The genomic signatures of evolutionary stasis.

Evolutionary stasis characterizes lineages that seldom speciate and show little phenotypic change over long stretches of geological time. Although lineages that appear to exhibit evolutionary stasis are often called living fossils, no single mechanism is thought to be responsible for their slow rates of morphological evolution and low species diversity. Some analyses of molecular evolutionary rates in a handful of living fossil lineages have indicated that these clades exhibit slow rates of genomic change. Here, we investigate mechanisms of evolutionary stasis using a dataset of 1,105 exons for 481 vertebrate species. We demonstrate that two ancient clades of ray-finned fishes classically called living fossils, gars and sturgeons, exhibit the lowest rates of molecular substitution in protein-coding genes among all jawed vertebrates. Comparably low rates of evolution are observed at fourfold degenerate sites in gars and sturgeons, implying a mechanism of stasis decoupled from selection that we speculate is linked to a highly effective DNA repair apparatus. We show that two gar species last sharing common ancestry over 100 million years ago produce morphologically intermediate and fertile hybrids in the wild. This makes gars the oldest naturally hybridizing divergence among eukaryotes and supports a theoretical prediction that slow rates of nucleotide substitution across the genome slow the accumulation of genetic incompatibilities, enabling hybridization across deeply divergent lineages and slowing the rate of speciation over geological timescales. Our results help establish molecular stasis as a barrier to speciation and phenotypic innovation and provide a mechanism to explain the low species diversity in living fossil lineages.

Lacustrine speciation associated with chromosomal inversion in a lineage of riverine fishes.

Geographic isolation is the primary driver of speciation in many vertebrate lineages. This trend is exemplified by North American darters, a clade of freshwater fishes where nearly all sister species pairs are allopatric and separated by millions of years of divergence. One of the only exceptions is the Lake Waccamaw endemic Etheostoma perlongum and its riverine sister species Etheostoma maculaticeps, which have no physical barriers to gene flow. Here we show that lacustrine speciation of E. perlongum is characterized by morphological and ecological divergence likely facilitated by a large chromosomal inversion. While E. perlongum is phylogenetically nested within the geographically widespread E. maculaticeps, there is a sharp genetic and morphological break coinciding with the lake-river boundary in the Waccamaw River system. Despite recent divergence, an active hybrid zone, and ongoing gene flow, analyses using a de novo reference genome reveal a 9 Mb chromosomal inversion with elevated divergence between E. perlongum and E. maculaticeps. This region exhibits striking synteny with known inversion supergenes in two distantly related fish lineages, suggesting deep evolutionary convergence of genomic architecture. Our results illustrate that rapid, ecological speciation with gene flow is possible even in lineages where geographic isolation is the dominant mechanism of speciation.

Phylogeography, hybridization, and species discovery in the Etheostoma nigrum complex (Percidae: Etheostoma: Boleosoma).

The history of riverine fish diversification is largely a product of geographic isolation. Physical barriers that reduce or eliminate gene flow between populations facilitate divergence via genetic drift and natural selection, eventually leading to speciation. For freshwater organisms, diversification is often the product of drainage basin rearrangements. In young clades where the history of isolation is the most recent, evolutionary relationships can resemble a tangled web. One especially recalcitrant group of freshwater fishes is the Johnny Darter (Etheostoma nigrum) species complex, where traditional taxonomy and molecular phylogenetics indicate a history of gene flow and conflicting inferences of species diversity. Here we assemble a genomic dataset using double digest restriction site associated DNA (ddRAD) sequencing and use phylogenomic and population genetic approaches to investigate the evolutionary history of the complex of species that includes E. nigrum, E. olmstedi, E. perlongum, and E. susanae. We reveal and validate several evolutionary lineages that we delimit as species, highlighting the need for additional work to formally describe the diversity of the Etheostoma nigrum complex. Our analyses also identify gene flow among recently diverged lineages, including one instance involving E. susanae, a localized and endangered species. Phylogeographic structure within the Etheostoma nigrum species complex coincides with major geologic events, such as parallel divergence in river basins during Pliocene inundation of the Atlantic coastal plain and multiple northward post-glacial colonization routes tracking river basin rearrangements. Our study serves as a nuanced example of how low dispersal rates coupled with geographic isolation among disconnected river systems in eastern North America has produced one of the world's freshwater biodiversity hotspots.

