Broken Ring Speciation in California Mygalomorph Spiders (Nemesiidae, Calisoga).

AbstractIdealized ring species, with approximately continuous gene flow around a geographic barrier but singular reproductive isolation at a ring terminus, are rare in nature. A broken ring species model preserves the geographic setting and fundamental features of an idealized model but accommodates varying degrees of gene flow restriction over complex landscapes through evolutionary time. Here we examine broken ring species dynamics in Calisoga spiders, which, like the classic ring species Ensatina salamanders, are distributed around the Central Valley of California. Using nuclear and mitogenomic data, we test key predictions of common ancestry, ringlike biogeography, biogeographic timing, population connectivity, and terminal overlap. We show that a ring complex of populations shares a single common ancestor, and from an ancestral area in the Sierra Nevada mountains, two distributional and phylogenomic arms encircle the Central Valley. Isolation by distance occurs along these distributional arms, although gene flow restriction is also evident. Where divergent lineages meet in the South Coast Ranges, we find rare lineage sympatry, without evidence for nuclear gene flow and with clear evidence for morphological and ecological divergence. We discuss general insights provided by broken ring species and how such a model could be explored and extended in other systems and future studies.

Microgeographic population structuring in a genus of California trapdoor spiders and discovery of an enigmatic new species (Euctenizidae: Promyrmekiaphila korematsui sp. nov.).

The recognition and delineation of cryptic species remains a perplexing problem in systematics, evolution, and species delimitation. Once recognized as such, cryptic species complexes provide fertile ground for studying genetic divergence within the context of phenotypic and ecological divergence (or lack thereof). Herein we document the discovery of a new cryptic species of trapdoor spider, Promyrmekiaphila korematsui sp. nov. Using subgenomic data obtained via target enrichment, we document the phylogeography of the California endemic genus Promyrmekiaphila and its constituent species, which also includes P. clathrata and P. winnemem. Based on these data we show a pattern of strong geographic structuring among populations but cannot entirely discount recent gene flow among populations that are parapatric, particularly for deeply diverged lineages within P. clathrata. The genetic data, in addition to revealing a new undescribed species, also allude to a pattern of potential phenotypic differentiation where species likely come into close contact. Alternatively, phenotypic cohesion among genetically divergent P. clathrata lineages suggests that some level of gene flow is ongoing or occurred in the recent past. Despite considerable field collection efforts over many years, additional sampling in potential zones of contact for both species and lineages is needed to completely resolve the dynamics of divergence in Promyrmekiaphila at the population-species interface.

Increasing Information Content and Diagnosability in Family-Level Classifications.

Higher-level classifications often must account for monotypic taxa representing depauperate evolutionary lineages and lacking synapomorphies of their better-known, well-defined sister clades. In a ranked (Linnean) or unranked (phylogenetic) classification system, discovering such a depauperate taxon does not necessarily invalidate the rank classification of sister clades. Named higher taxa must be monophyletic to be phylogenetically valid. Ranked taxa above the species level should also maximize information content, diagnosability, and utility (e.g., in biodiversity conservation). In spider classification, families are the highest rank that is systematically catalogued, and incertae sedis is not allowed. Consequently, it is important that family-level taxa be well defined and informative. We revisit the classification problem of Orbipurae, an unranked suprafamilial clade containing the spider families Nephilidae, Phonognathidae, and Araneidae sensu stricto. We argue that, to maximize diagnosability, information content, conservation utility, and practical taxonomic considerations, this "splitting" scheme is superior to its recently proposed alternative, which lumps these families together as Araneidae sensu lato. We propose to redefine Araneidae and recognize a monogeneric spider family, Paraplectanoididae fam. nov. to accommodate the depauperate lineage Paraplectanoides. We present new subgenomic data to stabilize Orbipurae topology which also supports our proposed family-level classification. Our example from spiders demonstrates why classifications must be able to accommodate depauperate evolutionary lineages, for example, Paraplectanoides. Finally, although clade age should not be a criterion to determine rank, other things being equal, comparable ages of similarly ranked taxa do benefit comparative biology. [Classification, family rank, phylogenomics, systematics, monophyly, spider phylogeny.].

