Genomic Consequences of Fragmentation in the Endangered Fennoscandian Arctic Fox (Vulpes lagopus).

Accelerating climate change is causing severe habitat fragmentation in the Arctic, threatening the persistence of many cold-adapted species. The Scandinavian arctic fox (Vulpes lagopus) is highly fragmented, with a once continuous, circumpolar distribution, it struggled to recover from a demographic bottleneck in the late 19th century. The future persistence of the entire Scandinavian population is highly dependent on the northernmost Fennoscandian subpopulations (Scandinavia and the Kola Peninsula), to provide a link to the viable Siberian population. By analyzing 43 arctic fox genomes, we quantified genomic variation and inbreeding in these populations. Signatures of genome erosion increased from Siberia to northern Sweden indicating a stepping-stone model of connectivity. In northern Fennoscandia, runs of homozygosity (ROH) were on average ~1.47-fold longer than ROH found in Siberia, stretching almost entire scaffolds. Moreover, consistent with recent inbreeding, northern Fennoscandia harbored more homozygous deleterious mutations, whereas Siberia had more in heterozygous state. This study underlines the value of documenting genome erosion following population fragmentation to identify areas requiring conservation priority. With the increasing fragmentation and isolation of Arctic habitats due to global warming, understanding the genomic and demographic consequences is vital for maintaining evolutionary potential and preventing local extinctions.

Genomic trajectories of a near-extinction event in the Chatham Island black robin.

BACKGROUND: Understanding the micro--evolutionary response of populations to demographic declines is a major goal in evolutionary and conservation biology. In small populations, genetic drift can lead to an accumulation of deleterious mutations, which will increase the risk of extinction. However, demographic recovery can still occur after extreme declines, suggesting that natural selection may purge deleterious mutations, even in extremely small populations. The Chatham Island black robin (Petroica traversi) is arguably the most inbred bird species in the world. It avoided imminent extinction in the early 1980s and after a remarkable recovery from a single pair, a second population was established and the two extant populations have evolved in complete isolation since then. Here, we analysed 52 modern and historical genomes to examine the genomic consequences of this extreme bottleneck and the subsequent translocation. RESULTS: We found evidence for two-fold decline in heterozygosity and three- to four-fold increase in inbreeding in modern genomes. Moreover, there was partial support for temporal reduction in total load for detrimental variation. In contrast, compared to historical genomes, modern genomes showed a significantly higher realised load, reflecting the temporal increase in inbreeding. Furthermore, the translocation induced only small changes in the frequency of deleterious alleles, with the majority of detrimental variation being shared between the two populations. CONCLUSION: Our results highlight the dynamics of mutational load in a species that recovered from the brink of extinction, and show rather limited temporal changes in mutational load. We hypothesise that ancestral purging may have been facilitated by population fragmentation and isolation on several islands for thousands of generations and may have already reduced much of the highly deleterious load well before human arrival and introduction of pests to the archipelago. The majority of fixed deleterious variation was shared between the modern populations, but translocation of individuals with low mutational load could possibly mitigate further fixation of high-frequency deleterious variation.

GenErode: a bioinformatics pipeline to investigate genome erosion in endangered and extinct species.

BACKGROUND: Many wild species have suffered drastic population size declines over the past centuries, which have led to 'genomic erosion' processes characterized by reduced genetic diversity, increased inbreeding, and accumulation of harmful mutations. Yet, genomic erosion estimates of modern-day populations often lack concordance with dwindling population sizes and conservation status of threatened species. One way to directly quantify the genomic consequences of population declines is to compare genome-wide data from pre-decline museum samples and modern samples. However, doing so requires computational data processing and analysis tools specifically adapted to comparative analyses of degraded, ancient or historical, DNA data with modern DNA data as well as personnel trained to perform such analyses. RESULTS: Here, we present a highly flexible, scalable, and modular pipeline to compare patterns of genomic erosion using samples from disparate time periods. The GenErode pipeline uses state-of-the-art bioinformatics tools to simultaneously process whole-genome re-sequencing data from ancient/historical and modern samples, and to produce comparable estimates of several genomic erosion indices. No programming knowledge is required to run the pipeline and all bioinformatic steps are well-documented, making the pipeline accessible to users with different backgrounds. GenErode is written in Snakemake and Python3 and uses Conda and Singularity containers to achieve reproducibility on high-performance compute clusters. The source code is freely available on GitHub ( https://github.com/NBISweden/GenErode ). CONCLUSIONS: GenErode is a user-friendly and reproducible pipeline that enables the standardization of genomic erosion indices from temporally sampled whole genome re-sequencing data.

