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Species recovery programs are increasingly using genomic data to measure neutral genetic diversity and calculate metrics like relatedness. While these measures can inform conservation management, determining the mechanisms underlying inbreeding depression requires information about functional genes associated with adaptive or maladaptive traits. Toll-like receptors (TLRs) are one family of functional genes, which play a crucial role in recognition of pathogens and activation of the immune system. Previously, these genes have been analysed using species-specific primers and PCR. Here, we leverage an existing short-read reference genome, whole-genome resequencing population data set, and bioinformatic tools to characterize TLR gene diversity in captive and wild tchūriwat'/tūturuatu/shore plover (Thinornis novaeseelandiae), a threatened bird endemic to Aotearoa New Zealand. Our results show that TLR gene diversity in tchūriwat'/tūturuatu is low, and forms two distinct captive and wild genetic clusters. The bioinformatic approach presented here has broad applicability to other threatened species with existing genomic resources in Aotearoa New Zealand and beyond.

SummaryCook’s Petrel Pterodroma cookii is an endemic New Zealand seabird that has experienced a large range decline since the arrival of humans and now only breeds on two offshore islands (Te Hauturu-o-Toi/Little Barrier Island and Whenua Hou/Codfish Island) at the extreme ends of its former distribution. Morphological, behavioural, and mitochondrial cytochrome oxidase 1 (CO1) sequence data led a previous study to recognise the two extant populations as distinct conservation management units. Here, we further examine the genetic relationship between the extant populations using two nuclear introns (β-fibint7 and PAX). Using one mitochondrial locus (CO1), we also investigate the past distribution of a single nucleotide polymorphism (SNP) that differentiates the modern populations using bone and museum skins sourced from within its former range across New Zealand’s North and South Islands. We found significant population genetic structure between the two extant Cook’s Petrel populations for one of the two nuclear introns (β-fibint7). The mitochondrial DNA CO1 analysis indicated that the SNP variant found in the Codfish Island population was formerly widely distributed across both the North and South Islands, whereas the Little Barrier Island variant was detected only in North Island samples. We argue that these combined data support the recognition of the extant populations as different subspecies. Previous names for these taxa exist, thus Cook’s Petrel from Little Barrier Island becomes Pterodroma cookii cookii and Cook’s Petrel from Codfish Island becomes P. c. orientalis. Furthermore, we suggest that both genetic and non-genetic data should be taken into consideration when planning future mainland translocations. Namely, any translocations on the South Island should be sourced from Codfish Island and future translocations on the North Island should continue to be sourced from Little Barrier Island only.

Conservation translocations—particularly those that weave diverse ways of knowing and seeing the world—promise to enhance species recovery and build ecosystem resilience. Yet, few studies to date have been led or co‐led by Indigenous peoples; or consider how centring Indigenous knowledge systems can lead to betterconservation translocation outcomes. In this Perspective, as Indigenous and non‐Indigenous researchers and practitioners working in partnership in Aotearoa New Zealand, we present a novel framework for co‐designing conservation translocations that centre Indigenous peoples and knowledge systems through Two‐Eyed Seeing. We apply this framework to Aotearoa New Zealand's threatened and underprioritized freshwater biodiversity. In particular, we highlight the co‐development of conservation translocations with Te Kōhaka o Tūhaitara and Te Nohoaka o Tukiauau that are weaving emerging genomic approaches into mātauraka Māori (Māori knowledge systems), including customary practices, processes and language. We envision the Two‐Eyed Seeing framework presented here will provide a critical point of reference for the co‐development of conservation translocations led or co‐led by Indigenous peoples elsewhere in the world to build more resilient biocultural heritage. A free Plain Language Summary can be found within the Supporting Information of this article.

Imperfect detection of individuals in threatened wild populations is common and can obscure real population trends when it is unaccounted for in population monitoring, and therefore impede conservation decision making. For many threatened insects, there is a lack of biological information or available long-term data to inform how best to practice data collection and population monitoring. Here, we inform the design of a long-term population density monitoring protocol for Brachaspis robustus, a Nationally Endangered grasshopper endemic to the Mackenzie Basin of New Zealand. We use removal sampling (repeated visual searches of a predefined area where any individuals found are temporarily removed to achieve successive depletion) during a single austral summer season (November to March) to rapidly quantify seasonal and demographic visual detectability. Juvenile instars dominated population composition in all months except December and males represented > 50% of monthly captures. Adult females were 2–3 times larger than adult males, and 79% of those captured were found during the first search of an area compared to only 52% of adult males. The odds of detecting an individual increased by 6% per 1 mm of body length. Removal sampling was found to be an effective method for rapidly informing future long-term monitoring design for a visually cryptic, threatened insect. Recommendations include monitoring adult females as an index of population size, restricting monitoring to when adult abundance peaks (November and December), and conducting multiple monitoring events within peak months to counter the effects of daily and seasonal variation and imperfect detection.

