Spatial variation in population genomic responses to over a century of anthropogenic change within a tidal marsh songbird.

Combating the current biodiversity crisis requires the accurate documentation of population responses to human-induced ecological change. However, our ability to pinpoint population responses to human activities is often limited to the analysis of populations studied well after the fact. Museum collections preserve a record of population responses to anthropogenic change that can provide critical baseline data on patterns of genetic diversity, connectivity, and population structure prior to the onset of human perturbation. Here, we leverage a spatially replicated time series of specimens to document population genomic responses to the destruction of nearly 90% of coastal habitats occupied by the Savannah sparrow (Passerculus sandwichensis) in California. We sequenced 219 sparrows collected from 1889 to 2017 across the state of California using an exome capture approach. Spatial-temporal analyses of genetic diversity found that the amount of habitat lost was not predictive of genetic diversity loss. Sparrow populations from southern California historically exhibited lower levels of genetic diversity and experienced the most significant temporal declines in genetic diversity. Despite experiencing the greatest levels of habitat loss, we found that genetic diversity in the San Francisco Bay area remained relatively high. This was potentially related to an observed increase in gene flow into the Bay Area from other populations. While gene flow may have minimized genetic diversity declines, we also found that immigration from inland freshwater-adapted populations into tidal marsh populations led to the erosion of divergence at loci associated with tidal marsh adaptation. Shifting patterns of gene flow through time in response to habitat loss may thus contribute to negative fitness consequences and outbreeding depression. Together, our results underscore the importance of tracing the genomic trajectories of multiple populations over time to address issues of fundamental conservation concern.

Blood Variation Implicates Respiratory Limits on Elevational Ranges of Andean Birds.

AbstractThe extent to which species ranges reflect intrinsic physiological tolerances is a major question in evolutionary ecology. To date, consensus has been hindered by the limited tractability of experimental approaches across most of the tree of life. Here, we apply a macrophysiological approach to understand how hematological traits related to oxygen transport shape elevational ranges in a tropical biodiversity hot spot. Along Andean elevational gradients, we measured traits that affect blood oxygen-carrying capacity-total and cellular hemoglobin concentration and hematocrit, the volume percentage of red blood cells-for 2,355 individuals of 136 bird species. We used these data to evaluate the influence of hematological traits on elevational ranges. First, we asked whether the sensitivity of hematological traits to changes in elevation is predictive of elevational range breadth. Second, we asked whether variance in hematological traits changed as a function of distance to the nearest elevational range limit. We found that birds showing greater hematological sensitivity had broader elevational ranges, consistent with the idea that a greater acclimatization capacity facilitates elevational range expansion. We further found reduced variation in hematological traits in birds sampled near their elevational range limits and at high absolute elevations, patterns consistent with intensified natural selection, reduced effective population size, or compensatory changes in other cardiorespiratory traits. Our findings suggest that constraints on hematological sensitivity and local genetic adaptation to oxygen availability promote the evolution of the narrow elevational ranges that underpin tropical montane biodiversity.

Natural history collections as a resource for conservation genomics: Understanding the past to preserve the future.

To avoid the worst outcomes of the current biodiversity crisis we need a deep understanding of population responses to human-induced ecological change. Rapidly expanding access to genomic resources for nonmodel taxa promises to play a unique role in meeting this goal. In particular, the increasing feasibility of sequencing DNA from historical specimens enables direct measures of population responses to the past century of anthropogenic change that will inform management strategies and refine projections of species responses to future environmental change. In this review, we discuss the methods that can be used to generate genome-scale data from the hundreds of millions of specimens housed in natural history collections around the world. We then highlight recent studies that utilize genomic data from specimens to address questions of fundamental importance to biodiversity conservation. Finally, we emphasize how traditional motivations of museum collectors, such as studies of geographic variation and community-wide inventories, provide unique opportunities for broad scale comparisons of genomic responses to anthropogenic change across time. We conclude that as sequencing technologies become increasingly accessible and more researchers take advantage of this resource, the importance of collections to the conservation of biodiversity will continue to grow.

A highly contiguous genome assembly for the California quail (Callipepla californica).

