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Many animals utilize self-built structures – so-called extended phenotypes – to enhance body functions, such as thermoregulation, prey capture or defence. Yet, it is unclear whether the evolution of animal constructions supplements or substitutes body functions. Here, using Austral brown spiders, we explored if the evolutionary loss and gain of silken webs as extended prey capture devices correlates with alterations in traits known to play an important role in predatory strikes - locomotor performance and leg spination. For this purpose, we combined the reconstruction of the phylogeny of the Austral marronoid clade of spiders based on UCE target sequence capture with the assembly of kinematic, morphological and ecological data. We found that in this group extreme locomotor performance, with running speeds of over 100 body lengths per second, evolved repeatedly – both in web builders and cursorial spiders. There was no correlation with running speed, and leg spination only poorly correlated, relative to the use of extended phenotypes, with all of these traits showing highly mosaic, independent evolutionary patterns. This indicates that the use of webs does not reduce the selective pressure on body functions involved in prey capture and defence per se.

Leptonetidae are rarely encountered spiders, usually associated with caves and mesic habitats, and are disjunctly distributed across the Holarctic. Data from ultraconserved elements (UCEs) were used in concatenated and coalescent-based analyses to estimate the phylogenetic history of the family. Our taxon sample included close outgroups, and 90% of described leptonetid genera, with denser sampling in North America and Mediterranean Europe. Two data matrices were assembled and analysed; the first ‘relaxed’ matrix includes the maximum number of loci and the second ‘strict’ matrix is limited to the same set of core orthologs but with flanking introns mostly removed. A molecular dating analysis incorporating fossil and geological calibration points was used to estimate divergence times, and dispersal–extinction–cladogenesis analysis (DEC) was used to infer ancestral distributions. Analysis of both data matrices using maximum likelihood and coalescent-based methods supports the monophyly of Archoleptonetinae and Leptonetinae. However, relationships among Archoleptonetinae, Leptonetinae, and Austrochiloidea are poorly supported and remain unresolved. Archoleptonetinae is elevated to family rank Archoleptonetidae (new rank) and Leptonetidae (new status) is restricted to include only members of the subfamily Leptonetinae; a taxonomic review with morphological diagnoses is provided for both families. Four well supported lineages within Leptonetidae (new status) are recovered: (1) the Calileptoneta group, (2) the Leptoneta group, (3) the Paraleptoneta group, and (4) the Protoleptoneta group. Most genera within Leptonetidae are monophyletic, although Barusia, Cataleptoneta, and Leptoneta include misplaced species and require taxonomic revision. The origin of Archoleptonetidae (new rank), Leptonetidae, and the four main lineages within Leptonetidae date to the Cretaceous. DEC analysis infers the Leptoneta and Paraleptoneta groups to have ancestral distributions restricted to Mediterranean Europe, whereas the Calileptoneta and Protoleptoneta groups include genera with ancestral distributions spanning eastern and western North America, Mediterranean Europe, and east Asia. Based on a combination of biology, estimated divergence times, and inferred ancestral distributions we hypothesise that Leptonetidae was once widespread across the Holarctic and their present distributions are largely the result of vicariance. Given the wide disjunctions between taxa, we broadly interpret the family as a Holarctic relict fauna and hypothesise that they were once part of the Boreotropical forest ecosystem.

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.

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 dataset 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, and uses modern computational tools ideally suited for species delimitation while also considering the biology and natural history of organisms to make more biologically informed species delimitation decisions. In principle, this approach is universally applicable for species delimitation of any taxon with genetic data, particularly for cryptic species.

A new genus and species of troglobitic Opiliones, Sinonychia martensi gen. n. & sp. n., is described and illustrated. This is the first report of the superfamily Travunioidea Absolon & Kratochvíl, 1932 from China. The new species is assigned to Cladonychiidae Hadži, 1935 on the basis of morphology of midgut and male genitalia, and can be easily distinguished from related genera by several characters.