Conservation management strategies for many highly threatened species include conservation breeding to prevent extinction and enhance recovery. Pairing decisions for these conservation breeding programmes can be informed by pedigree data to minimize relatedness between individuals in an effort to avoid inbreeding, maximize diversity and maintain evolutionary potential. However, conservation breeding programmes struggle to use this approach when pedigrees are shallow or incomplete. While genetic data (i.e., microsatellites) can be used to estimate relatedness to inform pairing decisions, emerging evidence indicates this approach may lack precision in genetically depauperate species, and more effective estimates will likely be obtained from genomic data (i.e., thousands of genome‐wide single nucleotide polymorphisms, or SNPs). Here, we compare relatedness estimates and subsequent pairing decisions using pedigrees, microsatellites and SNPs from whole‐genome resequencing approaches in two critically endangered birds endemic to New Zealand: kakī/black stilt (Himantopus novaezelandiae) and kākāriki karaka/orange‐fronted parakeet (Cyanoramphus malherbi). Our findings indicate that SNPs provide more precise estimates of relatedness than microsatellites when assessing empirical parent–offspring and full sibling relationships. Further, our results show that relatedness estimates and subsequent pairing recommendations using PMx are most similar between pedigree‐ and SNP‐based approaches. These combined results indicate that in lieu of robust pedigrees, SNPs are an effective tool for informing pairing decisions, which has important implications for many poorly pedigreed conservation breeding programmes worldwide.

Researchers have long debated which estimator of relatedness best captures the degree of relationship between two individuals. In the genomics era, this debate continues, with relatedness estimates being sensitive to the methods used to generate markers, marker quality, and levels of diversity in sampled individuals. Here, we compare six commonly used genome-based relatedness estimators (kinship genetic distance [KGD], Wang maximum likelihood [TrioML], Queller and Goodnight [R

Over the past 50 years conservation genetics has developed a substantive toolbox to inform species management. One of the most long-standing tools available to manage genetics - the pedigree - has been widely used to characterize diversity and maximize evolutionary potential in threatened populations. Now, with the ability to use high throughput sequencing (HTS) to estimate relatedness, inbreeding, and genome-wide functional diversity, some have asked whether it is warranted for conservation biologists to continue collecting and collating pedigrees for species management. In this perspective, we argue that pedigrees remain a relevant tool, and when combined with genomic data, create an invaluable resource for conservation genomic management. Genomic data can address pedigree pitfalls (e.g., founder relatedness, missing data, uncertainty), and in return robust pedigrees allow for more nuanced research design, including well-informed sampling strategies and quantitative analyses (e.g., heritability, linkage) to better inform genomic inquiry. We further contend that building and maintaining pedigrees provides an opportunity to strengthen trusted relationships among conservation researchers, practitioners, Indigenous Peoples, and local communities. Keywords: conservation genomics, quantitative genetics, pedigree, kinship,ex situ , in situ

Researchers have long debated which estimator of relatedness best captures the degree of relationship between two individuals. In the genomics era, this debate continues, with relatedness estimates being sensitive to the methods used to generate markers, marker quality, and levels of diversity in sampled individuals. Here, we compare six commonly used genome-based relatedness estimators (kinship genetic distance (KGD), Wang Maximum Likelihood (TrioML), Queller and Goodnight (Rxy), Kinship INference for Genome-wide association studies (KING-robust), and Pairwise Relatedness (RAB), allele-sharing co-ancestry (AS)) across five species bred in captivity–including three birds and two mammals–with varying degrees of reliable pedigree data, using reduced-representation and whole genome resequencing data. Genome-based relatedness estimates varied widely across estimators, sequencing methods, and species, yet the most consistent results for known first order relationships were found using Rxy, RAB, and AS. However, AS was found to be less consistently correlated with known pedigree relatedness than either Rxy or RAB. Our combined results indicate there is not a single genome-based estimator that is ideal across different species and data types. To determine the most appropriate genome-based relatedness estimator for each new dataset, we recommend assessing the relative: (1) correlation of candidate estimators with known relationships in the pedigree and (2) precision of candidate estimators with known first-order relationships. These recommendations are broadly applicable to conservation breeding programs, particularly where genome-based estimates of relatedness can complement and complete poorly pedigreed populations. Given a growing interest in the application of wild pedigrees, our results and are also applicable to in-situ wildlife management.

Increased capability in the conservation genomics community, combined with decreased sequencing costs, is providing new opportunities for the application of whole-genome sequence data to enhance species recovery. Indeed, assessments of genome-wide diversity based on SNP data are already informing the conservation management of threatened species around the world. However, SNP data alone may not sufficiently capture all of the information necessary for the effective conservation management of critically endangered species that suffer from severe inbreeding depression. For threatened species that require significant intervention, it is critical that we as conservation genomicists expand our repertoire to include assessments of functional diversity. Structural variants are a likely source of functional diversity, as their frequency and genomic context affect the dosage and regulation of gene expression through mechanisms that alter genome organization and impact fitness. In this future-focused Opinion, we leverage the existing literature - predominantly focused on model and agricultural species - to identify pan-genomic and chromosomic approaches for readily characterizing structural variants and to consider how integrating these into the conservation genomics toolbox will transform the way we manage some of the world’s most threatened species.