The California quail (Callipepla californica) is an iconic native bird of scrub and oak woodlands in California and the Baja Peninsula of Mexico. Here, we report a draft reference assembly for the species generated from PacBio HiFi long read and Omni-C chromatin-proximity sequencing data as part of the California Conservation Genomics Project (CCGP). Sequenced reads were assembled into 321 scaffolds totaling 1.08 Gb in length. Assembly metrics indicate a highly contiguous and complete assembly with a contig N50 of 5.5 Mb, scaffold N50 of 19.4 Mb, and BUSCO completeness score of 96.5%. Transposable elements (TEs) occupy 16.5% of the genome, more than previous Odontophoridae quail assemblies but in line with estimates of TE content for recent long-read assemblies of chicken and Peking duck. Together these metrics indicate that the present assembly is more complete than prior reference assemblies generated for Odontophoridae quail. This reference will serve as an essential resource for studies on local adaptation, phylogeography, and conservation genetics in this species of significant biological and recreational interest.

A highly contiguous reference genome for the Steller's jay (Cyanocitta stelleri).

The Steller's jay is a familiar bird of western forests from Alaska south to Nicaragua. Here, we report a draft reference assembly for the species generated from PacBio HiFi long-read and Omni-C chromatin-proximity sequencing data as part of the California Conservation Genomics Project (CCGP). Sequenced reads were assembled into 352 scaffolds totaling 1.16 Gb in length. Assembly metrics indicate a highly contiguous and complete assembly with a contig N50 of 7.8 Mb, scaffold N50 of 25.8 Mb, and BUSCO completeness score of 97.2%. Repetitive elements span 16.6% of the genome including nearly 90% of the W chromosome. Compared with high-quality assemblies from other members of the family Corvidae, the Steller's jay genome contains a larger proportion of repetitive elements than 4 crow species (Corvus), but a lower proportion of repetitive elements than the California scrub-jay (Aphelocoma californica). This reference genome will serve as an essential resource for future studies on speciation, local adaptation, phylogeography, and conservation genetics in this species of significant biological interest.

De novo assembly of a chromosome-level reference genome for the California Scrub-Jay, Aphelocoma californica.

We announce the assembly of the first de novo reference genome for the California Scrub-Jay (Aphelocoma californica). The genus Aphelocoma comprises four currently recognized species including many locally adapted populations across Mesoamerica and North America. Intensive study of Aphelocoma has revealed novel insights into the evolutionary mechanisms driving diversification in natural systems. Additional insights into the evolutionary history of this group will require continued development of high-quality, publicly available genomic resources. We extracted high molecular weight genomic DNA from a female California Scrub-Jay from northern California and generated PacBio HiFi long-read data and Omni-C chromatin conformation capture data. We used these data to generate a de novo partially phased diploid genome assembly, consisting of two pseudo-haplotypes, and scaffolded them using inferred physical proximity information from the Omni-C data. The more complete pseudo-haplotype assembly (arbitrarily designated "Haplotype 1") is 1.35 Gb in total length, highly contiguous (contig N50 = 11.53 Mb), and highly complete (BUSCO completeness score = 97%), with comparable scaffold sizes to chromosome-level avian reference genomes (scaffold N50 = 66.14 Mb). Our California Scrub-Jay assembly is highly syntenic with the New Caledonian Crow reference genome despite ~10 million years of divergence, highlighting the temporal stability of the avian genome. This high-quality reference genome represents a leap forward in publicly available genomic resources for Aphelocoma, and the family Corvidae more broadly. Future work using Aphelocoma as a model for understanding the evolutionary forces generating and maintaining biodiversity across phylogenetic scales can now benefit from a highly contiguous, in-group reference genome.

Genomic differentiation and local adaptation on a microgeographic scale in a resident songbird.

Elucidating forces capable of driving species diversification in the face of gene flow remains a key goal in evolutionary biology. Song sparrows, Melospiza melodia, occur as 25 subspecies in diverse habitats across North America, are among the continent's most widespread vertebrate species, and are exemplary of many highly variable species for which the conservation of locally adapted populations may be critical to their range-wide persistence. We focus here on six morphologically distinct subspecies resident in the San Francisco Bay region, including three salt-marsh endemics and three residents in upland and riparian habitats adjacent to the Bay. We used reduced-representation sequencing to generate 2,773 SNPs to explore genetic differentiation, spatial population structure, and demographic history. Clustering separated individuals from each of the six subspecies, indicating subtle differentiation at microgeographic scales. Evidence of limited gene flow and low nucleotide diversity across all six subspecies further supports a hypothesis of isolation among locally adapted populations. We suggest that natural selection for genotypes adapted to salt marsh environments and changes in demography over the past century have acted in concert to drive the patterns of diversification reported here. Our results offer evidence of microgeographic specialization in a highly polytypic bird species long discussed as a model of sympatric speciation and rapid adaptation, and they support the hypothesis that conserving locally adapted populations may be critical to the range-wide persistence of similarly highly variable species.

