Under the hood: Phylogenomics of hooded tick spiders (Arachnida, Ricinulei) uncovers discordance between morphology and molecules.

Ricinulei or hooded tick-spiders are a cryptic and ancient group of arachnids. The order consists of around 100 highly endemic extant species restricted to the Afrotropics and the Neotropics along with 22 fossil species. Their antiquity and low vagility make them an excellent group with which to interrogate biogeographic questions. To date, only four molecular analyses have been conducted on the group and they failed to resolve the relationships of the main lineages and even recovering the non-monophyly of the three genera. These studies were limited to a few Sanger loci or phylogenomic analyses with at most seven ingroup samples. To increase phylogenetic resolution in this little-understood and poorly studied group, we present the most comprehensive phylogenomic study of Ricinulei to date leveraging the Arachnida ultra-conserved element probe set. With a data set of 473 loci across 96 ingroup samples, analyses resolved a monophyletic Neotropical clade consisting of four main lineages. Two of them correspond to the current genera Cryptocellus and Pseudocellus while topology testing revealed one lineage to likely be a phylogenetic reconstruction artefact. The fourth lineage, restricted to Northwestern, Andean South America, is consistent with the Cryptocellus magnus group, likely corresponding to the historical genus Heteroricinoides. Since we did not sample the type species for this old genus, we do not formally re-erect Heteroricinoides but our data suggest the need for a thorough morphological re-examination of Neotropical Ricinulei.

Reference genome for the Mojave poppy bee (Perdita meconis), a specialist pollinator of conservation concern.

The Mojave poppy bee, Perdita meconis Griswold (Hymenoptera: Anthophila: Andrenidae), is a species of conservation concern that is restricted to the eastern Mojave Desert of North America. It is a specialist pollinator of two poppy genera, Arctomecon and Argemone (Papaveraceae), and is being considered for listing under the US Endangered Species Act along with one of its pollinator hosts, the Las Vegas bearpoppy (Arctomecon californica). Here, we present a near chromosome-level genome of the Mojave poppy bee to provide a genomic resource that will aid conservation efforts and future research. We isolated DNA from a single, small (<7 mm), male specimen collected using non-ideal preservation methods then performed whole-genome sequencing using PacBio HiFi technology. After quality and contaminant filtering, the final draft genome assembly is 327 Mb, with an N50 length of 17.5 Mb. Annotated repetitive elements compose 37.3% of the genome, although a large proportion (24.87%) of those are unclassified repeats. Additionally, we annotated 18,245 protein-coding genes and 19,433 transcripts. This genome represents one of only a few genomes from the large bee family Andrenidae and one of only a few genomes for pollinator specialists. We highlight both the potential of this genome as a resource for future research, and how high-quality genomes generated from small, non-ideal (in terms of preservation) specimens could facilitate biodiversity genomics.

Re-evaluating and dating myriapod diversification with phylotranscriptomics under a regime of dense taxon sampling.

