High genetic load without purging in caribou, a diverse species at risk.

High intra-specific genetic diversity is associated with adaptive potential, which is key for resilience to global change. However, high variation may also support deleterious alleles through genetic load, thereby increasing the risk of inbreeding depression if population sizes decrease. Purging of deleterious variation has been demonstrated in some threatened species. However, less is known about the costs of declines and inbreeding in species with large population sizes and high genetic diversity even though this encompasses many species globally that are expected to undergo population declines. Caribou is a species of ecological and cultural significance in North America with a wide distribution supporting extensive phenotypic variation but with some populations undergoing significant declines resulting in their at-risk status in Canada. We assessed intra-specific genetic variation, adaptive divergence, inbreeding, and genetic load across populations with different demographic histories using an annotated chromosome-scale reference genome and 66 whole-genome sequences. We found high genetic diversity and nine phylogenomic lineages across the continent with adaptive diversification of genes, but also high genetic load among lineages. We found highly divergent levels of inbreeding across individuals, including the loss of alleles by drift but not increased purging in inbred individuals, which had more homozygous deleterious alleles. We also found comparable frequencies of homozygous deleterious alleles between lineages regardless of nucleotide diversity. Thus, further inbreeding may need to be mitigated through conservation efforts. Our results highlight the "double-edged sword" of genetic diversity that may be representative of other species atrisk affected by anthropogenic activities.

Genomic population structure and inbreeding history of Lake Superior caribou.

Caribou (Rangifer tarandus) have experienced dramatic declines in both range and population size across Canada over the past century. Boreal caribou (R. t. caribou), 1 of the 12 Designatable Units, has lost approximately half of its historic range in the last 150 years, particularly along the southern edge of its distribution. Despite this overall northward contraction, some populations have persisted at the trailing range edge, over 150 km south of the continuous boreal caribou range in Ontario, along the coast and nearshore islands of Lake Superior. The population history of caribou along Lake Superior remains unclear. It appears that these caribou likely represent a remnant distribution at the trailing edge of the receding population of boreal caribou, but they may also exhibit local adaptation to the coastal environment. A better understanding of the population structure and history of caribou along Lake Superior is important for their conservation and management. Here, we use high-coverage whole genomes (N = 20) from boreal, eastern migratory, and barren-ground caribou sampled in Manitoba, Ontario, and Quebec to investigate population structure and inbreeding histories. We discovered that caribou from the Lake Superior range form a distinct group but also found some evidence of gene flow with the continuous boreal caribou range. Notably, caribou along Lake Superior demonstrated relatively high levels of inbreeding (measured as runs of homozygosity; ROH) and genetic drift, which may contribute to the differentiation observed between ranges. Despite inbreeding, caribou along Lake Superior retained high heterozygosity, particularly in genomic regions without ROH. These results suggest that they present distinct genomic characteristics but also some level of gene flow with the continuous range. Our study provides key insights into the genomics of the southernmost range of caribou in Ontario, beginning to unravel the evolutionary history of these small, isolated caribou populations.

A de novo genome assembly and annotation of the southern flying squirrel (Glaucomys volans).

Northern (Glaucomys sabrinus) and southern (Glaucomys volans) flying squirrels are widespread species distributed across North America. Northern flying squirrels are common inhabitants of the boreal forest, also occurring in coniferous forest remnants farther south, whereas the southern flying squirrel range is centered in eastern temperate woodlands. These two flying squirrel species exhibit a hybrid zone across a latitudinal gradient in an area of recent secondary contact. Glaucomys hybrid offspring are viable and can successfully backcross with either parental species, however, the fitness implications of such events are currently unknown. Some populations of G. sabrinus are endangered, and thus, interspecific hybridization is a key conservation concern in flying squirrels. To provide a resource for future studies to evaluate hybridization and possible introgression, we sequenced and assembled a de novo long-read genome from a G. volans individual sampled in southern Ontario, Canada, while four short-read genomes (two G. sabrinus and two G. volans, all from Ontario) were resequenced on Illumina platforms. The final genome assembly consisted of approximately 2.40 Gb with a scaffold N50 of 455.26 Kb. Benchmarking Universal Single-Copy Orthologs reconstructed 3,742 (91.2%) complete mammalian genes and genome annotation using RNA-Seq identified the locations of 19,124 protein-coding genes. The four short-read individuals were aligned to our reference genome to investigate the demographic history of the two species. A principal component analysis clearly separated resequenced individuals, while inferring population size history using the Pairwise Sequentially Markovian Coalescent model noted an approximate species split 1 million years ago, and a single, possibly recently introgressed individual.