Phylogeography and cohesion species delimitation of California endemic trapdoor spiders within the Aptostichus icenoglei sibling species complex (Araneae: Mygalomorphae: Euctenizidae).

Species delimitation is an imperative first step toward understanding Earth's biodiversity, yet what constitutes a species and the relative importance of the various processes by which new species arise continue to be debatable. Species delimitation in spiders has traditionally used morphological characters; however, certain mygalomorph spiders exhibit morphological homogeneity despite long periods of population-level isolation, absence of gene flow, and consequent high degrees of molecular divergence. Studies have shown strong geographic structuring and significant genetic divergence among several species complexes within the trapdoor spider genus Aptostichus, most of which are restricted to the California Floristic Province (CAFP) biodiversity hotspot. Specifically, the Aptostichus icenoglei complex, which comprises the three sibling species, A. barackobamai, A. isabella, and A. icenoglei, exhibits evidence of cryptic mitochondrial DNA diversity throughout their ranges in Northern, Central, and Southern California. Our study aimed to explicitly test species hypotheses within this assemblage by implementing a cohesion species-based approach. We used genomic-scale data (ultraconserved elements, UCEs) to first evaluate genetic exchangeability and then assessed ecological interchangeability of genetic lineages. Biogeographical analysis was used to assess the likelihood of dispersal versus vicariance events that may have influenced speciation pattern and process across the CAFP's complex geologic and topographic landscape. Considering the lack of congruence across data types and analyses, we take a more conservative approach by retaining species boundaries within A. icenoglei.

Light Environment Interacts with Visual Displays in a Species-Specific Manner in Multimodal-Signaling Wolf Spiders.

AbstractLight availability is highly variable, yet predictable, over various timescales and is expected to play an important role in the evolution of visual signals. Courtship displays of the wolf spider genus Schizocosa always involve the use of substrate-borne vibrations; however, there is substantial variation in the presence and complexity of visual displays among species. To gain insight into the role the light environment plays in the evolution of courtship displays, we tested the function of visual courtship signaling across distinct light environments in four species of Schizocosa that vary in their degree of ornamentation and dynamic visual signals. We ran mating and courtship trials at three light intensities (bright, dim, and dark) and tested the hypothesis that ornamentation interacts with light environment. We also examined each species' circadian activity patterns. The effects of the light environment on courtship and mating varied between species, as did circadian activity patterns. Our results suggest that femur pigmentation may have evolved for diurnal signaling, whereas tibial brushes may function to increase signal efficacy under dim light. Additionally, we found evidence for light-dependent changes in selection on male traits, illustrating that short-term changes in light intensity have the potential for strong effects on the dynamics of sexual selection.

Biogeography and eye size evolution of the ogre-faced spiders.

Net-casting spiders (Deinopidae) comprise a charismatic family with an enigmatic evolutionary history. There are 67 described species of deinopids, placed among three genera, Deinopis, Menneus, and Asianopis, that are distributed globally throughout the tropics and subtropics. Deinopis and Asianopis, the ogre-faced spiders, are best known for their giant light-capturing posterior median eyes (PME), whereas Menneus does not have enlarged PMEs. Molecular phylogenetic studies have revealed discordance between morphology and molecular data. We employed a character-rich ultra-conserved element (UCE) dataset and a taxon-rich cytochrome-oxidase I (COI) dataset to reconstruct a genus-level phylogeny of Deinopidae, aiming to investigate the group's historical biogeography, and examine PME size evolution. Although the phylogenetic results support the monophyly of Menneus and the single reduction of PME size in deinopids, these data also show that Deinopis is not monophyletic. Consequently, we formally transfer 24 Deinopis species to Asianopis; the transfers comprise all of the African, Australian, South Pacific, and a subset of Central American and Mexican species. Following the divergence of Eastern and Western deinopids in the Cretaceous, Deinopis/Asianopis dispersed from Africa, through Asia and into Australia with its biogeographic history reflecting separation of Western Gondwana as well as long-distance dispersal events.

Using natural history to guide supervised machine learning for cryptic species delimitation with genetic data.