Sampling impediments and paucity of suitable material for molecular analyses have precluded the study of speciation and radiation of deep-sea species in Antarctica. We analyzed barcodes together with genome-wide single nucleotide polymorphisms obtained from double digestion restriction site-associated DNA sequencing (ddRADseq) for species in the family Antarctophilinidae. We also reevaluated the fossil record associated with this taxon to provide further insights into the origin of the group. Novel approaches to identify distinctive genetic lineages, including unsupervised machine learning variational autoencoder plots, were used to establish species hypothesis frameworks. In this sense, three undescribed species and a complex of cryptic species were identified, suggesting allopatric speciation connected to geographic or bathymetric isolation. We further observed that the shallow waters around the Scotia Arc and on the continental shelf in the Weddell Sea present high endemism and diversity. In contrast, likely due to the glacial pressure during the Cenozoic, a deep-sea group with fewer species emerged expanding over great areas in the South-Atlantic Antarctic Ridge. Our study agrees on how diachronic paleoclimatic and current environmental factors shaped Antarctic communities both at the shallow and deep-sea levels, promoting Antarctica as the center of origin for numerous taxa such as gastropod mollusks Harvard University | Ref. NSF DEB-1754278 Fundación Ramón Areces | Ref. beca postdoctoral National Science Foundation | Ref. NSF ANT-1043749

The Opiliones superfamily Triaenonychoidea currently includes two families, the monogeneric New Zealand–endemic Synthetonychiidae Forster, 1954 and Triaenonychidae Sørensen, 1886, a diverse family distributed mostly throughout the temperate Gondwanan terranes, with ~110 genera and ~500 species and subspecies currently described. Traditionally, Triaenonychidae has been divided into subfamilies diagnosed by very few morphological characters largely derived from the troublesome ‘Roewerian system’ of morphology, and classifications based on this system led to many complications. Recent research within Triaenonychoidea using morphology and traditional multilocus data has shown multiple deeply divergent lineages, non-monophyly of Triaenonychidae, and non-monophyly of subfamilies, necessitating a revision based on phylogenomic data. We used sequence capture of ultraconserved elements across 164 samples to create a 50% taxon occupancy matrix with 704 loci. Using phylogenomic and morphological examinations, we explored family-level relationships within Triaenonychoidea, including describing two new families: (1) Lomanellidae Mendes & Derkarabetian, fam. nov., consisting of Lomanella Pocock, 1903, and a newly described genus Abaddon Derkarabetian & Baker, gen. nov. with one species, A. despoliator Derkarabetian, sp. nov.; and (2) the elevation to family of Buemarinoidae Karaman, 2019, consisting of Buemarinoa Roewer, 1956, Fumontana Shear, 1977, Flavonuncia Lawrence, 1959, and a newly described genus Turonychus Derkarabetian, Prieto & Giribet, gen. nov., with one species, T. fadriquei Derkarabetian, Prieto & Giribet, sp. nov. With our dataset we also explored phylogenomic relationships within Triaenonychidae with an extensive taxon set including samples representing ~80% of the genus-level diversity. Based on our results we (1) discuss systematics of this family including the historical use of subfamilies, (2) reassess morphology in the context of our phylogeny, (3) hypothesise placement for all unsampled genera, (4) highlight lineages most in need of taxonomic revision, and (5) provide an updated species-level checklist. Aside from describing new taxa, our study provides the phylogenomic context necessary for future evolutionary and systematic research across this diverse lineage. ZooBank Registration: urn:lsid:zoobank.org:pub:81683834-98AB-43AA-B25A-C28C6A404F41

