Remarkably High Repeat Content in the Genomes of Sparrows: The Importance of Genome Assembly Completeness for Transposable Element Discovery.

Transposable elements (TE) play critical roles in shaping genome evolution. Highly repetitive TE sequences are also a major source of assembly gaps making it difficult to fully understand the impact of these elements on host genomes. The increased capacity of long-read sequencing technologies to span highly repetitive regions promises to provide new insights into patterns of TE activity across diverse taxa. Here we report the generation of highly contiguous reference genomes using PacBio long-read and Omni-C technologies for three species of Passerellidae sparrow. We compared these assemblies to three chromosome-level sparrow assemblies and nine other sparrow assemblies generated using a variety of short- and long-read technologies. All long-read based assemblies were longer (range: 1.12 to 1.41 Gb) than short-read assemblies (0.91 to 1.08 Gb) and assembly length was strongly correlated with the amount of repeat content. Repeat content for Bell's sparrow (31.2% of genome) was the highest level ever reported within the order Passeriformes, which comprises over half of avian diversity. The highest levels of repeat content (79.2% to 93.7%) were found on the W chromosome relative to other regions of the genome. Finally, we show that proliferation of different TE classes varied even among species with similar levels of repeat content. These patterns support a dynamic model of TE expansion and contraction even in a clade where TEs were once thought to be fairly depauperate and static. Our work highlights how the resolution of difficult-to-assemble regions of the genome with new sequencing technologies promises to transform our understanding of avian genome evolution.

Ecological characteristics explain neutral genetic variation of three coastal sparrow species.

Eco-phylogeographic approaches to comparative population genetic analyses allow for the inclusion of intrinsic influences as drivers of intraspecific genetic structure. This insight into microevolutionary processes, including changes within a species or lineage, provides better mechanistic understanding of species-specific interactions and enables predictions of evolutionary responses to environmental change. In this study, we used single nucleotide polymorphisms (SNPs) identified from reduced representation sequencing to compare neutral population structure, isolation by distance (IBD), genetic diversity and effective population size (Ne) across three closely related and co-distributed saltmarsh sparrow species differing along a specialization gradient-Nelson's (Ammospiza nelsoni subvirgata), saltmarsh (A. caudacuta) and seaside sparrows (A. maritima maritima). Using an eco-phylogeographic lens within a conservation management context, we tested predictions about species' degree of evolutionary history and ecological specialization to tidal marshes, habitat, current distribution and population status on population genetic metrics. Population structure differed among the species consistent with their current distribution and habitat factors, rather than degree of ecological specialization: seaside sparrows were panmictic, saltmarsh sparrows showed hierarchical structure and Nelson's sparrows were differentiated into multiple, genetically distinct populations. Neutral population genetic theory and demographic/evolutionary history predicted patterns of genetic diversity and Ne rather than degree of ecological specialization. Patterns of population variation and evolutionary distinctiveness (Shapely metric) suggest different conservation measures for long-term persistence and evolutionary potential in each species. Our findings contribute to a broader understanding of the complex factors influencing genetic variation, beyond specialist-generalist status and support the role of an eco-phylogeographic approach in population and conservation genetics.

Plumage microorganism communities of tidal marsh sparrows.

Microorganism communities can shape host phenotype evolution but are often comprised of thousands of taxa with varied impact on hosts. Identification of taxa influencing host evolution relies on first describing microorganism communities and acquisition routes. Keratinolytic (keratin-degrading) microorganisms are hypothesized to be abundant in saltmarsh sediments and to contribute to plumage evolution in saltmarsh-adapted sparrows. Metabarcoding was used to describe plumage bacterial (16S rRNA) and fungal (ITS) communities in three sparrow species endemic to North America's Atlantic coast saltmarshes. Results describe limited within-species variability and moderate host species-level patterns in microorganism diversity and community composition. A small percentage of overall microorganism diversity was comprised of potentially keratinolytic microorganisms, warranting further functional studies. Distinctions between plumage and saltmarsh sediment bacteria, but not fungal, communities were detected, suggesting multiple bacterial acquisition routes and/or vertebrate host specialization. This research lays groundwork for future testing of causal links between microorganisms and avian host evolution.