The diversity of biological and ecological characteristics of organisms, and the underlying genetic patterns and processes of speciation, makes the development of universally applicable genetic species delimitation methods challenging. Many approaches, like those incorporating the multispecies coalescent, sometimes delimit populations and overestimate species numbers. This issue is exacerbated in taxa with inherently high population structure due to low dispersal ability, and in cryptic species resulting from nonecological speciation. These taxa present a conundrum when delimiting species: analyses rely heavily, if not entirely, on genetic data which over split species, while other lines of evidence lump. We showcase this conundrum in the harvester Theromaster brunneus, a low dispersal taxon with a wide geographic distribution and high potential for cryptic species. Integrating morphology, mitochondrial, and sub-genomic (double-digest RADSeq and ultraconserved elements) data, we find high discordance across analyses and data types in the number of inferred species, with further evidence that multispecies coalescent approaches over split. We demonstrate the power of a supervised machine learning approach in effectively delimiting cryptic species by creating a "custom" training data set derived from a well-studied lineage with similar biological characteristics as Theromaster. This novel approach uses known taxa with particular biological characteristics to inform unknown taxa with similar characteristics, using modern computational tools ideally suited for species delimitation. The approach also considers the natural history of organisms to make more biologically informed species delimitation decisions, and in principle is broadly applicable for taxa across the tree of life.

Phylogeny and secondary sexual trait evolution in Schizocosa wolf spiders (Araneae, Lycosidae) shows evidence for multiple gains and losses of ornamentation and species delimitation uncertainty.

Members of the Nearctic spider genus Schizocosa Chamberlin, 1904 have garnered much attention in behavioral studies and over many decades, a number of species have developed as model systems for investigating patterns of sexual selection and multimodal communication. Many of these studies have employed a comparative approach using putative, but not rigorously tested, sister species pairs that have distinctive morphological traits and attendant behaviors. Despite past emphasis on the efficacy of these presumably comparative-based studies of closely related species, generating a robust phylogenetic hypothesis for Schizocosa has been an ongoing challenge. Here, we apply a phylogenomic approach using anchored hybrid enrichment to generate a data set comprising over 400 loci representing a comprehensive taxonomic sample of 23 Nearctic Schizocosa. Our sampling also includes numerous outgroup lycosid genera that allow for a robust evaluation of genus monophyly. Based on analyses using concatenation and coalescent-based methods, we recover a well-supported phylogeny that infers the following: 1) The New World Schizocosa do not form a monophyletic group; 2) Previous hypotheses of North American species require reconsideration along with the composition of species groups; 3) Multiple longstanding model species are not genealogically exclusive and thus are not "good" species; 4) This updated phylogenetic framework establishes a new working paradigm for studying the evolution of characters associated with reproductive communication and mating. Ancestral character state reconstructions show a complex pattern of homoplasy that has likely obfuscated previous attempts to reconstruct relationships and delimit species. Important characters presumably related to sexual selection, such as foreleg pigmentation and dense bristle formation, have undergone repeated gain and loss events, many of which have led to increased morphological divergence between sister-species. Evaluation of these traits in a comparative framework illuminates how sexual selection and natural selection influence character evolution and provides a model for future studies of multimodal communication evolution and function.

Sister species diverge in modality-specific courtship signal form and function.

Understanding the relative importance of different sources of selection (e.g., the environment, social/sexual selection) on the divergence or convergence of reproductive communication can shed light on the origin, maintenance, or even disappearance of species boundaries. Using a multistep approach, we tested the hypothesis that two presumed sister species of wolf spider with overlapping ranges and microhabitat use, yet differing degrees of sexual dimorphism, have diverged in their reliance on modality-specific courtship signaling. We predicted that male Schizocosa crassipalpata (no ornamentation) rely predominantly on diet-dependent vibratory signaling for mating success. In contrast, we predicted that male S. bilineata (black foreleg brushes) rely on diet-dependent visual signaling. We first tested and corroborated the sister-species relationship between S. crassipalpata and S. bilineata using phylogenomic scale data. Next, we tested for species-specific, diet-dependent vibratory and visual signaling by manipulating subadult diet and subsequently quantifying adult morphology and mature male courtship signals. As predicted, vibratory signal form was diet-dependent in S. crassipalpata, while visual ornamentation (brush area) was diet-dependent in S. bilineata. We then compared the species-specific reliance on vibratory and visual signaling by recording mating across artificially manipulated signaling environments (presence/absence of each modality in a 2 × 2 full factorial design). In accordance with our diet dependence results for S. crassipalpata, the presence of vibratory signaling was important for mating success. In contrast, the light and vibratory environment interacted to influence mating success in S. bilineata, with vibratory signaling being important only in the absence of light. We found no differences in overall activity patterns. Given that these species overlap in much of their range and microhabitat use, we suggest that competition for signaling space may have led to the divergence and differential use of sensory modalities between these sister species.