Seasonal and elevational variation in glucose and glycogen in two songbird species.

Birds naturally maintain high glucose concentrations in the blood and tissues, even when relying on fat to meet the metabolic demands of flight or thermogenesis. One possibility is that high glucose levels might be needed to deal with these metabolic demands. Thus, we hypothesized that birds chronically exposed to colder temperatures and higher elevations have higher circulating glucose and tissue free glucose and glycogen compared to conspecifics living at warmer temperatures and lower elevations. Adult House Sparrows (Passer domesticus) and House Finches (Haemorhous mexicanus) were captured from Phoenix, AZ (340 m elevation), and Albuquerque, NM (1600 m elevation), during the summer and winter months. We measured plasma glucose, as well as free glucose and glycogen from multiple tissues. In general, high elevation and colder temperatures were associated with higher tissue glycogen and higher free glucose concentrations in the brain. These findings indicate that glucose and glycogen are subject to seasonal phenotypic flexibility as well as geographic variations that may relate to local food availability and abundance.

Divergent mitochondrial lineages arose within a large, panmictic population of the Savannah sparrow (Passerculus sandwichensis).

Unusual patterns of mtDNA diversity can reveal interesting aspects of a species' biology. However, making such inferences requires discerning among the many alternative scenarios that could underlie any given mtDNA pattern. Next-generation sequencing methods provide large, multilocus data sets with increased power to resolve unusual mtDNA patterns. A mtDNA-based phylogeography of the Savannah sparrow (Passerculus sandwichensis) previously identified two sympatric, but divergent (~2%) clades within the nominate subspecies group and a third clade that consisted of birds sampled from northwest Mexico. We revisited the phylogeography of this species using a population genomic data set to resolve the processes leading to the evolution of sympatric and divergent mtDNA lineages. We identified two genetic clusters in the genomic data set corresponding to (a) the nominate subspecies group and (b) northwestern Mexico birds. Following divergence, the nominate clade maintained a large, stable population, indicating that divergent mitochondrial lineages arose within a panmictic population. Simulations based on parameter estimates from this model further confirmed that this demographic history could produce observed levels of mtDNA diversity. Patterns of divergent, sympatric mtDNA lineages are frequently interpreted as admixture of historically isolated lineages. Our analyses reject this interpretation for Savannah sparrows and underscore the need for genomic data sets to resolve the evolutionary mechanisms behind anomalous, locus-specific patterns.

Detecting introgression despite phylogenetic uncertainty: The case of the South American siskins.

Genetic introgression among closely related species is a widespread phenomenon across the Tree of Life and could be an important source of adaptive variation during early stages of diversification. In particular, genomic studies have revealed that many rapidly radiating clades tend to have complex, reticulate evolutionary histories. Although rapid radiations appear to be susceptible to introgression, they present special challenges for its detection because formal tests require accurate phylogenies, and paradoxically, introgression itself may obscure evolutionary relationships. To address this methodological challenge, we assessed introgression in a recent, rapid avian radiation in the Andes, the South American siskins (Spinus). Using ~45,000 SNPs, we estimated the Spinus phylogeny using multiple analytical approaches and recovered four strongly conflicting topologies. We performed a series of complimentary introgression tests that included valid tests for each of the likely species trees. From the consilience of test results, we inferred multiple introgression events among Andean Spinus in a way that was robust to phylogenetic uncertainty in the species tree. Positive tests for introgression were corroborated by independent population structure and ancestral assignment analyses, as well as a striking geographic pattern of mitochondrial haplotype sharing among species. The methodological approach we describe could be applied using any genomewide data, including SNP data, for clades without fully resolvable species trees. Our discovery of multiple introgression events within the Andean radiation of Spinus siskins is consistent with an emerging paradigm, that introgression tends to accompany the early stages of diversification.