New extralimital breeding records of saltmarsh sparrows (Ammospiza caudacuta) and Nelson's sparrows (Ammospiza nelsoni) and their implications.

Saltmarsh (Ammospiza caudacuta) and Nelson's (A. nelsoni) sparrows are sister taxa that breed in tidal marshes along the coast of the Northeastern United States and Canada. The Saltmarsh Sparrow breeds from mid-coast Maine south to Virginia, while the Acadian Nelson's Sparrow breeds from the Canadian maritime provinces south to northern Massachusetts. Here, we present three extralimital observations of breeding Saltmarsh (n = 2) and Nelson's (n = 1) sparrows. In 2021 and 2022, we observed Saltmarsh Sparrow females attending nests at Mendall Marsh, ME, and Milbridge, ME, respectively, approximately 60 and 110 km beyond the documented northern extent of the Saltmarsh Sparrow breeding range. In 2022, we observed a breeding-condition male Nelson's sparrow singing in the upriver portion of a marsh on Cape Cod, Massachusetts, approximately 115 km beyond the previously documented southern extent of the Nelson's Sparrow breeding range. We confirmed morphological species identification using a panel of microsatellite DNA loci. Due to both the well-documented population declines of these species in the region and the intensity of sampling effort undertaken in recent years, we suggest that these observations likely are not indicative of range expansion. However, they do indicate that these 2 taxa have the capacity to use and successfully reproduce in marshes well beyond their established breeding limits. Our findings provide novel insight into the potential for these taxa to occur and successfully breed outside their documented breeding ranges. Given increased interest in their conservation, these results support the idea that management actions aimed at creating or maintaining nesting habitat across both species ranges could benefit both taxa.

A high-quality de novo genome assembly for clapper rail (Rallus crepitans).

The clapper rail (Rallus crepitans), of the family Rallidae, is a secretive marsh bird species that is adapted for high salinity habitats. They are very similar in appearance to the closely related king rail (R. elegans), but while king rails are limited primarily to freshwater marshes, clapper rails are highly adapted to tolerate salt marshes. Both species can be found in brackish marshes where they freely hybridize, but the distribution of their respective habitats precludes the formation of a continuous hybrid zone and secondary contact can occur repeatedly. This system, thus, provides unique opportunities to investigate the underlying mechanisms driving their differential salinity tolerance as well as the maintenance of the species boundary between the 2 species. To facilitate these studies, we assembled a de novo reference genome assembly for a female clapper rail. Chicago and HiC libraries were prepared as input for the Dovetail HiRise pipeline to scaffold the genome. The pipeline, however, did not recover the Z chromosome so a custom script was used to assemble the Z chromosome. We generated a near chromosome level assembly with a total length of 994.8 Mb comprising 13,226 scaffolds. The assembly had a scaffold N50 was 82.7 Mb, L50 of four, and had a BUSCO completeness score of 92%. This assembly is among the most contiguous genomes among the species in the family Rallidae. It will serve as an important tool in future studies on avian salinity tolerance, interspecific hybridization, and speciation.

Separating Proactive Conservation from Species Listing Decisions.

Proactive Conservation is a paradigm of natural resource management in the United States that encourages voluntary, collaborative efforts to restore species before they need to be protected through government regulations. This paradigm is widely used to conserve at-risk species today, and when used in conjunction with the Policy for Evaluation of Conservation Efforts (PECE), it allows for successful conservation actions to preclude listing of species under the Endangered Species Act (ESA). Despite the popularity of this paradigm, and recent flagship examples of its use (e.g., greater sage grouse, Centrocercus urophasianus), critical assessments of the outcomes of Proactive Conservation are lacking from the standpoint of species status and recovery metrics. Here, we provide such an evaluation, using the New England cottontail (Sylvilagus transitionalis), heralded as a success of Proactive Conservation efforts in the northeastern United States, as a case study. We review the history and current status of the species, based on the state of the science, in the context of the Conservation Initiative, and the 2015 PECE decision not to the list the species under the ESA. In addition to the impacts of the PECE decision on the New England cottontail conservation specifically, our review also evaluates the benefits and limits of the Proactive Conservation paradigm more broadly, and we make recommendations for its role in relation to ESA implementation for the future of at-risk species management. We find that the status and assurances for recovery under the PECE policy, presented at the time of the New England cottontail listing decision, were overly optimistic, and the status of the species has worsened in subsequent years. We suggest that use of PECE to avoid listing may occur because of the perception of the ESA as a punitive law and a misconception that it is a failure, although very few listed species have gone extinct. Redefining recovery to decouple it from delisting and instead link it to probability of persistence under recommended conservation measures would remove some of the stigma of listing, and it would strengthen the role of Species Status Assessments in endangered species conservation.