Genetic swamping resulting from interspecific hybridisation can increase extinction risk for threatened species. The development of high-throughput and reduced-representation genomic sequencing and analyses to generate large numbers of high resolution genomic markers has the potential to reveal introgression previously undetected using small numbers of genetic markers. However, few studies to date have implemented genomic tools to assess the extent of interspecific hybridisation in threatened species. Here we investigate the utility of genome-wide single nucleotide polymorphisms (SNPs) to detect introgression resulting from past interspecific hybridisation in one of the world’s rarest birds. Anthropogenic impacts have resulted in hybridisation and subsequent backcrossing of the critically endangered Aotearoa New Zealand endemic kakī (black stilts; Himantopus novaezelandiae) with the non-threatened self-introduced congeneric poaka (Aotearoa New Zealand population of pied stilts, Himantopus himantopus leucocephalus), yet genetic analyses with a limited set of microsatellite markers revealed no evidence of introgression of poaka genetic material in kakī, excluding one individual. We use genomic data for ∼63% of the wild adult kakī population to reassess the extent of introgression resulting from hybridisation between kakī and poaka. Consistent with previous genetic analyses, we detected no introgression from poaka into kakī. These collective results indicate that, for kakī, existing microsatellite markers provide a robust, cost-effective approach to detect cryptic hybrids. Further, for well-differentiated species, the use of genomic markers may not be required to detect admixed individuals.

We evaluate evidence that reducing mammalian predators benefits threatened and declining grasshoppers in the Mackenzie Basin, New Zealand. Long-term population trends of Sigaus minutus are investig...

Summary De-extinction sensu stricto is the resurrection of phenotypic traits once possessed by extinct species to create extant functional proxies. To realise the ecological benefit of de-extinction, self-sustaining (genetically viable) populations of functional proxies are required. It is often implied, yet rarely stated, that the genetic challenges associated with the survival and recovery of extant threatened species in an effort to conserve biodiversity are also relevant to the use of functional proxies of extinct species as a conversation tool. Here, we highlight the importance of prioritising evolutionary potential − the capacity to evolve (adapt) in response to environmental change − in populations of functional proxies. We use conservation genetic principles to describe impediments to the creation and maintenance of evolutionary potential as a series of potentially unavoidable genetic bottlenecks (pre-extinction, resurrection, captive, translocation). To give any successfully translocated populations of functional proxies the best chance to survive beyond the first few generations in the wild, we advocate the use of a holistic framework that includes the creation of sufficiently large, genetically diverse populations that harbour the ability to adapt to a changing environment. A lay summary is available for this article.

Conservation management strategies for many highly threatened species include conservation breeding to prevent extinction and enhance recovery. Pairing decisions for these conservation breeding programmes can be informed by pedigree data to minimise relatedness between individuals in an effort to avoid inbreeding, maximise diversity, and maintain evolutionary potential. However, conservation breeding programmes struggle to use this approach when pedigrees are shallow or incomplete. While genetic data (i.e., microsatellites) can be used to estimate relatedness to inform pairing decisions, emerging evidence indicates this approach lacks precision in genetically depauperate species, and more effective estimates will likely be obtained from genomic data (i.e., thousands of genome-wide single nucleotide polymorphisms, or SNPs). Here, we compare relatedness estimates using pedigree-, genetic-, and genomic-based approaches for making pairing decisions in two critically endangered birds endemic to New Zealand: kakī/black stilt (Himantopus novaezelandiae) and kākāriki karaka/orange-fronted parakeet (Cyanoramphus malherbi). Our findings indicate genetic-based estimates of relatedness are indeed the least precise when assessing known parent-offspring and full sibling relationships. Furthermore, our results show that relatedness estimates and subsequent pairing recommendations using PMx are most similar between pedigree- and genomic-based approaches. These combined results indicate that in lieu of robust pedigrees, SNPs are an effective tool for informing pairing decisions, which has exciting implications for many poorly pedigreed conservation breeding programmes worldwide.

The Masked Booby is a highly vagile, pantropical seabird of which up to six subspecies have been recognised: S. d. dactylatra, S. d. californica, S. d. personata, S. d. melanops, S. d. bedouti and S. d. tasmani. The genetic distinction of several S. dactylatra colonies has been previously investigated, but this has not yet been conducted for the Bedout Island population in Western Australia, which has been considered by some to be part of a distinct subspecies. Suspected population decline on Bedout Island has renewed interest in determining the extent to which genetic novelty might be threatened. To answer this question, morphological and mitochondrial control region sequence variation were used to determine the distinction of the Bedout Island population. Whereas the morphological measures were equivocal, six haplotypes were identified from 31 individuals, none of which were shared by individuals previously sampled from the Indo-Pacific. The Bedout Island haplotypes were most closely related to other haplotypes found in a distinct Indian Ocean haplogroup, but haplotype frequencies at Bedout Island differed significantly from all other. This indicates that the Bedout Island population rarely exchanges mitochondrial genes with any of the other Masked Booby colonies presently studied, which may reflect dependence on local recruitment for its persistence.

Several reviews in the past decade have heralded the benefits of embracing high-throughput sequencing technologies to inform conservation policy and the management of threatened species, but few have offered practical advice on how to expedite the transition from conservation genetics to conservation genomics. Here, we argue that an effective and efficient way to navigate this transition is to capitalize on emerging synergies between conservation genetics and primary industry (e.g., agriculture, fisheries, forestry and horticulture). Here, we demonstrate how building strong relationships between conservation geneticists and primary industry scientists is leading to mutually-beneficial outcomes for both disciplines. Based on our collective experience as collaborative New Zealand-based scientists, we also provide insight for forging these cross-sector relationships.

