Canine hyper-sociability structural variants associated with altered three-dimensional chromatin state.

Strong selection on complex traits can lead to skewed trait means and reduced trait variability in populations. An example of this phenomenon can be evidenced in allele frequency changes and skewed trait distributions driven by persistent human-directed selective pressures in domesticated species. Dog domestication is linked to several genomic variants; however, the functional impacts of these variants may not always be straightforward when found in non-coding regions of the genome. Four polymorphic transposable elements (TE) found within non-coding sites along a 5 Mb region on canine CFA6 have evolved due to directional selection associated with heightened human-directed hyper-sociability in domesticated dogs. We found that the polymorphic TE in intron 17 of the canine GTF2I gene, which was previously reported to be negatively correlated with canid human-directed hyper-sociability, is associated with altered chromatin looping and hence distinct cis-regulatory landscapes. We reported supporting evidence of an E2F1-DNA binding peak concordant with the altered loop and higher expression of GTF2I exon 18, indicative of alternative splicing. Globally, we discovered differences in pathways regulating the extra-cellular matrix with respect to TE copy number. Overall, we reported evidence suggesting an intriguing molecular convergence between the emergence of hypersocial behaviors in dogs and the same genes that, when hemizygous, produce human Williams Beuren Syndrome characterized by cranio-facial defects and heightened social behaviors. Our results additionally emphasize the often-overlooked potential role of chromatin architecture in social evolution.

Trapped between food, heat, and insects: Movement of moose (Alces alces) and exposure to flies in the boreal forest of Alaska.

Moose (Alces alces) in the boreal forest habitats of Alaska are unlike other northern ungulates because they tolerate high densities of flies (Diptera) even though flies cause wounds and infections during the warm summer months. Moose move to find food and to find relief from overheating (hyperthermia) but do they avoid flies? We used GPS collars to measure the rate of movement (m⋅h-1) and the time spent (min⋅day-1) by enclosed moose in four habitats: wetlands, black spruce, early seral boreal forest, and late seral boreal forest. Fly traps were used in each habitat to quantify spatio-temporal abundance. Average daily air temperatures increased into July when peak biomass of forage for moose was greatest in early seral boreal forest habitats (424.46 vs. 25.15 kg⋅ha-1 on average in the other habitats). Average daily air temperatures were 1.7°C cooler in black spruce than other habitats, but fly abundance was greatest in black spruce (approximately 4-fold greater on average than the other habitats). Moose increased their movement rate with counts of biting flies (mosquitoes, black flies, horse and deer flies), but not non-biting flies (coprophagous flies). However, as air temperature increased (above 14.7°C) moose spent more time in fly-abundant black spruce, than early seral boreal forest, showing great tolerance for mosquitoes. Warm summer temperatures appear to cause moose to trade-off foraging in fly-sparse habitats for resting and dissipating heat in shady, wet habitats with abundant flies that adversely affect the fitness of moose.

A genomic basis of vocal rhythm in birds.

Vocal rhythm plays a fundamental role in sexual selection and species recognition in birds, but little is known of its genetic basis due to the confounding effect of vocal learning in model systems. Uncovering its genetic basis could facilitate identifying genes potentially important in speciation. Here we investigate the genomic underpinnings of rhythm in vocal non-learning Pogoniulus tinkerbirds using 135 individual whole genomes distributed across a southern African hybrid zone. We find rhythm speed is associated with two genes that are also known to affect human speech, Neurexin-1 and Coenzyme Q8A. Models leveraging ancestry reveal these candidate loci also impact rhythmic stability, a trait linked with motor performance which is an indicator of quality. Character displacement in rhythmic stability suggests possible reinforcement against hybridization, supported by evidence of asymmetric assortative mating in the species producing faster, more stable rhythms. Because rhythm is omnipresent in animal communication, candidate genes identified here may shape vocal rhythm across birds and other vertebrates.

The value of hybrid genomes: Building two highly contiguous reference genome assemblies to advance Canis genomic studies.