Recent transcriptomic studies of myriapod phylogeny have been based on relatively small datasets with <40 myriapod terminals and variably supported or contradicted the traditional morphological groupings of Progoneata and Dignatha. Here we amassed a large dataset of 104 myriapod terminals, including multiple species for each of the four myriapod classes. Across the tree, most nodes are stable and well supported. Most analyses across a range of gene occupancy levels provide moderate to strong support for a deep split of Myriapoda into Symphyla + Pauropoda (=Edafopoda) and an uncontradicted grouping of Chilopoda + Diplopoda (=Pectinopoda nov.), as in other recent transcriptome-based analyses; no analysis recovers Progoneata or Dignatha as clades. As in all recent multi-locus and phylogenomic studies, chilopod interrelationships resolve with Craterostigmus excluded from Amalpighiata rather than uniting with other centipedes with maternal brood care in Phylactometria. Diplopod ordinal interrelationships are largely congruent with morphology-based classifications. Chilognathan clades that are not invariably advocated by morphologists include Glomerida + Glomeridesmida, such that the volvation-related characters of pill millipedes may be convergent, and Stemmiulida + Polydesmida more closely allied to Juliformia than to Callipodida + Chordeumatida. The latter relationship implies homoplasy in spinnerets and contradicts Nematophora. A time-tree with nodes calibrated by 25 myriapod and six outgroup fossil terminals recovers Cambrian-Ordovician divergences for the deepest splits in Myriapoda, Edafopoda and Pectinopoda, predating the terrestrial fossil record of myriapods as in other published chronograms, whereas age estimates within Chilopoda and Diplopoda overlap with or do not appreciably predate the calibration fossils. The grouping of Chilopoda and Diplopoda is recovered in all our analyses and is formalized as Pectinopoda nov., named for the shared presence of mandibular comb lamellae. New taxonomic proposals for Chilopoda based on uncontradicted clades are Tykhepoda nov. for the three blind families of Scolopendromorpha that share a "sieve-type" gizzard, and Taktikospina nov. for Scolopendromorpha to the exclusion of Mimopidae.

Comprehensive Species Sampling and Sophisticated Algorithmic Approaches Refute the Monophyly of Arachnida.

Deciphering the evolutionary relationships of Chelicerata (arachnids, horseshoe crabs, and allied taxa) has proven notoriously difficult, due to their ancient rapid radiation and the incidence of elevated evolutionary rates in several lineages. Although conflicting hypotheses prevail in morphological and molecular data sets alike, the monophyly of Arachnida is nearly universally accepted, despite historical lack of support in molecular data sets. Some phylotranscriptomic analyses have recovered arachnid monophyly, but these did not sample all living orders, whereas analyses including all orders have failed to recover Arachnida. To understand this conflict, we assembled a data set of 506 high-quality genomes and transcriptomes, sampling all living orders of Chelicerata with high occupancy and rigorous approaches to orthology inference. Our analyses consistently recovered the nested placement of horseshoe crabs within a paraphyletic Arachnida. This result was insensitive to variation in evolutionary rates of genes, complexity of the substitution models, and alternative algorithmic approaches to species tree inference. Investigation of sources of systematic bias showed that genes and sites that recover arachnid monophyly are enriched in noise and exhibit low information content. To test the impact of morphological data, we generated a 514-taxon morphological data matrix of extant and fossil Chelicerata, analyzed in tandem with the molecular matrix. Combined analyses recovered the clade Merostomata (the marine orders Xiphosura, Eurypterida, and Chasmataspidida), but merostomates appeared nested within Arachnida. Our results suggest that morphological convergence resulting from adaptations to life in terrestrial habitats has driven the historical perception of arachnid monophyly, paralleling the history of numerous other invertebrate terrestrial groups.

Converging on the orb: denser taxon sampling elucidates spider phylogeny and new analytical methods support repeated evolution of the orb web.

High throughput sequencing and phylogenomic analyses focusing on relationships among spiders have both reinforced and upturned long-standing hypotheses. Likewise, the evolution of spider webs-perhaps their most emblematic attribute-is being understood in new ways. With a matrix including 272 spider species and close arachnid relatives, we analyze and evaluate the relationships among these lineages using a variety of orthology assessment methods, occupancy thresholds, tree inference methods and support metrics. Our analyses include families not previously sampled in transcriptomic analyses, such as Symphytognathidae, the only araneoid family absent in such prior works. We find support for the major established spider lineages, including Mygalomorphae, Araneomorphae, Synspermiata, Palpimanoidea, Araneoidea and the Retrolateral Tibial Apophysis Clade, as well as the uloborids, deinopids, oecobiids and hersiliids Grade. Resulting trees are evaluated using bootstrapping, Shimodaira-Hasegawa approximate likelihood ratio test, local posterior probabilities and concordance factors. Using structured Markov models to assess the evolution of spider webs while accounting for hierarchically nested traits, we find multiple convergent occurrences of the orb web across the spider tree-of-life. Overall, we provide the most comprehensive spider tree-of-life to date using transcriptomic data and use new methods to explore controversial issues of web evolution, including the origins and multiple losses of the orb web.

