Endemic to Aotearoa New Zealand, the kākāpō is among the world’s rarest bird species with a current population of less than 250 individuals. As part of ongoing research efforts to support the conservation of this intensively managed species, we utilised a metatranscriptomics approach to elucidate, for the first time, the expression of antibiotic resistance genes within the kākāpō chick and adult gut microbiomes, further leveraging these data to describe the kākāpō virome. To determine differences among members of the kākāpō population, our data were obtained from birds encompassing different ages, sexes, geographic locations and antibiotic histories. We additionally analysed a time-series dataset following a single male kākāpō over the course of antibiotic treatment during a case of exudative cloacitis. There were significant differences between chicks and adult kākāpō in both the expression of antibiotic resistance genes and their viromes. Expression of these genes indicated potential resistance against 32 antibiotic classes, including 14 single classes of antibiotic and 18 multidrug classes. We identified 74 viral families, but no known avian-infecting viruses. Our case study of the single kākāpō during antibiotic treatment revealed notable changes in expression across time, with a reassuring lack of antibiotic resistance gene expression towards the end of the treatment, indicative of continued efficacy of antibiotic treatment. These novel data will help to inform conservation efforts for this enigmatic and unique bird species.

The k & amacr;k & amacr;p & omacr; (Strigops habroptilus) is a critically endangered, flightless parrot endemic to Aotearoa New Zealand. While an intensive recovery programme has led to considerable successes in k & amacr;k & amacr;p & omacr; conservation, management of this iconic species has been complicated since 2002 by the emergence of the inflammatory syndrome exudative cloacitis. Cloacitis is a significant threat to k & amacr;k & amacr;p & omacr; health yet its origin remains unknown. With recent increases in prevalence coupled with the first detection of cases originating on islands beyond where it was initially detected, understanding the risk factors that lead to this condition remains a high priority for k & amacr;k & amacr;p & omacr; conservation. Here we undertake a pathogen discovery approach incorporating metagenomic (DNA) and metatranscriptomic (RNA) sequencing of faecal and cloacal swab samples from affected and healthy k & amacr;k & amacr;p & omacr;. We report, for the first time, differentially higher transcript abundances from five bacterial taxa in cloacitis versus healthy birds, as well as abundant Clostridium sensu stricto 1 bacteria in severe cloacitis cases. Furthermore, affected birds exhibited significantly higher expression of hok/gef genes that encode for toxin proteins in bacteria including Escherichia coli, the dominant member of the k & amacr;k & amacr;p & omacr; microbiome. While not directly identifying specific risk factors, our results contribute to ongoing efforts to elucidate the underlying causes of exudative cloacitis in k & amacr;k & amacr;p & omacr;.

The critically endangered kākāpō ( Strigops habroptilus ) has suffered population declines due to habitat loss, hunting, and predation. Conservation efforts, including translocation to predator-free islands, have helped increase numbers of this flightless parrot from 51 individuals in 1995–142 in 2019. However, low fertility and high embryo mortality, likely due to genetic bottlenecks continue to hinder population growth. This is further aggravated by the kākāpō’s lek mating system, which allows only a minority of males to father a disproportionate number of offspring, resulting in 21% of non-reproductive males. The study aimed to enhance assisted reproduction techniques to assess male fertility, increase egg fertility, and ensure genetic diversity. Artificial insemination (AI) was used to mimic a second copulation, as females mating with multiple males show higher fertility rates. During the 2019 breeding season, semen collection was successful in 20 males and in 93.5% of 46 attempts using abdominal massage method and electric stimulation technique. Semen volume, colour, consistency, contamination, pH and the motility, concentration, viability and morphology of spermatozoa were analysed. Ejaculate volume ranged between 0.1 and 210 µl and the mean pH was 7.5 ± 0.4 ( x― ± SD). Average sperm viability was 87.4 ± 10.0% with a total motility of 60.9 ± 22.0% and a progressive motility of 28.3 ± 19.8%. AI was performed 15 times in 12 females, improving second clutch fertility (70% vs. 29.4% without AI). Egg fertility in the second clutch without AI was 29.41% (5/17) compared to 70% (14/20) after AI. Paternity testing confirmed AI offspring (four chicks of three females), including from two previously non-reproductive males, enriching the gene pool with rare alleles (e.g., genes from Fiordland founding population). This study demonstrates the value of assisted reproduction in conserving endangered avian species by improving reproductive success and preserving genetic diversity.