Previous studies of canid population and evolutionary genetics have relied on high-quality domestic dog reference genomes that have been produced primarily for biomedical and trait mapping studies in dog breeds. However, the absence of highly contiguous genomes from other Canis species like the gray wolf and coyote, that represent additional distinct demographic histories, may bias inferences regarding interspecific genetic diversity and phylogenetic relationships. Here, we present single haplotype de novo genome assemblies for the gray wolf and coyote, generated by applying the trio-binning approach to long sequence reads generated from the genome of a female first-generation hybrid produced from a gray wolf and coyote mating. The assemblies were highly contiguous, with contig N50 sizes of 44.6 and 42.0 Mb for the wolf and coyote, respectively. Genome scaffolding and alignments between the two Canis assemblies and published dog reference genomes showed near complete collinearity, with one exception: a coyote-specific chromosome fission of chromosome 13 and fusion of the proximal portion of that chromosome with chromosome 8, retaining the Canis-typical haploid chromosome number of 2n = 78. We evaluated mapping quality for previous RADseq data from 334 canids and found nearly identical mapping quality and patterns among canid species and regional populations regardless of the genome used for alignment (dog, coyote, or gray wolf). These novel wolf and coyote genome reference assemblies will be important resources for proper and accurate inference of Canis demography, taxonomic evaluation, and conservation genetics.

The gut microbiomes of Channel Island foxes and island spotted skunks exhibit fine-scale differentiation across host species and island populations.

California's Channel Islands are home to two endemic mammalian carnivores: island foxes (Urocyon littoralis) and island spotted skunks (Spilogale gracilis amphiala). Although it is rare for two insular terrestrial carnivores to coexist, these known competitors persist on both Santa Cruz Island and Santa Rosa Island. We hypothesized that examination of their gut microbial communities would provide insight into the factors that enable this coexistence, as microbial symbionts often reflect host evolutionary history and contemporary ecology. Using rectal swabs collected from island foxes and island spotted skunks sampled across both islands, we generated 16S rRNA amplicon sequencing data to characterize their gut microbiomes. While island foxes and island spotted skunks both harbored the core mammalian microbiome, host species explained the largest proportion of variation in the dataset. We further identified intraspecific variation between island populations, with greater differentiation observed between more specialist island spotted skunk populations compared to more generalist island fox populations. This pattern may reflect differences in resource utilization following fine-scale niche differentiation. It may further reflect evolutionary differences regarding the timing of intraspecific separation. Considered together, this study contributes to the growing catalog of wildlife microbiome studies, with important implications for understanding how eco-evolutionary processes enable the coexistence of terrestrial carnivores-and their microbiomes-in island environments.

Epigenetic changes to gene pathways linked to male fertility in ex situ black-footed ferrets.

Environmental variation can influence the reproductive success of species managed under human care and in the wild, yet the mechanisms underlying this phenomenon remain largely mysterious. Molecular mechanisms such as epigenetic modifiers are important in mediating the timing and progression of reproduction in humans and model organisms, but few studies have linked epigenetic variation to reproductive fitness in wildlife. Here, we investigated epigenetic variation in black-footed ferrets (Mustela nigripes), an endangered North American mammal reliant on ex situ management for survival and persistence in the wild. Despite similar levels of genetic diversity in human-managed and wild-born populations, individuals in ex situ facilities exhibit reproductive problems, such as poor sperm quality. Differences across these settings suggest that an environmentally driven decline in reproductive capacity may be occurring in this species. We examined the role of DNA methylation, one well-studied epigenetic modifier, in this emergent condition. We leveraged blood, testes, and semen samples from male black-footed ferrets bred in ex situ facilities and found tissue-type specificity in DNA methylation across the genome, although 1360 Gene Ontology terms associated with male average litter size shared functions across tissues. We then constructed gene networks of differentially methylated genomic sites associated with three different reproductive phenotypes to explore the putative biological impact of variation in DNA methylation. Sperm gene networks associated with average litter size and sperm count were functionally enriched for candidate genes involved in reproduction, development, and its regulation through transcriptional repression. We propose that DNA methylation plays an important role in regulating these reproductive phenotypes, thereby impacting the fertility of male ex situ individuals. Our results provide information into how DNA methylation may function in the alteration of reproductive pathways and phenotypes in artificial environments. These findings provide early insights to conservation hurdles faced in the protection of this rare species.