Population dynamics of caribou shaped by glacial cycles before the last glacial maximum.

Pleistocene glacial cycles influenced the diversification of high-latitude wildlife species through recurrent periods of range contraction, isolation, divergence, and expansion from refugia and subsequent admixture of refugial populations. We investigate population size changes and the introgressive history of caribou (Rangifer tarandus) in western Canada using 33 whole genome sequences coupled with larger-scale mitochondrial data. We found that a major population expansion of caribou occurred starting around 110,000 years ago (kya), the start of the last glacial period. Additionally, we found effective population sizes of some caribou reaching ~700,000 to 1,000,000 individuals, one of the highest recorded historical effective population sizes for any mammal species thus far. Mitochondrial analyses dated introgression events prior to the LGM dating to 20-30 kya and even more ancient at 60 kya, coinciding with colder periods with extensive ice coverage, further demonstrating the importance of glacial cycles and events prior to the LGM in shaping demographic history. Reconstructing the origins and differential introgressive history has implications for predictions on species responses under climate change. Our results have implications for other whole genome analyses using pairwise sequentially Markovian coalescent (PSMC) analyses, as well as highlighting the need to investigate pre-LGM demographic patterns to fully reconstruct the origin of species diversity, especially for high-latitude species.

Data on the first functionally-annotated de novo transcriptome assembly for North American flying squirrels (genus Glaucomys).

We report the first functionally-annotated de novo transcriptome assembly for North American flying squirrels (genus Glaucomys). RNA was extracted from tissue samples obtained from two northern flying squirrels and two southern flying squirrels sampled from Ontario, Canada, and sequenced on an Illumina paired-end sequencing platform. We reconstructed 702,228 Glaucomys transcripts using 193,323,120 sequence read pairs and captured sequence homologies, protein domains, and gene function classifications. Introgressive hybridization between northern (Glaucomys sabrinus) and southern flying squirrels (G. volans) has been observed in some areas of North America. However, existing molecular markers lack the resolution to discriminate late-generation introgressants and describe the extent to which hybridization influences the Glaucomys gene pool. These genomic resources can increase the resolution of molecular techniques used to examine the dynamics of the Glaucomys hybrid zone.

Considering Pleistocene North American wolves and coyotes in the eastern Canis origin story.

The evolutionary origins and hybridization patterns of Canis species in North America have been hotly debated for the past 30 years. Disentangling ancestry and timing of hybridization in Great Lakes wolves, eastern Canadian wolves, red wolves, and eastern coyotes are most often partitioned into a 2-species model that assigns all ancestry to gray wolves and/or coyotes, and a 3-species model that includes a third, North American evolved eastern wolf genome. The proposed models address recent or sometimes late Holocene hybridization events but have largely ignored potential Pleistocene era progenitors and opportunities for hybridization that may have impacted the current mixed genomes in eastern Canada and the United States. Here, we re-analyze contemporary and ancient mitochondrial DNA genomes with Bayesian phylogenetic analyses to more accurately estimate divergence dates among lineages. We combine that with a review of the literature on Late Pleistocene Canis distributions to: (a) identify potential Pleistocene progenitors to southern North American gray wolves and eastern wolves; and (b) illuminate opportunities for ancient hybridization events. Specifically, we propose that Beringian gray wolves (C. lupus) and extinct large wolf-like coyotes (C. latrans orcutti) are likely progenitors to Mexican and Plains gray wolves and eastern wolves, respectively, and may represent a potentially unrecognized source of introgressed genomic variation within contemporary Canis genomes. These events speak to the potential origins of contemporary genomes and provide a new perspective on Canis ancestry, but do not negate current conservation priorities of dwindling wolf populations with unique genomic signatures and key ecologically critical roles.

Spatial familial networks to infer demographic structure of wild populations.