Taxonomic Sampling and Rare Genomic Changes Overcome Long-Branch Attraction in the Phylogenetic Placement of Pseudoscorpions.

Long-branch attraction is a systematic artifact that results in erroneous groupings of fast-evolving taxa. The combination of short, deep internodes in tandem with long-branch attraction artifacts has produced empirically intractable parts of the Tree of Life. One such group is the arthropod subphylum Chelicerata, whose backbone phylogeny has remained unstable despite improvements in phylogenetic methods and genome-scale data sets. Pseudoscorpion placement is particularly variable across data sets and analytical frameworks, with this group either clustering with other long-branch orders or with Arachnopulmonata (scorpions and tetrapulmonates). To surmount long-branch attraction, we investigated the effect of taxonomic sampling via sequential deletion of basally branching pseudoscorpion superfamilies, as well as varying gene occupancy thresholds in supermatrices. We show that concatenated supermatrices and coalescent-based summary species tree approaches support a sister group relationship of pseudoscorpions and scorpions, when more of the basally branching taxa are sampled. Matrix completeness had demonstrably less influence on tree topology. As an external arbiter of phylogenetic placement, we leveraged the recent discovery of an ancient genome duplication in the common ancestor of Arachnopulmonata as a litmus test for competing hypotheses of pseudoscorpion relationships. We generated a high-quality developmental transcriptome and the first genome for pseudoscorpions to assess the incidence of arachnopulmonate-specific duplications (e.g., homeobox genes and miRNAs). Our results support the inclusion of pseudoscorpions in Arachnopulmonata (new definition), as the sister group of scorpions. Panscorpiones (new name) is proposed for the clade uniting Scorpiones and Pseudoscorpiones.

The Phylogeny and Evolution of the Flashiest of the Armored Harvestmen (Arachnida: Opiliones).

Gonyleptoidea, largely restricted to the Neotropics, constitutes the most diverse superfamily of Opiliones and includes the largest and flashiest representatives of this arachnid order. However, the relationships among its main lineages (families and subfamilies) and the timing of their origin are not sufficiently understood to explain how this tropical clade has been able to colonize the temperate zone. Here, we used transcriptomics and divergence time dating to investigate the phylogeny of Gonyleptoidea. Our results support the monophyly of Gonyleptoidea and all of its families with more than one species represented. Resolution within Gonyleptidae s.s. is achieved for many clades, but some subfamilies are not monophyletic (Gonyleptinae, Mitobatinae, and Pachylinae), requiring taxonomic revision. Our data show evidence for one colonization of today's temperate zone early in the history of Gonyleptidae, during the Paleogene, at a time when the Neotropical area extended poleward into regions now considered temperate. This provides a possible mechanism for the colonization of the extratropics by a tropical group following the Paleocene-Eocene Thermal Maximum, explaining how latitudinal diversity gradients can be established. Taxonomic acts: Ampycidae Kury 2003 is newly ranked as family; Neosadocus Mello-Leitão is transferred to Progonyleptoidellinae (new subfamilial assignment). [Arachnids; biogeography; phylogenomics; transcriptomics.].

Sequence capture phylogenomics of historical ethanol-preserved museum specimens: Unlocking the rest of the vault.

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.

Phylogenomic interrogation resolves the backbone of the Pseudoscorpiones tree of life.

