Mitochondrial genomes reveal mid-Pleistocene population divergence, and post-glacial expansion, in Australasian snapper (Chrysophrys auratus).

Glacial cycles play important roles in determining the phylogeographic structure of terrestrial species, however, relatively little is known about their impacts on the distribution of marine biota. This study utilised modern (n = 350) and ancient (n = 26) mitochondrial genomes from Australasian snapper (Chrysophrys auratus) sampled in New Zealand to assess their demographic and phylogeographic history. We also tested for changes in genetic diversity using the up to 750-year-old mitochondrial genomes from pre-European archaeological sites to assess the potential impacts of human exploitation. Nucleotide diversity and haplotype diversity was high (π = 0.005, h = 0.972). There was no significant change in nucleotide diversity over the last 750 years (p = 0.343), with no detectable loss of diversity as a result of indigenous and industrial-scale fishing activity. While there was no evidence for contemporary population structure (AMOVA, p = 0.764), phylogeographic analyses identified two distinct mitochondrial clades that diverged approximately 650,000 years ago during the mid-Pleistocene, suggesting the species experienced barriers to gene flow when sea levels dropped over 120 m during previous glacial maxima. An exponential population increase was also observed around 8000 years ago consistent with a post-glacial expansion, which was likely facilitated by increased ocean temperatures and rising sea levels. This study demonstrates that glacial cycles likely played an important role in the demographic history of C. auratus and adds to our growing understanding of how dynamic climatic changes have influenced the evolution of coastal marine species.

Description of the growth hormone gene of the Australasian snapper, Chrysophrys auratus, and associated intra- and interspecific genetic variation.

The growth hormone (GH) gene of the marine teleost, the Australasian snapper (Chrysophrys auratus), was identified and characterized from the reference genome showing it was approximately 5577 bp in length and consisted of six exons and five introns. Large polymorphic repeat regions were found in the first and third introns, and putative transcription factor binding sites were identified. Phylogenetic analysis of the GH genes of perciform fish showed largely conserved coding regions and highly variable noncoding regions among species. Despite some exon sequence variation and an amino acid deletion identified between C. auratus and its sister species Chrysophrys/Pagrus major, the amino acid sequences and putative secondary structures were largely conserved across the Sparidae. A population-level assessment of 99 samples caught at five separate coastal locations in New Zealand revealed six variable alleles at the intron 1 site of the C. auratus GH gene. A population genetic analysis suggested that C. auratus from the five sample locations were largely panmictic, with no evidence for departure from the Hardy-Weinberg equilibrium, and have a high level of heterozygosity. Overall these results suggest that the GH gene is largely conserved across the coding regions, but some variability could be detected.

Fifteen years of quantitative trait loci studies in fish: challenges and future directions.

Understanding the genetic basis of phenotypic variation is a major challenge in biology. Here, we systematically evaluate 146 quantitative trait loci (QTL) studies on teleost fish over the last 15 years to investigate (i) temporal trends and (ii) factors affecting QTL detection and fine-mapping. The number of fish QTL studies per year increased over the review period and identified a cumulative number of 3632 putative QTLs. Most studies used linkage-based mapping approaches and were conducted on nonmodel species with limited genomic resources. A gradual and moderate increase in the size of the mapping population and a sharp increase in marker density from 2011 onwards were observed; however, the number of QTLs and variance explained by QTLs changed only minimally over the review period. Based on these findings, we discuss the causative factors and outline how larger sample sizes, phenomics, comparative genomics, epigenetics and software development could improve both the quantity and quality of QTLs in future genotype-phenotype studies. Given that the technical limitations on DNA sequencing have mostly been overcome in recent years, a renewed focus on these and other study design factors will likely lead to significant improvements in QTL studies in the future.

Genome assembly and isoform analysis of a highly heterozygous New Zealand fisheries species, the tarakihi (Nemadactylus macropterus).

Although being some of the most valuable and heavily exploited wild organisms, few fisheries species have been studied at the whole-genome level. This is especially the case in New Zealand, where genomics resources are urgently needed to assist fisheries management. Here, we generated 55 Gb of short Illumina reads (92× coverage) and 73 Gb of long Nanopore reads (122×) to produce the first genome assembly of the marine teleost tarakihi [Nemadactylus macropterus (Forster, 1801)], a highly valuable fisheries species in New Zealand. An additional 300 Mb of Iso-Seq reads were obtained to assist in gene annotation. The final genome assembly was 568 Mb long with an N50 of 3.37 Mb. The genome completeness was high, with 97.8% of complete Actinopterygii Benchmarking Universal Single-Copy Orthologs. Heterozygosity values estimated through k-mer counting (1.00%) and bi-allelic SNPs (0.64%) were high compared with the same values reported for other fishes. Iso-Seq analysis recovered 91,313 unique transcripts from 15,515 genes (mean ratio of 5.89 transcripts per gene), and the most common alternative splicing event was intron retention. This highly contiguous genome assembly and the isoform-resolved transcriptome will provide a useful resource to assist the study of population genomics and comparative eco-evolutionary studies in teleosts and related organisms.