Chromosome-Level Genome Assembly of Chinese Sucker (Myxocyprinus asiaticus) Reveals Strongly Conserved Synteny Following a Catostomid-Specific Whole-Genome Duplication.

Fishes of the family Catostomidae ("suckers"; Teleostei: Cypriniformes) are hypothesized to have undergone an allopolyploidy event approximately 60 Ma. However, genomic evidence has previously been unavailable to assess this hypothesis. We sequenced and assembled the first chromosome-level catostomid genome, Chinese sucker (Myxocyprinus asiaticus), and present clear evidence of a catostomid-specific whole-genome duplication (WGD) event ("Cat-4R"). Our results reveal remarkably strong, conserved synteny since this duplication event, as well as between Myxocyprinus and an unduplicated outgroup, zebrafish (Danio rerio). Gene content and repetitive elements are also approximately evenly distributed across homeologous chromosomes, suggesting that both subgenomes retain some function, with no obvious bias in gene fractionation or subgenome dominance. The Cat-4R duplication provides another independent example of genome evolution following WGD in animals, in this case at the extreme end of conserved genome architecture over at least 25.2 Myr since the duplication. The M. asiaticus genome is a useful resource for researchers interested in understanding genome evolution following WGD in animals.

Accelerated Diversification Explains the Exceptional Species Richness of Tropical Characoid Fishes.

The Neotropics harbor the most species-rich freshwater fish fauna on the planet, but the timing of that exceptional diversification remains unclear. Did the Neotropics accumulate species steadily throughout their long history, or attain their remarkable diversity recently? Biologists have long debated the relative support for these museum and cradle hypotheses, but few phylogenies of megadiverse tropical clades have included sufficient taxa to distinguish between them. We used 1288 ultraconserved element loci spanning 293 species, 211 genera, and 21 families of characoid fishes to reconstruct a new, fossil-calibrated phylogeny and infer the most likely diversification scenario for a clade that includes a third of Neotropical fish diversity. This phylogeny implies paraphyly of the traditional delimitation of Characiformes because it resolves the largely Neotropical Characoidei as the sister lineage of Siluriformes (catfishes), rather than the African Citharinodei. Time-calibrated phylogenies indicate an ancient origin of major characoid lineages and reveal a much more recent emergence of most characoid species. Diversification rate analyses infer increased speciation and decreased extinction rates during the Oligocene at around 30 Ma during a period of mega-wetland formation in the proto-Orinoco-Amazonas. Three species-rich and ecomorphologically diverse lineages (Anostomidae, Serrasalmidae, and Characidae) that originated more than 60 Ma in the Paleocene experienced particularly notable bursts of Oligocene diversification and now account collectively for 68% of the approximately 2150 species of Characoidei. In addition to paleogeographic changes, we discuss potential accelerants of diversification in these three lineages. While the Neotropics accumulated a museum of ecomorphologically diverse characoid lineages long ago, this geologically dynamic region also cradled a much more recent birth of remarkable species-level diversity. [Biodiversity; Characiformes; macroevolution; Neotropics; phylogenomics; ultraconserved elements.].

Author Correction: Historical contingency shapes adaptive radiation in Antarctic fishes.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Revealing paraphyly and placement of extinct species within Epischura (Copepoda: Calanoida) using molecular data and quantitative morphometrics.

