Community-wide correlations between species richness, abundance and population genomic diversity in a freshwater biodiversity hotspot.

Understanding patterns of diversity across macro (e.g. species-level) and micro (e.g. molecular-level) scales can shed light on community function and stability by elucidating the abiotic and biotic drivers of diversity within ecological communities. We examined the relationships among taxonomic and genetic metrics of diversity in freshwater mussels (Bivalvia: Unionidae), an ecologically important and species-rich group in the southeastern United States. Using quantitative community surveys and reduced-representation genome sequencing across 22 sites in seven rivers and two river basins, we surveyed 68 mussel species and sequenced 23 of these species to characterize intrapopulation genetic variation. We tested for the presence of species diversity-abundance correlations (i.e. the more-individuals hypothesis, MIH), species-genetic diversity correlations (SGDCs) and abundance-genetic diversity correlations (AGDCs) across all sites to evaluate relationships between different metrics of diversity. Sites with greater cumulative multispecies density (a standardized metric of abundance) had a greater number of species, consistent with the MIH hypothesis. Intrapopulation genetic diversity was strongly associated with the density of most species, indicating the presence of AGDCs. However, there was no consistent evidence for SGDCs. Although sites with greater overall densities of mussels had greater species richness, sites with higher genetic diversity did not always exhibit positive correlations with species richness, suggesting that there are spatial and evolutionary scales at which the processes influencing community-level diversity and intraspecific diversity differ. Our work reveals the importance of local abundance as indicator (and possibly a driver) of intrapopulation genetic diversity.

Species limits and diversification of the Dendropsophus rubicundulus subgroup (Anura, Hylidae) in Neotropical savannas.

Understanding the processes that generate and maintain biodiversity at and below the species level is a central goal of evolutionary biology. Here we explore the spatial and temporal drivers of diversification of the treefrog subgroup Dendropsophus rubicundulus, a subgroup of the D. microcephalus species group, over periods of pronounced geological and climatic changes in the Neotropical savannas that they inhabit. This subgroup currently comprises 11 recognized species distributed across the Brazilian and Bolivian savannas, but the taxonomy has been in a state of flux, necessitating reexamination. Using newly generated single nucleotide polymorphism (SNP) data from restriction-site associated DNA sequencing (RADseq) and mitochondrial 16S sequence data for ∼150 specimens, we inferred phylogenetic relationships, tested species limits using a model-based approach, and estimated divergence times to gain insights into the geographic and climatic events that affected the diversification of this subgroup. Our results recognized at least nine species: D. anataliasiasi, D. araguaya, D. cerradensis, D. elianeae, D. jimi, D. rubicundulus, D. tritaeniatus, D. rozenmani, and D. sanborni. Although we did not collect SNP data for the latter two species, they are likely distinct based on mitochondrial data. In addition, we found genetic structure within the widespread species D. rubicundulus, which comprises three allopatric lineages connected by gene flow upon secondary contact. We also found evidence of population structure and perhaps undescribed diversity in D. elianeae, which warrants further study. The D. rubicundulus subgroup is estimated to have originated in the Late Miocene (∼5.45 million years ago), with diversification continuing through the Pliocene and Early Pleistocene, followed by the most recent divergence of D. rubicundulus lineages in the Middle Pleistocene. The epeirogenic uplift followed by erosion and denudation of the central Brazilian plateau throughout the Pliocene and Pleistocene, in combination with the increasing frequency and amplitude of climatic fluctuations during the Pleistocene, was important for generating and structuring diversity at or below the species level in the D. rubicundulus subgroup.

A new lineage of Galapagos giant tortoises identified from museum samples.