Mutation and evolutionary rates in adélie penguins from the antarctic.

Precise estimations of molecular rates are fundamental to our understanding of the processes of evolution. In principle, mutation and evolutionary rates for neutral regions of the same species are expected to be equal. However, a number of recent studies have shown that mutation rates estimated from pedigree material are much faster than evolutionary rates measured over longer time periods. To resolve this apparent contradiction, we have examined the hypervariable region (HVR I) of the mitochondrial genome using families of Adélie penguins (Pygoscelis adeliae) from the Antarctic. We sequenced 344 bps of the HVR I from penguins comprising 508 families with 915 chicks, together with both their parents. All of the 62 germline heteroplasmies that we detected in mothers were also detected in their offspring, consistent with maternal inheritance. These data give an estimated mutation rate (micro) of 0.55 mutations/site/Myrs (HPD 95% confidence interval of 0.29-0.88 mutations/site/Myrs) after accounting for the persistence of these heteroplasmies and the sensitivity of current detection methods. In comparison, the rate of evolution (k) of the same HVR I region, determined using DNA sequences from 162 known age sub-fossil bones spanning a 37,000-year period, was 0.86 substitutions/site/Myrs (HPD 95% confidence interval of 0.53 and 1.17). Importantly, the latter rate is not statistically different from our estimate of the mutation rate. These results are in contrast to the view that molecular rates are time dependent.

Phylogenetic relationships, origin and historical biogeography of the genus Sprattus (Clupeiformes: Clupeidae).

The genus Sprattus comprises five species of marine pelagic fishes distributed worldwide in antitropical, temperate waters. Their distribution suggests an ancient origin during a cold period of the earth's history. In this study, we evaluated this hypothesis and corroborated the non-monophyly of the genus Sprattus, using a phylogenetic approach based on DNA sequences of five mitochondrial genome regions. Sprattus sprattus is more closely related to members of the genus Clupea than to other Sprattus species. We also investigated the historical biogeography of the genus, with the phylogenetic tree showing two well-supported clades corresponding to the species distribution in each hemisphere. Time-calibrated phylogenetic analyses showed that an ancient divergence between Northern and Southern Hemispheres occurred at 55.8 MYBP, followed by a diversification in the Oligocene epoch in the Northern Hemisphere clade (33.8 MYBP) and a more recent diversification in the Southern Hemisphere clade (34.2 MYBP). Historical biogeography analyses indicated that the most recent common ancestor (MRCA) likely inhabited the Atlantic Ocean in the Southern Hemisphere. These results suggest that the ancestral population of the MRCA diverged in two populations, one was dispersed to the Northern Hemisphere and the other across the Southern Hemisphere. Given that the Eocene was the warmest epoch since the Paleogene, the ancestral populations would have crossed the tropics through deeper cooler waters, as proposed by the isothermal submergence hypothesis. The non-monophyly confirmed for the genus Sprattus indicates that its systematics should be re-evaluated.

DNA degradation in fish: Practical solutions and guidelines to improve DNA preservation for genomic research.

The more demanding requirements of DNA preservation for genomic research can be difficult to meet when field conditions limit the methodological approaches that can be used or cause samples to be stored in suboptimal conditions. Such limitations may increase rates of DNA degradation, potentially rendering samples unusable for applications such as genome-wide sequencing. Nonetheless, little is known about the impact of suboptimal sampling conditions. We evaluated the performance of two widely used preservation solutions (1. DESS: 20% DMSO, 0.25 M EDTA, NaCl saturated solution, and 2. Ethanol >99.5%) under a range of storage conditions over a three-month period (sampling at 1 day, 1 week, 2 weeks, 1 month, and 3 months) to provide practical guidelines for DNA preservation. DNA degradation was quantified as the reduction in average DNA fragment size over time (DNA fragmentation) because the size distribution of DNA segments plays a key role in generating genomic datasets. Tissues were collected from a marine teleost species, the Australasian snapper, Chrysophrys auratus. We found that the storage solution has a strong effect on DNA preservation. In DESS, DNA was only moderately degraded after three months of storage while DNA stored in ethanol showed high levels of DNA degradation already within 24 hr, making samples unsuitable for next-generation sequencing. Here, we conclude that DESS was the most promising solution when storing samples for genomic applications. We recognize that the best preservation protocol is highly dependent on the organism, tissue type, and study design. We highly recommend performing similar experiments before beginning a study. This study highlights the importance of testing sample preservation protocols and provides both practical and economical advice to improve DNA preservation when sampling for genome-wide applications.