Montane species endemic to the "sky islands" of the North American southwest were significantly impacted by changing climates during the Pleistocene. We combined mitochondrial and genomic data with species distribution modelling to determine whether Aphonopelma marxi, a large tarantula from the nearby Colorado Plateau, was similarly impacted by glacial climates. Genetic analyses revealed that the species comprises three main clades that diverged in the Pleistocene. A clade distributed along the Mogollon Rim appears to have persisted in place during glacial conditions, whereas the other two clades probably colonized central and northeastern portions of the species' range from refugia in canyons. Climate models support this hypothesis for the Mogollon Rim, but late glacial climate data appear too coarse to detect suitable areas in canyons. Locations of canyon refugia could not be inferred from genomic analyses due to missing data, encouraging us to explore the effect of missing loci in phylogeographical inferences using RADseq. Results from analyses with varying amounts of missing data suggest that samples with large amounts of missing data can still improve inferences, and the specific loci that are missing matters more than the number of missing loci. This study highlights the profound impact of Pleistocene climates on tarantulas endemic to the Colorado Plateau, as well as the mixed nature of the region's fauna. Some animals recently colonized from nearby deserts as glacial climates receded, whereas others, like tarantulas, appear to have persisted on the Mogollon Rim and in refugia associated with the region's famous river-cut canyons.

The common ancestor of spiders likely used silk to line burrows or make simple webs, with specialized spinning organs and aerial webs originating with the evolution of the megadiverse “true spiders” (Araneomorphae). The base of the araneomorph tree also concentrates the greatest number of changes in respiratory structures, a character system whose evolution is still poorly understood, and that might be related to the evolution of silk glands. Emphasizing a dense sampling of multiple araneomorph lineages where tracheal systems likely originated, we gathered genomic-scale data and reconstructed a phylogeny of true spiders. This robust phylogenomic framework was used to conduct maximum likelihood and Bayesian character evolution analyses for respiratory systems, silk glands, and aerial webs, based on a combination of original and published data. Our results indicate that in true spiders, posterior book lungs were transformed into morphologically similar tracheal systems six times independently, after the evolution of novel silk gland systems and the origin of aerial webs. From these comparative data, we put forth a novel hypothesis that early-diverging web-building spiders were faced with new energetic demands for spinning, which prompted the evolution of similar tracheal systems via convergence; we also propose tests of predictions derived from this hypothesis.[Book lungs; discrete character evolution; respiratory systems; silk; spider web evolution; ultraconserved elements.]

Triaenonychidae Sørensen in L. Koch, 1886 is a large family of Opiliones with ~480 described species broadly distributed across temperate forests in the Southern Hemisphere. However, it remains poorly understood taxonomically, as no comprehensive phylogenetic work has ever been undertaken. In this study we capitalise on samples largely collected by us during the last two decades and use Sanger DNA-sequencing techniques to produce a large phylogenetic tree with 300 triaenonychid terminals representing nearly 50% of triaenonychid genera and including representatives from all the major geographic areas from which they are known. Phylogenetic analyses using maximum likelihood and Bayesian inference methods recover the family as diphyletic, placing Lomanella Pocock, 1903 as the sister group to the New Zealand endemic family Synthetonychiidae Forster, 1954. With the exception of the Laurasian representatives of the family, all landmasses contain non-monophyletic assemblages of taxa. To determine whether this non-monophyly was the result of Gondwanan vicariance, ancient cladogenesis due to habitat regionalisation, or more recent over-water dispersal, we inferred divergence times. We found that most divergence times between landmasses predate Gondwanan breakup, though there has been at least one instance of transoceanic dispersal – to New Caledonia. In all, we identify multiple places in the phylogeny where taxonomic revision is needed, and transfer Lomanella outside of Triaenonychidae in order to maintain monophyly of the family.

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.