The Galapagos Archipelago is recognized as a natural laboratory for studying evolutionary processes. San Cristóbal was one of the first islands colonized by tortoises, which radiated from there across the archipelago to inhabit 10 islands. Here, we sequenced the mitochondrial control region from six historical giant tortoises from San Cristóbal (five long deceased individuals found in a cave and one found alive during an expedition in 1906) and discovered that the five from the cave are from a clade that is distinct among known Galapagos giant tortoises but closely related to the species from Española and Pinta Islands. The haplotype of the individual collected alive in 1906 is in the same clade as the haplotype in the contemporary population. To search for traces of a second lineage in the contemporary population on San Cristóbal, we closely examined the population by sequencing the mitochondrial control region for 129 individuals and genotyping 70 of these for both 21 microsatellite loci and >12,000 genome-wide single nucleotide polymorphisms [SNPs]. Only a single mitochondrial haplotype was found, with no evidence to suggest substructure based on the nuclear markers. Given the geographic and temporal proximity of the two deeply divergent mitochondrial lineages in the historical samples, they were likely sympatric, raising the possibility that the lineages coexisted. Without the museum samples, this important discovery of an additional lineage of Galapagos giant tortoise would not have been possible, underscoring the value of such collections and providing insights into the early evolution of this iconic radiation.

Demographic history and patterns of molecular evolution from whole genome sequencing in the radiation of Galapagos giant tortoises.

Whole genome sequencing provides deep insights into the evolutionary history of a species, including patterns of diversity, signals of selection, and historical demography. When applied to closely related taxa with a wealth of background knowledge, population genomics provides a comparative context for interpreting population genetic summary statistics and comparing empirical results with the expectations of population genetic theory. The Galapagos giant tortoises (Chelonoidis spp.), an iconic rapid and recent radiation, offer such an opportunity. Here, we sequenced whole genomes from three individuals of the 12 extant lineages of Galapagos giant tortoise and estimate diversity measures and reconstruct changes in coalescent rate over time. We also compare the number of derived alleles in each lineage to infer how synonymous and nonsynonymous mutation accumulation rates correlate with population size and life history traits. Remarkably, we find that patterns of molecular evolution are similar within individuals of the same lineage, but can differ significantly among lineages, reinforcing the evolutionary distinctiveness of the Galapagos giant tortoise species. Notably, differences in mutation accumulation among lineages do not align with simple population genetic predictions, suggesting that the drivers of purifying selection are more complex than is currently appreciated. By integrating results from earlier population genetic and phylogeographic studies with new findings from the analysis of whole genomes, we provide the most in-depth insights to date on the evolution of Galapagos giant tortoises, and identify discrepancies between expectation from population genetic theory and empirical data that warrant further scrutiny.

Temporal Mitogenomics of the Galapagos Giant Tortoise from Pinzón Reveals Potential Biases in Population Genetic Inference.

Empirical population genetic studies generally rely on sampling subsets of the population(s) of interest and of the nuclear or organellar genome targeted, assuming each is representative of the whole. Violations of these assumptions may impact population-level parameter estimation and lead to spurious inferences. Here, we used targeted capture to sequence the full mitochondrial genome from 123 individuals of the Galapagos giant tortoise endemic to Pinzón Island (Chelonoidis duncanensis) sampled at 2 time points pre- and postbottleneck (circa 1906 and 2014) to explicitly assess differences in diversity estimates and demographic reconstructions based on subsets of the mitochondrial genome versus the full sequences and to evaluate potential biases associated with diversity estimates and demographic reconstructions from postbottlenecked samples alone. Haplotypic diversities were equal between the temporal samples based on the full mitochondrial genome, but single gene estimates suggested either decreases or increases in diversity depending upon the region. Demographic reconstructions based on the full sequence were more similar between the temporal samples than those based on the control region alone, or a subset of 3 regions, where the trends in population size changes shifted in magnitude and direction between the temporal samples. In all cases, the estimated coalescent point was more distant for the historical than contemporary sample. In summary, our results empirically demonstrate the influence of sampling bias when interpreting population genetic patterns and punctuate the need for careful consideration of potentially conflicting evolutionary signal across the mitochondrial genome.

The evolution of phylogeographic data sets.