Origin, diversification, and systematics of the New Zealand skink fauna (Reptilia: Scincidae).

The diverse scincid lizard fauna of the largely submerged subcontinent of Zealandia (which incorporates New Zealand, New Caledonia, Lord Howe Island, Norfolk Island, and the Chatham Islands) forms a monophyletic lineage within the Eugongylus group of skinks. We use 4062 bp of mitochondrial (ND2, ND4, Cytochrome b, 12SrRNA, 16SrRNA) and nuclear (Rag-1) DNA sequence data to recover a molecular phylogeny for the New Zealand skink fauna, and investigate the origin and diversification of skinks in New Zealand. Our phylogeny includes 32 of the 33 extant described New Zealand skink species (Cyclodina and Oligosoma), the Lord Howe Island skink (C. lichenigera), and representatives from several New Caledonian genera. Neighbour-joining, Maximum Parsimony, Maximum Likelihood, and Bayesian phylogenetic analyses are used to demonstrate that the New Zealand skink species form a single monophyletic lineage, with C. lichenigera representing a closely related sister lineage to the New Zealand radiation. Our relaxed molecular clock analyses indicate that skinks colonised New Zealand in the early Miocene (16-22.6 mya), shortly after the 'Oligocene drowning' event (approximately 25 mya). We propose that skinks reached New Zealand from New Caledonia via long-distance overwater dispersal, with C. lichenigera persisting on volcanic islands along the Lord Howe Rise and Norfolk Ridge. Eight major genetic clades are evident within the New Zealand skink fauna, with the divergences among these clades during the early to mid-Miocene resulting in distinct open habitat, forest, and coastal radiations. Subsequent diversification in the late Miocene-Pliocene appears to coincide with tectonic activity along the Alpine Fault and the uplift of the Southern Alps. We were unable to resolve the phylogenetic affinities of O. suteri, New Zealand's only native oviparous skink. We use the phylogeny and topology tests to resolve several taxonomic issues and assess the taxonomic status of several suspected undescribed taxa. We complete a generic revision for the New Zealand skink fauna, placing C. lichenigera and all native New Zealand species into a single genus.

Phylogeographic history of the New Zealand stick insect Niveaphasma annulata (Phasmatodea) estimated from mitochondrial and nuclear loci.

We have assessed the utility of a single-copy nuclear locus and mitochondrial DNA (mtDNA) in a phylogeographic study of the New Zealand stick insect Niveaphasma annulata (Hutton). We amplified sequences from the mitochondrial cytochrome oxidase subunit I (COI) gene and the single-copy nuclear gene elongation factor-1alpha (EF1alpha) from 97 individuals. Allelic phase at the EF1alpha locus was determined using Denaturing Gradient Gel Electrophoresis. Phylogenetic analyses showed broad congruence between the geographic distribution of three major COI clades and EF1alpha alleles, which suggested that the phylogenetic patterns reflect population history rather than lineage sorting. However, the geographic boundaries of these clades were not always in exact agreement between the two loci. Our data indicate that Niveaphasma annulata was most likely separated into a number of refugia during Pleistocene glacial advances. Subsequent to glacial retreat these refugial populations have expanded and now form a number of zones of secondary contact. We contrast these patterns with those observed from other New Zealand taxa. Our study offers compelling evidence for the use of nuclear genes alongside mtDNA for future phylogeographic studies.

High-Density Linkage Map and QTLs for Growth in Snapper (Chrysophrys auratus).