The ability of viruses to emerge in new species is influenced by aspects of host biology and ecology, with some taxa harbouring a high diversity and abundance of viruses. However, how these factors shape virus diversity at the ecosystem scale is often unclear. To better understand the pattern and determinants of viral diversity within an ecosystem, and to describe the novel avian viruses infecting an individual avian community, we performed a metagenomic snapshot of the virome from the entire avian community on remote Pukenui/Anchor Island in Aotearoa New Zealand. Through total RNA sequencing of 18 bird species, we identified 50 avian viruses from 9 viral families, of which 96% were novel. Of note, passerines (perching birds) exhibited high viral abundance and diversity, with viruses found across all nine viral families identified. We also identified numerous viruses infecting seabirds on the Island, including megriviruses, hepaciviruses, and hepatoviruses, while parrots exhibited an extremely low diversity of avian viruses. Within passerines, closely related astroviruses and hepatoviruses, and multiple identical hepe-like viruses, were shared among host species. Phylogenetic reconciliation analysis of these viral groups revealed a mixture of co-divergence and cross-species transmission, with virus host-jumping relatively frequent among passerines. In contrast, there was no evidence for recent cross-species virus transmission in parrots or seabirds. The novel pegiviruses and a flavivirus identified here also pose intriguing questions regarding their origins, pathogenicity, and potential impact on vertebrate hosts. Overall, these results highlight the importance of understudied remote island ecosystems as refugia for novel viruses, as well as the intricate interplay between host ecology and behaviour in shaping viral communities.

The kākāpō is a critically endangered flightless parrot which suffers from exudative cloacitis, a debilitating disease resulting in inflammation of the vent margin or cloaca. Despite this disease emerging over 20 years ago, the cause of exudative cloacitis remains elusive. We used total RNA sequencing and metatranscriptomic analysis to characterise the infectome of lesions and cloacal swabs from nine kākāpō affected with exudative cloacitis, and compared this to cloacal swabs from 45 non-diseased kākāpō. We identified three bacterial species-Streptococcus gallolyticus, Enterococcus faecalis and Escherichia coli-as significantly more abundant in diseased kākāpō compared to healthy individuals. The genetic diversity observed in both S. gallolyticus and E. faecalis among diseased kākāpō suggests that these bacteria originate from exogenous sources rather than from kākāpō-to-kākāpō transmission. The presence of extraintestinal pathogenic E. coli (ExPEC)-associated virulence factors in the diseased kākāpō population suggests that E. coli may play a critical role in disease progression by facilitating iron acquisition and causing DNA damage in host cells, possibly in association with E. faecalis. No avian viral, fungal nor other parasitic species were identified. These results, combined with the consistent presence of one E. coli gnd sequence type across multiple diseased birds, suggest that this species may be the primary cause of exudative cloacitis. These findings shed light on possible causative agents of exudative cloacitis, and offer insights into the interplay of microbial factors influencing the disease.

The information contained in population genomic data can tell us much about the past ecology and evolution of species. We leveraged detailed phenotypic and genomic data of nearly all living kākāpō to understand the evolution of its feather color polymorphism. The kākāpō is an endangered and culturally significant parrot endemic to Aotearoa New Zealand, and the green and olive feather colorations are present at similar frequencies in the population. The presence of such a neatly balanced color polymorphism is remarkable because the entire population currently numbers less than 250 birds, which means it has been exposed to severe genetic drift. We dissected the color phenotype, demonstrating that the two colors differ in their light reflectance patterns due to differential feather structure. We used quantitative genomics methods to identify two genetic variants whose epistatic interaction can fully explain the species’ color phenotype. Our genomic forward simulations show that balancing selection might have been pivotal to establish the polymorphism in the ancestrally large population, and to maintain it during population declines that involved a severe bottleneck. We hypothesize that an extinct apex predator was the likely agent of balancing selection, making the color polymorphism in the kākāpō a “ghost of selection past.”