Pseudoscorpiones, with nearly 3700 described species, are an ancient and globally distributed group of arachnids with a fossil record dating back to the Middle Devonian. Previous attempts to reconstruct their phylogenetic history have used morphology or a few amplicons, mostly of rRNAs and mitochondrial genes, which have not been able to completely resolve family-level relationships nor the earliest nodes in the pseudoscorpion tree-those which are most informative about the origins of key characters like venoms and silk. Here we undertake a phylogenetic approach using 41 pseudoscorpion transcriptomes and a series of analyses that account for many of the common pitfalls faced in large phylogenomic analyses. All analyses, using concatenation methods and coalescent approaches, supported monophyly of Iocheirata (the venomous pseudoscorpions), which diversified mostly during the Mesozoic, but paraphyly of Epiocheirata, with a sister group relationship of Feaelloidea to Iocheirata, with Chthonioidea as their sister group. These three main lineages were established during the mid-to-late Paleozoic. Our phylogenetic scheme is consistent with the prior hypothesis that the lack of venom in Pseudoscorpiones is plesiomorphic and not a synapomorphy of Epiocheirata. Based on the results of this study, a new classification is proposed for Pseudoscorpiones including the following new nomenclatural and taxonomic acts: the new suborders Palaeosphyronida Harvey and Atoposphyronida Harvey for Dracochelidae and Feaelloidea, respectively; the newly recognized superfamily Garypinoidea for Garypinidae and Larcidae; the revised rank for Lechytiidae and Tridenchthoniidae, which are regarded as subfamilies of Chthoniidae; the revised rank for Tridenchthoniini and Verrucadithini which are regarded as tribes of Tridenchthoniinae; and the elevation of Hesperolpiinae as a distinct family, Hesperolpiidae.

Rounding up the usual suspects: a standard target-gene approach for resolving the interfamilial phylogenetic relationships of ecribellate orb-weaving spiders with a new family-rank classification (Araneae, Araneoidea).

We test the limits of the spider superfamily Araneoidea and reconstruct its interfamilial relationships using standard molecular markers. The taxon sample (363 terminals) comprises for the first time representatives of all araneoid families, including the first molecular data of the family Synaphridae. We use the resulting phylogenetic framework to study web evolution in araneoids. Araneoidea is monophyletic and sister to Nicodamoidea rank. n. Orbiculariae are not monophyletic and also include the RTA clade, Oecobiidae and Hersiliidae. Deinopoidea is paraphyletic with respect to a lineage that includes the RTA clade, Hersiliidae and Oecobiidae. The cribellate orb-weaving family Uloboridae is monophyletic and is sister group to a lineage that includes the RTA Clade, Hersiliidae and Oecobiidae. The monophyly of most Araneoidea families is well supported, with a few exceptions. Anapidae includes holarchaeids but the family remains diphyletic even if Holarchaea is considered an anapid. The orb-web is ancient, having evolved by the early Jurassic; a single origin of the orb with multiple "losses" is implied by our analyses. By the late Jurassic, the orb-web had already been transformed into different architectures, but the ancestors of the RTA clade probably built orb-webs. We also find further support for a single origin of the cribellum and multiple independent losses. The following taxonomic and nomenclatural changes are proposed: the cribellate and ecribellate nicodamids are grouped in the superfamily Nicodamoidea rank n. (Megadictynidae rank res. and Nicodamidae stat. n.). Araneoidea includes 17 families with the following changes: Araneidae is re-circumscribed to include nephilines, Nephilinae rank res., Arkyidae rank n., Physoglenidae rank n., Synotaxidae is limited to the genus Synotaxus, Pararchaeidae is a junior synonym of Malkaridae (syn. n.), Holarchaeidae of Anapidae (syn. n.) and Sinopimoidae of Linyphiidae (syn. n.).

Molecular phylogenetic analysis of "pirate spiders" (Araneae, Mimetidae) with the description of a new African genus and the first report of maternal care in the family.