Characterizing the genetic variation underlying phenotypic traits is a central objective in biological research. This research has been hampered in the past by the limited genomic resources available for most non-model species. However, recent advances in sequencing technologies and related genotyping methods are rapidly changing this. Here we report the use of genome-wide SNP data from the ecologically and commercially important marine fish species Chrysophrys auratus (snapper) to 1) construct the first linkage map for this species, 2) scan for growth QTL, and 3) search for putative candidate genes in the surrounding QTL regions. The newly constructed linkage map contained ∼11K SNP markers and is one of the densest maps to date in the fish family Sparidae. Comparisons with genome scaffolds of the recently assembled snapper genome indicated that marker placement was mostly consistent between the scaffolds and linkage map (R = 0.7), but that at fine scales (< 5 cM) some precision limitations occurred. Of the 24 linkage groups, which likely reflect the 24 chromosomes of this species, three were found to contain QTL with genome-wide significance for growth-related traits. A scan of 13 candidate growth genes located the growth hormone, myogenin, and parvalbumin genes within 5.3, 9.6, and 25.0 cM of these QTL, respectively. The linkage map and QTL found in this study will advance the investigation of genome structure and aquaculture breeding efforts in this and related species.

Domestication and Temperature Modulate Gene Expression Signatures and Growth in the Australasian Snapper Chrysophrys auratus.

Identifying genes and pathways involved in domestication is critical to understand how species change in response to human-induced selection pressures, such as increased temperatures. Given the profound influence of temperature on fish metabolism and organismal performance, a comparison of how temperature affects wild and domestic strains of snapper is an important question to address. We experimentally manipulated temperature conditions for F1-hatchery and wild Australasian snapper (Chrysophrys auratus) for 18 days to mimic seasonal extremes and measured differences in growth, white muscle RNA transcription and hematological parameters. Over 2.2 Gb paired-end reads were assembled de novo for a total set of 33,017 transcripts (N50 = 2,804). We found pronounced growth and gene expression differences between wild and domesticated individuals related to global developmental and immune pathways. Temperature-modulated growth responses were linked to major pathways affecting metabolism, cell regulation and signaling. This study is the first step toward gaining an understanding of the changes occurring in the early stages of domestication, and the mechanisms underlying thermal adaptation and associated growth in poikilothermic vertebrates. Our study further provides the first transcriptome resources for studying biological questions in this non-model fish species.

Unlocking the potential of ancient fish DNA in the genomic era.

Fish are the most diverse group of vertebrates, fulfil important ecological functions and are of significant economic interest for aquaculture and wild fisheries. Advances in DNA extraction methods, sequencing technologies and bioinformatic applications have advanced genomic research for nonmodel organisms, allowing the field of fish ancient DNA (aDNA) to move into the genomics era. This move is enabling researchers to investigate a multitude of new questions in evolutionary ecology that could not, until now, be addressed. In many cases, these new fields of research have relevance to evolutionary applications, such as the sustainable management of fisheries resources and the conservation of aquatic animals. Here, we focus on the application of fish aDNA to (a) highlight new research questions, (b) outline methodological advances and current challenges, (c) discuss how our understanding of fish ecology and evolution can benefit from aDNA applications and (d) provide a future perspective on how the field will help answer key questions in conservation and management. We conclude that the power of fish aDNA will be unlocked through the application of continually improving genomic resources and methods to well-chosen taxonomic groups represented by well-dated archaeological samples that can provide temporally and/or spatially extensive data sets.

A SINE of restricted gene flow across the Alpine Fault: phylogeography of the New Zealand common skink (Oligosoma nigriplantare polychroma).

New Zealand has experienced a complex climatic and geological history since the Pliocene. Thus, identifying the processes most important in having driven the evolution of New Zealand's biota has proven difficult. Here we examine the phylogeography of the New Zealand common skink (Oligosoma nigriplantare polychroma) which is distributed throughout much of New Zealand and crosses many putative biogeographical boundaries. Using mitochondrial DNA sequence data, we revealed five geographically distinct lineages that are highly differentiated (pairwise Phi(ST) 0.54-0.80). The phylogeographical pattern and inferred age of the lineages suggests Pliocene mountain building along active fault lines promoted their divergence 3.98-5.45 million years ago. A short interspersed nuclear element (SINE) polymorphism in the myosin gene intron (MYH-2) confirmed a pattern of restricted gene flow between lineages on either side of the mountain ranges associated with the Alpine Fault that runs southwest to northeast across the South Island of New Zealand. An analysis of molecular variance confirmed that approximately 40% of the genetic differentiation in O. n. polychroma is distributed across this major fault line. The straits between the main islands of New Zealand accounted for much less of the variation found within O. n. polychroma, most likely due to the repeated existence of landbridges between islands during periods of the Pleistocene that allowed migration. Overall, our findings reveal the relative roles of different climatic and geological processes, and in particular, demonstrate the importance of the Alpine Fault in the evolution of New Zealand's biota.