Empirical phylogeographic studies have progressively sampled greater numbers of loci over time, in part motivated by theoretical papers showing that estimates of key demographic parameters improve as the number of loci increases. Recently, next-generation sequencing has been applied to questions about organismal history, with the promise of revolutionizing the field. However, no systematic assessment of how phylogeographic data sets have changed over time with respect to overall size and information content has been performed. Here, we quantify the changing nature of these genetic data sets over the past 20 years, focusing on papers published in Molecular Ecology. We found that the number of independent loci, the total number of alleles sampled and the total number of single nucleotide polymorphisms (SNPs) per data set has improved over time, with particularly dramatic increases within the past 5 years. Interestingly, uniparentally inherited organellar markers (e.g. animal mitochondrial and plant chloroplast DNA) continue to represent an important component of phylogeographic data. Single-species studies (cf. comparative studies) that focus on vertebrates (particularly fish and to some extent, birds) represent the gold standard of phylogeographic data collection. Based on the current trajectory seen in our survey data, forecast modelling indicates that the median number of SNPs per data set for studies published by the end of the year 2016 may approach ~20,000. This survey provides baseline information for understanding the evolution of phylogeographic data sets and underscores the fact that development of analytical methods for handling very large genetic data sets will be critical for facilitating growth of the field.

Lineage fusion in Galápagos giant tortoises.

Although many classic radiations on islands are thought to be the result of repeated lineage splitting, the role of past fusion is rarely known because during these events, purebreds are rapidly replaced by a swarm of admixed individuals. Here, we capture lineage fusion in action in a Galápagos giant tortoise species, Chelonoidis becki, from Wolf Volcano (Isabela Island). The long generation time of Galápagos tortoises and dense sampling (841 individuals) of genetic and demographic data were integral in detecting and characterizing this phenomenon. In C. becki, we identified two genetically distinct, morphologically cryptic lineages. Historical reconstructions show that they colonized Wolf Volcano from Santiago Island in two temporally separated events, the first estimated to have occurred ~199 000 years ago. Following arrival of the second wave of colonists, both lineages coexisted for approximately ~53 000 years. Within that time, they began fusing back together, as microsatellite data reveal widespread introgressive hybridization. Interestingly, greater mate selectivity seems to be exhibited by purebred females of one of the lineages. Forward-in-time simulations predict rapid extinction of the early arriving lineage. This study provides a rare example of reticulate evolution in action and underscores the power of population genetics for understanding the past, present and future consequences of evolutionary phenomena associated with lineage fusion.

Ecological coassociations influence species' responses to past climatic change: an example from a Sonoran Desert bark beetle.

Ecologically interacting species may have phylogeographical histories that are shaped both by features of their abiotic landscape and by biotic constraints imposed by their coassociation. The Baja California peninsula provides an excellent opportunity to examine the influence of abiotic vs. biotic factors on patterns of diversity in plant-insect species.This is because past climatic and geological changes impacted the genetic structure of plants quite differently to that of codistributed free-living animals (e.g. herpetofauna and small mammals). Thus, 'plant-like' patterns should be discernible in host-specific insect herbivores. Here, we investigate the population history of a monophagous bark beetle, Araptus attenuatus, and consider drivers of phylogeographical patterns in the light of previous work on its host plant, Euphorbia lomelii. Using a combination of phylogenetic, coalescent-simulation-based and exploratory analyses of mitochondrial DNA sequences and nuclear genotypic data, we found that the evolutionary history of A. attenuatus exhibits similarities to its host plant that are attributable to both biotic and abiotic processes. Southward range expansion and recent colonization of continental Sonora from the Baja peninsula appear to be unique to this taxon pair and probably reflect influences of the host plant. On the other hand, abiotic factors with landscape level influences on a diverse suite of codistributed arid-adapted taxa, such as Plio- and Pleistocene-aged marine incursions in the region, also left genetic signatures in beetle and host plant populations. Superimposed on these similarities, bark beetle-specific patterns and processes were also evident: our data revealed two secondarily sympatric,reproductively isolated genetic lineages, as well as a previously unrecognized mid peninsular warm desert refuge. Taken together, this work illustrates that the evolutionary history of species-specific insect herbivores may represent a mosaic of influences,including­but not limited to­those imposed by the host plant.

Genetic signatures of lineage fusion closely resemble population decline.

