museum specimens and recent target capture sequencing techniques to sequence both Ultra-Conserved Elements (UCE) and exonic regions for lineages that span the modern spiders, Araneomorphae, with a focus on Palpimanoidea. While many previous studies have used target capture techniques on dried museum specimens (for example, skins, pinned insects), this study includes specimens that were collected over the last two decades and stored in 70% ethanol at room temperature. Our findings support the utility of target capture methods for examining deep relationships within Araneomorphae: sequences from both UCE and exonic loci were important for resolving relationships; a monophyletic Palpimanoidea was recovered in many analyses and there was strong support for family and generic-level palpimanoid relationships. Ancestral character state reconstructions reveal that the highly modified carapace observed in mecysmaucheniids and archaeids has evolved independently.

Ancient DNA tracks the mainland extinction and island survival of the Tasmanian devil

AbstractAimThe Tasmanian devil (Sarcophilus harrisii), currently restricted to the island of Tasmania, was found over most of the Australian mainland prior to its extinction ~3,000 years ago. Recent debate has focused on the roles of humans, climate change and dingoes as drivers of the mainland extinction. Determining past genetic diversity and population dynamics of both populations is a fundamental component to understand why the species went extinct on mainland Australia, but survived in Tasmania. Here, we investigate the phylogeography and demographic history of the Tasmanian devil across southern Australia over the last ~30k years.LocationAustralia.TaxonTasmanian devil (Sarcophilus harrisii).MethodsWe used complete and partial mitochondrial DNA (mtDNA) genomes from 202 devils representing the extinct mainland (n = 17) and the extant Tasmanian (n = 185) populations to investigate the population dynamics of southern mainland and Tasmanian devils. The samples were sub‐fossil bones, historical museum specimens and modern tissue samples, dating from the present to 17k years before present. Using summary statistics, frequentist inference and Bayesian phylogenetic analysis we explored whether levels of genetic diversity were similar, and if the southern mainland experienced a gradual rather than an abrupt decline prior to its extinction.ResultsMtDNA genomes from mainland devils suggest that this population was larger and had more genetic diversity than the Tasmanian population. Directly dated samples indicates that the southern mainland population expanded after the last glacial maximum and remained stable until its extinction. The Tasmanian population has much lower diversity and descends from a single mtDNA lineage ~3,000 years ago. The recent origin for all Tasmanian mtDNA diversity is concordant with a previously documented late‐Holocene population bottleneck and is broadly contemporaneous with the extinction of the southern mainland population.Main conclusionsThis pattern shows striking similarity to the demographic history of thylacines, suggesting that a shared factor initiated population declines in both species on the southern mainland and in Tasmania. El Niño Southern Oscillation (ENSO)‐related climate change is the only factor common to both mainland Australia and Tasmania. Additional, direct or indirect, pressures from humans and/or dingoes on the mainland may have ultimately resulted in their extinction.

Rapid Expansion of Banded Killifish Fundulus diaphanus across Northern Illinois: Dramatic Recovery or Invasive Species?

The distribution of the Illinois state-threatened Banded Killifish Fundulus diaphanus has remained largely unchanged in Illinois from 1880 to 2000, being restricted mainly to the northeastern corner of the state. One population identified as Western Banded Killifish F. d. menona has remained stable in the glacial lakes region along the southeastern Wisconsin–northeastern Illinois border. Starting in 2001 a second population began to spread and become more common along the Lake Michigan shoreline. From there the population expanded through the Chicago Area Waterway System into the Des Plaines River and eventually the Illinois River. Historical museum specimens from this region are identified as F. d. menona, but recent specimens are identified as hybrids between F. d. menona and Eastern Banded Killifish F. d. diaphanus. Subsequently, a third population appeared in the Mississippi River near the mouth of the Rock River in 2009 and has spread from there. These individuals are also identified as F. d. menona. The rapid expansion of Banded Killifish from Lake Michigan to the Illinois River appears to be an invasion of the Eastern subspecies F. d. diaphanus and the subsequent hybridization with the native Western subspecies. It is unknown where the Banded Killifish in the Mississippi River originated, possibly from populations 160 kilometers upstream or human introductions. As the Illinois River and Mississippi River populations continue to expand their ranges, their ecological impacts are unknown at this time.

Genome Sequencing of Museum Specimens Reveals Rapid Changes in the Genetic Composition of Honey Bees in California.

The western honey bee, Apis mellifera, is an enormously influential pollinator in both natural and managed ecosystems. In North America, this species has been introduced numerous times from a variety of different source populations in Europe and Africa. Since then, feral populations have expanded into many different environments across their broad introduced range. Here, we used whole genome sequencing of historical museum specimens and newly collected modern populations from California (USA) to analyze the impact of demography and selection on introduced populations during the past 105 years. We find that populations from both northern and southern California exhibit pronounced genetic changes, but have changed in different ways. In northern populations, honey bees underwent a substantial shift from western European to eastern European ancestry since the 1960s, whereas southern populations are dominated by the introgression of Africanized genomes during the past two decades. Additionally, we identify an isolated island population that has experienced comparatively little change over a large time span. Fine-scale comparison of different populations and time points also revealed SNPs that differ in frequency, highlighting a number of genes that may be important for recent adaptations in these introduced populations.