Mercury exposure of tidal marsh songbirds in the northeastern United States and its association with nest survival.

The biogeochemistry of tidal marsh sediments facilitates the transformation of mercury (Hg) into the biologically available form methylmercury (MeHg), resulting in elevated Hg exposures to tidal marsh wildlife. Saltmarsh and Acadian Nelson's sparrows (Ammospiza caudacutua and A. nelsoni subvirgatus, respectively) exclusively inhabit tidal marshes, potentially experiencing elevated risk to Hg exposure, and have experienced range-wide population declines. To characterize spatial and temporal variation of Hg exposure in these species, we sampled total mercury (THg) in blood collected from 9 populations spanning 560 km of coastline, including individuals resampled within and among years. Using concurrent nesting studies, we tested whether THg was correlated with nest survival probabilities, an index of fecundity. Blood THg ranged from 0.074-3.373 µg/g ww across 170 samples from 127 individuals. We detected high spatial variability in Hg exposure, observing differences of more than 45-fold across all individuals and 8-fold in mean blood THg among all study plots, including 4-fold between study plots within 4 km. Intraindividual changes in blood Hg exposure did not vary systematically in time but were considerable, varying by up to 2-fold within and among years. Controlling for both species differences and maximum water level, the dominant driver of fecundity in this system, nest survival probability decreased by 10% across the full range of female blood THg concentrations observed. We conclude that Hg has the potential to impair songbird reproduction, potentially exacerbating known climate-change driven population declines from sea-level rise in saltmarsh and Acadian Nelson's sparrows.

Meta-analysis: Congruence of genomic and phenotypic differentiation across diverse natural study systems.

Linking genotype to phenotype is a primary goal for understanding the genomic underpinnings of evolution. However, little work has explored whether patterns of linked genomic and phenotypic differentiation are congruent across natural study systems and traits. Here, we investigate such patterns with a meta-analysis of studies examining population-level differentiation at subsets of loci and traits putatively responding to divergent selection. We show that across the 31 studies (88 natural population-level comparisons) we examined, there was a moderate (R 2 = 0.39) relationship between genomic differentiation (F ST ) and phenotypic differentiation (P ST ) for loci and traits putatively under selection. This quantitative relationship between P ST and F ST for loci under selection in diverse taxa provides broad context and cross-system predictions for genomic and phenotypic adaptation by natural selection in natural populations. This context may eventually allow for more precise ideas of what constitutes "strong" differentiation, predictions about the effect size of loci, comparisons of taxa evolving in nonparallel ways, and more. On the other hand, links between P ST and F ST within studies were very weak, suggesting that much work remains in linking genomic differentiation to phenotypic differentiation at specific phenotypes. We suggest that linking genotypes to specific phenotypes can be improved by correlating genomic and phenotypic differentiation across a spectrum of diverging populations within a taxon and including wide coverage of both genomes and phenomes.

Sperm length divergence as a potential prezygotic barrier in a passerine hybrid zone.