Integrative species delimitation reveals cryptic diversity in the southern Appalachian Antrodiaetus unicolor (Araneae: Antrodiaetidae) species complex.

Although species delimitation can be highly contentious, the development of reliable methods to accurately ascertain species boundaries is an imperative step in cataloguing and describing Earth's quickly disappearing biodiversity. Spider species delimitation remains largely based on morphological characters; however, many mygalomorph spider populations are morphologically indistinguishable from each other yet have considerable molecular divergence. The focus of our study, the Antrodiaetus unicolor species complex containing two sympatric species, exhibits this pattern of relative morphological stasis with considerable genetic divergence across its distribution. A past study using two molecular markers, COI and 28S, revealed that A. unicolor is paraphyletic with respect to A. microunicolor. To better investigate species boundaries in the complex, we implement the cohesion species concept and use multiple lines of evidence for testing genetic exchangeability and ecological interchangeability. Our integrative approach includes extensively sampling homologous loci across the genome using a RADseq approach (3RAD), assessing population structure across their geographic range using multiple genetic clustering analyses that include structure, principal components analysis and a recently developed unsupervised machine learning approach (Variational Autoencoder). We evaluate ecological similarity by using large-scale ecological data for niche-based distribution modelling. Based on our analyses, we conclude that this complex has at least one additional species as well as confirm species delimitations based on previous less comprehensive approaches. Our study demonstrates the efficacy of genomic-scale data for recognizing cryptic species, suggesting that species delimitation with one data type, whether one mitochondrial gene or morphology, may underestimate true species diversity in morphologically homogenous taxa with low vagility.

A stable phylogenomic classification of Travunioidea (Arachnida, Opiliones, Laniatores) based on sequence capture of ultraconserved elements.

Molecular phylogenetics has transitioned into the phylogenomic era, with data derived from next-generation sequencing technologies allowing unprecedented phylogenetic resolution in all animal groups, including understudied invertebrate taxa. Within the most diverse harvestmen suborder, Laniatores, most relationships at all taxonomic levels have yet to be explored from a phylogenomics perspective. Travunioidea is an early-diverging lineage of laniatorean harvestmen with a Laurasian distribution, with species distributed in eastern Asia, eastern and western North America, and south-central Europe. This clade has had a challenging taxonomic history, but the current classification consists of ~77 species in three families, the Travuniidae, Paranonychidae, and Nippononychidae. Travunioidea classification has traditionally been based on structure of the tarsal claws of the hind legs. However, it is now clear that tarsal claw structure is a poor taxonomic character due to homoplasy at all taxonomic levels. Here, we utilize DNA sequences derived from capture of ultraconserved elements (UCEs) to reconstruct travunioid relationships. Data matrices consisting of 317-677 loci were used in maximum likelihood, Bayesian, and species tree analyses. Resulting phylogenies recover four consistent and highly supported clades; the phylogenetic position and taxonomic status of the enigmatic genus Yuria is less certain. Based on the resulting phylogenies, a revision of Travunioidea is proposed, now consisting of the Travuniidae, Cladonychiidae, Paranonychidae (Nippononychidae is synonymized), and the new family Cryptomastridae Derkarabetian & Hedin, fam. n., diagnosed here. The phylogenetic utility and diagnostic features of the intestinal complex and male genitalia are discussed in light of phylogenomic results, and the inappropriateness of the tarsal claw in diagnosing higher-level taxa is further corroborated.

Cryptic elevational zonation in trapdoor spiders (Araneae, Antrodiaetidae, Aliatypus janus complex) from the California southern Sierra Nevada.