Accurate interpretation of the genetic signatures of past demographic events is crucial for reconstructing evolutionary history. Lineage fusion (complete merging, resulting in a single panmictic population) is a special case of secondary contact that is seldom considered. Here, the circumstances under which lineage fusion can be distinguished from population size constancy, growth, bottleneck, and decline were investigated. Multi-locus haplotype data were simulated under models of lineage fusion with different divergence versus sampling lag times (D:L ratios). These pseudo-observed datasets also differed in their allocation of a fixed amount of sequencing resources (number of sampled alleles, haplotype length, number of loci). Distinguishability of lineage fusion versus each of 10 untrue non-fusion scenarios was quantified based on six summary statistics (neutrality tests). Some datasets were also analyzed using extended Bayesian skyline plots. Results showed that signatures of lineage fusion very closely resemble those of decline-high distinguishability was generally limited to the most favorable scenario (D:L = 9), using the most sensitive summary statistics (F S and Z nS), coupled with the optimal sequencing resource allocation (maximizing number of loci). Also, extended Bayesian skyline plots often erroneously inferred population decline. Awareness of the potential for lineage fusion to carry the hallmarks of population decline is critical.

Nonrecombining genes in a recombination environment: the Drosophila "dot" chromosome.

Rate of recombination is a powerful variable affecting several aspects of molecular variation and evolution. A nonrecombining portion of the genome of most Drosophila species, the "dot" chromosome or F element, exhibits very low levels of variation and unusual codon usage. One lineage of Drosophila, the willistoni/saltans groups, has the F element fused to a normally recombining E element. Here, we present polymorphism data for genes on the F element in two Drosophila willistoni and one D. insularis populations, genes previously studied in D. melanogaster. The D. willistoni populations were known to be very low in inversion polymorphism, thus minimizing the recombination suppression effect of inversions. We first confirmed, by in situ hybridization, that D. insularis has the same E + F fusion as D. willistoni, implying this was a monophyletic event. A clear gradient in codon usage exists along the willistoni F element, from the centromere distally to the fusion with E; estimates of recombination rates parallel this gradient and also indicate D. insularis has greater recombination than D. willistoni. In contrast to D. melanogaster, genes on the F element exhibit moderate levels of nucleotide polymorphism not distinguishable from two genes elsewhere in the genome. Although some linkage disequilibrium (LD) was detected between polymorphic sites within genes (generally <500 bp apart), no long-range LD between F element loci exists in the two willistoni group species. In general, the distribution of allele frequencies of F element genes display the typical pattern of expectations of neutral variation at equilibrium. These results are consistent with the hypothesis that recombination allows the accumulation of nucleotide variation as well as allows selection to act on synonymous codon usage. It is estimated that the fusion occurred ∼20 Mya and while the F element in the willistoni lineage has evolved "normal" levels and patterns of nucleotide variation, equilibrium may not have been reached for codon usage.

The effect of sampling density and study area size on landscape genetics inferences for the Mississippi slimy salamander (Plethodon mississippi).

In landscape genetics, it is largely unknown how choices regarding sampling density and study area size impact inferences upon which habitat features impede vs. facilitate gene flow. While it is recommended that sampling locations be spaced no further apart than the average individual's dispersal distance, for low-mobility species, this could lead to a challenging number of sampling locations, or an unrepresentative study area. We assessed the effects of sampling density and study area size on landscape genetic inferences for a dispersal-limited amphibian, Plethodon mississippi, via analysis of nested datasets. Microsatellite-based genetic distances among individuals were divided into three datasets representing sparse sampling across a large study area, dense sampling across a small study area, or sparse sampling across the same small study area. These datasets were a proxy for gene flow (i.e., the response variable) in maximum-likelihood population effects models that assessed the nature and strength of their relationship with each of five land-use classes (i.e., potential predictor variables). Comparisons of outcomes were based on the rank order of effect, sign of effect (i.e., gene flow resistance vs. facilitation), spatial scale of effect, and functional relationship with gene flow. The best-fit model for each dataset had the same sign of effect for hardwood forests, manmade structures, and pine forests, indicating the impacts of these land-use classes on dispersal and gene flow in P. mississippi are robust to sampling scheme. Contrasting sampling densities led to a different inferred functional relationship between agricultural areas and gene flow. Study area size appeared to influence the scale of effect of manmade structures and the sign of effect of pine forests. Our findings provided evidence for an influence of sampling density, study area size, and sampling effort upon inferences. Accordingly, we recommend iterative subsampling of empirical datasets and continued investigation into the sensitivities of landscape genetic analyses using simulations.