The saltmarsh sparrow Ammospiza caudacuta and Nelson's sparrow A. nelsoni differ in ecological niche, mating behavior, and plumage, but they hybridize where their breeding distributions overlap. In this advanced hybrid zone, past interbreeding and current backcrossing result in substantial genomic introgression in both directions, although few hybrids are currently produced in most locations. However, because both species are nonterritorial and have only brief male-female interactions, it is difficult to determine to what extent assortative mating explains the low frequency of hybrid offspring. Since females often copulate with multiple males, a role of sperm as a postcopulatory prezygotic barrier appears plausible. Here, we show that sperm length differs between the two species in the hybrid zone, with low among-male variation consistent with strong postcopulatory sexual selection on sperm cells. We hypothesize that divergence in sperm length may constitute a reproductive barrier between species, as sperm length co-evolves with the size of specialized female sperm storage tubules. Sperm does not appear to act as a postzygotic barrier, as sperm from hybrids was unexceptional.

Patterns of introgression vary within an avian hybrid zone.

BACKGROUND: Exploring hybrid zone dynamics at different spatial scales allows for better understanding of local factors that influence hybrid zone structure. In this study, we tested hypotheses about drivers of introgression at two spatial scales within the Saltmarsh Sparrow (Ammospiza caudacuta) and Nelson's Sparrow (A. nelsoni) hybrid zone. Specifically, we evaluated the influence of neutral demographic processes (relative species abundance), natural selection (exogenous environmental factors and genetic incompatibilities), and sexual selection (assortative mating) in this mosaic hybrid zone. By intensively sampling adults (n = 218) and chicks (n = 326) at two geographically proximate locations in the center of the hybrid zone, we determined patterns of introgression on a fine scale across sites of differing habitat. We made broadscale comparisons of patterns from the center with those of prior studies in the southern edge of the hybrid zone. RESULTS: A panel of fixed SNPs (135) identified from ddRAD sequencing was used to calculate a hybrid index and determine genotypic composition/admixture level of the populations. Another panel of polymorphic SNPs (589) was used to assign paternity and reconstruct mating pairs to test for sexual selection. On a broad-scale, patterns of introgression were not explained by random mating within marshes. We found high rates of back-crossing and similarly low rates of recent-generation (F1/F2) hybrids in the center and south of the zone. Offspring genotypic proportions did not meet those expected from random mating within the parental genotypic distribution. Additionally, we observed half as many F1/F2 hybrid female adults than nestlings, while respective male groups showed no difference, in support of Haldane's Rule. The observed proportion of interspecific mating was lower than expected when accounting for mate availability, indicating assortative mating was limiting widespread hybridization. On a fine spatial scale, we found variation in the relative influence of neutral and selective forces between inland and coastal habitats, with the smaller, inland marsh influenced primarily by neutral demographic processes, and the expansive, coastal marsh experiencing higher selective pressures in the form of natural (exogenous and endogenous) and sexual selection. CONCLUSIONS: Multiple drivers of introgression, including neutral and selective pressures (exogenous, endogenous, and sexual selection), are structuring this hybrid zone, and their relative influence is site and context-dependent.

Spatiotemporal landscape genetics: Investigating ecology and evolution through space and time.

Genetic time-series data from historical samples greatly facilitate inference of past population dynamics and species evolution. Yet, although climate and landscape change are often touted as post-hoc explanations of biological change, our understanding of past climate and landscape change influences on evolutionary processes is severely hindered by the limited application of methods that directly relate environmental change to species dynamics through time. Increased integration of spatiotemporal environmental and genetic data will revolutionize the interpretation of environmental influences on past population processes and the quantification of recent anthropogenic impacts on species, and vastly improve prediction of species responses under future climate change scenarios, yielding widespread revelations across evolutionary biology, landscape ecology and conservation genetics. This review encourages greater use of spatiotemporal landscape genetic analyses that explicitly link landscape, climate and genetic data through time by providing an overview of analytical approaches for integrating historical genetic and environmental data in five key research areas: population genetic structure, demography, phylogeography, metapopulation connectivity and adaptation. We also include a tabular summary of key methodological information, suggest approaches for mitigating the particular difficulties in applying these techniques to ancient DNA and palaeoclimate data, and highlight areas for future methodological development.

Divergent selection and drift shape the genomes of two avian sister species spanning a saline-freshwater ecotone.