Epischura (Calanoida: Temoridae) is a Holarctic group of copepods serving important ecological roles, but it is difficult to study because of small range sizes of individual species and widespread distribution of the genus. This genus includes Tertiary relicts, some endemic to single, isolated lakes and can play major roles in unique ecosystems like Lakes Baikal and Tahoe. We present the first molecular and morphological analysis of Epischura that reveals their spatio-temporal evolutionary history. Morphological measurements of mandibles and genetics estimated phylogenetic relationships among all species represented in Epischura, including E. massachusettsensis, whose extinction status is of concern. Analyses used three gene regions for six previously unsequenced species to infer highly-resolved and well-supported phylogenies confirming a split between Siberian and North American species. Previously published age estimates and sequence data from broad taxonomic sampling of calanoid copepods estimated divergence times between the two Epischura groups. Divergence time estimates for Epischura were consistent with earlier molecular clock estimates and late-Miocene cooling events. Additionally, we provide the first taxonomically broad estimates of divergence times within Calanoida. The paraphyletic nature of the genus Epischura (and the family Temoridae) is apparent and requires the resurrection of the genus Epischurella (Smirnov, 1936) to describe the Siberian species.

Historical contingency shapes adaptive radiation in Antarctic fishes.

Adaptive radiation illustrates links between ecological opportunity, natural selection and the generation of biodiversity. Central to adaptive radiation is the association between a diversifying lineage and the evolution of phenotypic variation that facilitates the use of new environments or resources. However, is not clear whether adaptive evolution or historical contingency is more important for the origin of key phenotypic traits in adaptive radiation. Here we use targeted sequencing of >250,000 loci across 46 species to examine hypotheses concerning the origin and diversification of key traits in the adaptive radiation of Antarctic notothenioid fishes. Contrary to expectations of adaptive evolution, we show that notothenioids experienced a punctuated burst of genomic diversification and evolved key skeletal modifications before the onset of polar conditions in the Southern Ocean. We show that diversifying selection in pathways associated with human skeletal dysplasias facilitates ecologically important variation in buoyancy among Antarctic notothenioid species, and demonstrate the sufficiency of altered trip11, col1a2 and col1a1a function in zebrafish (Danio rerio) to phenocopy skeletal reduction in Antarctic notothenioids. Rather than adaptation being driven by the cooling of the Antarctic, our results highlight the role of historical contingency in shaping the adaptive radiation of notothenioids. Understanding the historical and environmental context for the origin of key traits in adaptive radiations extends beyond reconstructing events that result in evolutionary innovation, as it also provides a context in forecasting the effects of climate change on the stability and evolvability of natural populations.

Phylogenomic Signatures of Ancient Introgression in a Rogue Lineage of Darters (Teleostei: Percidae).

Evolutionary history is typically portrayed as a branching phylogenetic tree, yet not all evolution proceeds in a purely bifurcating manner. Introgressive hybridization is one process that results in reticulate evolution. Most known examples of genome-wide introgression occur among closely related species with relatively recent common ancestry; however, we present evidence for ancient hybridization and genome-wide introgression between major stem lineages of darters, a species-rich clade of North American freshwater fishes. Previous attempts to resolve the relationships of darters have been confounded by the uncertain phylogenetic resolution of the lineage Allohistium. In this study, we investigate the phylogenomics of darters, specifically the relationships of Allohistium, through analyses of approximately 30,000 RADseq loci sampled from 112 species. Our phylogenetic inferences are based on traditional approaches in combination with strategies that accommodate reticulate evolution. These analyses result in a novel phylogenetic hypothesis for darters that includes ancient introgression between Allohistium and other two major darter lineages, minimally occurring 20 million years ago. Darters offer a compelling case for the necessity of incorporating phylogenetic networks in reconstructing the evolutionary history of diversification in species-rich lineages. We anticipate that the growing wealth of genomic data for clades of non-model organisms will reveal more examples of ancient hybridization, eventually requiring a re-evaluation of how evolutionary history is visualized and utilized in macroevolutonary investigations.

Phylogenetic analysis of Antarctic notothenioids illuminates the utility of RADseq for resolving Cenozoic adaptive radiations.