Two captive-reared birds of wild origin presumed to be Kakī/Black Stilt (Himantopus novaezelandiae) were found to display plumage atypical of Kakī. We have combined genetic and non-genetic data to test the hypothesis that these birds are a product of brood parasitism by “non-kakī” [i.e. Poaka/Pied Stilt (Himantopus himantopus leucocephalus) or Kakī–Poaka hybrids]. We found that these atypically plumed birds have cytochrome b haplotypes and microsatellite alleles that could not be attributed to the putative Kakī parents associated with the nest, thus providing the first evidence for brood parasitism in Kakī.

Statistically robust monitoring of threatened populations is essential for effective conservation management because the population trend data that monitoring generates is often used to make decisions about when and how to take action. Despite representing the highest proportion of threatened animals globally, the development of best practice methods for monitoring populations of threatened insects is relatively uncommon. Traditionally, population trend data for the Nationally Endangered New Zealand grasshopper Brachaspis robustus has been determined by counting all adults and nymphs seen on a single ~1.5 km transect searched once annually. This method lacks spatial and temporal replication, both of which are essential to overcome detection errors in highly cryptic species like B. robustus. It also provides no information about changes in the grasshopper’s distribution throughout its range. Here, we design and test new population density and site occupancy monitoring protocols by comparing a) comprehensive plot and transect searches at one site and b) transect searches at two sites representing two different habitats (gravel road and natural riverbed) occupied by the species across its remaining range. Using power analyses, we determined a) the number of transects, b) the number of repeated visits and c) the grasshopper demographic to count to accurately detect long term change in relative population density. To inform a monitoring protocol design to track trends in grasshopper distribution, we estimated the probability of detecting an individual with respect to a) search area, b) weather and c) the grasshopper demographic counted at each of the two sites. Density estimates from plots and transects did not differ significantly. Population density monitoring was found to be most informative when large adult females present in early summer were used to index population size. To detect a significant change in relative density with power > 0.8 at the gravel road habitat, at least seventeen spatial replicates (transects) and four temporal replicates (visits) were required. Density estimates at the natural braided river site performed poorly and likely require a much higher survey effort. Detection of grasshopper presence was highest (pg > 0.6) using a 100 m x 1 m transect at both sites in February under optimal (no cloud) conditions. At least three visits to a transect should be conducted per season for distribution monitoring. Monitoring protocols that inform the management of threatened species are crucial for better understanding and mitigation of the current global trends of insect decline. This study provides an exemplar of how appropriate monitoring protocols can be developed for threatened insect species.

It is now over 10 years since the first avian genome [International Chicken Genome Sequencing Consortium, 2004] and the first complete avian karyotype [Masabanda et al., 2004] were both published; however, until 2014, avian cytogenetics has focused heavily on descriptive studies [e.g. Griffin et al., 2007, 2008; Skinner et al., 2009; Volker et al., 2010] with less attention to its functional relevance. Last year, however, saw 2 landmark efforts in the chromosomal studies of birds: a special issue of Chromosome Research in April and the announcement of recently completed sequences of multiple new avian genomes in Science and the BMC journals (taking the total number sequenced to over 50) in December. Studying the chromosomes of birds is, perhaps for the first time, telling us more about avian biology, function and evolution than it ever has... Conclusions. The most recent advances in avian cytogenetics have culminated in great promise not only for the study of bird karyotypes, but also for providing insight into the mechanisms of chromosome evolution in general. New avenues for investigation include gene regulation; for instance, it will become necessary to map accurately the physical location of polyadenylation and transcription start sites, important reference points that define promoters and post-transcriptional regulation. It will also become possible to sequence full-length transcripts, to allow accurate identification of alternate splicing events and their controlling elements. The ENCODE (Encyclopedia of DNA Elements) project has helped to define functional elements of the human genome, including those aforementioned as well as other chromatin signals, e.g. active chromatin, enhancers, insulators, methylation domains, etc. An effort of agENCODE is underway to include agriculturally important birds such as chicken, turkey, duck, quail, and perhaps ostrich. The study of cytogenetics will be essential here in helping to define higher-order structures in nuclear organization that show regulatory interactions within and between chromosomes. Finally, reconstruction of evolutionary events allows us to study genome organization and function not only in extant but, by extrapolation, in extinct species also. Reconstruction of avian-reptilian ancestral karyotypes will allow us to define chromosomal rearrangements in long-dead species that have captured the public imagination. Here be dragons!

Males of many animal species fight to establish social dominance and control access to females so that they have more opportunities to reproduce than their competitors. Males with lower social status will struggle to directly compete for mates, thus they attempt to mate with females by stealth. This often leads to more than one male mating with the same female so that the sperm from each male end up competing to fertilise that female’s eggs, a phenomenon known as sperm competition. Males suspend their sperm in a fluid to make a mixture known as semen. It has been shown that, compared to high status males, low status males will produce higher quality semen that contains greater numbers of faster swimming sperm, giving them an advantage in sperm competition. Growing evidence from several species indicates that males can quickly adjust how fast their sperm swim in response to social cues that signal changing risks of sperm competition. However, how these rapid adjustments occur remains largely unknown, and whether they alter a male’s reproductive success against a competitor has seldom been examined. Chinook salmon usually live in the North Pacific Ocean but they swim up rivers in North America and Asia to reproduce. They have also been introduced to several other countries including New Zealand where they are farmed commercially. The fish are highly prized by sport fishermen and are also of cultural significance to certain groups of indigenous people in North America. Barlett et al. studied the semen of chinook salmon, undertaking a series of experiments in which males switched between high and low social status. The experiments show that, as predicted, the sperm of males that changed from high to low social status started to swim faster. These changes in speed were caused by the fluid in the semen and altered the number of eggs that the male’s sperm fertilised when competing against sperm from another male. In their natural range some populations of chinook salmon are declining due to overfishing combined with habitat loss and alteration. The findings of Barlett et al. contribute to a better understanding of how this fish species reproduces, which may lead to the introduction of measures that help natural populations to recover or help to improve commercial farming. Improved knowledge of how the fluid in semen affects sperm activity may also have important consequences for our wider understanding of male fertility in humans and other animals.