Contribution of a mutational hot spot to hemoglobin adaptation in high-altitude Andean house wrens.

A key question in evolutionary genetics is why certain mutations or certain types of mutation make disproportionate contributions to adaptive phenotypic evolution. In principle, the preferential fixation of particular mutations could stem directly from variation in the underlying rate of mutation to function-altering alleles. However, the influence of mutation bias on the genetic architecture of phenotypic evolution is difficult to evaluate because data on rates of mutation to function-altering alleles are seldom available. Here, we report the discovery that a single point mutation at a highly mutable site in the ß(A)-globin gene has contributed to an evolutionary change in hemoglobin (Hb) function in high-altitude Andean house wrens (Troglodytes aedon). Results of experiments on native Hb variants and engineered, recombinant Hb mutants demonstrate that a nonsynonymous mutation at a CpG dinucleotide in the ß(A)-globin gene is responsible for an evolved difference in Hb-O2 affinity between high- and low-altitude house wren populations. Moreover, patterns of genomic differentiation between high- and low-altitude populations suggest that altitudinal differentiation in allele frequencies at the causal amino acid polymorphism reflects a history of spatially varying selection. The experimental results highlight the influence of mutation rate on the genetic basis of phenotypic evolution by demonstrating that a large-effect allele at a highly mutable CpG site has promoted physiological differentiation in blood O2 transport capacity between house wren populations that are native to different elevations.

The dual role of Andean topography in primary divergence: functional and neutral variation among populations of the hummingbird, Metallura tyrianthina.

BACKGROUND: The ridges and valleys of the Andes create physical barriers that limit animal dispersal and cause deterministic local variation in rainfall. This has resulted in physical isolation of animal populations and variation in habitats, each of which has likely contributed to the evolution of high species diversity in the region. However, the relative influences of geographic isolation, ecoclimatic conditions, and their potential interactions remain poorly understood. To address this, we compared patterns of genetic and morphological diversity in Peruvian populations of the hummingbird Metallura tyrianthina. RESULTS: Phylogenetic and variation partitioning analyses showed that geographic isolation rather than climatic dissimilarity explained the greatest proportion of genetic variance. In contrast, bill length variation was explained by climatic seasonality, but not by genetic divergence. We found that mutation-scaled migration rate (m) between persistently humid and semi-humid environments was nearly 20 times higher when the habitats were contiguous (m = 39.9) than when separated by a barrier, the Cordillera de Vilcanota (m = 2.1). Moreover, the population experiencing more gene flow exhibited a lesser degree of bill length divergence despite similar differences in climate. CONCLUSIONS: Geographic isolation is necessary for genetic divergence. Ecological differences, represented here by climate characteristics, are necessary for functional divergence. Gene flow appears to hinder the evolution of functional traits toward local adaptive optima. This suggests that functional diversification requires geographic isolation followed or accompanied by a shift in ecological conditions. Andean topography causes both isolation and climatic variation, underscoring its dual role in biotic diversification.

Integrating spatial analyses of genomic and physiological data to understand avian responses to environmental change.

Projected rates of climate change over the next century are expected to force species to shift ranges, adapt or acclimate to evade extinction. Predicting which of these scenarios may be most likely is a central challenge for conserving biodiversity in the immediate future. Modeling frameworks that take advantage of intraspecific variation across environmental gradients can be particularly important for meeting this challenge. While these space-for-time approaches are essential for climatic and genomic modeling approaches, mechanistic models that incorporate ecological physiology data into assessing species vulnerabilities rarely include intraspecific variation. A major reason for this gap is the general lack of empirical data on intraspecific geographic variation in avian physiological traits. In this review, we outline the evidence for and processes shaping geographic variation in avian traits. We use the example of evaporative water loss to underscore the lack of research on geographic variation even in traits central to cooling costs in birds. We next demonstrate how shifting the focus of avian physiological research to intraspecific variation can facilitate greater integration with emerging genomics approaches. Finally, we outline important next steps for an integrative approach to advance understanding of avian physiological adaptation within species. Addressing the knowledge gaps outlined in this review will contribute to an improved predictive framework that synthesizes environmental, morphological, physiological, and genomic data to assess species specific vulnerabilities to a warming planet.