Weak spatial-genetic structure in a native invasive, the southern pine beetle (Dendroctonus frontalis), across the eastern United States.

The southern pine beetle, Dendroctonus frontalis, is a native pest of pine trees that has recently expanded its range into the northeastern United States. Understanding its colonization, dispersal, and connectivity will be critical for mitigating negative economic and ecological impacts in the newly invaded areas. Characterization of spatial-genetic structure can contribute to this; however, previous studies have reached different conclusions about regional population genetic structure, with one study reporting a weak east-west pattern, and the most recent reporting an absence of structure. Here we systematically assessed several explanations for the absence of spatial-genetic structure. To do this, we developed nine new microsatellite markers and combined them with an existing 24-locus data matrix for the same individuals. We then reanalyzed this full dataset alongside datasets in which certain loci were omitted with the goal of creating more favorable signal to noise ratios. We also partitioned the data based on the sex of D. frontalis individuals, and then employed a broad suite of genotypic clustering and isolation-by-distance (IBD) analyses. We found that neither inadequate information content in the molecular marker set, nor unfavorable signal-to-noise ratio, nor insensitivity of the analytical approaches could explain the absence of structure. Regardless of dataset composition, there was little evidence for clusters (i.e., distinct geo-genetic groups) or clines (i.e., gradients of increasing allele frequency differences over larger geographic distances), with one exception: significant IBD was repeatedly detected using an individual-based measure of relatedness whenever datasets included males (but not for female-only datasets). This is strongly indicative of broad-scale female-biased dispersal, which has not previously been reported for D. frontalis, in part owing to logistical limitations of direct approaches (e.g., capture-mark-recapture). Weak spatial-genetic structure suggests long-distance connectivity and that gene flow is high, but additional research is needed to understand range expansion and outbreak dynamics in this species using alternate approaches.

Nuclear gene phylogeography using PHASE: dealing with unresolved genotypes, lost alleles, and systematic bias in parameter estimation.

BACKGROUND: A widely-used approach for screening nuclear DNA markers is to obtain sequence data and use bioinformatic algorithms to estimate which two alleles are present in heterozygous individuals. It is common practice to omit unresolved genotypes from downstream analyses, but the implications of this have not been investigated. We evaluated the haplotype reconstruction method implemented by PHASE in the context of phylogeographic applications. Empirical sequence datasets from five non-coding nuclear loci with gametic phase ascribed by molecular approaches were coupled with simulated datasets to investigate three key issues: (1) haplotype reconstruction error rates and the nature of inference errors, (2) dataset features and genotypic configurations that drive haplotype reconstruction uncertainty, and (3) impacts of omitting unresolved genotypes on levels of observed phylogenetic diversity and the accuracy of downstream phylogeographic analyses. RESULTS: We found that PHASE usually had very low false-positives (i.e., a low rate of confidently inferring haplotype pairs that were incorrect). The majority of genotypes that could not be resolved with high confidence included an allele occurring only once in a dataset, and genotypic configurations involving two low-frequency alleles were disproportionately represented in the pool of unresolved genotypes. The standard practice of omitting unresolved genotypes from downstream analyses can lead to considerable reductions in overall phylogenetic diversity that is skewed towards the loss of alleles with larger-than-average pairwise sequence divergences, and in turn, this causes systematic bias in estimates of important population genetic parameters. CONCLUSIONS: A combination of experimental and computational approaches for resolving phase of segregating sites in phylogeographic applications is essential. We outline practical approaches to mitigating potential impacts of computational haplotype reconstruction on phylogeographic inferences. With targeted application of laboratory procedures that enable unambiguous phase determination via physical isolation of alleles from diploid PCR products, relatively little investment of time and effort is needed to overcome the observed biases.