There is contentious debate surrounding the merits of de-extinction as a biodiversity conservation tool. Here, we use extant analogues to predict conservation actions for potential de-extinction candidate species from New Zealand and the Australian state of New South Wales, and use a prioritization protocol to predict the impacts of reintroducing and maintaining populations of these species on conservation of extant threatened species. Even using the optimistic assumptions that resurrection of species is externally sponsored, and that actions for resurrected species can share costs with extant analogue species, public funding for conservation of resurrected species would lead to fewer extant species that could be conserved, suggesting net biodiversity loss. If full costs of establishment and maintenance for resurrected species populations were publicly funded, there could be substantial sacrifices in extant species conservation. If conservation of resurrected species populations could be fully externally sponsored, there could be benefits to extant threatened species. However, such benefits would be outweighed by opportunity costs, assuming such discretionary money could directly fund conservation of extant species. Potential sacrifices in conservation of extant species should be a crucial consideration in deciding whether to invest in de-extinction or focus our efforts on extant species.

Beak and feather disease virus (BFDV) infections are often fatal to both captive and wild parrot populations. Its recent discovery in a wild population of native red-fronted parakeets has raised concerns for the conservation of native parrots, all of which are threatened or endangered. The question of a recent introduction versus a native genotype of the virus poses different conservation-management challenges, and thus, a clear understanding of the molecular phylogeny of BDFV is a crucial step towards integrated management planning. This study represents the first comprehensive attempt to screen New Zealand's endangered and threatened psittacines systematically for BFDV. We sampled and screened kakapos (Strigops habroptilus), kakas (Nestor meridionalis), keas (N. notabilis), Chatham parakeets (Cyanoramphus forbesi), Malherbe's parakeets (Cyanoramphus malherbi), yellow-crowned parakeets (C. auriceps) and red-fronted parakeets (Cyanoramphus novaezelandiae), as well as eastern rosellas (Platycercus eximius), an introduced species that is now common throughout the North Island, for BFDV. Out of all species and populations sampled (786 individuals), we found 16 BFDV-positive red-fronted parakeets from Little Barrier Island/Hauturu, seven eastern rosellas from the Auckland region, and eight yellow-crowned parakeets from the Eglinton Valley in the South Island. The full genomes of the viral isolates from the red-fronted parakeets share 95-97 % sequence identity to those from the invasive eastern rosellas and 92.7-93.4 % to those isolates from the South Island yellow-crowned parakeets. The yellow-crowned parakeet BFDV isolates share 92-94 % sequence identity with those from eastern rosellas. The low level of diversity among all BFDV isolates from red-fronted parakeets could suggest a more recent infection among these birds compared to the yellow-crowned parakeets, whereas the diversity in the eastern rosellas indicates a much more established infection. Pro-active screening and monitoring of BFDV infection rates in aviaries as well as in wild populations are necessary to limit the risk of transmission among threatened and endangered parrot populations in New Zealand.

For threatened species with small captive populations, it is advisable to incorporate conservation management strategies that minimize inbreeding in an effort to avoid inbreeding depression. Using multilocus microsatellite genotype data, we found a significant negative relationship between genetic relatedness (inbreeding) and reproductive success (fitness) in a captive population of the critically endangered Black Stilt or KakīHimantopus novaezelandiae. In an effort to avoid inbreeding depression in this iconic New Zealand endemic, we recommend re-pairing closely related captive birds with less related individuals and pairing new captive birds with distantly related individuals.

Threatened species recovery programmes benefit from incorporating genomic data into conservation management strategies to enhance species recovery. However, a lack of readily available genomic resources, including conspecific reference genomes, often limits the inclusion of genomic data. Here, we investigate the utility of closely related high-quality reference genomes for single nucleotide polymorphism (SNP) discovery using the critically endangered kakī/black stilt (Himantopus novaezelandiae) and four Charadriiform reference genomes as proof of concept. We compare diversity estimates (i.e., nucleotide diversity, individual heterozygosity, and relatedness) based on kakī SNPs discovered from genotyping-by-sequencing and whole genome resequencing reads mapped to conordinal (killdeer, Charadrius vociferus), confamilial (pied avocet, Recurvirostra avosetta), congeneric (pied stilt, Himantopus himantopus) and conspecific reference genomes. Results indicate that diversity estimates calculated from SNPs discovered using closely related reference genomes correlate significantly with estimates calculated from SNPs discovered using a conspecific genome. Congeneric and confamilial references provide higher correlations and more similar measures of nucleotide diversity, individual heterozygosity, and relatedness. While conspecific genomes may be necessary to address other questions in conservation, SNP discovery using high-quality reference genomes of closely related species is a cost-effective approach for estimating diversity measures in threatened species.