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.

Maintenance of local adaptation despite gene flow in a coastal songbird.

Adaptation to local environments is common in widespread species and the basis of ecological speciation. The song sparrow (Melospiza melodia) is a widespread, polytypic passerine that occurs in shrubland habitats throughout North America. We examined the population structure of two parapatric subspecies that inhabit different environments: the Atlantic song sparrow (M. m. atlantica), a coastal specialist, and the eastern song sparrow (M. m. melodia), a shrubland generalist. These populations lacked clear mitochondrial population structure, yet coastal birds formed a distinct nuclear genetic cluster. We found weak overall genomic differentiation between these subspecies, suggesting either recent divergence, extensive gene flow, or a combination thereof. There was a steep genetic cline at the transition to coastal habitats, consistent with isolation by environment, not isolation by distance. A phenotype under divergent selection, bill size, varied with the amount of coastal ancestry in transitional areas, but larger bill size was maintained in coastal habitats regardless of ancestry, further supporting a role for selection in the maintenance of these subspecies. Demographic modeling suggested a divergence history of limited gene flow followed by secondary contact, which has emerged as a common theme in adaptive divergence across taxa.

The influence of spatially heterogeneous anthropogenic change on bill size evolution in a coastal songbird.

Natural history collections provide an unparalleled resource for documenting population responses to past anthropogenic change. However, in many cases, traits measured on specimens may vary temporally in response to a number of different anthropogenic pressures or demographic processes. While teasing apart these different drivers is challenging, approaches that integrate analyses of spatial and temporal series of specimens can provide a robust framework for examining whether traits exhibit common responses to ecological variation in space and time. We applied this approach to analyze bill morphology variation in California Savannah Sparrows (Passerculus sandwichensis). We found that bill surface area increased in birds from higher salinity tidal marshes that are hotter and drier. Only the coastal subspecies, alaudinus, exhibited a significant increase in bill size through time. As with patterns of spatial variation, alaudinus populations occupying higher salinity tidal marshes that have become warmer and drier over the past century exhibited the greatest increases in bill surface area. We also found a significant negative correlation between bill surface area and total evaporative water loss (TEWL) and estimated that observed increases in bill size could result in a reduction of up to 16.2% in daily water losses. Together, these patterns of spatial and temporal variation in bill size were consistent with the hypothesis that larger bills are favored in freshwater-limited environments as a mechanism of dissipating heat, reducing reliance on evaporative cooling, and increasing water conservation. With museum collections increasingly being leveraged to understand past responses to global change, this work highlights the importance of considering the influence of many different axes of anthropogenic change and of integrating spatial and temporal analyses to better understand the influence of specific human impacts on population change over time.

Population history and the selective landscape shape patterns of osmoregulatory trait divergence in tidal marsh Savannah sparrows (Passerculus sandwichensis).

A persistent challenge in making associations between phenotypic and environmental variation is understanding how ecological factors and demographic history interact to shape adaptive outcomes. Evaluating the degree to which conspecific populations exposed to similar environmental pressures respond in parallel provides a powerful framework for addressing this challenge. We took this comparative approach with multiple populations of Savannah sparrows (Passerculus sandwichensis) found in tidal marshes along the Pacific coast of North America. The high salinities characterizing tidal marshes select for increased osmoregulatory performance and salinity tolerance. We collected data on physiological traits associated with osmoregulatory performance from 10 tidal marsh and three freshwater-adapted interior populations to evaluate the degree of parallel divergence across populations. All traits showed differences in the magnitude of divergence, but only total evaporative water loss (TEWL) showed differences in the direction of divergence. The drivers of these differences in both the magnitude and direction of divergence varied among traits. For kidney morphology and TEWL, patterns of divergence were best explained by variation in immigration rate from interior populations. Maximum temperature was the best predictor of variation in urine excretion ability, and both gene flow and temperature contributed to variation in plasma osmolality. Finally, analysis of multitrait divergence patterns indicated that differences in the direction of divergence were best explained by population genetic structure, whereas differences in the magnitude of divergence were explained by environmental differences. Together these results show that the influences of demography and the selective landscape can manifest themselves differently across functionally integrated traits.

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.