Transposons in the Williams-Beuren Syndrome Critical Region are Associated with Social Behavior in Assistance Dogs.

A strong signature of selection in the domestic dog genome is found in a five-megabase region of chromosome six in which four structural variants derived from transposons have previously been associated with human-oriented social behavior, such as attentional bias to social stimuli and social interest in strangers. To explore these genetic associations in more phenotypic detail-as well as their role in training success in a specialized assistance dog program-we genotyped 1001 assistance dogs from Canine Companions for Independence®, including both successful graduates and dogs released from the training program for behaviors incompatible with their working role. We collected phenotypes on each dog using puppy-raiser questionnaires, trainer questionnaires, and both cognitive and behavioral tests. Using Bayesian mixed models, we found strong associations (95% credibility intervals excluding zero) between genotypes and certain behavioral measures, including separation-related problems, aggression when challenged or corrected, and reactivity to other dogs. Furthermore, we found moderate differences in the genotypes of dogs who graduated versus those who did not; insertions in GTF2I showed the strongest association with training success (ß = 0.23, CI95% = - 0.04, 0.49), translating to an odds-ratio of 1.25 for one insertion. Our results provide insight into the role of each of these four transposons in canine sociability and may inform breeding and training practices for working dog organizations. Furthermore, the observed importance of the gene GTF2I supports the emerging consensus that variation in GTF2I genotypes and expression have important consequences for social behavior broadly.

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.

Patterns of reproduction and autozygosity distinguish the breeding from non-breeding gray wolves of Yellowstone National Park.

For species of management concern, accurate estimates of inbreeding and associated consequences on reproduction are crucial for predicting their future viability. However, few studies have partitioned this aspect of genetic viability with respect to reproduction in a group-living social mammal. We investigated the contributions of foundation stock lineages, putative fitness consequences of inbreeding, and genetic diversity of the breeding versus non-reproductive segment of the Yellowstone National Park gray wolf population. Our dataset spans 25 years and seven generations since reintroduction, encompassing 152 nuclear families and 329 litters. We found over 87% of the pedigree foundation genomes persisted and report influxes of allelic diversity from two translocated wolves from a divergent source in Montana. As expected for group-living species, mean kinship significantly increased over time but with minimal loss of observed heterozygosity. Strikingly, the reproductive portion of the population carried a significantly lower genome-wide inbreeding coefficients, autozygosity, and more rapid decay for linkage disequilibrium relative to the non-breeding population. Breeding wolves had significantly longer lifespans and lower inbreeding coefficients than non-breeding wolves. Our model revealed that the number of litters was negatively significantly associated with heterozygosity (R=-0.11). Our findings highlight genetic contributions to fitness, and the importance of the reproductively active individuals in a population to counteract loss of genetic variation in a wild, free-ranging social carnivore. It is crucial for managers to mitigate factors that significantly reduce effective population size and genetic connectivity, which supports the dispersion of genetic variation that aids in rapid evolutionary responses to environmental challenges.

Response to Hansen Wheat et al.: Additional analysis further supports the early emergence of cooperative communication in dogs compared to wolves raised with more human exposure.

Here, we address Hansen Wheat et al.'s commentary in this journal in response to Salomons et al. Current Biology, 31(14), 3137-3144.E11, (2021). We conduct additional analyses in response to Hansen Wheat et al.'s two main questions. First, we examine the claim that it was the move to a human home environment which enabled the dog puppies to outperform the wolf puppies in gesture comprehension tasks. We show that the youngest dog puppies who had not yet been individually placed in raisers' homes were still highly skilled, and outperformed similar-aged wolf puppies who had higher levels of human interaction. Second, we address the claim that willingness to approach a stranger can explain the difference between dog and wolf pups' ability to succeed in gesture comprehension tasks. We explain the various controls in the original study that render this explanation insufficient, and demonstrate via model comparison that the covariance of species and temperament also make this parsing impossible. Overall, our additional analyses and considerations support the domestication hypothesis as laid out by Salomons et al. Current Biology, 31(14), 3137-3144.E11, (2021).