Hybridization facilitated by human activities has dramatically altered the evolutionary trajectories of threatened taxa around the globe. Whereas introduced mammalian predators and widespread habitat loss and degradation clearly imperil the recovery and survival of the New Zealand endemic black stilt or kak o( Himantopus novaezelandiae), the risk associated with hybridization between this critically endangered endemic and its self-introduced congener, the pied stilt or poaka (Himantopus himantopus leucocephalus) is less clear. Here, we combine Bayesian admixture analyses of microsatellite data with mitochondrial DNA sequence data to assess the levels of hybridization and introgression between kak o and poaka. We show that birds classified as hybrids on the basis of adult plumage are indeed of hybrid origin and that hybridization between kak o and poaka is both extensive and bidirectional. Despite this, we found almost no evidence for introgression from poaka to kak o, thus negating the popular belief that kak o represent a hybrid swarm. To our knowledge, ours represents the first comprehensive study to document a lack of widespread introgression for a species at risk despite a recent history of extensive bidirectional human-induced hybridization. We attribute this rather surprising result, in part, to reduced reproductive success in female hybrids combined with a transient male-biased kak o sex ratio. To maximize the evolutionary potential of kak o, we use these data to recommend conservation management activities aimed to maintain the genetic integrity and to maximize the genetic diversity of this iconic rare bird.

To test the hypothesis that both physical and ecological barriers to gene flow drive population differentiation in tropical seabirds, we surveyed mitochondrial control region variation in 242 brown boobies (Sula leucogaster), which prefer inshore habitat, and 271 red-footed boobies (S. sula), which prefer pelagic habitat. To determine the relative influence of isolation and gene flow on population structure, we used both traditional methods and a recently developed statistical method based on coalescent theory and Bayesian inference (Isolation with Migration). We found that global population genetic structure was high in both species, and that female-mediated gene flow among ocean basins apparently has been restricted by major physical barriers including the Isthmus of Panama, and the periodic emergence of the Sunda and Sahul Shelves in Southeast Asia. In contrast, the evolutionary history of populations within ocean basins differed markedly between the two species. In brown boobies, we found high levels of population genetic differentiation and limited gene flow among colonies, even at spatial scales as small as 500 km. Although red-footed booby colonies were also genetically differentiated within ocean basins, coalescent analyses indicated that populations have either diverged in the face of ongoing gene flow, or diverged without gene flow but recently made secondary contact. Regardless, gene flow among red-footed booby populations was higher than among brown booby populations. We suggest that these contrasting patterns of gene flow within ocean basins may be explained by the different habitat preferences of brown and red-footed boobies.

Ancient DNA has revolutionized the way in which evolutionary biologists research both extinct and extant taxa, from the inference of evolutionary history to the resolution of taxonomy. Here, we present, to our knowledge, the first study to report the rediscovery of an ‘extinct’ avian taxon, the Tasman booby ( Sula tasmani ), using classical palaeontological data combined with ancient and modern DNA data. Contrary to earlier work, we show an overlap in size between fossil and modern birds in the North Tasman Sea (classified currently as S. tasmani and Sula dactylatra fullagari , respectively). In addition, we show that Holocene fossil birds have mitochondrial control region sequences that are identical to those found in modern birds. These results indicate that the Tasman booby is not an extinct taxon: S. dactylatra fullagari O'Brien & Davies, 1990 is therefore a junior synonym of Sula tasmani van Tets, Meredith, Fullagar & Davidson, 1988 and all North Tasman Sea boobies should be known as S. d. tasmani . In addition to reporting the rediscovery of an extinct avian taxon, our study highlights the need for researchers to be cognizant of multidisciplinary approaches to understanding taxonomy and past biodiversity.

Translocation of individuals among extant populations is an important tool in species conservation that allows managers to supplement dwindling populations and potentially alleviate the deleterious effects of inbreeding. Ideal translocation strategy should consider historical relationships among existing populations to avoid potential disruption of population subdivision and local adaptation. Here, we examine mitochondrial sequence variation in the endangered blue duck Hymenolaimus malacorhynchos, a New Zealand endemic riverine specialist, to facilitate informed decision making in future translocations. Behavioural observations suggest that blue duck dispersal is limited and may result in genetic structure within and between regional populations. We analysed 894 base pairs of mitochondrial control region in 78 adult blue ducks sampled from 11 river catchments across the species’ range (representing four regions in the North Island and three regions in the South Island) and found strong and significant genetic structure both within and among islands. These results, combined with a 2.0% sequence divergence between islands, indicates that North Island and South Island blue ducks should be treated as separate management units. The relationship between genetic differentiation and geographic distance for blue ducks on the South Island conformed to an “isolation by distance” pattern. Overall, we recommend that translocations of blue ducks should not be made between the North and the South Islands and those within each island should be restricted to neighbouring catchments.

