Richard N. Holdaway

The half-life of DNA in bone: measuring decay kinetics in 158 dated fossils

Claims of extreme survival of DNA have emphasized the need for reliable models of DNA degradation through time. By analysing mitochondrial DNA (mtDNA) from 158 radiocarbon-dated bones of the extinct New Zealand moa, we confirm empirically a long-hypothesized exponential decay relationship. The average DNA half-life within this geographically constrained fossil assemblage was estimated to be 521 years for a 242 bp mtDNA sequence, corresponding to a per nucleotide fragmentation rate (k) of 5.50 × 10–6 per year. With an effective burial temperature of 13.1°C, the rate is almost 400 times slower than predicted from published kinetic data of in vitro DNA depurination at pH 5. Although best described by an exponential model (R2 = 0.39), considerable sample-to-sample variance in DNA preservation could not be accounted for by geologic age. This variation likely derives from differences in taphonomy and bone diagenesis, which have confounded previous, less spatially constrained attempts to study DNA decay kinetics. Lastly, by calculating DNA fragmentation rates on Illumina HiSeq data, we show that nuclear DNA has degraded at least twice as fast as mtDNA. These results provide a baseline for predicting long-term DNA survival in bone.

A working list of breeding bird species of the New Zealand region at first human contact

Abstract We present an annotated working list of the bird species breeding in New Zealand during the late Pleistocene and Holocene, up to the time of human contact. New Zealand is defined as including the three main islands and the surrounding smaller islands, plus outlying island groups from Norfolk Island in the northwest, the Kermadec, Chatham, Bounty, Antipodes, Campbell, Auckland, Snares, to Macquarie Islands, but excluding islands south of Macquarie Island and the Ross Dependency. Inclusions or exclusions of species from the list were based on specified criteria. We include only species with a breeding population and not vagrants that occur in New Zealand but which breed elsewhere. Species with validly published names were included if there was fossil evidence for a breeding population before human contact. Species with a breeding population at the time of European contact were included unless contrary evidence from the fossil record indicates that they actually colonised after human settlement. Species without a fossil record were included if a breeding population exists on a relatively undisturbed island within the New Zealand archipelago as defined above. Species now present on the main islands were excluded if they are absent from all well‐documented fossil faunas. Species were excluded from the breeding fauna and treated as vagrants where sustained breeding has not been demonstrated. The phylogenetic species concept is applied both to fossil and to living taxa. The late Quaternary fossil record of birds in New Zealand is excellent, and the contribution of extinct taxa to the total list is understood at least as well as that of the surviving taxa. Many taxa presently recognised at subspecific level are treated here as full species. Twelve extinct species whose former presence is known from fossil evidence, but for which no description has been published, are listed under informal species designations. Taxonomic considerations limited the extent to which the main list could reflect present understanding of the diversity of the avifauna; some undescribed species are at present subsumed under one species name. Where previous taxonomic publications provide precedence, available names at the species‐level have been used. A supplementary hypothetical species list includes all nomenclatural changes signalled in extensive annotations to the main list. In this list we accept 245 species in 110 genera representing 46 families; 176 species were endemic to the archipelago. Preliminary biogeographic analyses based on the composition of the supplementary list show that there were four separate regional faunas: a northern subtropical fauna (Norfolk, Kermadecs); the major fauna of the main islands (North, South, Stewart, and offshore islands); a Chathams fauna (Chatham Islands only); and a subantarctic fauna on the southern islands. Species with wider distributions formed link groups. The origin and compositions of the regional avifaunas and their endemic species differ with their geographic position, climate, and proximity to source faunas. Instances of speciation in groups such as the Coenocoiypha snipe and Petroica flycatchers, and adaptive radiations in groups including moa and acanthisittid wrens, show that there are many avenues for research on the rate of evolution in island and mainland populations of New Zealand birds and that there are large gaps in knowledge of even common taxa. A brief case study demonstrates the inadequacies of using species lists that do not include Holocene fossil species. Species‐area curves based on the total fauna differ substantially from those developed in previous studies based on incomplete, or biased, lists. Pleistocene glaciations caused the pattern of distribution of species on the main islands to change in concert with vegetation changes. Other possible effects include the elimination of warm climate species early in the cooling phase more than 1 million years ago, the speciation in groups including waders and parrots as new habitats (e.g., braided riverbeds and alpine areas) appeared, and the appearance regularly during the Pleistocene of islands that were potential staging points for colonisation of the Chatham Islands. For at least the past 100 000 years, until 2000 years ago, the fauna appears to have been very stable in composition, despite strong cyclic fluctuations in climate and vegetation. The effects of extinctions within the past 2000 years on the composition of the present fauna include the elimination of most of the endemic taxa from all but the subantarctic faunas. Only 169 species of the original late Holocene breeding fauna survive. The extinctions have resulted in a strong bias towards marine and coastal taxa in the present avifauna, in contrast to the balanced representation of terrestrial and marine species in the Pleistocene and Holocene fauna. The importance of systematic studies and the determination of the status of island populations to conservation and basic ornithological research is emphasised. The systematic status of many New Zealand birds is poorly known at present.