We used non-invasive real-time genomic approaches to monitor one of the last surviving populations of the critically endangered kākāpō (Strigops habroptilus). We first established an environmental DNA metabarcoding protocol to identify the distribution of kākāpō and other vertebrate species in a highly localized manner using soil samples. Harnessing real-time nanopore sequencing and the high-quality kākāpō reference genome, we then extracted species-specific DNA from soil. We combined long read-based haplotype phasing with known individual genomic variation in the kākāpō population to identify the presence of individuals, and confirmed these genomically informed predictions through detailed metadata on kākāpō distributions. This study shows that individual identification is feasible through nanopore sequencing of environmental DNA, with important implications for future efforts in the application of genomics to the conservation of rare species, potentially expanding the application of real-time environmental DNA research from monitoring species distribution to inferring fitness parameters such as genomic diversity and inbreeding.

The information contained in population genomic data can tell us much about the past ecology and evolution of species. We leveraged detailed phenotypic and genomic data of nearly all living kākāpō to understand the evolution of its remarkable feather color polymorphism. The kākāpō is an endangered and culturally significant parrot endemic to Aotearoa New Zealand, and the green and olive feather colorations are present at similar frequencies in the population. The presence of such a neatly balanced color polymorphism is remarkable because the entire population currently numbers less than 250 birds, which means it has been exposed to severe genetic drift. We dissected the color phenotype, demonstrating that the two colors differ in their light reflectance patterns due to differential feather structure. We used quantitative genomics methods to identify two genetic variants whose epistatic interaction can fully explain the species’ color phenotype. Our genomic forward simulations show that balancing selection might have been pivotal to establish the polymorphism in the ancestrally large population, and to maintain it during population declines that involved a severe bottleneck. We hypothesize that an extinct apex predator was the likely agent of balancing selection, making the color polymorphism in the kākāpō a “ghost of selection past”.

The critically endangered kākāpō is a flightless, nocturnal parrot endemic to Aotearoa New Zealand. During the 2019 breeding season multiple kākāpō on Whenua Hou, an island with one of the largest kākāpō populations, contracted the respiratory fungal disease aspergillosis. Infection is caused by members of the genus Aspergillus which can seriously impact avian health and is often fatal to wild and domestic birds. We sought to understand how Aspergillus influenced the gut mycobiota of kākāpō chicks and surrounding nest litter communities over a 10-week period. This study was conducted in parallel with another longitudinal project utilising the same samples where we investigated the influence of management practice on development of gut bacterial communities in kākāpō chicks. Ultimately, aspergillosis infection had no significant impact on the fungal communities of kākāpō chicks or nest litter, and nor did regular management practices (i.e. artificially removing faecal material from nests). Age and time, however, were significantly associated with variation in the mycobiotas of chick and nest samples. Overall, this study represents our first glimpse of the kākāpō gut mycobiota and provides essential information and future research directions for the conservation of this rare species.

Background Animal conservation often requires intensive management actions to improve reproductive output, yet any adverse effects of these may not be immediately apparent, particularly in threatened species with small populations and long lifespans. Hand-rearing is an example of a conservation management strategy which, while boosting populations, can cause long-term demographic and behavioural problems. It is used in the recovery of the critically endangered kākāpō (Strigops habroptilus), a flightless parrot endemic to New Zealand, to improve the slow population growth that is due to infrequent breeding, low fertility and low hatching success. Methods We applied Bayesian mixed models to examine whether hand-rearing and other factors were associated with clutch fertility in kākāpō. We used projection predictive variable selection to compare the relative contributions to fertility from the parents’ rearing environment, their age and previous copulation experience, the parental kinship, and the number of mates and copulations for each clutch. We also explored how the incidence of repeated copulations and multiple mates varied with kākāpō density. Results The rearing status of the clutch father and the number of mates and copulations of the clutch mother were the dominant factors in predicting fertility. Clutches were less likely to be fertile if the father was hand-reared compared to wild-reared, but there was no similar effect for mothers. Clutches produced by females copulating with different males were more likely to be fertile than those from repeated copulations with one male, which in turn had a higher probability of fertility than those from a single copulation. The likelihood of multiple copulations and mates increased with female:male adult sex ratio, perhaps as a result of mate guarding by females. Parental kinship, copulation experience and age all had negligible associations with clutch fertility. Conclusions These results provide a rare assessment of factors affecting fertility in a wild threatened bird species, with implications for conservation management. The increased fertility due to multiple mates and copulations, combined with the evidence for mate guarding and previous results of kākāpō sperm morphology, suggests that an evolutionary mechanism exists to optimise fertility through sperm competition in kākāpō. The high frequency of clutches produced from single copulations in the contemporary population may therefore represent an unnatural state, perhaps due to too few females. This suggests that opportunity for sperm competition should be maximised by increasing population densities, optimising sex ratios, and using artificial insemination. The lower fertility of hand-reared males may result from behavioural defects due to lack of exposure to conspecifics at critical development stages, as seen in other taxa. This potential negative impact of hand-rearing must be balanced against the short-term benefits it provides.

