Grey wolf genomic history reveals a dual ancestry of dogs.

The grey wolf (Canis lupus) was the first species to give rise to a domestic population, and they remained widespread throughout the last Ice Age when many other large mammal species went extinct. Little is known, however, about the history and possible extinction of past wolf populations or when and where the wolf progenitors of the present-day dog lineage (Canis familiaris) lived1-8. Here we analysed 72 ancient wolf genomes spanning the last 100,000 years from Europe, Siberia and North America. We found that wolf populations were highly connected throughout the Late Pleistocene, with levels of differentiation an order of magnitude lower than they are today. This population connectivity allowed us to detect natural selection across the time series, including rapid fixation of mutations in the gene IFT88 40,000-30,000 years ago. We show that dogs are overall more closely related to ancient wolves from eastern Eurasia than to those from western Eurasia, suggesting a domestication process in the east. However, we also found that dogs in the Near East and Africa derive up to half of their ancestry from a distinct population related to modern southwest Eurasian wolves, reflecting either an independent domestication process or admixture from local wolves. None of the analysed ancient wolf genomes is a direct match for either of these dog ancestries, meaning that the exact progenitor populations remain to be located.

A Hen in the Wolf Den: A Pathobiont Tale.

Disruption of the gut microbiota is thought to contribute to disease onset in individuals with a genetic predisposition to autoimmunity. In a recent issue of Science, Manfredo Vieira et al. (2018) identify translocation of the gut commensal Enterococcus gallinarum into the liver as a trigger for the autoimmune disease systemic lupus erythematous.

Dire wolves were the last of an ancient New World canid lineage.

Dire wolves are considered to be one of the most common and widespread large carnivores in Pleistocene America1, yet relatively little is known about their evolution or extinction. Here, to reconstruct the evolutionary history of dire wolves, we sequenced five genomes from sub-fossil remains dating from 13,000 to more than 50,000 years ago. Our results indicate that although they were similar morphologically to the extant grey wolf, dire wolves were a highly divergent lineage that split from living canids around 5.7 million years ago. In contrast to numerous examples of hybridization across Canidae2,3, there is no evidence for gene flow between dire wolves and either North American grey wolves or coyotes. This suggests that dire wolves evolved in isolation from the Pleistocene ancestors of these species. Our results also support an early New World origin of dire wolves, while the ancestors of grey wolves, coyotes and dholes evolved in Eurasia and colonized North America only relatively recently.

Ancient genomes reveal over two thousand years of dingo population structure.

Dingoes are culturally and ecologically important free-living canids whose ancestors arrived in Australia over 3,000 B.P., likely transported by seafaring people. However, the early history of dingoes in Australia-including the number of founding populations and their routes of introduction-remains uncertain. This uncertainty arises partly from the complex and poorly understood relationship between modern dingoes and New Guinea singing dogs, and suspicions that post-Colonial hybridization has introduced recent domestic dog ancestry into the genomes of many wild dingo populations. In this study, we analyzed genome-wide data from nine ancient dingo specimens ranging in age from 400 to 2,746 y old, predating the introduction of domestic dogs to Australia by European colonists. We uncovered evidence that the continent-wide population structure observed in modern dingo populations had already emerged several thousand years ago. We also detected excess allele sharing between New Guinea singing dogs and ancient dingoes from coastal New South Wales (NSW) compared to ancient dingoes from southern Australia, irrespective of any post-Colonial hybrid ancestry in the genomes of modern individuals. Our results are consistent with several demographic scenarios, including a scenario where the ancestry of dingoes from the east coast of Australia results from at least two waves of migration from source populations with varying affinities to New Guinea singing dogs. We also contribute to the growing body of evidence that modern dingoes derive little genomic ancestry from post-Colonial hybridization with other domestic dog lineages, instead descending primarily from ancient canids introduced to Sahul thousands of years ago.

Recovery of a marine keystone predator transforms terrestrial predator-prey dynamics.