In social species, reproductive success and rates of dispersal vary among individuals resulting in spatially structured populations. Network analyses of familial relationships may provide insights on how these parameters influence population-level demographic patterns. These methods, however, have rarely been applied to genetically derived pedigree data from wild populations.Here, we use parent-offspring relationships to construct familial networks from polygamous boreal woodland caribou (Rangifer tarandus caribou) in Saskatchewan, Canada, to inform recovery efforts. We collected samples from 933 individuals at 15 variable microsatellite loci along with caribou-specific primers for sex identification. Using network measures, we assess the contribution of individual caribou to the population with several centrality measures and then determine which measures are best suited to inform on the population demographic structure. We investigate the centrality of individuals from eighteen different local areas, along with the entire population.We found substantial differences in centrality of individuals in different local areas, that in turn contributed differently to the full network, highlighting the importance of analyzing networks at different scales. The full network revealed that boreal caribou in Saskatchewan form a complex, interconnected familial network, as the removal of edges with high betweenness did not result in distinct subgroups. Alpha, betweenness, and eccentricity centrality were the most informative measures to characterize the population demographic structure and for spatially identifying areas of highest fitness levels and family cohesion across the range. We found varied levels of dispersal, fitness, and cohesion in family groups. Synthesis and applications: Our results demonstrate the value of different network measures in assessing genetically derived familial networks. The spatial application of the familial networks identified individuals presenting different fitness levels, short- and long-distance dispersing ability across the range in support of population monitoring and recovery efforts.

Functional genetic diversity of domestic and wild American mink (Neovison vison).

The release of domestic organisms to the wild threatens biodiversity because the introduction of domestic genes through interbreeding can negatively impact wild conspecifics via outbreeding depression. In North America, farmed American mink (Neovison vison) frequently escape captivity, yet the impact of these events on functional genetic diversity of wild mink populations is unclear. We characterized domestic and wild mink in Ontario at 17 trinucleotide microsatellites located in functional genes thought to be associated with traits affected by domestication. We found low functional genetic diversity in both mink types, as only four of 17 genes were variable, yet allele frequencies varied widely between captive and wild populations. To determine whether allele frequencies of wild populations were affected by geographic location, we performed redundancy analysis and spatial analysis of principal components on three polymorphic loci (AR, ATN1 and IGF-1). We found evidence to suggest domestic release events are affecting the functional genetic diversity of wild mink, as sPCA showed clear distinctions between wild individuals near mink farms and those located in areas without mink farms. This is further substantiated through RDA, where spatial location was associated with genetic variation of AR, ATN1 and IGF1.

An assessment of sampling designs using SCR analyses to estimate abundance of boreal caribou.

Accurately estimating abundance is a critical component of monitoring and recovery of rare and elusive species. Spatial capture-recapture (SCR) models are an increasingly popular method for robust estimation of ecological parameters. We provide an analytical framework to assess results from empirical studies to inform SCR sampling design, using both simulated and empirical data from noninvasive genetic sampling of seven boreal caribou populations (Rangifer tarandus caribou), which varied in range size and estimated population density. We use simulated population data with varying levels of clustered distributions to quantify the impact of nonindependence of detections on density estimates, and empirical datasets to explore the influence of varied sampling intensity on the relative bias and precision of density estimates. Simulations revealed that clustered distributions of detections did not significantly impact relative bias or precision of density estimates. The genotyping success rate of our empirical dataset (n = 7,210 samples) was 95.1%, and 1,755 unique individuals were identified. Analysis of the empirical data indicated that reduced sampling intensity had a greater impact on density estimates in smaller ranges. The number of captures and spatial recaptures was strongly correlated with precision, but not absolute relative bias. The best sampling designs did not differ with estimated population density but differed between large and small ranges. We provide an efficient framework implemented in R to estimate the detection parameters required when designing SCR studies. The framework can be used when designing a monitoring program to minimize effort and cost while maximizing effectiveness, which is critical for informing wildlife management and conservation.

Spatial and environmental influences on selection in a clock gene coding trinucleotide repeat in Canada lynx (Lynx canadensis).

Clock genes exhibit substantial control over gene expression and ultimately life-histories using external cues such as photoperiod, and are thus likely to be critical for adaptation to shifting seasonal conditions and novel environments as species redistribute their ranges under climate change. Coding trinucleotide repeats (cTNRs) are found within several clock genes, and may be interesting targets of selection due to their containment within exonic regions and elevated mutation rates. Here, we conduct inter-specific characterization of the NR1D1 cTNR between Canada lynx and bobcat, and intra-specific spatial and environmental association analyses of neutral microsatellites and our functional cTNR marker, to investigate the role of selection on this locus in Canada lynx. We report signatures of divergent selection between lynx and bobcat, with the potential for hybrid-mediated gene flow in the area of range overlap. We also provide evidence that this locus is under selection across Canada lynx in eastern Canada, with both spatial and environmental variables significantly contributing to the explained variation, after controlling for neutral population structure. These results suggest that cTNRs may play an important role in the generation of functional diversity within some mammal species, and allow for contemporary rates of adaptation in wild populations in response to environmental change. We encourage continued investment into the study of cTNR markers to better understand their broader relevance to the evolution and adaptation of mammals.