Origin and post-colonization evolution of the Chatham Islands skink (Oligosoma nigriplantare nigriplantare).

Island ecosystems provide an opportunity to examine a range of evolutionary and ecological processes. The Chatham Islands are an isolated archipelago situated approximately 800 km east of New Zealand. Geological evidence indicates that the Chatham Islands re-emerged within the last 1-4 million years, following a prolonged period of marine inundation, and therefore the resident flora and fauna is the result of long-distance overwater dispersal. We examine the origin and post-colonization evolution of the Chatham Islands skink, Oligosoma nigriplantare nigriplantare, the sole reptile species occurring on the archipelago. We sampled O. n. nigriplantare from across nine islands within the Chatham Islands group, and representative samples from across the range of its closest relative, the New Zealand mainland common skink (Oligosoma nigriplantare polychroma). Our mitochondrial sequence data indicate that O. n. nigriplantare diverged from O. n. polychroma 5.86-7.29 million years ago. This pre-dates the emergence date for the Chatham Islands, but indicates that O. n. nigriplantare colonized the Chatham Islands via overwater dispersal on a single occasion. Despite the substantial morphological variability evident in O. n. nigriplantare, only relatively shallow genetic divergences (maximum divergence approximately 2%) were found across the Chatham Islands. Our analyses (haplotypic diversity, Phi(ST), analysis of molecular variance, and nested clade phylogeographical analysis) indicated restricted gene flow in O. n. nigriplantare resulting in strong differentiation between islands. However, the restrictions to gene flow might have only arisen recently as there was also a significant pattern of isolation by distance, possibly from when the Chatham Islands were a single landmass during Pleistocene glacial maxima when sea levels were lower. The level of genetic and morphological divergence between O. n. nigriplantare and O. n. polychroma might warrant their recognition as distinct species.

Phylogeography of two New Zealand lizards: McCann's skink (Oligosoma maccanni) and the brown skink (O. zelandicum).

The New Zealand skink fauna has proven to be an ideal taxonomic group in which to examine the impact of climatic and geological processes on the evolution of the New Zealand biota since the Pliocene. Here we examine the phylogeography of McCann's skink (Oligosoma maccanni) in order to gain insight into the relative contribution of Pliocene and Pleistocene processes on patterns of genetic structure in the South Island biota, and investigate the phylogeography of the brown skink (O. zelandicum) to examine whether Cook Strait landbridges facilitated geneflow between the North and South Islands in the late-Pleistocene. We obtained mitochondrial DNA sequence data (ND2 and ND4; 1282bp) from across the range of both species. We examined the phylogeographic patterns evident in each species using Neighbour-Joining, Maximum Likelihood and Bayesian methods. We found substantial phylogeographic structure within O. maccanni, with seven distinct clades identified. Divergences among clades are estimated to have occurred during the Pliocene. Populations in the Otago/Southland region (south of the Waitaki River valley) formed a well-supported lineage within O. maccanni. A substantial genetic break was evident between populations in east and west Otago, either side of the Nevis-Cardrona fault system, while north-south genetic breaks were evident within the Canterbury region. Within-clade divergences in O. maccanni appear to have occurred during the mid- to late-Pleistocene. Shimodaira-Hasegawa topology tests indicated that the 'Garston' skink is not genetically distinct from O. maccanni. There was only relatively minor phylogeographic structure within O. zelandicum, with divergences among populations occurring during the mid- to late-Pleistocene. Our genetic data supports a single colonisation of the North Island by O. zelandicum from the South Island, with the estimated timing of this event (0.46mya) consistent with the initial formation of Cook Strait.

A retrospective approach to testing the DNA barcoding method.

A decade ago, DNA barcoding was proposed as a standardised method for identifying existing species and speeding the discovery of new species. Yet, despite its numerous successes across a range of taxa, its frequent failures have brought into question its accuracy as a short-cut taxonomic method. We use a retrospective approach, applying the method to the classification of New Zealand skinks as it stood in 1977 (primarily based upon morphological characters), and compare it to the current taxonomy reached using both morphological and molecular approaches. For the 1977 dataset, DNA barcoding had moderate-high success in identifying specimens (78-98%), and correctly flagging specimens that have since been confirmed as distinct taxa (77-100%). But most matching methods failed to detect the species complexes that were present in 1977. For the current dataset, there was moderate-high success in identifying specimens (53-99%). For both datasets, the capacity to discover new species was dependent on the methodological approach used. Species delimitation in New Zealand skinks was hindered by the absence of either a local or global barcoding gap, a result of recent speciation events and hybridisation. Whilst DNA barcoding is potentially useful for specimen identification and species discovery in New Zealand skinks, its error rate could hinder the progress of documenting biodiversity in this group. We suggest that integrated taxonomic approaches are more effective at discovering and describing biodiversity.