Ancient mitochondrial and modern whole genomes unravel massive genetic diversity loss during near extinction of Alpine ibex.

Population bottlenecks can have dramatic consequences for the health and long-term survival of a species. Understanding of historic population size and standing genetic variation prior to a contraction allows estimating the impact of a bottleneck on the species' genetic diversity. Although historic population sizes can be modelled based on extant genomics, uncertainty is high for the last 10-20 millenia. Hence, integrating ancient genomes provides a powerful complement to retrace the evolution of genetic diversity through population fluctuations. Here, we recover 15 high-quality mitogenomes of the once nearly extinct Alpine ibex spanning 8601 BP to 1919 CE and combine these with 60 published modern whole genomes. Coalescent demography simulations based on modern whole genomes indicate population fluctuations coinciding with the last major glaciation period. Using our ancient and historic mitogenomes, we investigate the more recent demographic history of the species and show that mitochondrial haplotype diversity was reduced to a fifth of the prebottleneck diversity with several highly differentiated mitochondrial lineages having coexisted historically. The main collapse of mitochondrial diversity coincides with elevated human population growth during the last 1-2 kya. After recovery, one lineage was spread and nearly fixed across the Alps due to recolonization efforts. Our study highlights that a combined approach integrating genomic data of ancient, historic and extant populations unravels major long-term population fluctuations from the emergence of a species through its near extinction up to the recent past.

Integrating multi-taxon palaeogenomes and sedimentary ancient DNA to study past ecosystem dynamics.

Ancient DNA (aDNA) has played a major role in our understanding of the past. Important advances in the sequencing and analysis of aDNA from a range of organisms have enabled a detailed understanding of processes such as past demography, introgression, domestication, adaptation and speciation. However, to date and with the notable exception of microbiomes and sediments, most aDNA studies have focused on single taxa or taxonomic groups, making the study of changes at the community level challenging. This is rather surprising because current sequencing and analytical approaches allow us to obtain and analyse aDNA from multiple source materials. When combined, these data can enable the simultaneous study of multiple taxa through space and time, and could thus provide a more comprehensive understanding of ecosystem-wide changes. It is therefore timely to develop an integrative approach to aDNA studies by combining data from multiple taxa and substrates. In this review, we discuss the various applications, associated challenges and future prospects of such an approach.

Ancient and modern genomes unravel the evolutionary history of the rhinoceros family.

Only five species of the once-diverse Rhinocerotidae remain, making the reconstruction of their evolutionary history a challenge to biologists since Darwin. We sequenced genomes from five rhinoceros species (three extinct and two living), which we compared to existing data from the remaining three living species and a range of outgroups. We identify an early divergence between extant African and Eurasian lineages, resolving a key debate regarding the phylogeny of extant rhinoceroses. This early Miocene (∼16 million years ago [mya]) split post-dates the land bridge formation between the Afro-Arabian and Eurasian landmasses. Our analyses also show that while rhinoceros genomes in general exhibit low levels of genome-wide diversity, heterozygosity is lowest and inbreeding is highest in the modern species. These results suggest that while low genetic diversity is a long-term feature of the family, it has been particularly exacerbated recently, likely reflecting recent anthropogenic-driven population declines.

Genomic and fitness consequences of inbreeding in an endangered carnivore.

Reduced fitness through genetic drift and inbreeding is a major threat to small and isolated populations. Although previous studies have generally used genetically verified pedigrees to document effects of inbreeding and gene flow, these often fail to capture the whole inbreeding history of the species. By assembling a draft arctic fox (Vulpes lagopus) genome and resequencing complete genomes of 23 additional foxes born before and after a well-documented immigration event in Scandinavia, we here look into the genomic consequences of inbreeding and genetic rescue. We found a difference in genome-wide diversity, with 18% higher heterozygosity and 81% lower FROH in immigrant F1 compared to native individuals. However, more distant descendants of immigrants (F2, F3) did not show the same pattern. We also found that foxes with lower inbreeding had higher probability to survive their first year of life. Our results demonstrate the important link between genetic variation and fitness as well as the transient nature of genetic rescue. Moreover, our results have implications in conservation biology as they demonstrate that inbreeding depression can effectively be detected in the wild by a genomic approach.

Genomic insights into the conservation status of the world's last remaining Sumatran rhinoceros populations.