A simulation-based approach to assess the power of trend detection in high- and low-frequency water quality records.

To provide more precise understanding of water quality changes, continuous sampling is being used more in surface water quality monitoring networks. However, it remains unclear how much improvement continuous monitoring provides over spot sampling, in identifying water quality changes over time. This study aims (1) to assess our ability to detect trends using water quality data of both high and low frequencies and (2) to assess the value of using high-frequency data as a surrogate to help detect trends in other constituents. Statistical regression models were used to identify temporal trends and then to assess the trend detection power of high-frequency (15 min) and low-frequency (monthly) data for turbidity and electrical conductivity (EC) data collected across Victoria, Australia. In addition, we developed surrogate models to simulate five sediment and nutrients constituents from runoff, turbidity and EC. A simulation-based statistical approach was then used to the compare the power to detect trends between the low- and high-frequency water quality records. Results show that high-frequency sampling shows clear benefits in trend detection power for turbidity, EC, as well as simulated sediment and nutrients, especially over short data periods. For detecting a 1% annual trend with 5 years of data, up to 97% and 94% improvements on the trend detection probability are offered by high-frequency data compared with monthly data, for turbidity and EC, respectively. Our results highlight the benefits of upgrading monitoring networks with wider application of high-frequency sampling.

The role of introgression and ecotypic parallelism in delineating intraspecific conservation units.

Parallel evolution can occur through selection on novel mutations, standing genetic variation or adaptive introgression. Uncovering parallelism and introgressed populations can complicate management of threatened species as parallelism may have influenced conservation unit designations and admixed populations are not generally considered under legislations. We examined high coverage whole-genome sequences of 30 caribou (Rangifer tarandus) from across North America and Greenland, representing divergent intraspecific lineages, to investigate parallelism and levels of introgression contributing to the formation of ecotypes. Caribou are split into four subspecies and 11 extant conservation units, known as designatable units (DUs), in Canada. Using genomes from all four subspecies and six DUs, we undertake demographic reconstruction and confirm two previously inferred instances of parallel evolution in the woodland subspecies and uncover an additional instance of parallelism of the eastern migratory ecotype. Detailed investigations reveal introgression in the woodland subspecies, with introgressed regions found spread throughout the genomes encompassing both neutral and functional sites. Our investigations using whole genomes highlight the difficulties in unequivocally demonstrating parallelism through adaptive introgression in nonmodel species with complex demographic histories, with standing variation and introgression both potentially involved. Additionally, the impact of parallelism and introgression on conservation policy for management units needs to be considered in general, and the caribou designations will need amending in light of our results. Uncovering and decoupling parallelism and differential patterns of introgression will become prevalent with the availability of comprehensive genomic data from nonmodel species, and we highlight the need to incorporate this into conservation unit designations.

Climate connectivity of the bobcat in the Great Lakes region.

The Great Lakes and the St. Lawrence River are imposing barriers for wildlife, and the additive effect of urban and agricultural development that dominates the lower Great Lakes region likely further reduces functional connectivity for many terrestrial species. As the climate warms, species will need to track climate across these barriers. It is important therefore to investigate land cover and bioclimatic hypotheses that may explain the northward expansion of species through the Great Lakes. We investigated the functional connectivity of a vagile generalist, the bobcat, as a representative generalist forest species common to the region. We genotyped tissue samples collected across the region at 14 microsatellite loci and compared different landscape hypotheses that might explain the observed gene flow or functional connectivity. We found that the Great Lakes and the additive influence of forest stands with either low or high canopy cover and deep lake-effect snow have disrupted gene flow, whereas intermediate forest cover has facilitated gene flow. Functional connectivity in southern Ontario is relatively low and was limited in part by the low amount of forest cover. Pathways across the Great Lakes were through the Niagara region and through the Lower Peninsula of Michigan over the Straits of Mackinac and the St. Marys River. These pathways are important routes for bobcat range expansion north of the Great Lakes and are also likely pathways that many other mobile habitat generalists must navigate to track the changing climate. The extent to which species can navigate these routes will be important for determining the future biodiversity of areas north of the Great Lakes.

Spatial differences in genetic diversity and northward migration suggest genetic erosion along the boreal caribou southern range limit and continued range retraction.