Sores of boreal moose reveal a previously unknown genetic lineage of parasitic nematode within the genus Onchocerca.

Long-standing reports of open sores on the hind legs of moose (Alces alces) have been recorded in Alaska (as well as Canada, Europe, and Michigan), eliciting concerns about causes and infection. We used histological and genomic methods to investigate the sores from 20 adult moose on the Kenai Peninsula, Alaska. We paired this with thermal imagery and molt scoring of adult moose to further describe sore formation and understand its timing. Severe, ulcerative and eosinophilic dermatitis was found in all moose with sores present, and microfilariae within intraepidermal pustules were additionally found in four samples. Genetic analysis of sores from moose revealed a previously unknown genetic lineage of Onchocerca. Adult moose molt and lose their barrier of protection against flies in June and July during peak fly activity, leaving them vulnerable and allowing the development of sores. In summary, our results indicate that the cause for the sores on the hindleg of moose is a previously unknown genetic lineage of Onchocerca, probably transmitted by black flies, in timing with the molt cycle of adult moose. These sores leave moose exposed to pathogens, making them vulnerable, and challenging their health and fitness.

An IS-mediated, RecA-dependent, bet-hedging strategy in Burkholderia thailandensis.

Adaptation to fluctuating environmental conditions is difficult to achieve. Phase variation mechanisms can overcome this difficulty by altering genomic architecture in a subset of individuals, creating a phenotypically heterogeneous population with subpopulations optimized to persist when conditions change, or are encountered, suddenly. We have identified a phase variation system in Burkholderia thailandensis that generates a genotypically and phenotypically heterogeneous population. Genetic analyses revealed that RecA-mediated homologous recombination between a pair of insertion sequence (IS) 2-like elements duplicates a 208.6 kb region of DNA that contains 157 coding sequences. RecA-mediated homologous recombination also resolves merodiploids, and hence copy number of the region is varied and dynamic within populations. We showed that the presence of two or more copies of the region is advantageous for growth in a biofilm, and a single copy is advantageous during planktonic growth. While IS elements are well known to contribute to evolution through gene inactivation, polar effects on downstream genes, and altering genomic architecture, we believe that this system represents a rare example of IS element-mediated evolution in which the IS elements provide homologous sequences for amplification of a chromosomal region that provides a selective advantage under specific growth conditions, thereby expanding the lifestyle repertoire of the species.

Bacterial populations are often diverse, even when originating from a single cell. This diversity helps microbes survive in fluctuating environmental conditions by increasing the odds of population survival. For example, if environmental conditions change such that only a subpopulation with unique abilities survives, the entire population will be saved. Genomes are naturally dynamic. For example, mobile sections of DNA, called transposable elements, can change their position within a genome. If a transposable element jumps into a gene, it can harm the cell. But if it moves into a different site, it may provide an organism with new features that can help it survive. Most organisms contain multiple copies of transposable elements in their DNA. For example, a subtype of the soil bacterium Burkholderia thailandensis, strain E264, has two identical transposable elements that book-end a region of DNA that contains 157 genes. Lowrey et al. studied this bacterial strain in different environmental conditions to find out more. The experiments revealed that in growing populations of E264, some bacteria had one copy of the region, while others had two or three. In a rich environment, most bacteria had just one copy of the region. However, when grown in challenging conditions, most bacteria contained two or three copies of the region. Moreover, bacteria required at least two copies to form dense communities known as biofilms, which are advantageous for bacterial survival in challenging conditions. Bacteria with only one copy, however, were better adapted to a free-swimming lifestyle. Lowrey et al. further showed that the DNA repair system was required for duplicating the region. Usually, this system finds and recombines identical DNA sequences to repair broken DNA. However, if two identical DNA sequences (a pair of transposable elements) are present, the repair system can recombine them during DNA replication, resulting in the duplication of the DNA between the identical sequences. The same system also reduces the copy number of the region from three or two to just one. Since the repair system is constantly working and DNA recombination is always occurring at a low level, B. thailandensis E264 maintains a genetically diverse population with bacteria containing different copy numbers of the region. This diversity ensures that the strain survives in fluctuating environmental conditions. Transposable elements are hotspots of evolution. They are known to interrupt genes and shrink genomes. Lowrey et al. showed that transposable elements also influence evolution by providing DNA sequences that the DNA repair system can use to duplicate DNA. This process of duplicating genes is more frequent than random genetic mutations, expediting adaptation.