The systematics of sitticine jumping spiders is reviewed, with a focus on the Palearctic and Nearctic regions, in order to revise their generic classification, clarify the species of one region (Canada), and study their chromosomes. A genome-wide molecular phylogeny of 23 sitticine species, using more than 700 loci from the arachnid Ultra-Conserved Element (UCE) probeset, confirms the Neotropical origins of sitticines, whose basal divergence separates thenew subtribeAillutticina (a group of five Neotropical genera) from the subtribe Sitticina (five genera of Eurasia and the Americas). The phylogeny shows that most Eurasian sitticines form a relatively recent and rapid radiation, which we unite into the genusAttulusSimon, 1868, consisting of the subgeneraSitticusSimon, 1901 (seven described species),Attulus(41 described species), andSittilongPrószyński, 2017 (one species). Five species ofAttulusoccur natively in North America, presumably through dispersals back from the Eurasian radiation, but an additional three species were more recently introduced from Eurasia.Attus palustrisPeckham & Peckham, 1883 is considered to be a full synonym ofEuophrys floricolaC. L. Koch, 1837 (not a distinct subspecies).Attus sylvestrisEmerton, 1891 is removed from synonymy and recognized as a senior synonym ofSitticus magnusChamberlin & Ivie, 1944. Thus, the five nativeAttulusin North America areAttulus floricola,A. sylvestris,A. cutleri,A. striatus, andA. finschi. The other sitticines of Canada and the U.S.A. are placed in separate genera, all of which arose from a Neotropical radiation includingJollasSimon, 1901 andTomisF.O.Pickard-Cambridge, 1901: (1)AttinellaBanks, 1905 (A. dorsata,A. concolor,A. juniperi), (2)Tomis(T. welchi), and (3)SittisaxPrószyński, 2017 (S. ranieri). All Neotropical and Caribbean “Sitticus” are transferred to eitherJollas(12 species total) orTomis(14 species).Attinella(three species) andTomisare both removed from synonymy withSitticus; the synonymy ofSitticus cabellensisPrószyński, 1971 withPseudattulus kratochviliCaporiacco, 1947 is restored;PseudattulusCaporiacco, 1947 is synonymized withTomis. Six generic names are newly synonymized withAttulusand one withAttinella. Two Neotropical species are described as new,Jollas cupreussp. nov.andTomis manabitasp. nov.Forty-six new combinations are established and three are restored. Three species synonymies are restored, one is new, and two are rejected. Across this diversity of species is a striking diversification of chromosome complements, with X-autosome fusions occurring at least four times to produce neo-Y sex chromosome systems (X1X2Y and X1X2X3Y), some of which (Sittisax ranieriandS. saxicola) are sufficiently derived as to no longer preserve the simple traces of ancestral X material. The correlated distribution of neo-Y and a base autosome number of 28 suggests that neo-Y origins occurred preferentially in lineages with the presence of an extra pair of autosomes.

Natural history collections play a crucial role in biodiversity research, and museum specimens are increasingly being incorporated into modern genetics-based studies. Sequence capture methods have proven incredibly useful for phylogenomics, providing the additional ability to sequence historical museum specimens with highly degraded DNA, which until recently have been deemed less valuable for genetic work. The successful sequencing of ultraconserved elements (UCEs) from historical museum specimens has been demonstrated on multiple tissue types including dried bird skins, formalin-fixed squamates and pinned insects. However, no study has thoroughly demonstrated this approach for historical ethanol-preserved museum specimens. Alongside sequencing of "fresh" specimens preserved in >95% ethanol and stored at -80°C, we used extraction techniques specifically designed for degraded DNA coupled with sequence capture protocols to sequence UCEs from historical museum specimens preserved in 70%-80% ethanol and stored at room temperature, the standard for such ethanol-preserved museum collections. Across 35 fresh and 15 historical museum samples of the arachnid order Opiliones, an average of 345 UCE loci were included in phylogenomic matrices, with museum samples ranging from six to 495 loci. We successfully demonstrate the inclusion of historical ethanol-preserved museum specimens in modern sequence capture phylogenomic studies, show a high frequency of variant bases at the species and population levels, and from off-target reads successfully recover multiple loci traditionally sequenced in multilocus studies including mitochondrial loci and nuclear rRNA loci. The methods detailed in this study will allow researchers to potentially acquire genetic data from millions of ethanol-preserved museum specimens held in collections worldwide.