Sea otters (Enhydra lutris) and wolves (Canis lupus) are two apex predators with strong and cascading effects on ecosystem structure and function. After decades of recovery from near extirpation, their ranges now overlap, allowing sea otters and wolves to interact for the first time in the scientific record. We intensively studied wolves during 2015 to 2021 in an island system colonized by sea otters in the 2000s and by wolves in 2013. After wolf colonization, we quantified shifts in foraging behavior with DNA metabarcoding of 689 wolf scats and stable isotope analyses, both revealing a dietary switch from Sitka black-tailed deer (Odocoileus hemionus), the terrestrial in situ primary prey, to sea otters. Here we show an unexpected result of the reintroduction and restoration of sea otters, which became an abundant marine subsidy for wolves following population recovery. The availability of sea otters allowed wolves to persist and continue to reproduce, subsequently nearly eliminating deer. Genotypes from 390 wolf scats and telemetry data from 13 wolves confirmed island fidelity constituting one of the highest known wolf population densities and upending standardly accepted wolf density predictions based on ungulate abundance. Whereas marine subsidies in other systems are generally derived from lower trophic levels, here an apex nearshore predator became a key prey species and linked nearshore and terrestrial food webs in a recently deglaciated and rapidly changing ecosystem. These results underscore that species restoration may serve as an unanticipated nutrient pathway for recipient ecosystems even resulting in cross-boundary subsidy cascades.

Whole-genome resequencing of temporally stratified samples reveals substantial loss of haplotype diversity in the highly inbred Scandinavian wolf population.

Genetic drift can dramatically change allele frequencies in small populations and lead to reduced levels of genetic diversity, including loss of segregating variants. However, there is a shortage of quantitative studies of how genetic diversity changes over time in natural populations, especially on genome-wide scales. Here, we analyzed whole-genome sequences from 76 wolves of a highly inbred Scandinavian population, founded by only one female and two males, sampled over a period of 30 yr. We obtained chromosome-level haplotypes of all three founders and found that 10%-24% of their diploid genomes had become lost after about 20 yr of inbreeding (which approximately corresponds to five generations). Lost haplotypes spanned large genomic regions, as expected from the amount of recombination during this limited time period. Altogether, 160,000 SNP alleles became lost from the population, which may include adaptive variants as well as wild-type alleles masking recessively deleterious alleles. Although not sampled, we could indirectly infer that the two male founders had megabase-sized runs of homozygosity and that all three founders showed significant haplotype sharing, meaning that there were on average only 4.2 unique haplotypes in the six copies of each autosome that the founders brought into the population. This violates the assumption of unrelated founder haplotypes often made in conservation and management of endangered species. Our study provides a novel view of how whole-genome resequencing of temporally stratified samples can be used to visualize and directly quantify the consequences of genetic drift in a small inbred population.

Wolf attacks predict far-right voting.

Does the return of large carnivores affect voting behavior? We study this question through the lens of wolf attacks on livestock. Sustained environmental conservation has allowed the wolf (Canis lupus) to make an impressive and unforeseen comeback across Central Europe in recent years. While lauded by conservationists, local residents often see the wolf as a threat to economic livelihoods, particularly those of farmers. As populists appear to exploit such sentiments, the wolf's reemergence is a plausible source for far-right voting behavior. To test this hypothesis, we collect fine-grained spatial data on wolf attacks and construct a municipality-level panel in Germany. Using difference-in-differences models, we find that wolf attacks are accompanied by a significant rise in far-right voting behavior, while the Green party, if anything, suffers electoral losses. We buttress this finding using local-level survey data, which confirms a link between wolf attacks and negative sentiment toward environmental protection. To explore potential mechanisms, we analyze Twitter posts, election manifestos, and Facebook ads to show that far-right politicians frame the wolf as a threat to economic livelihoods.

Focused ion beam-SEM 3D study of osteodentin in the teeth of the Atlantic wolfish Anarhichas lupus.