The role of species divergence due to ecologically based divergent selection-or ecological speciation-in generating and maintaining biodiversity is a central question in evolutionary biology. Comparison of the genomes of phylogenetically related taxa spanning a selective habitat gradient enables discovery of divergent signatures of selection and thereby provides valuable insight into the role of divergent ecological selection in speciation. Tidal marsh ecosystems provide tractable opportunities for studying organisms' adaptations to selective pressures that underlie ecological divergence. Sharp environmental gradients across the saline-freshwater ecotone within tidal marshes present extreme adaptive challenges to terrestrial vertebrates. Here, we sequence 20 whole genomes of two avian sister species endemic to tidal marshes-the saltmarsh sparrow (Ammospiza caudacutus) and Nelson's sparrow (A. nelsoni)-to evaluate the influence of selective and demographic processes in shaping genome-wide patterns of divergence. Genome-wide divergence between these two recently diverged sister species was notably high (genome-wide F ST = 0.32). Against a background of high genome-wide divergence, regions of elevated divergence were widespread throughout the genome, as opposed to focused within islands of differentiation. These patterns may be the result of genetic drift resulting from past tidal march colonization events in conjunction with divergent selection to different environments. We identified several candidate genes that exhibited elevated divergence between saltmarsh and Nelson's sparrows, including genes linked to osmotic regulation, circadian rhythm, and plumage melanism-all putative candidates linked to adaptation to tidal marsh environments. These findings provide new insights into the roles of divergent selection and genetic drift in generating and maintaining biodiversity.

Novel signals of adaptive genetic variation in northwestern Atlantic cod revealed by whole-genome sequencing.

Selection can create complex patterns of adaptive differentiation among populations in the wild that may be relevant to management. Atlantic cod in the Northwest Atlantic are at a fraction of their historical abundance and a lack of recovery within the Gulf of Maine has created concern regarding the misalignment of fisheries management structures with biological population structure. To address this and investigate genome-wide patterns of variation, we used low-coverage sequencing to perform a region-wide, whole-genome analysis of fine-scale population structure. We sequenced 306 individuals from 20 sampling locations in U.S. and Canadian waters, including the major spawning aggregations in the Gulf of Maine in addition to spawning aggregations from Georges Bank, southern New England, the eastern Scotian Shelf, and St. Pierre Bank. With genotype likelihoods estimated at almost 11 million loci, we found large differences in haplotype frequencies of previously described chromosomal inversions between Canadian and U.S. sampling locations and also among U.S. sampling locations. Our whole-genome resolution also revealed novel outlier peaks, some of which showed significant genetic differentiation among sampling locations. Comparisons between allochronic winter- and spring-spawning populations revealed highly elevated relative (FST ) and absolute (dxy ) genetic differentiation near genes involved in reproduction, particularly genes associated with the brain-pituitary-gonadal axis, which likely control timing of spawning, contributing to prezygotic isolation. We also found genetic differentiation associated with heat shock proteins and other genes of functional relevance, with complex patterns that may point to multifaceted selection pressures and local adaptation among spawning populations. We provide a high-resolution picture of U.S. Atlantic cod population structure, revealing greater complexity than is currently recognized in management. Our genome-scan approach likely underestimates the full suite of adaptive differentiation among sampling locations. Nevertheless, it should inform the revision of stock boundaries to preserve adaptive genetic diversity and evolutionary potential of cod populations.

Framework for quantifying population responses to disturbance reveals that coastal birds are highly resilient to hurricanes.

Changes in the frequency and severity of extreme weather may introduce new threats to species that are already under stress from gradual habitat loss and climate change. We provide a probabilistic framework that quantifies potential threats by applying concepts from ecological resilience to single populations. Our approach uses computation to compare disturbance-impacted projections to a population's normal range of variation, quantifying the full range of potential impacts. We illustrate this framework with projection models for coastal birds, which are commonly depicted as vulnerable to disturbances, especially hurricanes and oil spills. We found that populations of coastal specialists are resilient to extreme disturbances, with high resistance to the effects of short-term reductions in vital rates and recovery within 20 years. Applying the general framework presented here across disturbance-prone species and ecosystems would improve understanding of population resilience and generate specific projections of resilience that are needed for effective conservation planning.

Genomics of rapid ecological divergence and parallel adaptation in four tidal marsh sparrows.