One major challenge to delimiting species with genetic data is successfully differentiating population structure from species-level divergence, an issue exacerbated in taxa inhabiting naturally fragmented habitats. Many fields of science are now using machine learning, and in evolutionary biology supervised machine learning has recently been used to infer species boundaries. These supervised methods require training data with associated labels. Conversely, unsupervised machine learning (UML) uses inherent data structure and does not require user-specified training labels, potentially providing more objectivity in species delimitation. In the context of integrative taxonomy, we demonstrate the utility of three UML approaches (random forests, variational autoencoders, t-distributed stochastic neighbor embedding) for species delimitation in an arachnid taxon with high population genetic structure (Opiliones, Laniatores, Metanonychus). We find that UML approaches successfully cluster samples according to species-level divergences and not high levels of population structure, while model-based validation methods severely over-split putative species. UML offers intuitive data visualization in two-dimensional space, the ability to accommodate various data types, and has potential in many areas of systematic and evolutionary biology. We argue that machine learning methods are ideally suited for species delimitation and may perform well in many natural systems and across taxa with diverse biological characteristics.

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.

One major challenge to delimiting species with genetic data is successfully differentiating species divergences from population structure, with some current methods biased towards overestimating species numbers. Many fields of science are now utilizing machine learning (ML) approaches, and in systematics and evolutionary biology, supervised ML algorithms have recently been incorporated to infer species boundaries. However, these methods require the creation of training data with associated labels. Unsupervised ML, on the other hand, uses the inherent structure in data and hence does not require any user-specified training labels, thus providing a more objective approach to species delimitation. In the context of integrative taxonomy, we demonstrate the utility of three unsupervised ML approaches, specifically random forests, variational autoencoders, and t-distributed stochastic neighbor embedding, for species delimitation utilizing a short-range endemic harvestman taxon (Laniatores,Metanonychus). First, we combine mitochondrial data with examination of male genitalic morphology to identify a priori species hypotheses. Then we use single nucleotide polymorphism data derived from sequence capture of ultraconserved elements (UCEs) to test the efficacy of unsupervised ML algorithms in successfully identifying a priori species, comparing results to commonly used genetic approaches. Finally, we use two validation methods to assess a priori species hypotheses using UCE data. We find that unsupervised ML approaches successfully cluster samples according to species level divergences and not to high levels of population structure, while standard model-based validation methods over-split species, in some instances suggesting that all sampled individuals are distinct species. Moreover, unsupervised ML approaches offer the benefits of better data visualization in two-dimensional space and the ability to accommodate various data types. We argue that ML methods may be better suited for species delimitation relative to currently used model-based validation methods, and that species delimitation in a truly integrative framework provides more robust final species hypotheses relative to separating delimitation into distinct “discovery” and “validation” phases. Unsupervised ML is a powerful analytical approach that can be incorporated into many aspects of systematic biology, including species delimitation. Based on results of our empirical dataset, we make several taxonomic changes including description of a new species.

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 genusYuriais 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.

Here we show that the most venomous spiders in the world are phylogenetically misplaced. Australian atracine spiders (family Hexathelidae), including the notorious Sydney funnel-web spider Atrax robustus, produce venom peptides that can kill people. Intriguingly, eastern Australian mouse spiders (family Actinopodidae) are also medically dangerous, possessing venom peptides strikingly similar to Atrax hexatoxins. Based on the standing morphology-based classification, mouse spiders are hypothesized distant relatives of atracines, having diverged over 200 million years ago. Using sequence-capture phylogenomics, we instead show convincingly that hexathelids are non-monophyletic, and that atracines are sister to actinopodids. Three new mygalomorph lineages are elevated to the family level, and a revised circumscription of Hexathelidae is presented. Re-writing this phylogenetic story has major implications for how we study venom evolution in these spiders, and potentially genuine consequences for antivenom development and bite treatment research. More generally, our research provides a textbook example of the applied importance of modern phylogenomic research. Fil: Hedin, Marshal. San Diego State University; Estados Unidos Fil: Derkarabetian, Shahan. University of California; Estados Unidos. San Diego State University; Estados Unidos Fil: Ramirez, Martin Javier. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Museo Argentino de Ciencias Naturales “Bernardino Rivadavia”; Argentina Fil: Vink, Cor. Canterbury Museum Christchurch; Nueva Zelanda Fil: Bond, Jason E.. Auburn University; Estados Unidos