The gut microbiota plays an essential role in host health that has important implications for the conservation management of threatened wildlife. While factors such as diet, medication, and habitat are known to shape the microbiota, our understanding of the entirety of factors, including the complex role of the host genomic background, remains incomplete. Our research on the gut microbiota of the critically endangered kākāpō ( Strigops habroptilus ), a flightless parrot species endemic to Aotearoa New Zealand, represents, to our knowledge, the first study to describe the gastrointestinal bacterial diversity for virtually an entire species and to assess the relationship between gut microbiota and host genomic diversity in a highly threatened population. Here we report a 16S rRNA gene-based analysis of kākāpō faecal samples representing the gut microbiota for 84% of kākāpō (n = 133). This survey was then leveraged with exceptional metadata to tease apart the impact of host genomic diversity and factors such as sex, diet, antibiotic treatment, disease status, habitat, and time of sampling on the kākāpō gut microbiota, with sex being the only covariate significantly associated with gut microbiota diversity. We find evidence of a highly polygenic genomic architecture of the gut microbiota and further identify putative associations between gut bacterial diversity and functional biological pathways related to intestinal homeostasis, inflammation, immune response and metabolism. This improved understanding of the kākāpō gut microbiota – and its relationship with host genomics – can directly benefit kākāpō management and conservation by providing new insights into the role of the gut microbiome in kākāpō health and disease mitigation. Overall, we anticipate that an integration of microbiome studies in conservation research and management will improve our understanding of how the concept of One Health with its implications for human, animal and environmental welfare can be achieved.

Virus transmission between host species underpins disease emergence. Both host phylogenetic relatedness and aspects of their ecology, such as species interactions and predator-prey relationships, may govern rates and patterns of cross-species virus transmission and hence zoonotic risk. To address the impact of host phylogeny and ecology on virus diversity and evolution, we characterized the virome structure of a relatively isolated island ecological community in Fiordland, New Zealand, that are linked through a food web. We show that phylogenetic barriers that inhibited cross-species virus transmission occurred at the level of host phyla (between the Chordata, Arthropoda and Streptophyta) as well as at lower taxonomic levels. By contrast, host ecology, manifest as predator-prey interactions and diet, had a smaller influence on virome composition, especially at higher taxonomic levels. The virus-host community comprised a 'small world' network, in which hosts with a high diversity of viruses were more likely to acquire new viruses, and generalist viruses that infect multiple hosts were more likely to infect additional species compared to host specialist viruses. Such a highly connected ecological community increases the likelihood of cross-species virus transmission, particularly among closely related species, and suggests that host generalist viruses present the greatest risk of disease emergence.

The kākāpō is a critically endangered, intensively managed, long-lived nocturnal parrot endemic to Aotearoa New Zealand. We generated and analyzed whole-genome sequence data for nearly all individuals living in early 2018 (169 individuals) to generate a high-quality species-wide genetic variant callset. We leverage extensive long-term metadata to quantify genome-wide diversity of the species over time and present new approaches using probabilistic programming, combined with a phenotype dataset spanning five decades, to disentangle phenotypic variance into environmental and genetic effects while quantifying uncertainty in small populations. We find associations for growth, disease susceptibility, clutch size, and egg fertility within genic regions previously shown to influence these traits in other species. Finally, we generate breeding values to predict phenotype and illustrate that active management over the past 45 years has maintained both genome-wide diversity and diversity in breeding values, and hence, evolutionary potential. We provide new pathways for informing future conservation management decisions for kākāpō, including prioritizing individuals for translocation and monitoring individuals with poor growth or high disease risk. Overall, by explicitly addressing the challenge of small sample size, we provide a template for the inclusion of genomic data that will be transformational for species recovery efforts around the globe.