Genetic diversity is positively associated with fine-scale momentary abundance of an invasive ant.

Many introduced species become invasive despite genetic bottlenecks that should, in theory, decrease the chances of invasion success. By contrast, population genetic bottlenecks have been hypothesized to increase the invasion success of unicolonial ants by increasing the genetic similarity between descendent populations, thus promoting co-operation. We investigated these alternate hypotheses in the unicolonial yellow crazy ant, Anoplolepis gracilipes, which has invaded Arnhem Land in Australia's Northern Territory. We used momentary abundance as a surrogate measure of invasion success, and investigated the relationship between A. gracilipes genetic diversity and its abundance, and the effect of its abundance on species diversity and community structure. We also investigated whether selected habitat characteristics contributed to differences in A. gracilipes abundance, for which we found no evidence. Our results revealed a significant positive association between A. gracilipes genetic diversity and abundance. Invaded communities were less diverse and differed in structure from uninvaded communities, and these effects were stronger as A. gracilipes abundance increased. These results contradict the hypothesis that genetic bottlenecks may promote unicoloniality. However, our A. gracilipes study population has diverged since its introduction, which may have obscured evidence of the bottleneck that would likely have occurred on arrival. The relative importance of genetic diversity to invasion success may be context dependent, and the role of genetic diversity may be more obvious in the absence of highly favorable novel ecological conditions.

Genetic effects of harvest on wild animal populations.

Human harvest of animals in the wild occurs in terrestrial and aquatic habitats throughout the world and is often intense. Harvest has the potential to cause three types of genetic change: alteration of population subdivision, loss of genetic variation, and selective genetic changes. To sustain the productivity of harvested populations, it is crucial to incorporate genetic considerations into management. Nevertheless, it is not necessary to disentangle genetic and environmental causes of phenotypic changes to develop management plans for individual species. We recommend recognizing that some genetic change due to harvest is inevitable. Management plans should be developed by applying basic genetic principles combined with molecular genetic monitoring to minimize harmful genetic change.

Evidence for specificity of psittacine beak and feather disease viruses among avian hosts.

Beak and feather disease is a major avian disease of both captive and wild parrot and cockatoo populations. Clinical signs include beak elongation and abnormal growth, together with weight loss and in some individuals the disease is fatal. We investigated the relationship between viral genotypes and their hosts in order to test for a positive association between distinct viral genomes and avian species. Specifically, we used the polymerase chain reaction (PCR) to amplify and sequence a 605-nucleotide (nt) segment of a coding region in the Beak and Feather Disease Virus (BFDV) genome. Feather and blood samples from 25 caged birds representing 10 species were assayed and the BFDV was detected in 21 samples from New Zealand. A phylogenetic analysis of DNA sequences from 17 specimens together with previously published sequences from Australian "isolates" revealed three lineages present in New Zealand. One viral lineage was found in six cockatoos representing two species (designated CT), a second lineage was detected in a budgerigar (designated BG), and a third was found in 10 lorikeets representing seven species (designated LK). This distinctive clustering pattern of viral sequences with groups of psittacine species indicates a genotypic association between the virus and these hosts.

Ancient DNA enables timing of the pleistocene origin and holocene expansion of two adélie penguin lineages in antarctica.

The timing of divergent events in history is one of the central goals of contemporary evolutionary biology. Such studies are however dependent on accurate evolutionary rates. Recent developments in ancient DNA analysis enable the estimation of more accurate evolutionary rates and therefore more accurate timing of divergence events. Consequently, this leads to a better understanding of changes in populations through time. We use an evolutionary rate calculated from ancient DNA of Adélie penguins (Pygoscelis adeliae) to time divergent events in their history. We report the presence of two distinct and highly variable mitochondrial DNA lineages and track changes in these lineages through space and time. When the ancient DNA and the phylogenetic rates are used to estimate the time of origin of the lineages, two very different estimates resulted. In addition, these same rates provide very different estimates of the time of expansion of these lineages. We suggest that the rate calculated from ancient DNA is more consistent with the glacial history of Antarctica and requires fewer assumptions than does a narrative based on the phylogenetic rate. Finally, we suggest that our study indicates an important new role for ancient DNA studies in the timing of divergent events in history.