Morphological, mitochondrial, and nuclear phylogenomic data were combined to address phylogenetic and species delimitation questions in cave-limitedCicurinaspiders from central Texas. Special effort was focused on specimens and cave locations in the San Antonio region (Bexar County), home to four eyeless species listed as US Federally Endangered. Sequence capture experiments resulted in the recovery of ~200–400 homologous ultra-conserved element (UCE) nuclear loci across taxa, and nearly complete COI mitochondrial DNA sequences from the same set of individuals. Some of these nuclear and mitochondrial sequences were recovered from “standard” museum specimens without special preservation of DNA material, including museum specimens preserved in the 1990s. Multiple phylogenetic analyses of the UCE data agree in the recovery of two major lineages of eyelessCicurinain Texas. These lineages also differ in mitochondrial clade membership, female genitalic morphology, degree of troglomorphy (as measured by relative leg length), and are mostly allopatric across much of Texas. Rare sympatry was confirmed in Bexar County, where members of the two major clades sometimes co-exist in the same karst feature. Both nuclear phylogenomic and mitochondrial data indicate the existence of undescribed species from the San Antonio region, although further sampling and collection of adult specimens is needed to explicitly test these hypotheses. Our data support the two following species synonymies (CicurinaveniiGertsch, 1992 =CicurinamadlaGertsch, 1992;CicurinaloftiniCokendolpher, 2004 =CicurinavesperaGertsch, 1992), formally proposed here. Overall, our taxonomy-focused research has many important conservation implications, and again highlights the fundamental importance of robust taxonomy in conservation research.

Live whole specimens of two species of the harvestman Superfamily Travunioidea Absolon & Kratchovil 1932 from the eastern United States, eight species from the western United States, six morphospecies of the family Triaenonychidae Sorensen 1886 from New Zealand, and specimens of the phylogenetically early-diverging North American triaenonychid Fumontana deprehendor Shear 1977 were extracted in methanol, and the solvent analyzed for components from their defensive secretions. The components were then mapped on a recent phylogeny of the taxa. In both eastern cladonychiid species, Erebomaster flavescens Cope 1872 and Theromaster brunneus (Banks 1902), the major component found was anabaseine, an alkaloid related to nicotine. In the western species, Paranonychus brunneus (Banks 1893), Cryptomaster leviathan Briggs 1969, Speleomaster lexi Briggs 1974, S. pecki Briggs 1974, Speleonychia sengeri Briggs 1974, Metanonychus idahoensis Briggs 1971, Briggsus flavescens (Briggs 1971) and Sclerobunus nondimorphicus Briggs 1971, the major component was N,N-dimethylphenylethylamine, implying that the travunioids from the two regions represent different phyletic lines. The secretions of the triaenonychid species, members of the genera Soerensenella Pocock 1903 and Nuncia Loman 1902, were dominated by 4-methyl-3-hexanone, and that of F. deprehendor by phenol. The completely different chemistry of the two taxa, Travunioidea and Triaenonychidae, implies significant phylogenetic differences, and the presence of phenol in F. deprehendor may suggest a long period of separate evolution for this species.

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 this applied 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 confounded these attempts. 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 genomic 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, even down to the level of species 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.

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.

The southern Rocky Mountains and adjacent Intermontane Plateau Highlands region of western North America is a geographically diverse area with an active geologic history. Given the topological complexity and extensive geologic activity, organisms inhabiting this region are expected to show some degree of morphological and genetic divergence, especially populations found on the southern montane 'sky islands' of this region. Here we examine the phylogeographic history and diversification of a montane forest inhabiting harvestmen, Sclerobunus robustus, using a combination of genetic and morphological data. Divergence time estimates indicate that much of the diversification within and between major groups S. robustus predate the Pleistocene glacial cycles. The most widespread subspecies, Sclerobunus robustus robustus, is recovered as six genetically distinct, geographically cohesive mitochondrial phylogroups. Gene tree data for a single nuclear gene reveals congruent, albeit slightly more conservative, patterns of genetic divergence. Despite high levels of genetic divergence throughout their distribution, phylogroups show extreme conservation in somatic and reproductive morphology. This uncoupling of morphological and genetic differentiation may be due to morphological conservatism associated with a conserved microhabitat preference. Based on these data, it is obvious that S. robustus has undergone some level of cryptic diversification.