Small populations are often exposed to high inbreeding and mutational load that can increase the risk of extinction. The Sumatran rhinoceros was widespread in Southeast Asia, but is now restricted to small and isolated populations on Sumatra and Borneo, and most likely extinct on the Malay Peninsula. Here, we analyse 5 historical and 16 modern genomes from these populations to investigate the genomic consequences of the recent decline, such as increased inbreeding and mutational load. We find that the Malay Peninsula population experienced increased inbreeding shortly before extirpation, which possibly was accompanied by purging. The populations on Sumatra and Borneo instead show low inbreeding, but high mutational load. The currently small population sizes may thus in the near future lead to inbreeding depression. Moreover, we find little evidence for differences in local adaptation among populations, suggesting that future inbreeding depression could potentially be mitigated by assisted gene flow among populations.

African and Asian leopards are highly differentiated at the genomic level.

Leopards are the only big cats still widely distributed across the continents of Africa and Asia. They occur in a wide range of habitats and are often found in close proximity to humans. But despite their ubiquity, leopard phylogeography and population history have not yet been studied with genomic tools. Here, we present population-genomic data from 26 modern and historical samples encompassing the vast geographical distribution of this species. We find that Asian leopards are broadly monophyletic with respect to African leopards across almost their entire nuclear genomes. This profound genetic pattern persists despite the animals' high potential mobility, and despite evidence of transfer of African alleles into Middle Eastern and Central Asian leopard populations within the last 100,000 years. Our results further suggest that Asian leopards originated from a single out-of-Africa dispersal event 500-600 thousand years ago and are characterized by higher population structuring, stronger isolation by distance, and lower heterozygosity than African leopards. Taxonomic categories do not take into account the variability in depth of divergence among subspecies. The deep divergence between the African subspecies and Asian populations contrasts with the much shallower divergence among putative Asian subspecies. Reconciling genomic variation and taxonomy is likely to be a growing challenge in the genomics era.

Population genomics of the critically endangered kakapo.

The kakapo 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 kakapo, 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 kakapo indicate that present-day island kakapo 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 kakapo breeding and recovery and more generally into the application of genetic tools in conservation efforts for endangered species.

Competitive mapping allows for the identification and exclusion of human DNA contamination in ancient faunal genomic datasets.

BACKGROUND: After over a decade of developments in field collection, laboratory methods and advances in high-throughput sequencing, contamination remains a key issue in ancient DNA research. Currently, human and microbial contaminant DNA still impose challenges on cost-effective sequencing and accurate interpretation of ancient DNA data. RESULTS: Here we investigate whether human contaminating DNA can be found in ancient faunal sequencing datasets. We identify variable levels of human contamination, which persists even after the sequence reads have been mapped to the faunal reference genomes. This contamination has the potential to affect a range of downstream analyses. CONCLUSIONS: We propose a fast and simple method, based on competitive mapping, which allows identifying and removing human contamination from ancient faunal DNA datasets with limited losses of true ancient data. This method could represent an important tool for the ancient DNA field.

Population genomics reveals lack of greater white-fronted introgression into the Swedish lesser white-fronted goose.

Interspecific introgression is considered a potential threat to endangered taxa. One example where this has had a major impact on conservation policy is the lesser white-fronted goose (LWfG). After a dramatic decline in Sweden, captive breeding birds were released between 1981-1999 with the aim to reinforce the population. However, the detection of greater white-fronted goose (GWfG) mitochondrial DNA in the LWfG breeding stock led to the release program being dismantled, even though the presence of GWfG introgression in the actual wild Swedish LWfG population was never documented. To examine this, we sequenced the complete genomes of 21 LWfG birds from the Swedish, Russian and Norwegian populations, and compared these with genomes from other goose species, including the GWfG. We found no evidence of interspecific introgression into the wild Swedish LWfG population in either nuclear genomic or mitochondrial data. Moreover, Swedish LWfG birds are genetically distinct from the Russian and Norwegian populations and display comparatively low genomic diversity and high levels of inbreeding. Our findings highlight the utility of genomic approaches in providing scientific evidence that can help improve conservation management as well as policies for breeding and reinforcement programmes.

Consequences of past climate change and recent human persecution on mitogenomic diversity in the arctic fox.