Theory suggests that different taxa having colonized a similar, challenging environment will show parallel or lineage-specific adaptations to shared selection pressures, but empirical examples of parallel evolution in independent taxa are exceedingly rare. We employed comparative genomics to identify parallel and lineage-specific responses to selection within and among four species of North American sparrows that represent four independent, post-Pleistocene colonization events by an ancestral, upland subspecies and a derived salt marsh specialist. We identified multiple cases of parallel adaptation in these independent comparisons following salt marsh colonization, including selection of 12 candidate genes linked to osmoregulation. In addition to detecting shared genetic targets of selection across multiple comparisons, we found many novel, species-specific signatures of selection, including evidence of selection of loci associated with both physiological and behavioral mechanisms of osmoregulation. Demographic reconstructions of all four species highlighted their recent divergence and small effective population sizes, as expected given their rapid radiation into saline environments. Our results highlight the interplay of both shared and lineage-specific selection pressures in the colonization of a biotically and abiotically challenging habitat and confirm theoretical expectations that steep environmental clines can drive repeated and rapid evolutionary diversification in birds.

Plasticity in nesting adaptations of a tidal marsh endemic bird.

If individuals can perceive and manage risks, they may alter their behaviors based on prior experience. This expectation may apply to nest site selection of breeding birds, for which adaptive behavioral responses may enhance fitness. Birds that nest in tidal marshes have adapted to the challenges posed primarily by periodic, monthly tidal flooding and secondarily by predation. We investigated adaptive responses in nesting behavior of the saltmarsh sparrow (Ammospiza caudacutus), an obligate tidal-marsh-breeding bird, using 536 nests monitored across 5 years. Using linear mixed effects models, we tested whether nest characteristics differed among nests that were successful, depredated, or flooded, and we investigated whether females made changes in nest structure and placement according to outcome of their previous nesting attempt. Nest characteristics differed among females with different nest fates. Fledged and depredated nests were built higher in the vegetation and in higher elevation areas of the marsh than those that flooded. Successful nests had more canopy cover and were comprised of a lower proportion of high marsh vegetation (Spartina patens) than those that were flooded or depredated. Females with nests that failed due to flooding constructed subsequent nests higher in the vegetation and at higher elevation than those that were successful in their prior attempt, consistent with a response to previous experience. Eighty-five percent of females renested within the average home range core area distance (77 m), indicating a high degree of nest placement fidelity. Females for which nests were depredated in their prior nesting attempt renested at a greater distance than females for which the previous nesting attempts were successful. Our findings suggest saltmarsh sparrows exhibit plasticity in nesting behavior, which may be important for balancing selective pressures in a dynamic environment. This plasticity, however, is insufficient to enable them to adapt to the increased flooding predicted with sea-level rise.

Bidirectional adaptive introgression between two ecologically divergent sparrow species.

Natural hybrid zones can be used to dissect the mechanisms driving key evolutionary processes by allowing us to identify genomic regions important for establishing reproductive isolation and that allow for transfer of adaptive variation. We leverage whole-genome data in a system where two bird species, the saltmarsh (Ammospiza caudacuta) and Nelson's (A. nelsoni) sparrow, hybridize despite their relatively high background genetic differentiation and past ecological divergence. Adaptive introgression is plausible in this system because Nelson's sparrows are recent colonists of saltwater marshes, compared to the specialized saltmarsh sparrow that has a longer history of saltmarsh adaptation. Comparisons among whole-genome sequences of 34 individuals from allopatric and sympatric populations show that ongoing gene flow is shaping the genomic landscape, with allopatric populations exhibiting genome-wide FST estimates close to double of that observed in sympatry. We characterized patterns of introgression across the genome and identify regions that exhibit biased introgression into hybrids from one parental species. These regions offer compelling candidates for genes related to tidal marsh adaptations suggesting that adaptive introgression may be an important consequence of hybridization. These findings highlight the value of considering the landscapes of both genome-wide introgression and divergence when characterizing the evolutionary forces that drive speciation.

Microsatellite marker development from next-generation sequencing in the New England cottontail (Sylvilagus transitionalis) and cross-amplification in the eastern cottontail (S. floridanus).