Landscape modelling of gene flow: improved power using conditional genetic distance derived from the topology of population networks.

Landscape genetics is a burgeoning field of interest that focuses on how site-specific factors influence the distribution of genetic variation and the genetic connectivity of individuals and populations. In this manuscript, we focus on two methodological extensions for landscape genetic analyses: the use of conditional genetic distance (cGD) derived from population networks and the utility of extracting potentially confounding effects caused by correlations between phylogeographic history and contemporary ecological factors. Individual-based simulations show that when describing the spatial distribution of genetic variation, cGD consistently outperforms the traditional genetic distance measure of linearized F(ST) under both 1- and 2-dimensional stepping stone models and Cavalli-Sforza and Edward's chord distance D(c) in 1-dimensional landscapes. To show how to identify and extract the effects of phylogeographic history prior to embarking on landscape genetic analyses, we use nuclear genotypic data from the Sonoran desert succulent Euphorbia lomelii (Euphrobiaceae), for which a detailed phylogeographic history has previously been determined. For E. lomelii, removing the effect of phylogeographic history significantly influences our ability to infer both the identity and the relative importance of spatial and bio-climatic variables in subsequent landscape genetic analyses. We close by discussing the utility of cGD in landscape genetic analyses.

Inference of population history by coupling exploratory and model-driven phylogeographic analyses.

Understanding the nature, timing and geographic context of historical events and population processes that shaped the spatial distribution of genetic diversity is critical for addressing questions relating to speciation, selection, and applied conservation management. Cladistic analysis of gene trees has been central to phylogeography, but when coupled with approaches that make use of different components of the information carried by DNA sequences and their frequencies, the strength and resolution of these inferences can be improved. However, assessing concordance of inferences drawn using different analytical methods or genetic datasets, and integrating their outcomes, can be challenging. Here we overview the strengths and limitations of different types of genetic data, analysis methods, and approaches to historical inference. We then turn our attention to the potentially synergistic interactions among widely-used and emerging phylogeographic analyses, and discuss some of the ways that spatial and temporal concordance among inferences can be assessed. We close this review with a brief summary and outlook on future research directions.

Correction: Hyseni, C.; Garrick, R.C. Ecological Drivers of Species Distributions and Niche Overlap for Three Subterranean Termite Species in the Southern Appalachian Mountains, USA. Insects 2019, 10, 33.

It has recently come to our attention that two of the environmental rasters we used for analyses inour study [1] were mislabeled in a raster processing pipeline [...].

Ecological Drivers of Species Distributions and Niche Overlap for Three Subterranean Termite Species in the Southern Appalachian Mountains, USA.

In both managed and unmanaged forests, termites are functionally important members of the dead-wood-associated (saproxylic) insect community. However, little is known about regional-scale environmental drivers of geographic distributions of termite species, and how these environmental factors impact co-occurrence among congeneric species. Here we focus on the southern Appalachian Mountains-a well-known center of endemism for forest biota-and use Ecological Niche Modeling (ENM) to examine the distributions of three species of Reticulitermes termites (i.e., R. flavipes, R. virginicus, and R. malletei). To overcome deficiencies in public databases, ENMs were underpinned by field-collected high-resolution occurrence records coupled with molecular taxonomic species identification. Spatial overlap among areas of predicted occurrence of each species was mapped, and aspects of niche similarity were quantified. We also identified environmental factors that most strongly contribute to among-species differences in occupancy. Overall, we found that R. flavipes and R. virginicus showed significant niche divergence, which was primarily driven by dry-season precipitation. Also, all three species were most likely to co-occur in the mid-latitudes of the study area (i.e., northern Alabama and Georgia, eastern Tennessee and western North Carolina), which is an area of considerable topographic complexity. This work provides important baseline information for follow-up studies of local-scale drivers of these species' distributions. It also identifies specific geographic areas where future assessments of the frequency of true syntopy vs. micro-allopatry, and associated interspecific competitive interactions, should be focused.

The role of glacial-interglacial climate change in shaping the genetic structure of eastern subterranean termites in the southern Appalachian Mountains, USA.