The evolutionary history of the extinct ratite moa and New Zealand Neogene paleogeography

The ratite moa (Aves: Dinornithiformes) were a speciose group of massive graviportal avian herbivores that dominated the New Zealand (NZ) ecosystem until their extinction ≈600 years ago. The phylogeny and evolutionary history of this morphologically diverse order has remained controversial since their initial description in 1839. We synthesize mitochondrial phylogenetic information from 263 subfossil moa specimens from across NZ with morphological, ecological, and new geological data to create the first comprehensive phylogeny, taxonomy, and evolutionary timeframe for all of the species of an extinct order. We also present an important new geological/paleogeographical model of late Cenozoic NZ, which suggests that terrestrial biota on the North and South Island landmasses were isolated for most of the past 20–30 Ma. The data reveal that the patterns of genetic diversity within and between different moa clades reflect a complex history following a major marine transgression in the Oligocene, affected by marine barriers, tectonic activity, and glacial cycles. Surprisingly, the remarkable morphological radiation of moa appears to have occurred much more recently than previous early Miocene (ca. 15 Ma) estimates, and was coincident with the accelerated uplift of the Southern Alps just ca. 5–8.5 Ma. Together with recent fossil evidence, these data suggest that the recent evolutionary history of nearly all of the iconic NZ terrestrial biota occurred principally on just the South Island.

Identification of microsatellites from an extinct moa species using high-throughput (454) sequence data.

Genetic variation in microsatellites is rarely examined in the field of ancient DNA (aDNA) due to the low quantity of nuclear DNA in the fossil record together with the lack of characterized nuclear markers in extinct species. 454 sequencing platforms provide a new high-throughput technology capable of generating up to 1 gigabases per run as short (200-400-bp) read lengths. 454 data were generated from the fossil bone of an extinct New Zealand moa (Aves: Dinornithiformes). We identified numerous short tandem repeat (STR) motifs, and here present the successful isolation and characterization of one polymorphic microsatellite (Moa_MS2). Primers designed to flank this locus amplified all three moa species tested here. The presented method proved to be a fast and efficient way of identifying microsatellite markers in ancient DNA templates and, depending on biomolecule preservation, has the potential of enabling high-resolution population genetic studies of extinct taxa. As sequence read lengths of the 454 platforms and its competitors (e.g., the SOLEXA and SOLiD platforms) increase, this approach will become increasingly powerful in identifying microsatellites in extinct (and extant) organisms, and will afford new opportunities to study past biodiversity and extinction processes.

Fossil avian eggshell preserves ancient DNA

Owing to exceptional biomolecule preservation, fossil avian eggshell has been used extensively in geochronology and palaeodietary studies. Here, we show, to our knowledge, for the first time that fossil eggshell is a previously unrecognized source of ancient DNA (aDNA). We describe the successful isolation and amplification of DNA from fossil eggshell up to 19 ka old. aDNA was successfully characterized from eggshell obtained from New Zealand (extinct moa and ducks), Madagascar (extinct elephant birds) and Australia (emu and owl). Our data demonstrate excellent preservation of the nucleic acids, evidenced by retrieval of both mitochondrial and nuclear DNA from many of the samples. Using confocal microscopy and quantitative PCR, this study critically evaluates approaches to maximize DNA recovery from powdered eggshell. Our quantitative PCR experiments also demonstrate that moa eggshell has approximately 125 times lower bacterial load than bone, making it a highly suitable substrate for high-throughput sequencing approaches. Importantly, the preservation of DNA in Pleistocene eggshell from Australia and Holocene deposits from Madagascar indicates that eggshell is an excellent substrate for the long-term preservation of DNA in warmer climates. The successful recovery of DNA from this substrate has implications in a number of scientific disciplines; most notably archaeology and palaeontology, where genotypes and/or DNA-based species identifications can add significantly to our understanding of diets, environments, past biodiversity and evolutionary processes.