With increasing human activities and associated landscape changes, distributions of terrestrial mammals become fragmented. These changes in distribution are often associated with reduced population sizes and loss of genetic connectivity and diversity (i.e., genetic erosion) which may further diminish a species' ability to respond to changing environmental conditions and lead to local population extinctions. We studied threatened boreal caribou (Rangifer tarandus caribou) populations across their distribution in Ontario/Manitoba (Canada) to assess changes in genetic diversity and connectivity in areas of high and low anthropogenic activity. Using data from >1,000 caribou and nine microsatellite loci, we assessed population genetic structure, genetic diversity, and recent migration rates using a combination of network and population genetic analyses. We used Bayesian clustering analyses to identify population genetic structure and explored spatial and temporal variation in those patterns by assembling networks based on R ST and F ST as historical and contemporary genetic edge distances, respectively. The Bayesian clustering analyses identified broad-scale patterns of genetic structure and closely aligned with the R ST network. The F ST network revealed substantial contemporary genetic differentiation, particularly in areas presenting contemporary anthropogenic disturbances and habitat fragmentation. In general, relatively lower genetic diversity and greater genetic differentiation were detected along the southern range limit, differing from areas in the northern parts of the distribution. Moreover, estimation of migration rates suggested a northward movement of animals away from the southern range limit. The patterns of genetic erosion revealed in our study suggest ongoing range retraction of boreal caribou in central Canada.

The Caribou (Rangifer tarandus) Genome.

Rangifer tarandus, known as caribou or reindeer, is a widespread circumpolar species which presents significant variability in their morphology, ecology, and genetics. A genome was sequenced from a male boreal caribou (R. t. caribou) from Manitoba, Canada. Both paired end and Chicago libraries were constructed and sequenced on Illumina platforms. The final assembly consists of approximately 2.205 Gb, and has a scaffold N50 of 11.765 Mb. BUSCO (Benchmarking Universal Single-Copy Orthologs) reconstructed 3820 (93.1%) complete mammalian genes, and genome annotation identified the locations of 33,177 protein-coding genes. An alignment to the bovine genome was carried out, indicating sequence coverage on all bovine chromosomes. A high-quality reference genome will be invaluable for evolutionary research and for conservation efforts for the species. Further information about the genome, including a FASTA file of the assembly and the annotation files, is available on our caribou genome website. Raw sequence data is available at the National Centre for Biotechnology Information (NCBI), under the BioProject accession number PRJNA549927.

Partitioning drivers of spatial genetic variation for a continuously distributed population of boreal caribou: Implications for management unit delineation.

Isolation by distance (IBD) is a natural pattern not readily incorporated into theoretical models nor traditional metrics for differentiating populations, although clinal genetic differentiation can be characteristic of many wildlife species. Landscape features can also drive population structure additive to baseline IBD resulting in differentiation through isolation-by-resistance (IBR). We assessed the population genetic structure of boreal caribou across western Canada using nonspatial (STRUCTURE) and spatial (MEMGENE) clustering methods and investigated the relative contribution of IBD and IBR on genetic variation of 1,221 boreal caribou multilocus genotypes across western Canada. We further introduced a novel approach to compare the partitioning of individuals into management units (MU) and assessed levels of genetic connectivity under different MU scenarios. STRUCTURE delineated five genetic clusters while MEMGENE identified finer-scale differentiation across the study area. IBD was significant and did not differ for males and females both across and among detected genetic clusters. MEMGENE landscape analysis further quantified the proportion of genetic variation contributed by IBD and IBR patterns, allowing for the relative importance of spatial drivers, including roads, water bodies, and wildfires, to be assessed and incorporated into the characterization of population structure for the delineation of MUs. Local population units, as currently delineated in the boreal caribou recovery strategy, do not capture the genetic variation and connectivity of the ecotype across the study area. Here, we provide the tools to assess fine-scale spatial patterns of genetic variation, partition drivers of genetic variation, and evaluate the best management options for maintaining genetic connectivity. Our approach is highly relevant to vagile wildlife species that are of management and conservation concern and demonstrate varying degrees of IBD and IBR with clinal spatial genetic structure that challenges the delineation of discrete population boundaries.

Parallel evolution of site-specific changes in divergent caribou lineages.