The eastern subterranean termite, Reticulitermes flavipes, currently inhabits previously glaciated regions of the northeastern U.S., as well as the unglaciated southern Appalachian Mountains and surrounding areas. We hypothesized that Pleistocene climatic fluctuations have influenced the distribution of R. flavipes, and thus the evolutionary history of the species. We estimated contemporary and historical geographic distributions of R. flavipes by constructing Species Distribution Models (SDM). We also inferred the evolutionary and demographic history of the species using mitochondrial (cytochrome oxidase I and II) and nuclear (endo-beta-1,4-glucanase) DNA sequence data. To do this, genetic populations were delineated using Bayesian spatial-genetic clustering, competing hypotheses about population divergence were assessed using approximate Bayesian computation (ABC), and changes in population size were estimated using Bayesian skyline plots. SDMs identified areas in the north with suitable habitat during the transition from the Last Interglacial to the Last Glacial Maximum, as well as an expanding distribution from the mid-Holocene to the present. Genetic analyses identified three geographically cohesive populations, corresponding with northern, central, and southern portions of the study region. Based on ABC analyses, divergence between the Northern and Southern populations was the oldest, estimated to have occurred 64.80 thousand years ago (kya), which corresponds with the timing of available habitat in the north. The Central and Northern populations diverged in the mid-Holocene, 8.63 kya, after which the Central population continued to expand. Accordingly, phylogeographic patterns of R. flavipes in the southern Appalachians appear to have been strongly influenced by glacial-interglacial climate change. OPEN RESEARCH BADGES: This article has been awarded Open Materials, Open Data Badges. All materials and data are publicly accessible via the Open Science Framework at https://doi.org/10.5061/dryad.5hr7f31.

Two Divergent Genetic Lineages within the Horned Passalus Beetle, Odontotaenius disjunctus (Coleoptera: Passalidae): An Emerging Model for Insect Behavior, Physiology, and Microbiome Research.

The horned passalus (Odontotaenius disjunctus) is one of the most extensively studied saproxylic beetles in the eastern United States. For several decades this species has been the subject of investigations into the behaviors associated with subsociality as well as physiological responses to stress, and, most recently, the composition of its gut microbiome has been closely examined. However, no published study to date has characterized this beetle's broad-scale population genetic structure. Here, we conducted intensive geographic sampling throughout the southern Appalachian Mountains and surrounding areas and then assessed mitochondrial DNA (mtDNA) sequence variation among individuals. Unexpectedly, we discovered two divergent, yet broadly sympatric, mtDNA clades. Indeed, the magnitude of divergence between- vs. within-clades ranged from 5.9 to 7.5×, depending on the dataset under consideration, and members of the two lineages were often syntopic (i.e., found in the same rotting log). Given the potential implications for past and future studies on behavior, physiology, and the gut microbiome, we developed a simple cost-efficient molecular assay (i.e., polymerase chain reaction restriction fragment length polymorphism; PCR-RFLP) to rapidly determine mtDNA clade membership of O. disjunctus individuals. We suggest that the evolutionary processes that gave rise to the emergence and persistence of divergent sympatric lineages reported here warrant investigation, as this type of spatial-genetic pattern appears to be rare among southern Appalachian forest invertebrates.

Giant tortoise genomes provide insights into longevity and age-related disease.

Giant tortoises are among the longest-lived vertebrate animals and, as such, provide an excellent model to study traits like longevity and age-related diseases. However, genomic and molecular evolutionary information on giant tortoises is scarce. Here, we describe a global analysis of the genomes of Lonesome George-the iconic last member of Chelonoidis abingdonii-and the Aldabra giant tortoise (Aldabrachelys gigantea). Comparison of these genomes with those of related species, using both unsupervised and supervised analyses, led us to detect lineage-specific variants affecting DNA repair genes, inflammatory mediators and genes related to cancer development. Our study also hints at specific evolutionary strategies linked to increased lifespan, and expands our understanding of the genomic determinants of ageing. These new genome sequences also provide important resources to help the efforts for restoration of giant tortoise populations.