The relative roles of ecological niche conservatism versus niche divergence in promoting montane speciation remains an important topic in biogeography. Here, our aim was to test whether lineage diversification in a species complex of trapdoor spiders corresponds with riverine barriers or with an ecological gradient associated with elevational tiering. Aliatypus janus was sampled from throughout its range, with emphasis on populations in the southern Sierra Nevada Mountains of California. We collected multi-locus genetic data to generate a species tree for A. janus and its close relatives. Coalescent based hypothesis tests were conducted to determine if genetic breaks within A. janus conform to riverine barriers. Ecological niche models (ENM) under current and Last Glacial Maximum (LGM) conditions were generated and hypothesis tests of niche conservatism and divergence were performed. Coalescent analyses reveal deeply divergent genetic lineages within A. janus, likely corresponding to cryptic species. Two primary lineages meet along an elevational gradient on the western slopes of the southern Sierra Nevada Mountains. ENMs under both current and LGM conditions indicate that these groups occupy largely non-overlapping niches. ENM hypothesis testing rejected niche identity between the two groups, and supported a sharp ecological gradient occurring where the groups meet. However, the niche similarity test indicated that the two groups may not inhabit different background niches. The Sierra Nevada Mountains provide a natural laboratory for simultaneously testing ecological niche divergence and conservatism and their role in speciation across a diverse range of taxa. Aliatypus janus represents a species complex with cryptic lineages that may have diverged due to parapatric speciation along an ecological gradient, or been maintained by the evolution of ecological niche differences following allopatric speciation.

Inactivation of thermogenic UCP1 as a historical contingency in multiple placental mammal clades.

Mitochondrial uncoupling protein 1 (UCP1) is essential for nonshivering thermogenesis in brown adipose tissue and is widely accepted to have played a key thermoregulatory role in small-bodied and neonatal placental mammals that enabled the exploitation of cold environments. We map ucp1 sequences from 133 mammals onto a species tree constructed from a ~51-kb sequence alignment and show that inactivating mutations have occurred in at least 8 of the 18 traditional placental orders, thereby challenging the physiological importance of UCP1 across Placentalia. Selection and timetree analyses further reveal that ucp1 inactivations temporally correspond with strong secondary reductions in metabolic intensity in xenarthrans and pangolins, or in six other lineages coincided with a ~30 million-year episode of global cooling in the Paleogene that promoted sharp increases in body mass and cladogenesis evident in the fossil record. Our findings also demonstrate that members of various lineages (for example, cetaceans, horses, woolly mammoths, Steller's sea cows) evolved extreme cold hardiness in the absence of UCP1-mediated thermogenesis. Finally, we identify ucp1 inactivation as a historical contingency that is linked to the current low species diversity of clades lacking functional UCP1, thus providing the first evidence for species selection related to the presence or absence of a single gene product.

Comparative performance of double-digest RAD sequencing across divergent arachnid lineages.

Next-generation sequencing technologies now allow researchers of non-model systems to perform genome-based studies without the requirement of a (often unavailable) closely related genomic reference. We evaluated the role of restriction endonuclease (RE) selection in double-digest restriction-site-associated DNA sequencing (ddRADseq) by generating reduced representation genome-wide data using four different RE combinations. Our expectation was that RE selections targeting longer, more complex restriction sites would recover fewer loci than RE with shorter, less complex sites. We sequenced a diverse sample of non-model arachnids, including five congeneric pairs of harvestmen (Opiliones) and four pairs of spiders (Araneae). Sample pairs consisted of either conspecifics or closely related congeneric taxa, and in total 26 sample pair analyses were tested. Sequence demultiplexing, read clustering and variant calling were performed in the pyRAD program. The 6-base pair cutter EcoRI combined with methylated site-specific 4-base pair cutter MspI produced, on average, the greatest numbers of intra-individual loci and shared loci per sample pair. As expected, the number of shared loci recovered for a sample pair covaried with the degree of genetic divergence, estimated with cytochrome oxidase I sequences, although this relationship was non-linear. Our comparative results will prove useful in guiding protocol selection for ddRADseq experiments on many arachnid taxa where reference genomes, even from closely related species, are unavailable.

High phylogenetic utility of an ultraconserved element probe set designed for Arachnida.