Ear mite infection restructures otic microbial networks in conservation-reliant Santa Catalina Island foxes (Urocyon littoralis catalinae).

Ceruminous gland tumours are highly prevalent in the ear canals of Santa Catalina Island foxes (Urocyon littoralis catalinae). Previous work suggests that tumours may result from a combination of ectoparasites, disruption of the host-associated microbiome, and host immunopathology. More specifically, ear mite infection has been associated with broad-scale microbial dysbiosis marked by secondary bacterial infection with the opportunistic pathogen Staphylococcus pseudintermedius. Together, ear mites and S. pseudintermedius probably sustain chronic inflammation and promote conditions suitable for tumour development. In the present study, we expanded upon this framework by constructing otic microbial community networks for mite-infected and uninfected foxes sampled in 2017-2019. Across sampling years, we observed consistent signatures of microbial dysbiosis in mite-infected ear canals, including reduced microbial diversity and shifted abundance towards S. pseudintermedius. Network analysis further revealed that mite infection disrupts overall community structure. In mite-infected networks, interaction strengths between taxa were generally weaker, and numerous subnetworks disappeared altogether. We also found that two strains of S. pseudintermedius connected to the main network, suggesting that multistrain biofilm formation may be occurring. In contrast, S. pseudintermedius is peripheral in the uninfected network, with its only connections including a second strain of S. pseudintermedius and the possible competitor Acinetobacter rhizosphaerae. Finally, the lineup of potential keystone taxa shifted across disease states. Fusobacteria spp., a carcinogenesis-promoting microbe, assumed a keystone role in the mite-infected community. Considered together, these findings provide insights into how mite infection may destabilize the microbiome and ultimately contribute to tumour development in this island endemic species.

Changes in the rearing environment cause reorganization of molecular networks associated with DNA methylation.

Disentangling the interaction between the genetic basis and environmental context underlying phenotypic variation is critical for understanding organismal evolution. Environmental change, such as increased rates of urbanization, can induce shifts in phenotypic plasticity with some individuals adapting to city life while others are displaced. A key trait that can facilitate adaptation is the degree at which animals respond to stressors. This stress response, which includes elevation of baseline circulating concentrations of glucocorticoids, has a heritable component and exhibits intra- and inter-individual variation. However, the mechanisms behind this variability and whether they might be responsible for adaptation to different environments are not known. Variation in DNA methylation can be a potential mechanism that mediates environmental effects on the stress response, as early-life stressors increase glucocorticoid concentrations and change adult phenotype. We used an inter- and intra-environmental cross-foster experiment to analyse the contribution of DNA methylation to early-life phenotypic variation. We found that at hatching, urban house wren (Troglodytes aedon) offspring had higher methylation frequencies compared with their rural counterparts. We also observed age-related patterns in offspring methylation, indicating the developmental effects of the rearing environment on methylation. At fledgling, differential methylation analyses showed that cellular respiration genes were differentially methylated in broods of different origins and behavioural and metabolism genes were differentially methylated in broods of different rearing environments. Lastly, hyper-methylation of a single gene (CNTNAP2) is associated with decreased glucocorticoid levels and the rearing environment. These differential methylation patterns linked to a specific physiological phenotype suggest that DNA methylation may be a mechanism by which individuals adjust to novel environments during their lifespan. Characterizing genetic and environmental influences on methylation is critical for understanding the role of epigenetic mechanisms in evolutionary adaptation.