Notothenioids are a clade of ∼120 species of marine fishes distributed in extreme southern hemisphere temperate near-shore habitats and in the Southern Ocean surrounding Antarctica. Over the past 25 years, molecular and morphological approaches have redefined hypotheses of relationships among notothenioid lineages as well as their relationships among major lineages of percomorph teleosts. These phylogenies provide a basis for investigation of mechanisms of evolutionary diversification within the clade and have enhanced our understanding of the notothenioid adaptive radiation. Despite extensive efforts, there remain several questions concerning the phylogeny of notothenioids. In this study, we deploy DNA sequences of ∼100,000 loci obtained using RADseq to investigate the phylogenetic relationships of notothenioids and to assess the utility of RADseq loci for lineages that exhibit divergence times ranging from the Paleogene to the Quaternary. The notothenioid phylogenies inferred from the RADseq loci provide unparalleled resolution and node support for several long-standing problems including, (1) relationships among species of Trematomus, (2) resolution of Indonotothenia cyanobrancha as the sister lineage of Trematomus, (3) the deep paraphyly of Nototheniidae, (4) the paraphyly of Lepidonotothen s.l., (5) paraphyly of Artedidraco, and 6) the monophyly of the Bathydraconidae. Assessment of site rates demonstrates that RADseq loci are similar to mtDNA protein coding genes and exhibit peak phylogenetic informativeness at the time interval during which the major Antarctic notothenioid lineages originated and diversified. In addition to providing a well-resolved phylogenetic hypothesis for notothenioids, our analyses quantify the predicted utility of RADseq loci for Cenozoic phylogenetic inferences.

A genomic assessment of species boundaries and hybridization in a group of highly polymorphic anoles (distichus species complex).

Delimiting young species is one of the great challenges of systematic biology, particularly when the species in question exhibit little morphological divergence. Anolis distichus, a trunk anole with more than a dozen subspecies that are defined primarily by dewlap color, may actually represent several independent evolutionary lineages. To test this, we utilized amplified fragment length polymorphisms (AFLP) genome scans and genetic clustering analyses in conjunction with a coalescent-based species delimitation method. We examined a geographically widespread set of samples and two heavily sampled hybrid zones. We find that genetic divergence is associated with a major biogeographic barrier, the Hispaniolan paleo-island boundary, but not with dewlap color. Additionally, we find support for hypotheses regarding colonization of two Hispaniolan satellite islands and the Bahamas from mainland Hispaniola. Our results show that A. distichus is composed of seven distinct evolutionary lineages still experiencing a limited degree of gene flow. We suggest that A. distichus merits taxonomic revision, but that dewlap color cannot be relied upon as the primary diagnostic character.

The role of heritable and dietary factors in the sexual signal of a Hispaniolan Anolis lizard, Anolis distichus.

The diversity of sexual signals is astounding, and divergence in these traits is believed to be associated with the early stages of speciation. An increasing number of studies also suggest a role for natural selection in driving signal divergence for effective transmission in heterogeneous environments. Both speciation and adaptive divergence, however, are contingent on the sexual signal being heritable, yet this often remains assumed and untested. It is particularly critical that the heritability of carotenoid-based sexual signals is investigated because such traits may instead be phenotypically plastic indicators of an individual's quality that exhibit no or little heritable variation. We present the first study to investigate the relative contribution of genetic and environmental factors to the striking diversity of dewlap color and pattern in Anolis lizards. Using a breeding experiment with Anolis distichus populations exhibiting different dewlap phenotypes, we raise F1 offspring in a common garden experiment to assess whether dewlap color is inherited. We follow this with carotenoid supplementation to investigate the influence of dietary pigments to dewlap color variation. We find significant differences in several aspects of dewlap color and pattern to persist to the F1 generation (fathers: N = 19; F1 males: N = 50; P < 0.01) with no change in dewlap phenotype with carotenoid supplementation (N = 52; P > 0.05). These results strongly support that genetic differences underlie dewlap color variation, thereby satisfying a key requirement of natural selection. Our findings provide an important stepping-stone to understanding the evolution of an incredibly diverse signal important for sexual selection and species recognition.