The palette of mineralized tissues in fish is wide, and this is particularly apparent in fish dentin. While the teeth of all vertebrates except fish contain a single dentinal tissue type, called orthodentin, dentin in the teeth of fish can be one of several different tissue types. The most common dentin type in fish is orthodentin. Orthodentin is characterized by several key structural features that are fundamentally different from those of bone and from those of osteodentin. Osteodentin, the second-most common dentin type in fish (based on the tiny fraction of fish species out of ∼30,000 extant fish species in which tooth structure was so far studied), is found in most Selachians (sharks and rays) as well as in several teleost species, and is structurally different from orthodentin. Here we examine the hypothesis that osteodentin is similar to anosteocytic bone tissue in terms of its micro- and nano-structure. We use Focused Ion Beam-Scanning Electron Microscopy (FIB/SEM), as well as several other high-resolution imaging techniques, to characterize the 3D architecture of the three main components of osteodentin (denteons, inter-denteonal matrix, and the transition zone between them). We show that the matrix of osteodentin, although acellular, is extremely similar to mammalian osteonal bone matrix, both in general morphology and in the three-dimensional nano-arrangement of its mineralized collagen fibrils. We also document the presence of a complex network of nano-channels, similar to such networks recently described in bone. Finally, we document the presence of strings of hyper-mineralized small 'pearls' which surround the denteonal canals, and characterize their structure.

Aspen recovery in northern Yellowstone: A comment on Brice et al. (2021).

Aspen sapling recruitment increased as browsing by elk decreased, following the 1995-96 reintroduction of wolves in Yellowstone National Park. We address claims by Brice et al. (2021) that previous studies exaggerated recent aspen recovery. We conclude that their results actually supported previous work showing a trophic cascade benefiting aspen.

Non-random sampling measures the occurrence but not the strength of a textbook trophic cascade.

Although sampling the five tallest young aspen in a stand is useful for detecting the occurrence of any aspen recruitment, this technique overestimates the population response of aspen to wolf reintroduction. Our original conclusion that random sampling described a trophic cascade that was weaker than the one described by non-random sampling is unchanged.

Long-term Small Population Size, Deleterious Variation, and Altitude Adaptation in the Ethiopian Wolf, a Severely Endangered Canid.

Ethiopian wolves, a canid species endemic to the Ethiopian Highlands, have been steadily declining in numbers for decades. Currently, out of 35 extant species, it is now one of the world's most endangered canids. Most conservation efforts have focused on preventing disease, monitoring movements and behavior, and assessing the geographic ranges of sub-populations. Here, we add an essential layer by determining the Ethiopian wolf's demographic and evolutionary history using high-coverage (∼40×) whole-genome sequencing from 10 Ethiopian wolves from the Bale Mountains. We observe exceptionally low diversity and enrichment of weakly deleterious variants in the Ethiopian wolves in comparison with two North American gray wolf populations and four dog breeds. These patterns are consequences of long-term small population size, rather than recent inbreeding. We infer the demographic history of the Ethiopian wolf and find it to be concordant with historic records and previous genetic analyses, suggesting Ethiopian wolves experienced a series of both ancient and recent bottlenecks, resulting in a census population size of fewer than 500 individuals and an estimated effective population size of approximately 100 individuals. Additionally, long-term small population size may have limited the accumulation of strongly deleterious recessive mutations. Finally, as the Ethiopian wolves have inhabited high-altitude areas for thousands of years, we searched for evidence of high-altitude adaptation, finding evidence of positive selection at a transcription factor in a hypoxia-response pathway [CREB-binding protein (CREBBP)]. Our findings are pertinent to continuing conservation efforts and understanding how demography influences the persistence of deleterious variation in small populations.

Multiple Origins and Genomic Basis of Complex Traits in Sighthounds.