Quaternary fossil faunas from caves in Takaka Valley and on Takaka Hill, northwest Nelson, South Island, New Zealand

The late Quaternary fossil vertebrate faunas from 43 caves in Oligocene limestones and Ordovician marbles in the Takaka Valley and on Takaka Hill, northwest Nelson, New Zealand, are described and discussed. Depositional environments are described and interpreted. Major sites, including Ngarua Cave, Hawkes Cave, Kairuru Cave, Hobsons Tomo, and Irvines Tomo are described in detail. Many sites on Takaka Hill have been damaged by casual collectors since their discovery around 1900. Most sites were pitfall traps, but some deposits had been redistributed by water. Two deposits were attributed to an accumulation of material from pellets ejected by laughing owls (Sceloglaux albifacies), and of these the spectacularly rich Predator Cave site provided a large sample of small vertebrates. The fossil faunas included 42 species of land snails, three species of leiopelmatid frog, a tuatara, three species of geckoes, one or more species of skink, at least 58 (including two introduced) species of bird, three species of b...

A spatio‐temporal model for the invasion of the New Zealand archipelago by the Pacific rat Rattus exulans

New Zealand is at the southern limits of the human‐extended geographic range of the Pacific rat Rattus exulans On the two main islands, radiocarbon dates on rat bones from natural sites show that rat populations were established more than 1000 years before permanent human settlement, presumably during transient visits by Polynesian voyagers Both main islands were colonised after these first contacts, but offshore and outlying islands were not reached by rats until after Polynesian settlement about 700 years B P Chatham Island was not colonised by Pacific rats until about 650 years B P I present a model that relates the time of first appearance of rats in the fossil record and the exploitation of native fauna to the pattern of spread of the rat through the archipelago I hypothesise that the stepwise spread of the rat through the archipelago is mirrored by the pattern of reduction and extinction of indigenous fauna vulnerable to rat predation The 1000 year delay between the arrival of rats and permanent hum...

Quaternary fossil faunas, overlapping taphonomies, and palaeofaunal reconstruction in North Canterbury, South Island, New Zealand

This paper describes the late Quaternary fossil fauna from the area within a 10 km radius of Waikari, North Canterbury, New Zealand. Fossils from a pitfall deposit (Waikari Cave), ten predator sites attributed to laughing owls Sceloglaux albifacies (notably Ardenest, Gowan Hills Owl site, and P. Lambs Owl site), five swamp sites (notably Pyramid Valley and Glencrieff), and three archaeological sites, contributed most of the data. A few specimens came from coalluvial deposits in rock shelters. A small fauna is described from alluvial deposits along Home Creek, near Waipara. The age of these faunas includes the Late Pleistocene (Otira Glacial) for the Home Creek fauna, Late Glacial — early Holocene for Glencrieff, and Late Holocene for Pyramid Valley swamp, Waikari Cave, and all predator sites. Some of the predator sites accumulated fauna until late in the nineteenth century. The total avian fauna for the Waikari region (including Glenmark) comprises 65 indigenous and 4 introduced species of birds. In addi...

Ancient DNA Provides New Insights into the Evolutionary History of New Zealand's Extinct Giant Eagle

Prior to human settlement 700 years ago New Zealand had no terrestrial mammals—apart from three species of bats—instead, approximately 250 avian species dominated the ecosystem. At the top of the food chain was the extinct Haasts eagle, Harpagornis moorei. H. moorei (10–15 kg; 2–3 m wingspan) was 30%–40% heavier than the largest extant eagle (the harpy eagle, Harpia harpyja), and hunted moa up to 15 times its weight. In a dramatic example of morphological plasticity and rapid size increase, we show that the H. moorei was very closely related to one of the worlds smallest extant eagles, which is one-tenth its mass. This spectacular evolutionary change illustrates the potential speed of size alteration within lineages of vertebrates, especially in island ecosystems.

Non-Destructive Sampling of Ancient Insect DNA

Background A major challenge for ancient DNA (aDNA) studies on insect remains is that sampling procedures involve at least partial destruction of the specimens. A recent extraction protocol reveals the possibility of obtaining DNA from past insect remains without causing visual morphological damage. We test the applicability of this protocol on historic museum beetle specimens dating back to AD 1820 and on ancient beetle chitin remains from permafrost (permanently frozen soil) dating back more than 47,000 years. Finally, we test the possibility of obtaining ancient insect DNA directly from non-frozen sediments deposited 3280-1800 years ago - an alternative approach that also does not involve destruction of valuable material. Methodology/Principal Findings The success of the methodological approaches are tested by PCR and sequencing of COI and 16S mitochondrial DNA (mtDNA) fragments of 77–204 base pairs (-bp) in size using species-specific and general insect primers. Conclusion/Significance The applied non-destructive DNA extraction method shows promising potential on insect museum specimens of historical age as far back as AD 1820, but less so on the ancient permafrost-preserved insect fossil remains tested, where DNA was obtained from samples up to ca. 26,000 years old. The non-frozen sediment DNA approach appears to have great potential for recording the former presence of insect taxa not normally preserved as macrofossils and opens new frontiers in research on ancient biodiversity.