Here we present the results of a large-scale bioinformatics annotation of non-coding RNA loci in 48 avian genomes. Our approach uses probabilistic models of hand-curated families from the Rfam database to infer conserved RNA families within each avian genome. We supplement these annotations with predictions from the tRNA annotation tool, tRNAscan-SE and microRNAs from miRBase. We identify 34 lncRNA-associated loci that are conserved between birds and mammals and validate 12 of these in chicken. We report several intriguing cases where a reported mammalian lncRNA, but not its function, is conserved. We also demonstrate extensive conservation of classical ncRNAs (e.g., tRNAs) and more recently discovered ncRNAs (e.g., snoRNAs and miRNAs) in birds. Furthermore, we describe numerous "losses" of several RNA families, and attribute these to either genuine loss, divergence or missing data. In particular, we show that many of these losses are due to the challenges associated with assembling avian microchromosomes. These combined results illustrate the utility of applying homology-based methods for annotating novel vertebrate genomes.

To test the hypothesis that nonphysical barriers to gene flow play a role in the divergence of low-latitude seabird populations, we applied phylogeographic methods to mitochondrial control region sequence variation in a global sample of masked boobies ( Sula dactylatra ). In accord with previous studies, we found that Indo-Pacific and Atlantic haplotypes form two divergent lineages, excluding one haplotype previously attributed to secondary contact between the Indian Ocean and the Caribbean Sea. Within the Indo-Pacific and the Atlantic, we found a relatively large number of haplotypes, many of which were unique to a single population. Although haplotypes from most populations were found in more than one higher-level clade, nested clade analysis revealed a significant association between clades and geography for the majority of higher-level clades, most often interpreted as a consequence of isolation by distance. We found low levels of gene flow within Indo-Pacific and Atlantic populations, and a significant correlation between gene flow and geographical distance among Indo-Pacific populations. We estimate that Indo-Pacific masked boobies experienced rapid population growth ∼ 180 000 years ago and that the majority of Indo-Pacific and Atlantic populations diverged within the last ∼ ∼ ∼ 115 000 years. These combined data suggest that the predominant pattern between Indo-Pacific and Atlantic populations is long-term isolation by physical barriers to gene flow. In contrast, populations within these regions appear to have diverged despite few obvious physical barriers to gene flow, perhaps as a consequence of limited natal dispersal combined with local adaptation and/or genetic drift.

To further test the hypothesis that the Isthmus of Panama is a major barrier to gene flow in pantropical seabirds, we applied phylogeographic methods to mitochondrial control sequence variation in masked booby (Sula dactylatra) populations on either side of the Isthmus of Panama and the southern tip of Africa. In accord with Steeves et al. (2003), we found that all Caribbean masked boobies with the ‘secondary contact’ cytochrome b haplotype (m-B) shared a control region haplotype (Sd_100), which grouped with Indian‐ Pacific haplotypes and not Caribbean‐Atlantic haplotypes. In addition, Sd_100 was more closely related to control region haplotypes in the Indian Ocean than in the Pacific. We also found that the ‘secondary contact’ birds diverged more recently from extant populations in the Indian Ocean than in the Pacific. Thus, it appears that these masked boobies did not breach the Isthmus of Panama. Rather, birds likely dispersed around the southern tip of Africa during favourable oceanographic conditions in the Pleistocene.

We examined mitochondrial cytochrome b sequence variation in masked Sula dactylatra, red-footed S. sula, and brown S. leucogaster boobies sampled from islands in the central and eastern Pacific Ocean and in the Caribbean Sea. Each species showed a different phylogeographic pattern. Whereas haplotypes in masked and red-footed boobies were shared across the central and eastern Pacific (i.e., across the Eastern Pacific Basin), brown booby haplotypes were not shared across the Eastern Pacific Basin. Although most masked booby haplotypes from the Pacific were distinct from those in the Caribbean, one haplotype was shared across the Isthmus of Panama. Red-footed and brown boobies, however, did not share haplotypes across the Isthmus of Panama. We estimate that divergence of these regional populations occurred within the last 560,000 years. Thus, the Isthmus of Panama and the Eastern Pacific Basin (albeit to a lesser degree) appear to have played a role in the diversification of these species.

Cyanoramphus malherbi is a critically endangered endemic parakeet, or kākāriki, restricted to three mainland valleys in North Canterbury and four translocated populations on predator-free offshore islands of New Zealand. Using genomic next generation sequencing to identify microsatellites, 16,497 sequenced fragments were obtained. Primers were designed for 35 of 62 detected loci: 26 amplified successfully and 18 were polymorphic. Sequence homology to three assembled avian genomes was used to assign five loci to five different autosomes. The number of alleles per locus ranged from 2 to 7 with observed heterozygosities ranging between 0.09 and 0.70. All 18 loci readily cross-amplified in Cyanoramphus auriceps, indicating the utility of these markers for conservation genetic management of both species.

Although the majority of eastern North Pacific (ENP) gray whales migrate to feeding grounds in the Bering and Chukchi Seas, some terminate the migration in more southerly areas such as British Columbia (BC). Long-term sighting studies in Clayoquot Sound (CS), BC, indicate that approximately 35–50 individuals exhibit long-term fidelity to this site. To determine the sex composition (based on genetic sexing) of CS gray whales and to assess whether matrilineal site-fidelity occurs in CS, we collected skin biopsy samples from 16 CS individuals (‘residents’) and 41 samples from other areas (representative of the overall population in the ENP: ‘non-residents’). A total of 27 polymorphic sites defined 24 haploytpes among the 57 samples sequenced for HV1 of the mtDNA control region. The nucleotide and haplotype diversities of these samples were 0.017 (SE = 0.0012) and 0.94 (SE = 0.0019), respectively. Neighbor-joining analysis revealed five lineages each of which contained haplotypes that were observed in both residents and non-residents. Residents did not differ significantly from non-residents, and no significant sex-ratio bias was found. These data suggest a level of diversity that is inconsistent with a severe historical bottleneck, and given the available sample size, do not indicate matrilineally directed fidelity to Clayoquot Sound.