Sighthounds, a distinctive group of hounds comprising numerous breeds, have their origins rooted in ancient artificial selection of dogs. In this study, we performed genome sequencing for 123 sighthounds, including one breed from Africa, six breeds from Europe, two breeds from Russia, and four breeds and 12 village dogs from the Middle East. We gathered public genome data of five sighthounds and 98 other dogs as well as 31 gray wolves to pinpoint the origin and genes influencing the morphology of the sighthound genome. Population genomic analysis suggested that sighthounds originated from native dogs independently and were comprehensively admixed among breeds, supporting the multiple origins hypothesis of sighthounds. An additional 67 published ancient wolf genomes were added for gene flow detection. Results showed dramatic admixture of ancient wolves in African sighthounds, even more than with modern wolves. Whole-genome scan analysis identified 17 positively selected genes (PSGs) in the African population, 27 PSGs in the European population, and 54 PSGs in the Middle Eastern population. None of the PSGs overlapped in the three populations. Pooled PSGs of the three populations were significantly enriched in "regulation of release of sequestered calcium ion into cytosol" (gene ontology: 0051279), which is related to blood circulation and heart contraction. In addition, ESR1, JAK2, ADRB1, PRKCE, and CAMK2D were under positive selection in all three selected groups. This suggests that different PSGs in the same pathway contributed to the similar phenotype of sighthounds. We identified an ESR1 mutation (chr1: g.42,177,149 T > C) in the transcription factor (TF) binding site of Stat5a and a JAK2 mutation (chr1: g.93,277,007 T > A) in the TF binding site of Sox5. Functional experiments confirmed that the ESR1 and JAK2 mutation reduced their expression. Our results provide new insights into the domestication history and genomic basis of sighthounds.

Genomic Underpinnings of Population Persistence in Isle Royale Moose.

Island ecosystems provide natural laboratories to assess the impacts of isolation on population persistence. However, most studies of persistence have focused on a single species, without comparisons to other organisms they interact with in the ecosystem. The case study of moose and gray wolves on Isle Royale allows for a direct contrast of genetic variation in isolated populations that have experienced dramatically differing population trajectories over the past decade. Whereas the Isle Royale wolf population recently declined nearly to extinction due to severe inbreeding depression, the moose population has thrived and continues to persist, despite having low genetic diversity and being isolated for ∼120 years. Here, we examine the patterns of genomic variation underlying the continued persistence of the Isle Royale moose population. We document high levels of inbreeding in the population, roughly as high as the wolf population at the time of its decline. However, inbreeding in the moose population manifests in the form of intermediate-length runs of homozygosity suggestive of historical inbreeding and purging, contrasting with the long runs of homozygosity observed in the smaller wolf population. Using simulations, we confirm that substantial purging has likely occurred in the moose population. However, we also document notable increases in genetic load, which could eventually threaten population viability over the long term. Overall, our results demonstrate a complex relationship between inbreeding, genetic diversity, and population viability that highlights the use of genomic datasets and computational simulation tools for understanding the factors enabling persistence in isolated populations.

Tracing Eastern Wolf Origins From Whole-Genome Data in Context of Extensive Hybridization.

Southeastern Canada is inhabited by an amalgam of hybridizing wolf-like canids, raising fundamental questions regarding their taxonomy, origins, and timing of hybridization events. Eastern wolves (Canis lycaon), specifically, have been the subject of significant controversy, being viewed as either a distinct taxonomic entity of conservation concern or a recent hybrid of coyotes (C. latrans) and grey wolves (C. lupus). Mitochondrial DNA analyses show some evidence of eastern wolves being North American evolved canids. In contrast, nuclear genome studies indicate eastern wolves are best described as a hybrid entity, but with unclear timing of hybridization events. To test hypotheses related to these competing findings we sequenced whole genomes of 25 individuals, representative of extant Canadian wolf-like canid types of known origin and levels of contemporary hybridization. Here we present data describing eastern wolves as a distinct taxonomic entity that evolved separately from grey wolves for the past ∼67,000 years with an admixture event with coyotes ∼37,000 years ago. We show that Great Lakes wolves originated as a product of admixture between grey wolves and eastern wolves after the last glaciation (∼8,000 years ago) while eastern coyotes originated as a product of admixture between "western" coyotes and eastern wolves during the last century. Eastern wolf nuclear genomes appear shaped by historical and contemporary gene flow with grey wolves and coyotes, yet evolutionary uniqueness remains among eastern wolves currently inhabiting a restricted range in southeastern Canada.