Arachnida is an ancient, diverse and ecologically important animal group that contains a number of species of interest for medical, agricultural and engineering applications. Despite their importance, many aspects of the arachnid tree of life remain unresolved, hindering comparative approaches to arachnid biology. Biologists have made considerable efforts to resolve the arachnid phylogeny; yet, limited and challenging morphological characters, as well as a dearth of genetic resources, have hindered progress. Here, we present a genomic toolkit for arachnids featuring hundreds of conserved DNA regions (ultraconserved elements or UCEs) that allow targeted sequencing of any species in the arachnid tree of life. We used recently developed capture probes designed from conserved regions of available arachnid genomes to enrich a sample of loci from 32 diverse arachnids. Sequence capture returned an average of 487 UCE loci for all species, with a range from 170 to 722. Phylogenetic analysis of these UCEs produced a highly resolved arachnid tree with relationships largely consistent with recent transcriptome-based phylogenies. We also tested the phylogenetic informativeness of UCE probes within the spider, scorpion and harvestman orders, demonstrating the utility of these markers at shallower taxonomic scales and suggesting that these loci will be useful for species-level differences. This probe set will open the door to phylogenomic and population genomic studies across the arachnid tree of life, enabling systematics, species delimitation, species discovery and conservation of these diverse arthropods.

Population genomic evidence for multiple Pliocene refugia in a montane-restricted harvestman (Arachnida, Opiliones, Sclerobunus robustus) from the southwestern United States.

The integration of ecological niche modelling into phylogeographic analyses has allowed for the identification and testing of potential refugia under a hypothesis-based framework, where the expected patterns of higher genetic diversity in refugial populations and evidence of range expansion of nonrefugial populations are corroborated with empirical data. In this study, we focus on a montane-restricted cryophilic harvestman, Sclerobunus robustus, distributed throughout the heterogeneous Southern Rocky Mountains and Intermontane Plateau of southwestern North America. We identified hypothetical refugia using ecological niche models (ENMs) across three time periods, corroborated these refugia with population genetic methods using double-digest RAD-seq data and conducted population-level phylogenetic and divergence dating analyses. ENMs identify two large temporally persistent regions in the mid-latitude highlands. Genetic patterns support these two hypothesized refugia with higher genetic diversity within refugial populations and evidence for range expansion in populations found outside hypothesized refugia. Phylogenetic analyses identify five to six genetically divergent, geographically cohesive clades of S. robustus. Divergence dating analyses suggest that these separate refugia date to the Pliocene and that divergence between clades pre-dates the late Pleistocene glacial cycles, while diversification within clades was likely driven by these cycles. Population genetic analyses reveal effects of both isolation by distance (IBD) and isolation by environment (IBE), with IBD more important in the continuous mountainous portion of the distribution, while IBE was stronger in the populations inhabiting the isolated sky islands of the south. Using model-based coalescent approaches, we find support for postdivergence migration between clades from separate refugia.

A new monster from southwest Oregon forests: Cryptomaster behemoth sp. n. (Opiliones, Laniatores, Travunioidea).

The monotypic genus Cryptomaster Briggs, 1969 was described based on individuals from a single locality in southwestern Oregon. The described species Cryptomaster leviathan Briggs, 1969 was named for its large body size compared to most travunioid Laniatores. However, as the generic name suggests, Cryptomaster are notoriously difficult to find, and few subsequent collections have been recorded for this genus. Here, we increase sampling of Cryptomaster to 15 localities, extending their known range from the Coast Range northeast to the western Cascade Mountains of southern Oregon. Phylogenetic analyses of mitochondrial and nuclear DNA sequence data reveal deep phylogenetic breaks consistent with independently evolving lineages. We use discovery and validation species delimitation approaches to generate and test species hypotheses, including a coalescent species delimitation method to test multi-species hypotheses. For delimited species, we use light microscopy and SEM to discover diagnostic morphological characters. Although Cryptomaster has a small geographic distribution, this taxon is consistent with other short-range endemics in having deep phylogenetic breaks indicative of species level divergences. Herein we describe Cryptomaster behemoth sp. n., and provide morphological diagnostic characters for identifying Cryptomaster leviathan and Cryptomaster behemoth.

Inactivation of C4orf26 in toothless placental mammals.