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.

Alpha taxonomy, and specifically the delimitation of species, is becoming increasingly objective and integrative. The use of coalescent-based methods applied to genetic data is providing new tools for the discovery and delimitation of species. Here, we use an integrative approach via a combination of discovery-based multivariate morphological analyses to detect potential new species. These potential species are then used as a priori species in hypothesis-driven validation analyses with genetic data. This research focuses on the harvestmen genus Sclerobunus found throughout the mountainous regions of western North America. Based on our analyses, we conduct a revision of Sclerobunus resulting in synonymy of Cyptobunus with Sclerobunus including transfer of S. cavicolens comb. nov. and elevation of both subspecies of S. ungulatus: S. ungulatus comb. nov. and S. madhousensis comb. nov., stat. nov. The three subspecies of S. robustus are elevated, S. robustus, S. glorietus stat. nov., and S. idahoensis stat. nov. Additionally, five new species of Sclerobunus are described from New Mexico and Colorado, including S. jemez sp. nov., S. klomax sp. nov., S. skywalkeri sp. nov., S. speoventus sp. nov., and S. steinmanni sp. nov. Several of the newly described species are single-cave endemics, and our findings suggest that further exploration of western North American cave habitats will likely yield additional new species.

BACKGROUND: Many cave-dwelling animal species display similar morphologies (troglomorphism) that have evolved convergent within and among lineages under the similar selective pressures imposed by cave habitats. Here we study such ecomorphological evolution in cave-dwelling Sclerobuninae harvestmen (Opiliones) from the western United States, providing general insights into morphological homoplasy, rates of morphological change, and the temporal context of cave evolution. METHODOLOGY/PRINCIPAL FINDINGS: We gathered DNA sequence data from three independent gene regions, and combined these data with Bayesian hypothesis testing, morphometrics analysis, study of penis morphology, and relaxed molecular clock analyses. Using multivariate morphometric analysis, we find that phylogenetically unrelated taxa have convergently evolved troglomorphism; alternative phylogenetic hypotheses involving less morphological convergence are not supported by Bayesian hypothesis testing. In one instance, this morphology is found in specimens from a high-elevation stony debris habitat, suggesting that troglomorphism can evolve in non-cave habitats. We discovered a strong positive relationship between troglomorphy index and relative divergence time, making it possible to predict taxon age from morphology. Most of our time estimates for the origin of highly-troglomorphic cave forms predate the Pleistocene. CONCLUSIONS/SIGNIFICANCE: While several regions in the eastern and central United States are well-known hotspots for cave evolution, few modern phylogenetic studies have addressed the evolution of cave-obligate species in the western United States. Our integrative studies reveal the recurrent evolution of troglomorphism in a perhaps unexpected geographic region, at surprisingly deep time depths, and in sometimes surprising habitats. Because some newly discovered troglomorphic populations represent undescribed species, our findings stress the need for further biological exploration, integrative systematic research, and conservation efforts in western US cave habitats.

The harvestman species Sclerobunus parvus was described by Roewer (1931) from the Queen Charlotte Islands, British Columbia, Canada. Some forty years later, Briggs (1971) revised the Triaenonychidae of North America, but missed including Roewer’s species, which had not been mentioned in the literature since its description. Briggs (1971) recognized two subfamilies in North America, Triaenonychinae Sørensen 1886 (Briggs attributed the subfamily name to Pocock, but according to the International Code of Zoological Nomenclature, Sørensen’s original proposal of the family name included the nominate subfamily) and Paranonychinae Briggs 1971. Paranonychinae included two new genera, Metanonychus Briggs 1971 and Paranonychus Briggs 1971. The latter genus was based on Sclerobunus brunneus Banks 1893, a commonly occurring species distributed from Clackamas County, Oregon, north to Atka Island, Alaska (Briggs 1971).