A map of canine sequence variation relative to a Greenland wolf outgroup.

For over 15 years, canine genetics research relied on a reference assembly from a Boxer breed dog named Tasha (i.e., canFam3.1). Recent advances in long-read sequencing and genome assembly have led to the development of numerous high-quality assemblies from diverse canines. These assemblies represent notable improvements in completeness, contiguity, and the representation of gene promoters and gene models. Although genome graph and pan-genome approaches have promise, most genetic analyses in canines rely upon the mapping of Illumina sequencing reads to a single reference. The Dog10K consortium, and others, have generated deep catalogs of genetic variation through an alignment of Illumina sequencing reads to a reference genome obtained from a German Shepherd Dog named Mischka (i.e., canFam4, UU_Cfam_GSD_1.0). However, alignment to a breed-derived genome may introduce bias in genotype calling across samples. Since the use of an outgroup reference genome may remove this effect, we have reprocessed 1929 samples analyzed by the Dog10K consortium using a Greenland wolf (mCanLor1.2) as the reference. We efficiently performed remapping and variant calling using a GPU-implementation of common analysis tools. The resulting call set removes the variability in genetic differences seen across samples and breed relationships revealed by principal component analysis are not affected by the choice of reference genome. Using this sequence data, we inferred the history of population sizes and found that village dog populations experienced a 9-13 fold reduction in historic effective population size relative to wolves.

Fear of the human 'super predator' in native marsupials and introduced deer in Australia.

Recent experiments have demonstrated that carnivores and ungulates in Africa, Asia, Europe and North America fear the human 'super predator' far more than other predators. Australian mammals have been a focus of research on predator naiveté because it is suspected they show atypical antipredator responses. To experimentally test if mammals in Australia also most fear humans, we quantified the responses of four native marsupials (eastern grey kangaroo, Bennett's wallaby, Tasmanian pademelon, common brushtail possum) and introduced fallow deer to playbacks of predator (human, dog, Tasmanian devil, wolf) or non-predator control (sheep) vocalizations. Native marsupials most feared the human 'super predator', fleeing humans 2.4 times more often than the next most frightening predator (dogs), and being most, and significantly, vigilant to humans. These results demonstrate that native marsupials are not naïve to the peril humans pose, substantially expanding the taxonomic and geographic scope of the growing experimental evidence that wildlife worldwide generally perceive humans as the planet's most frightening predator. Introduced fallow deer fled humans, but not more than other predators, which we suggest may result from their being introduced. Our results point to both challenges concerning marsupial conservation and opportunities for exploiting fear of humans as a wildlife management tool.

Demographic history shapes North American gray wolf genomic diversity and informs species' conservation.

Effective population size estimates are critical information needed for evolutionary predictions and conservation decisions. This is particularly true for species with social factors that restrict access to breeding or experience repeated fluctuations in population size across generations. We investigated the genomic estimates of effective population size along with diversity, subdivision, and inbreeding from 162,109 minimally filtered and 81,595 statistically neutral and unlinked SNPs genotyped in 437 grey wolf samples from North America collected between 1986 and 2021. We found genetic structure across North America, represented by three distinct demographic histories of western, central, and eastern regions of the continent. Further, grey wolves in the northern Rocky Mountains have lower genomic diversity than wolves of the western Great Lakes and have declined over time. Effective population size estimates revealed the historical signatures of continental efforts of predator extermination, despite a quarter century of recovery efforts. We are the first to provide molecular estimates of effective population size across distinct grey wolf populations in North America, which ranged between Ne ~ 275 and 3050 since early 1980s. We provide data that inform managers regarding the status and importance of effective population size estimates for grey wolf conservation, which are on average 5.2-9.3% of census estimates for this species. We show that while grey wolves fall above minimum effective population sizes needed to avoid extinction due to inbreeding depression in the short term, they are below sizes predicted to be necessary to avoid long-term risk of extinction.

The vomeronasal system of the wolf (Canis lupus signatus): The singularities of a wild canid.