Genetic diversity and family groups detected in a coyote population with red wolf ancestry on Galveston Island, Texas.

BACKGROUND: Hybridization can be a conservation concern if genomic introgression leads to the loss of an endangered species' unique genome, or when hybrid offspring are sterile or less fit than their parental species. Yet hybridization can also be an adaptive management tool if rare populations are inbred and have reduced genetic variation, and there is the opportunity to enhance genetic variation through hybridization. The red wolf (Canis rufus) is a critically endangered wolf endemic to the eastern United States, where all extant red wolves are descended from 14 founders which has led to elevated levels of inbreeding over time. Red wolves were considered extirpated from the wild by 1980, but before they disappeared, they interbred with encroaching coyotes creating a genetically admixed population of canids along coastal Texas and Louisiana. In 2018, a genetic study identified individuals on Galveston Island, Texas with significant amounts of red wolf ancestry. We collected 203 fecal samples from Galveston for a more in-depth analysis of this population to identify the amount of red wolf ancestry present and potential mechanisms that support retention of red wolf ancestry on the landscape. RESULTS: We identified 24 individual coyotes from Galveston Island and 8 from mainland Texas with greater than 10% red wolf ancestry. Two of those individuals from mainland Texas had greater than 50% red wolf ancestry estimates. Additionally, this population had 5 private alleles that were absent in the North American reference canid populations used in this study, which included 107 southeastern coyotes, 19 captive red wolves, and 38 gray wolves, possibly representing lost red wolf genetic variation. We also identified several individuals on Galveston Island and the mainland of Texas that retained a unique red wolf mitochondrial haplotype present in the red wolf founding population. On Galveston Island, we identified a minimum of four family groups and found coyotes on the island to be highly related, but not genetically depauperate. We did not find clear associations between red wolf ancestry estimates and landscape features, such as open green space or developed areas. CONCLUSION: Our results confirm the presence of substantial red wolf ancestry persisting on Galveston Island and adjacent mainland Texas. This population has the potential to benefit future red wolf conservation efforts through novel reproductive techniques and possibly through de-introgression strategies, with the goals of recovering extinct red wolf genetic variation and reducing inbreeding within the species.

Disease outbreaks select for mate choice and coat color in wolves.

We know much about pathogen evolution and the emergence of new disease strains, but less about host resistance and how it is signaled to other individuals and subsequently maintained. The cline in frequency of black-coated wolves (Canis lupus) across North America is hypothesized to result from a relationship with canine distemper virus (CDV) outbreaks. We tested this hypothesis using cross-sectional data from wolf populations across North America that vary in the prevalence of CDV and the allele that makes coats black, longitudinal data from Yellowstone National Park, and modeling. We found that the frequency of CDV outbreaks generates fluctuating selection that results in heterozygote advantage that in turn affects the frequency of the black allele, optimal mating behavior, and black wolf cline across the continent.

Coyotes in New York City Carry Variable Genomic Dog Ancestry and Influence Their Interactions with Humans.

Coyotes are ubiquitous on the North American landscape as a result of their recent expansion across the continent. They have been documented in the heart of some of the most urbanized cities, such as Chicago, Los Angeles, and New York City. Here, we explored the genomic composition of 16 coyotes in the New York metropolitan area to investigate genomic demography and admixture for urban-dwelling canids in Queens County, New York. We identified moderate-to-high estimates of relatedness among coyotes living in Queens (r = 0.0-0.5) and adjacent neighborhoods, suggestive of a relatively small population. Although we found low background levels of domestic-dog ancestry across most coyotes in our sample (5%), we identified a male suspected to be a first-generation coyote-dog hybrid with 46% dog ancestry, as well as his two putative backcrossed offspring that carried approximately 25% dog ancestry. The male coyote-dog hybrid and one backcrossed offspring each carried two transposable element insertions that are associated with human-directed hypersociability in dogs and gray wolves. An additional, unrelated coyote with little dog ancestry also carried two of these insertions. These genetic patterns suggest that gene flow from domestic dogs may become an increasingly important consideration as coyotes continue to inhabit metropolitan regions.