We investigate the phylogeny of "pirate spiders" (Mimetidae), a family of araneophagic spiders known for their use of aggressive mimicry as a foraging strategy, but poorly understood phylogenetically. Relationships are inferred by including molecular data from six loci for 92 mimetid terminals spanning four genera, and 119 outgroups representing 12 families. Phylogenetic analyses based on parsimony, maximum-likelihood and Bayesian approaches, as well as static and dynamic homology, robustly support monophyly of Mimetidae and a sister-group relationship to a clade comprising Tetragnathidae + Arkyidae. Relationships among the mimetid genera are largely congruent across methods, as follows: (Gelanor (Ero (Anansi n. gen. (Australomimetus, Mimetus)))). Diversification of Mimetidae is estimated to be around 114 Ma, in the Early Cretaceous. In light of the results of our phylogenetic analyses, we erect Anansi n. gen. to include a clade of mimetids from West Africa that contains at least four species, including the newly described A. luki n. sp. We present the first report of maternal care in Mimetidae based on novel field observations.

Taxonomic revision of the Neotropical pirate spiders of the genus Gelanor Thorell, 1869 (Araneae, Mimetidae) with the description of five new species.

We revise the Neotropical spider genus Gelanor Thorell, 1869 (Mimetidae). Gelanor is distributed from northeast Mexico to southern Uruguay , from sea level to 1,600 m. We describe five new species of Gelanor and report eleven new synonymies. Gelanor is here circumscribed to include ten species: Gelanor fortuna new species, Gelanor juruti new species, Gelanor moyobamba new species, Gelanor siquirres new species, Gelanor waorani new species, Gelanor altithorax Keyserling, 1893 (= Gelanor lanei Soares, 1941 new synonymy), Gelanor consequus O. P.-Cambridge, 1902 (= Gelanor depressus Chickering, 1956 new synonymy, Gelanor gertschi Chickering, 1947 new synonymy and Gelanor heraldicus Petrunkevitch, 1925 new synonymy), Gelanor innominatus Chamberlin, 1916, Gelanor latus (Keyserling, 1881) (= Gelanor mixtus O. P.-Cambridge, 1899 new synonymy, Gelanor mabelae Chickering, 1947 new synonymy, Gelanor ornatus Schenkel, 1953 new synonymy and Gelanor proximus Mello-Leitão, 1929 new synonymy) and Gelanor zonatus (C.L. Koch, 1845) (= Gelanor distinctus O-P. Cambridge, 1899 new synonymy, Gelanor insularis Mello-Leitão, 1929 new synonymy and Gelanor obscurus Mello-Leitão, 1929 new synonymy). In addition, we describe for the first time the males of G. altithorax and G. consequus. Species descriptions are provided for all ten species in the genus, together with a compilation of available data, including type specimens, type localities and morphological diagnoses. Light and electron microscope images and updated data on known geographical distributions, are also provided. We also discuss the phylogenetic placement of Gelanor in Mimetidae.

Forest refugia in Western and Central Africa as 'museums' of Mesozoic biodiversity.

The refugial speciation model, or 'species pump', is widely accepted in the context of tropical biogeography and has been advocated as an explanation for present species distributions in tropical Western and Central Africa. In order to test this hypothesis, a phylogeny of the cryptic arachnid order Ricinulei, based on four nuclear and mitochondrial DNA markers, was inferred. This ancient clade of litter-dwelling arthropods, endemic to the primary forests of Western and Central Africa and the Neotropics, might provide insights into the mode and tempo of evolution in Africa. Twenty-six African ricinuleid specimens were sampled from eight countries spanning the distribution of Ricinulei on the continent, and analysed together with Neotropical samples plus other arachnid outgroups. The phylogenetic and molecular dating results suggest that Ricinulei diversified in association with the fragmentation of Gondwana. The early diversification of Ricinoides in Western and Central Africa around 88 (±33) Ma fits old palaeogeographical events better than recent climatic fluctuations. Unlike most recent molecular studies, these results agree with fossil evidence, suggesting that refugia may have acted as 'museums' conserving ancient diversity rather than as engines generating diversity during successive episodes of climatic fluctuation in Africa.