Abstract Background The critically endangered kākāpō is a flightless, nocturnal parrot endemic to Aotearoa New Zealand. Recent efforts to describe the gastrointestinal microbial community of this threatened herbivore revealed a low-diversity microbiota that is often dominated by Escherichia-Shigella bacteria. Given the importance of associated microbial communities to animal health, and increasing appreciation of their potential relevance to threatened species conservation, we sought to better understand the development of this unusual gut microbiota profile. To this end, we conducted a longitudinal analysis of faecal material collected from kākāpō chicks during the 2019 breeding season, in addition to associated nest litter material. Results Using an experimental approach rarely seen in studies of threatened species microbiota, we evaluated the impact of a regular conservation practice on the developing kākāpō microbiota, namely the removal of faecal material from nests. Artificially removing chick faeces from nests had negligible impact on bacterial community diversity for either chicks or nests (p > 0.05). However, the gut microbiota did change significantly over time as chick age increased (p < 0.01), with an increasing relative abundance of Escherichia-Shigella coli over the study period and similar observations for the associated nest litter microbiota (p < 0.01). Supplementary feeding substantially altered gut bacterial diversity of kākāpō chicks (p < 0.01), characterised by a significant increase in Lactobacillus bacteria. Conclusions Overall, chick age and hand rearing conditions had the most marked impact on faecal bacterial communities. Similarly, the surrounding nest litter microbiota changed significantly over time since a kākāpō chick was first placed in the nest, though we found no evidence that removal of faecal material influenced the bacterial communities of either litter or faecal samples. Taken together, these observations will inform ongoing conservation and management of this most enigmatic of bird species.

Kākāpō (Strigops habroptilus) are critically endangered nocturnal parrots endemic to New Zealand. Exudative cloacitis is a disease only affecting the breeding population of Kākāpō on Whenua Hou (Codfish Island), for which a consistent primary pathogenic organism involved has not been identified. This epidemiological study was conducted to identify the environmental factors contributing to the initiation of this disease in Kākāpō by 1) producing and describing a case series; 2) mapping the geographic distribution of exudative cloacitis cases; 3) investigating the chemical characteristics of Kākāpō roosting sites; and 4) assessing the effects of climatic factors on the incidence of exudative cloacitis each year. Soil samples from the Kākāpō roost sites and other areas of the Whenua Hou were examined for pH, ammonium, and moisture contents. From 2002 to 2017, 22 sporadic cases of exudative cloacitis have been diagnosed and the disease distribution on Whenua Hou overlaps the Kākāpō distribution. A mixed group of adults and juveniles was affected and there was no evidence of spatial or temporal clustering of the disease. Current findings on the chemical characteristics of Kākāpō roosting sites do not show any evidence that these factors are involved in the initiation of the exudative cloacitis. Nevertheless, the results suggest that the ammonium and moisture levels of the roosts are worthy of more detailed study in future cases. We were not able to demonstrate any significant influence of broad measures of climate on the incidence of exudative cloacitis on Whenua Hou. Prospective data collection would help for a complete epidemiological investigation of this disease in future cases.

Kākāpō are a critically endangered species of parrots restricted to a few islands off the coast of New Zealand. Kākāpō are very closely monitored, especially during nesting seasons. In 2019, during a highly successful nesting season, an outbreak of aspergillosis affected 21 individuals and led to the deaths of 9, leaving a population of only 211 kākāpō. In monitoring this outbreak, cultures of aspergillus were grown, and genome sequenced. These sequences demonstrate that, very unusually for an aspergillus outbreak, a single strain of aspergillus caused the outbreak. This strain was found on two islands, but only one had an outbreak of aspergillosis; indicating that the strain was necessary, but not sufficient, to cause disease. Our analysis provides an understanding of the 2019 outbreak and provides potential ways to manage such events in the future.