Previous studies have reported inactivated copies of six enamel-related genes (AMBN, AMEL, AMTN, ENAM, KLK4, MMP20) and one dentin-related gene (DSPP) in one or more toothless vertebrates and/or vertebrates with enamelless teeth, thereby providing evidence that these genes are enamel or tooth-specific with respect to their critical functions that are maintained by natural selection. Here, we employ available genome sequences for edentulous and enamelless mammals to evaluate the enamel specificity of four genes (WDR72, SLC24A4, FAM83H, C4orf26) that have been implicated in amelogenesis imperfecta, a condition in which proper enamel formation is abrogated during tooth development. Coding sequences for WDR72, SCL24A4, and FAM83H are intact in four edentulous taxa (Chinese pangolin, three baleen whales) and three taxa (aardvark, nine-banded armadillo, Hoffmann's two-toed sloth) with enamelless teeth, suggesting that these genes have critical functions beyond their involvement in tooth development. By contrast, genomic data for C4orf26 reveal inactivating mutations in pangolin and bowhead whale as well as evidence for deletion of this gene in two minke whale species. Hybridization capture of exonic regions and PCR screens provide evidence for inactivation of C4orf26 in eight additional baleen whale species. However, C4orf26 is intact in all three species with enamelless teeth that were surveyed, as well as in 95 additional mammalian species with enamel-capped teeth. Estimates of selection intensity suggest that dN/dS ratios on branches leading to taxa with enamelless teeth are similar to the dN/dS ratio on branches leading to taxa with enamel-capped teeth. Based on these results, we conclude that C4orf26 is tooth-specific, but not enamel-specific, with respect to its essential functions that are maintained by natural selection. A caveat is that an alternative splice site variant, which translates exon 3 in a different reading frame, is putatively functional in Catarrhini and may have evolved an additional role in this primate clade.

Multilocus sequence data reveal dozens of putative cryptic species in a radiation of endemic Californian mygalomorph spiders (Araneae, Mygalomorphae, Nemesiidae).

We use mitochondrial and multi-locus nuclear DNA sequence data to infer both species boundaries and species relationships within California nemesiid spiders. Higher-level phylogenetic data show that the California radiation is monophyletic and distantly related to European members of the genus Brachythele. As such, we consider all California nemesiid taxa to belong to the genus Calisoga Chamberlin, 1937. Rather than find support for one or two taxa as previously hypothesized, genetic data reveal Calisoga to be a species-rich radiation of spiders, including perhaps dozens of species. This conclusion is supported by multiple mitochondrial barcoding analyses, and also independent analyses of nuclear data that reveal general genealogical congruence. We discovered three instances of sympatry, and genetic data indicate reproductive isolation when in sympatry. An examination of female reproductive morphology does not reveal species-specific characters, and observed male morphological differences for a subset of putative species are subtle. Our coalescent species tree analysis of putative species lays the groundwork for future research on the taxonomy and biogeographic history of this remarkable endemic radiation.

Hemocyanin gene family evolution in spiders (Araneae), with implications for phylogenetic relationships and divergence times in the infraorder Mygalomorphae.

Hemocyanins are multimeric copper-containing hemolymph proteins involved in oxygen binding and transport in all major arthropod lineages. Most arachnids have seven primary subunits (encoded by paralogous genes a-g), which combine to form a 24-mer (4×6) quaternary structure. Within some spider lineages, however, hemocyanin evolution has been a dynamic process with extensive paralog duplication and loss. We have obtained hemocyanin gene sequences from numerous representatives of the spider infraorders Mygalomorphae and Araneomorphae in order to infer the evolution of the hemocyanin gene family and estimate spider relationships using these conserved loci. Our hemocyanin gene tree is largely consistent with the previous hypotheses of paralog relationships based on immunological studies, but reveals some discrepancies in which paralog types have been lost or duplicated in specific spider lineages. Analyses of concatenated hemocyanin sequences resolved deep nodes in the spider phylogeny and recovered a number of clades that are supported by other molecular studies, particularly for mygalomorph taxa. The concatenated data set is also used to estimate dates of higher-level spider divergences and suggests that the diversification of extant mygalomorphs preceded that of extant araneomorphs. Spiders are diverse in behavior and respiratory morphology, and our results are beneficial for comparative analyses of spider respiration. Lastly, the conserved hemocyanin sequences allow for the inference of spider relationships and ancient divergence dates.