Reviving ghost alleles: Genetically admixed coyotes along the American Gulf Coast are critical for saving the endangered red wolf.

The last known red wolves were captured in southwestern Louisiana and eastern Texas in 1980 to establish a captive breeding population. Before their extirpation, gene flow with coyotes resulted in the persistence of endangered red wolf genetic variation in local coyote populations. We assessed genomic ancestry and morphology of coyotes in southwestern Louisiana. We detected that 38 to 62% of the coyote genomes contained red wolf ancestry acquired in the past 30 years and have an admixture profile similar to that of the canids captured before the extirpation of red wolves. We further documented a positive correlation between ancestry and weight. Our findings highlight the importance of hybrids and admixed genomes as a reservoir of endangered species ancestry for innovative conservation efforts. Together, this work presents an unprecedented system that conservation can leverage to enrich the recovery program of an endangered species.

The effects of age, sex, weight, and breed on canid methylomes.

Unlike genomes, which are static throughout the lifespan of an organism, DNA methylomes are dynamic. To study these dynamics, we developed quantitative models that measure the effect of multiple factors on DNA methylomes including, age, sex, weight, and genetics. We conducted our study in canids, which prove to be an ideal species to assess epigenetic moderators due to their extreme variability in size and well-characterized genetic structure. We collected buccal swabs from 217 canids (207 domestic dogs and 10 grey wolves) and used targeted bisulphite sequencing to measure methylomes. We also measured genotypes at over one thousand single nucleotide polymorphisms (SNPs). As expected, we found that DNA methylomes are strongly associated with age, enabling the construction of epigenetic clocks. However, we also identify novel associations between methylomes and sex, weight, and sterilization status, leading to accurate models that predict these factors. Methylomes are also affected by genetics, and we observe multiple associations between SNP loci and methylated CpGs. Finally, we show that several factors moderate the relationship between epigenetic ages and real ages, such as body weight, which increases epigenetic ageing. In conclusion, we demonstrate that the plasticity of DNA methylomes is impacted by myriad genetics and physiological factors, and that DNA methylation biomarkers are accurate predictors of age, sex and sterilization status.

Natural and human-driven selection of a single non-coding body size variant in ancient and modern canids.

Domestic dogs (Canis lupus familiaris) are the most variable-sized mammalian species on Earth, displaying a 40-fold size difference between breeds.1 Although dogs of variable size are found in the archeological record,2-4 the most dramatic shifts in body size are the result of selection over the last two centuries, as dog breeders selected and propagated phenotypic extremes within closed breeding populations.5 Analyses of over 200 domestic breeds have identified approximately 20 body size genes regulating insulin processing, fatty acid metabolism, TGFß signaling, and skeletal formation.6-10 Of these, insulin-like growth factor 1 (IGF1) predominates, controlling approximately 15% of body size variation between breeds.8 The identification of a functional mutation associated with IGF1 has thus far proven elusive.6,10,11 Here, to identify and elucidate the role of an ancestral IGF1 allele in the propagation of modern canids, we analyzed 1,431 genome sequences from 13 species, including both ancient and modern canids, thus allowing us to define the evolutionary history of both ancestral and derived alleles at this locus. We identified a single variant in an antisense long non-coding RNA (IGF1-AS) that interacts with the IGF1 gene, creating a duplex. While the derived mutation predominates in both modern gray wolves and large domestic breeds, the ancestral allele, which predisposes to small size, was common in small-sized breeds and smaller wild canids. Our analyses demonstrate that this major regulator of canid body size nearly vanished in Pleistocene wolves, before its recent resurgence resulting from human-imposed selection for small-sized breed dogs.