Eight polymorphic microsatellite primer pairs were developed for the critically endangered New Zealand black stilt, Himantopus novaezelandiae, representing the first microsatellite markers available for birds in the family Recurvirostridae. The number of alleles ranged from two to four per locus. Observed and expected heterozygosities ranged from 0.30 to 0.80 and from 0.37 to 0.70, respectively. All eight loci were polymorphic in the related species Himantopus himantopus leucocephalus, indicating these primer pairs may be useful for additional taxa in the globally distributed genus Himantopus.

Habitat utilization and prey species of Vancouver Island gray whales were investigated by (1) summarizing 26 yr of distribution and feeding data and (2) conducting intensive observations in Clayoquot Sound, Vancouver Island, from 1989 to 1996. Whale distribution and movements were monitored from March to November through systematic boat surveys and whale-watch sighting programs. Prey species were collected by suction hose and plankton net or determined through analysis of fecal samples. Gray whales utilized virtually all of the southern west coast of Vancouver Island over the 26-yr observation period. Distribution, prey species, and feeding behavior showed marked variability during any one season and between years. Some feeding areas were used on an annual basis, others with >10-yr intervals between use. Feeding occurred in shallow sand or mud bays, eel grass beds, kelp beds, in the open water column, and at the surface. Young whales appeared to utilize habitat and prey species differently than adults. Main prey species included herring eggs/larvae (Clupea harengus pallasi), crab larvae (Cancer magister megalops, Pachycbeles spp. zoea), mysids (Holmesimysis sculpta, Neomysis rayii, Acanthomysis spp.), amphipods (Ampelisca spp., Atylus borealis), and ghost shrimp (Callianassa californiensis). The definition and relative importance of specific feeding grounds and the study of human impacts on this population are complicated by its broad and variable use of habitat and prey species.

Pelagic seabirds are highly mobile, reducing the likelihood of allopatric speciation where disruption of gene flow between populations is caused by physically insurmountable, extrinsic barriers. Spatial segregation during the non-breeding season appears to provide an intrinsic barrier to gene flow among seabird populations that otherwise occupy nearby or overlapping regions during breeding, but how this is achieved remains unclear. Here we show that the two genetically distinct populations of Cook's petrel (Pterodroma cookii) exhibit transequatorial separation of non-breeding ranges at contemporary (ca. 2-3 yrs) and historical (ca. 100 yrs) time scales. Segregation during the non-breeding season per se appears as an unlikely barrier to gene flow. Instead we provide evidence that habitat specialization during the non-breeding season is associated with breeding asynchrony which, in conjunction with philopatry, restricts gene flow. Habitat specialization during breeding and non-breeding likely promotes evolutionary divergence between these two populations via local adaptation.

Hybridization facilitated by human activities has dramatically altered the evolutionary trajectories of threatened taxa around the globe. Whereas introduced mammalian predators and widespread habitat loss and degradation clearly imperil the recovery and survival of the New Zealand endemic black stilt or kakī (Himantopus novaezelandiae), the risk associated with hybridization between this critically endangered endemic and its self-introduced congener, the pied stilt or poaka (Himantopus himantopus leucocephalus) is less clear. Here, we combine Bayesian admixture analyses of microsatellite data with mitochondrial DNA sequence data to assess the levels of hybridization and introgression between kakī and poaka. We show that birds classified as hybrids on the basis of adult plumage are indeed of hybrid origin and that hybridization between kakī and poaka is both extensive and bidirectional. Despite this, we found almost no evidence for introgression from poaka to kakī, thus negating the popular belief that kakī represent a hybrid swarm. To our knowledge, ours represents the first comprehensive study to document a lack of widespread introgression for a species at risk despite a recent history of extensive bidirectional human-induced hybridization. We attribute this rather surprising result, in part, to reduced reproductive success in female hybrids combined with a transient male-biased kakī sex ratio. To maximize the evolutionary potential of kakī, we use these data to recommend conservation management activities aimed to maintain the genetic integrity and to maximize the genetic diversity of this iconic rare bird.

Pitman and Jehl (1998) recently argued that Masked Boobies (formerly Sula dactylatra granti) breeding on the Nazca Plate in the eastern Pacific Ocean are morphologically and ecologically distinct from other Masked Boobies and may represent a full species. The American Ornithologists' Union subsequently elevated that subspecies to a full species: the Nazca Booby (S. granti). To evaluate that change in classification, we compared sequence variation in the mitochondrial cytochrome-b gene among 75 Nazca Boobies and 37 Masked Boobies representing three subspecies from the central and eastern Pacific and Atlantic oceans. Results indicated strong differentiation of cytochrome-b variation among taxa. Sequences constituted three distinct groups: Nazca Boobies, Masked Boobies from the central and eastern Pacific (S. d. personata and S. d. californica), and Masked Boobies (S. d. dactylatra) from the Caribbean and Atlantic. Those three groups probably diverged within a very short period, 400,000–500,000 year...