Background The Aotearoa New Zealand takahē ( Porphyrio hochstetteri ), once thought to be extinct, is a nationally threatened flightless rail under intensive conservation management. While there has been previous research into disease-related microbes in takahē, little is known about the microbes present in the gastrointestinal tract. Given the importance of gut-associated microbes to herbivore nutrition and immunity, knowledge of these communities is likely to be of considerable conservation value. Here we examined the gut microbiotas of 57 takahē at eight separate locations across Aotearoa New Zealand. Results Faecal samples, taken as a proxy for the hindgut bacterial community, were subjected to 16S rRNA gene amplicon sequencing using Illumina MiSeq. Phylogenetic analysis of > 2200 amplicon sequence variants (ASVs) revealed nine main bacterial phyla ( Acidobacteriota, Actinobacteriota , Bacteroidota , Campilobacterota, Firmicutes , Fusobacteriota , Planctomycetota, Proteobacteria , and Verrucomicrobiota ) that accounted for the majority of sequence reads. Location was a significant effect ( p value < 0.001, 9999 permutations) that accounted for 32% of the observed microbiota variation. One ASV, classified as Lactobacillus aviarius , was present in all samples at an average relative abundance of 17% (SD = 23.20). There was strong evidence ( p = 0.002) for a difference in the abundance of the genus Lactobacillus between locations. A common commensal bacterium previously described in takahē, Campylobacter spp., was also detected in most faecal samples. Conclusions Location plays a pivotal role in the observed variation among takahē gut bacterial communities and is potentially due to factors such as supplemental feeding and medical treatment experienced by birds housed in captivity at one of the eight sampled sites. These data present a first glimpse of the previously unexplored takahē gut microbiota and provide a baseline for future microbiological studies and conservation efforts.

In response to anthropogenic threats, conservation translocations are increasingly used to combat species' population and range declines. However, moving animals outside of their current distribution can mean introducing them to novel conditions, even in the case of reintroductions to formerly inhabited areas due to ecosystem changes following extirpation. This exposure to novel conditions introduces uncertainty that can undermine decision making for species conservation. Here we propose two strategies, which we define as conservative and extrapolative, for approaching and managing novelty and the resulting uncertainty in conservation translocations. Conservative strategies are characterised by the avoidance and removal of novel conditions as much as possible, whereas extrapolative strategies are more experimental, allowing exposure to novel conditions and monitoring outcomes to increase understanding of a species' ecology. As each strategy carries specific risks and opportunities, they will be applicable in different scenarios. Extrapolative strategies suit species in recovery which can afford some experimental management, or species facing novel and emerging threats which require less traditional translocations, such as assisted colonisations. We provide examples, applying our framework to two endemic New Zealand species with long histories of translocation management: tuatara (Sphenodon punctatus), a reptile and takahē (Porphyrio hochstetteri), a flightless bird.

High-quality and complete reference genome assemblies are fundamental for the application of genomics to biology, disease, and biodiversity conservation. However, such assemblies are available for only a few non-microbial species 1–4 . To address this issue, the international Genome 10K (G10K) consortium 5,6 has worked over a five-year period to evaluate and develop cost-effective methods for assembling highly accurate and nearly complete reference genomes. Here we present lessons learned from generating assemblies for 16 species that represent six major vertebrate lineages. We confirm that long-read sequencing technologies are essential for maximizing genome quality, and that unresolved complex repeats and haplotype heterozygosity are major sources of assembly error when not handled correctly. Our assemblies correct substantial errors, add missing sequence in some of the best historical reference genomes, and reveal biological discoveries. These include the identification of many false gene duplications, increases in gene sizes, chromosome rearrangements that are specific to lineages, a repeated independent chromosome breakpoint in bat genomes, and a canonical GC-rich pattern in protein-coding genes and their regulatory regions. Adopting these lessons, we have embarked on the Vertebrate Genomes Project (VGP), an international effort to generate high-quality, complete reference genomes for all of the roughly 70,000 extant vertebrate species and to help to enable a new era of discovery across the life sciences.

The kākāpō is a flightless parrot endemic to New Zealand. Once common in the archipelago, only 201 individuals remain today, most of them descending from an isolated island population. We report the first genome-wide analyses of the species, including a high-quality genome assembly for kākāpō, one of the first chromosome-level reference genomes sequenced by the Vertebrate Genomes Project (VGP). We also sequenced and analyzed 35 modern genomes from the sole surviving island population and 14 genomes from the extinct mainland population. While theory suggests that such a small population is likely to have accumulated deleterious mutations through genetic drift, our analyses on the impact of the long-term small population size in kākāpō indicate that present-day island kākāpō have a reduced number of harmful mutations compared to mainland individuals. We hypothesize that this reduced mutational load is due to the island population having been subjected to a combination of genetic drift and purging of deleterious mutations, through increased inbreeding and purifying selection, since its isolation from the mainland ∼10,000 years ago. Our results provide evidence that small populations can survive even when isolated for hundreds of generations. This work provides key insights into kākāpō breeding and recovery and more generally into the application of genetic tools in conservation efforts for endangered species.