Historical Genomes Reveal the Genomic Consequences of Recent Population Decline in Eastern Gorillas

Many endangered species have experienced severe population declines within the last centuries. However, despite concerns about negative fitness effects resulting from increased genetic drift and inbreeding, there is a lack of empirical data on genomic changes in conjunction with such declines. Here, we use whole genomes recovered from century old historical museum specimens to quantify the genomic consequences of small population size in the critically endangered Grauer’s and mountain gorillas. We find a reduction of genetic diversity and increase in inbreeding and genetic load in the severely bottlenecked Grauer’s gorilla, whereas the small but relatively stable mountain gorilla has experienced little genomic change during the last century. These results suggest that species histories as well as the rate of demographic change may influence how population declines affect genome diversity.

Phylogeography of southern brown and golden bandicoots: implications for the taxonomy and distribution of endangered subspecies and species

Southern brown (Isoodon obesulus) and golden (Isoodon auratus) bandicoots are iconic Australian marsupials that have experienced dramatic declines since European settlement. Conservation management programs seek to protect the remaining populations; however, these programs are impeded by major taxonomic uncertainties. We investigated the history of population connectivity to inform subspecies and species boundaries through a broad-scale phylogeographic and population genetic analysis of Isoodon taxa. Our analyses reveal a major east–west phylogeographic split within I. obesulus/I. auratus, supported by both mtDNA and nuclear gene analyses, which is not coincident with the current species or subspecies taxonomy. In the eastern lineage, all Tasmanian samples formed a distinct monophyletic haplotype group to the exclusion of all mainland samples, indicative of long-term isolation of this population from mainland Australia and providing support for retention of the subspecific status of the Tasmanian population (I. o. affinis). Analyses further suggest that I. o. obesulus is limited to south-eastern mainland Australia, representing a significant reduction in known range. However, the analyses provide no clear consensus on the taxonomic status of bandicoot populations within the western lineage, with further analyses required, ideally incorporating data from historical museum specimens to fill distributional gaps.

Intersection of historical museum collections and modern systematics: a relict population of the Arctic nudibranch Dendronotus velifer G.O. Sars, 1878 in a Swedish fjord

Based on morphological, bathymetric and molecular data comparing recently collected Arctic and North Atlantic specimens with morphological and bathymetrical data on historical museum specimens, a unique relict population of the deep-water mollusc Dendronotus velifer G.O. Sars, 1878 (Gastropoda: Nudibranchia) is shown to have existed in the deepest section of the Swedish Gullmar Fjord (the only true silled fjord in Sweden) at least until the middle of the 20th century. This population is more than 1500 km away from the nearest point in the species’ distributional range in the Arctic Ocean today. Using an integrative approach incorporating the data mentioned above, including genetic distances, from recently collected specimens taken from the Arctic Ocean, D. velifer is validated and its species status is restored, for the first time in more than a century after being regarded as a junior synonym of D. robustus. The bathymetric data for historical and recently collected specimens of D. velifer demonstrate significant differences compared to the shallow-water species D. robustus. The findings support the necessity of a stronger protection for the unique marine habitats of the Gullmar Fjord.

Historical museum specimens reveal the loss of genetic and morphological diversity due to local extinctions in the endangered water chestnut Trapa natans L. (Lythraceae) from the southern Alpine lake area

Freshwater aquatic plants have increased extinction risks due to strong human pressure and the patchy nature of their habitats. However, their unclear population structure frequently hinders conservation planning. To investigate population subdivision and risks to long-term survival of such species, we combined genetic, morphometric and biogeographical approaches to the example of the endangered water chestnut Trapa natans (Lythraceae) from the southern Alpine lake area (Insubria). Amplified fragment length polymorphism (AFLP) of seven extant local stands revealed similar and intermediate levels of genetic diversity, of which c. 97% was partitioned within lakes. Thus, no signs of strong genetic drift and associated loss of genetic diversity were found, despite a reduction of c. 52% of local populations since the early 19th century. Nuclear ribosomal sequences (ITS1-5.8S rRNA-ITS2) combined with a morphometric study of nuts (based on fresh and historical museum material) revealed the presence of two genetically and morphologically slightly distinct lineages, one of which went extinct during the 20th century after a prolonged period of hyper-eutrophication. Taken together, our results indicate the current presence of one large Insubric Trapa population. To prevent genetic risks to survival associated with small population size and increasing fragmentation due to past extinctions, freshwater managers should preserve the large census sizes still present in many Insubric lakes and reduce eutrophication.

Museum Genomics Confirms that the Lord Howe Island Stick Insect Survived Extinction

The Lord Howe Island stick insect, Dryococelus australis, was once common on the island but was driven to extinction after the arrival of ship rats in the early 20th century [1, 2]. It was thought to be extinct for decades, until a tiny population of similar-looking stick insects was discovered 20km away, on the islet of Ball's Pyramid, in 2001 [2]. Individuals from this population are currently being reared in Australia and elsewhere in the world, with the eventual goal of recolonizing Lord Howe Island [3]. Recent surveys of the wild population on Ball's Pyramid suggest that it is among the world's rarest species. However, there are significant morphological differences between Ball's Pyramid and museum specimens, and there has never been a genetic confirmation of the rediscovered population's species identity. Because Dryococelus is monotypic, there are also no known extant relatives for comparison. Using shotgun genomic data from the Ball's Pyramid population, we assembled a draft genome and the complete mitochondrial genome. We found that the genome is massive, over 4 Gb in size, and is most likely hexaploid. We re-sequenced mitochondrial genomes from historic museum specimens collected on Lord Howe Island before the extinction event. Sequence divergence between the two populations is less than 1% and is within the range of intraspecific differences between the museum specimens, suggesting that they are conspecific and that D.australis has successfully evaded extinction so far. This work highlights the importance of museum collections for taxonomic validation in the context of ongoing conservation efforts.