OBJECTIVE: The New England cottontail (Sylvilagus transitionalis) is a species of high conservation priority in the Northeastern United States, and was a candidate for federal listing under the Endangered Species Act until a recent decision determined that conservation actions were sufficient to preclude listing. The aim of this study was to develop a suite of microsatellite loci to guide future research efforts such as the analysis of population genetic structure, genetic variation, dispersal, and genetic mark-recapture population estimation. RESULTS: Thirty-five microsatellite markers containing tri- and tetranucleotide sequences were developed from shotgun genomic sequencing of tissue from S. transitionalis, S. obscurus, and S. floridanus. These loci were screened in n = 33 wild S. transitionalis sampled from a population in eastern Massachusetts, USA. Thirty-two of the 35 loci were polymorphic with 2-6 alleles, and observed heterozygosities of 0.06-0.82. All loci conformed to Hardy-Weinberg Equilibrium proportions and there was no evidence of linkage disequilibrium or null alleles. Primers for 33 of the 35 loci amplified DNA extracted from n = 6 eastern cottontail (S. floridanus) samples, of which nine revealed putative species-diagnostic alleles. These loci will provide a useful tool for conservation genetics investigations of S. transitionalis and a potential diagnostic species assay for differentiating sympatric eastern and New England cottontails.

Subspecies delineation amid phenotypic, geographic and genetic discordance in a songbird.

Understanding the processes that drive divergence within and among species is a long-standing goal in evolutionary biology. Traditional approaches to assessing differentiation rely on phenotypes to identify intra- and interspecific variation, but many species express subtle morphological gradients in which boundaries among forms are unclear. This intraspecific variation may be driven by differential adaptation to local conditions and may thereby reflect the evolutionary potential within a species. Here, we combine genetic and morphological data to evaluate intraspecific variation within the Nelson's (Ammodramus nelsoni) and salt marsh (Ammodramus caudacutus) sparrow complex, a group with populations that span considerable geographic distributions and a habitat gradient. We evaluated genetic structure among and within five putative subspecies of A. nelsoni and A. caudacutus using a reduced-representation sequencing approach to generate a panel of 1929 SNPs among 69 individuals. Although we detected morphological differences among some groups, individuals sorted along a continuous phenotypic gradient. In contrast, the genetic data identified three distinct clusters corresponding to populations that inhabit coastal salt marsh, interior freshwater marsh and coastal brackish-water marsh habitats. These patterns support the current species-level recognition but do not match the subspecies-level taxonomy within each species-a finding which may have important conservation implications. We identified loci exhibiting patterns of elevated divergence among and within these species, indicating a role for local selective pressures in driving patterns of differentiation across the complex. We conclude that this evidence for adaptive variation among subspecies warrants the consideration of evolutionary potential and genetic novelty when identifying conservation units for this group.

Seasonal fecundity is not related to geographic position across a species' global range despite a central peak in abundance.

The range of a species is determined by the balance of its demographic rates across space. Population growth rates are widely hypothesized to be greatest at the geographic center of the species range, but indirect empirical support for this pattern using abundance as a proxy has been mixed, and demographic rates are rarely quantified on a large spatial scale. Therefore, the texture of how demographic rates of a species vary over its range remains an open question. We quantified seasonal fecundity of populations spanning the majority of the global range of a single species, the saltmarsh sparrow (Ammodramus caudacutus), which demonstrates a peak of abundance at the geographic center of its range. We used a novel, population projection method to estimate seasonal fecundity inclusive of seasonal and spatial variation in life history traits that contribute to seasonal fecundity. We replicated our study over 3 years, and compared seasonal fecundity to latitude and distance among plots. We observed large-scale patterns in some life history traits that contribute to seasonal fecundity, such as an increase in clutch size with latitude. However, we observed no relationship between latitude and seasonal fecundity. Instead, fecundity varied greatly among plots separated by as little as 1 km. Our results do not support the hypothesis that demographic rates are highest at the geographic and abundance center of a species range, but rather they suggest that local drivers strongly influence saltmarsh sparrow fecundity across their global range.