Ancient DNA provides a powerful means to investigate the timing, rate and extent of population declines caused by extrinsic factors, such as past climate change and human activities. One species probably affected by both these factors is the arctic fox, which had a large distribution during the last glaciation that subsequently contracted at the start of the Holocene. More recently, the arctic fox population in Scandinavia went through a demographic bottleneck owing to human persecution. To investigate the consequences of these processes, we generated mitogenome sequences from a temporal dataset comprising Pleistocene, historical and modern arctic fox samples. We found no evidence that Pleistocene populations in mid-latitude Europe or Russia contributed to the present-day gene pool of the Scandinavian population, suggesting that postglacial climate warming led to local population extinctions. Furthermore, during the twentieth-century bottleneck in Scandinavia, at least half of the mitogenome haplotypes were lost, consistent with a 20-fold reduction in female effective population size. In conclusion, these results suggest that the arctic fox in mainland Western Europe has lost genetic diversity as a result of both past climate change and human persecution. Consequently, it might be particularly vulnerable to the future challenges posed by climate change. This article is part of a discussion meeting issue 'The past is a foreign country: how much can the fossil record actually inform conservation?'

Complete genomes of two extinct New Zealand passerines show responses to climate fluctuations but no evidence for genomic erosion prior to extinction.

Human intervention, pre-human climate change (or a combination of both), as well as genetic effects, contribute to species extinctions. While many species from oceanic islands have gone extinct due to direct human impacts, the effects of pre-human climate change and human settlement on the genomic diversity of insular species and the role that loss of genomic diversity played in their extinctions remains largely unexplored. To address this question, we sequenced whole genomes of two extinct New Zealand passerines, the huia (Heteralocha acutirostris) and South Island kokako (Callaeas cinereus). Both species showed similar demographic trajectories throughout the Pleistocene. However, the South Island kokako continued to decline after the last glaciation, while the huia experienced some recovery. Moreover, there was no indication of inbreeding resulting from recent mating among closely related individuals in either species. This latter result indicates that population fragmentation associated with forest clearing by Maori may not have been strong enough to lead to an increase in inbreeding and exposure to genomic erosion. While genomic erosion may not have directly contributed to their extinctions, further habitat fragmentation and the introduction of mammalian predators by Europeans may have been an important driver of extinction in huia and South Island kokako.

Full Mitogenomes in the Critically Endangered Kakapo Reveal Major Post-Glacial and Anthropogenic Effects on Neutral Genetic Diversity.

Understanding how species respond to population declines is a central question in conservation and evolutionary biology. Population declines are often associated with loss of genetic diversity, inbreeding and accumulation of deleterious mutations, which can lead to a reduction in fitness and subsequently contribute to extinction. Using temporal approaches can help us understand the effects of population declines on genetic diversity in real time. Sequencing pre-decline as well as post-decline mitogenomes representing all the remaining mitochondrial diversity, we estimated the loss of genetic diversity in the critically endangered kakapo (Strigops habroptilus). We detected a signal of population expansion coinciding with the end of the Pleistocene last glacial maximum (LGM). Also, we found some evidence for northern and southern lineages, supporting the hypothesis that the species may have been restricted to isolated northern and southern refugia during the LGM. We observed an important loss of neutral genetic diversity associated with European settlement in New Zealand but we could not exclude a population decline associated with Polynesian settlement in New Zealand. However, we did not find evidence for fixation of deleterious mutations. We argue that despite high pre-decline genetic diversity, a rapid and range-wide decline combined with the lek mating system, and life-history traits of kakapo contributed to a rapid loss of genetic diversity following severe population declines.

Genome-Based Sexing Provides Clues about Behavior and Social Structure in the Woolly Mammoth.

While present-day taxa are valuable proxies for understanding the biology of extinct species, it is also crucial to examine physical remains in order to obtain a more comprehensive view of their behavior, social structure, and life histories [1, 2]. For example, information on demographic parameters such as age distribution and sex ratios in fossil assemblages can be used to accurately infer socioecological patterns (e.g., [3]). Here we use genomic data to determine the sex of 98 woolly mammoth (Mammuthus primigenius) specimens in order to infer social and behavioral patterns in the last 60,000 years of the species' existence. We report a significant excess of males among the identified samples (69% versus 31%; p < 0.0002). We argue that this male bias among mammoth remains is best explained by males more often being caught in natural traps that favor preservation. We hypothesize that this is a consequence of social structure in proboscideans, which is characterized by matriarchal hierarchy and sex segregation. Without the experience associated with living in a matriarchal family group, or a bachelor group with an experienced bull, young or solitary males may have been more prone to die in natural traps where good preservation is more likely.