Wolves, akin to their fellow canids, extensively employ chemical signals for various aspects of communication, including territory maintenance, reproductive synchronisation and social hierarchy signalling. Pheromone-mediated chemical communication operates unconsciously among individuals, serving as an innate sensory modality that regulates both their physiology and behaviour. Despite its crucial role in the life of the wolf, there is a lacuna in comprehensive research on the neuroanatomical and physiological underpinnings of chemical communication within this species. This study investigates the vomeronasal system (VNS) of the Iberian wolf, simultaneously probing potential alterations brought about by dog domestication. Our findings demonstrate the presence of a fully functional VNS, vital for pheromone-mediated communication, in the Iberian wolf. While macroscopic similarities between the VNS of the wolf and the domestic dog are discernible, notable microscopic differences emerge. These distinctions include the presence of neuronal clusters associated with the sensory epithelium of the vomeronasal organ (VNO) and a heightened degree of differentiation of the accessory olfactory bulb (AOB). Immunohistochemical analyses reveal the expression of the two primary families of vomeronasal receptors (V1R and V2R) within the VNO. However, only the V1R family is expressed in the AOB. These findings not only yield profound insights into the VNS of the wolf but also hint at how domestication might have altered neural configurations that underpin species-specific behaviours. This understanding holds implications for the development of innovative strategies, such as the application of semiochemicals for wolf population management, aligning with contemporary conservation goals.

Relatedness-based mate choice and female philopatry: inbreeding trends of wolf packs in a human-dominated landscape.

Inbreeding can reduce offspring fitness and has substantial implications for the genetic diversity and long-term viability of populations. In social cooperative canids, inbreeding is conditioned by the geographic proximity between opposite-sex kin outside natal groups and the presence of related individuals in neighbouring groups. Consequently, challenges in moving into other regions where the species is present can also affect inbreeding rates. These can be particularly problematic in areas of high human density, where movement can be restricted, even for highly vagile species. In this study, we investigate the socio-ecological dynamics of Iberian wolf packs in the human-dominated landscape of Alto Minho, in northwest Portugal, where wolves exhibit a high prevalence of short-distance dispersal and limited gene flow with neighbouring regions. We hypothesise that mating occurs regardless of relatedness, resulting in recurrent inbreeding due to high kin encounter rates. Using data from a 10-year non-invasive genetic monitoring programme and a combination of relatedness estimates and genealogical reconstructions, we describe genetic diversity, mate choice, and dispersal strategies among Alto Minho packs. In contrast with expectations, our findings reveal relatedness-based mate choice, low kin encounter rates, and a reduced number of inbreeding events. We observed a high prevalence of philopatry, particularly among female breeders, with the most common breeding strategy involving the pairing of a philopatric female with an unrelated immigrant male. Overall, wolves were not inbred, and temporal changes in genetic diversity were not significant. Our findings are discussed, considering the demographic trend of wolves in Alto Minho and its human-dominated landscape.

A first exploratory comparison of the behaviour of wolves (Canis lupus) and wolf-dog hybrids in captivity.

Extensive introgression of genes from domesticated taxa may be a serious threat for the genomic integrity and adaptability of wild populations. Grey wolves (Canis lupus) are especially vulnerable to this phenomenon, but there are no studies yet assessing the potential behavioural effects of dog-introgression in wolves. In this study, we conducted a first systematic comparison of admixed (N = 11) and non-admixed (N = 14) wolves in captivity, focusing on their reaction to unfamiliar humans and novel objects, and the cohesiveness of their social groups. When exposed to unfamiliar humans in the experimental task, wolves were more vigilant, fearful and aggressive than admixed wolves, and less likely to approach humans, but also more likely to spend time in human proximity. When exposed to novel objects, wolves were more aggressive than admixed wolves, less likely to spend time in object proximity, and more likely to interact with objects, but also less vigilant and as fearful as admixed wolves. Finally, social networks were more cohesive in wolves than in admixed wolves. Although caution is needed when comparing groups of captive individuals with different life experiences, our study suggests that dog admixture may lead to important behavioural changes in wolves, with possible implications for conservation strategies.