The parallel evolution of phenotypes or traits within or between species provides important insight into the basic mechanisms of evolution. Genetic and genomic advances have allowed investigations into the genetic underpinnings of parallel evolution and the independent evolution of similar traits in sympatric species. Parallel evolution may best be exemplified among species where multiple genetic lineages, descended from a common ancestor, colonized analogous environmental niches, and converged on a genotypic or phenotypic trait. Modern North American caribou (Rangifer tarandus) originated from three ancestral sources separated during the Last Glacial Maximum (LGM): the Beringian-Eurasian lineage (BEL), the North American lineage (NAL), and the High Arctic lineage (HAL). Historical introgression between the NAL and the BEL has been found throughout Ontario and eastern Manitoba. In this study, we first characterized the functional differentiation in the cytochrome-b (cytB) gene by identifying nonsynonymous changes. Second, the caribou lineages were used as a direct means to assess site-specific parallel changes among lineages. There was greater functional diversity within the NAL despite the BEL having greater neutral diversity. The patterns of amino acid substitutions occurring within different lineages supported the parallel evolution of cytB amino acid substitutions suggesting different selective pressures among lineages. This study highlights the independent evolution of identical amino acid substitutions within a wide-ranging mammal species that have diversified from different ancestral haplogroups and where ecological niches can invoke parallel evolution.

Multi-species genetic connectivity in a terrestrial habitat network.

BACKGROUND: Habitat fragmentation reduces genetic connectivity for multiple species, yet conservation efforts tend to rely heavily on single-species connectivity estimates to inform land-use planning. Such conservation activities may benefit from multi-species connectivity estimates, which provide a simple and practical means to mitigate the effects of habitat fragmentation for a larger number of species. To test the validity of a multi-species connectivity model, we used neutral microsatellite genetic datasets of Canada lynx (Lynx canadensis), American marten (Martes americana), fisher (Pekania pennanti), and southern flying squirrel (Glaucomys volans) to evaluate multi-species genetic connectivity across Ontario, Canada. RESULTS: We used linear models to compare node-based estimates of genetic connectivity for each species to point-based estimates of landscape connectivity (current density) derived from circuit theory. To our knowledge, we are the first to evaluate current density as a measure of genetic connectivity. Our results depended on landscape context: habitat amount was more important than current density in explaining multi-species genetic connectivity in the northern part of our study area, where habitat was abundant and fragmentation was low. In the south however, where fragmentation was prevalent, genetic connectivity was correlated with current density. Contrary to our expectations however, locations with a high probability of movement as reflected by high current density were negatively associated with gene flow. Subsequent analyses of circuit theory outputs showed that high current density was also associated with high effective resistance, underscoring that the presence of pinch points is not necessarily indicative of gene flow. CONCLUSIONS: Overall, our study appears to provide support for the hypothesis that landscape pattern is important when habitat amount is low. We also conclude that while current density is proportional to the probability of movement per unit area, this does not imply increased gene flow, since high current density tends to be a result of neighbouring pixels with high cost of movement (e.g., low habitat amount). In other words, pinch points with high current density appear to constrict gene flow.

Signatures of selection in mammalian clock genes with coding trinucleotide repeats: Implications for studying the genomics of high-pace adaptation.

Climate change is predicted to affect the reproductive ecology of wildlife; however, we have yet to understand if and how species can adapt to the rapid pace of change. Clock genes are functional genes likely critical for adaptation to shifting seasonal conditions through shifts in timing cues. Many of these genes contain coding trinucleotide repeats, which offer the potential for higher rates of change than single nucleotide polymorphisms (SNPs) at coding sites, and, thus, may translate to faster rates of adaptation in changing environments. We characterized repeats in 22 clock genes across all annotated mammal species and evaluated the potential for selection on repeat motifs in three clock genes (NR1D1,CLOCK, and PER1) in three congeneric species pairs with different latitudinal range limits: Canada lynx and bobcat (Lynx canadensis and L. rufus), northern and southern flying squirrels (Glaucomys sabrinus and G. volans), and white-footed and deer mouse (Peromyscus leucopus and P. maniculatus). Signatures of positive selection were found in both the interspecific comparison of Canada lynx and bobcat, and intraspecific analyses in Canada lynx. Northern and southern flying squirrels showed differing frequencies at common CLOCK alleles and a signature of balancing selection. Regional excess homozygosity was found in the deer mouse at PER1 suggesting disruptive selection, and further analyses suggested balancing selection in the white-footed mouse. These preliminary signatures of selection and the presence of trinucleotide repeats within many clock genes warrant further consideration of the importance of candidate gene motifs for adaptation to climate change.