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.

Vestigial organs alter fossil placements in an ancient group of terrestrial chelicerates.

Vestigial organs provide a link between ancient and modern traits and therefore have great potential to resolve the phylogeny of contentious fossils that bear features not seen in extant species. Here we show that extant daddy-longlegs (Arachnida, Opiliones), a group once thought to possess only one pair of eyes, in fact additionally retain a pair of vestigial median eyes and a pair of vestigial lateral eyes. Neuroanatomical gene expression surveys of eye-patterning transcription factors, opsins, and other structural proteins in the daddy-longlegs Phalangium opilio show that the vestigial median and lateral eyes innervate regions of the brain positionally homologous to the median and lateral eye neuropils, respectively, of chelicerate groups like spiders and horseshoe crabs. Gene silencing of eyes absent shows that the vestigial eyes are under the control of the retinal determination gene network. Gene silencing of dachshund disrupts the lateral eyes, but not the median eyes, paralleling loss-of-function phenotypes in insect models. The existence of lateral eyes in extant daddy-longlegs bears upon the placement of the oldest harvestmen fossils, a putative stem group that possessed both a pair of median eyes and a pair of lateral eyes. Phylogenetic analysis of harvestman relationships with an updated understanding of lateral eye incidence resolved the four-eyed fossil group as a member of the extant daddy-longlegs suborder, which in turn resulted in older estimated ages of harvestman diversification. This work underscores that developmental vestiges in extant taxa can influence our understanding of character evolution, placement of fossils, and inference of divergence times.

Evolution of the Spider Homeobox Gene Repertoire by Tandem and Whole Genome Duplication.

Gene duplication generates new genetic material that can contribute to the evolution of gene regulatory networks and phenotypes. Duplicated genes can undergo subfunctionalization to partition ancestral functions and/or neofunctionalization to assume a new function. We previously found there had been a whole genome duplication (WGD) in an ancestor of arachnopulmonates, the lineage including spiders and scorpions but excluding other arachnids like mites, ticks, and harvestmen. This WGD was evidenced by many duplicated homeobox genes, including two Hox clusters, in spiders. However, it was unclear which homeobox paralogues originated by WGD versus smaller-scale events such as tandem duplications. Understanding this is a key to determining the contribution of the WGD to arachnopulmonate genome evolution. Here we characterized the distribution of duplicated homeobox genes across eight chromosome-level spider genomes. We found that most duplicated homeobox genes in spiders are consistent with an origin by WGD. We also found two copies of conserved homeobox gene clusters, including the Hox, NK, HRO, Irx, and SINE clusters, in all eight species. Consistently, we observed one copy of each cluster was degenerated in terms of gene content and organization while the other remained more intact. Focussing on the NK cluster, we found evidence for regulatory subfunctionalization between the duplicated NK genes in the spider Parasteatoda tepidariorum compared to their single-copy orthologues in the harvestman Phalangium opilio. Our study provides new insights into the relative contributions of multiple modes of duplication to the homeobox gene repertoire during the evolution of spiders and the function of NK genes.

Congruence between ultraconserved element-based matrices and phylotranscriptomic datasets in the scorpion Tree of Life.

Scorpions are ancient and historically renowned for their potent venom. Traditionally, the systematics of this group of arthropods was supported by morphological characters, until recent phylogenomic analyses (using RNAseq data) revealed most of the higher-level taxa to be non-monophyletic. While these phylogenomic hypotheses are stable for almost all lineages, some nodes have been hard to resolve due to minimal taxonomic sampling (e.g. family Chactidae). In the same line, it has been shown that some nodes in the Arachnid Tree of Life show disagreement between hypotheses generated using transcritptomes and other genomic sources such as the ultraconserved elements (UCEs). Here, we compared the phylogenetic signal of transcriptomes vs. UCEs by retrieving UCEs from new and previously published scorpion transcriptomes and genomes, and reconstructed phylogenies using both datasets independently. We reexamined the monophyly and phylogenetic placement of Chactidae, sampling an additional chactid species using both datasets. Our results showed that both sets of genome-scale datasets recovered highly similar topologies, with Chactidae rendered paraphyletic owing to the placement of Nullibrotheas allenii. As a first step toward redressing the systematics of Chactidae, we establish the family Anuroctonidae (new family) to accommodate the genus Anuroctonus.

The Impact of Whole Genome Duplication on the Evolution of the Arachnids.

The proliferation of genomic resources for Chelicerata in the past 10 years has revealed that the evolution of chelicerate genomes is more dynamic than previously thought, with multiple waves of ancient whole genome duplications affecting separate lineages. Such duplication events are fascinating from the perspective of evolutionary history because the burst of new gene copies associated with genome duplications facilitates the acquisition of new gene functions (neofunctionalization), which may in turn lead to morphological novelties and spur net diversification. While neofunctionalization has been invoked in several contexts with respect to the success and diversity of spiders, the overall impact of whole genome duplications on chelicerate evolution and development remains imperfectly understood. The purpose of this review is to examine critically the role of whole genome duplication on the diversification of the extant arachnid orders, as well as assess functional datasets for evidence of subfunctionalization or neofunctionalization in chelicerates. This examination focuses on functional data from two focal model taxa: the spider Parasteatoda tepidariorum, which exhibits evidence for an ancient duplication, and the harvestman Phalangium opilio, which exhibits an unduplicated genome. I show that there is no evidence that taxa with genome duplications are more successful than taxa with unduplicated genomes. I contend that evidence for sub- or neofunctionalization of duplicated developmental patterning genes in spiders is indirect or fragmentary at present, despite the appeal of this postulate for explaining the success of groups like spiders. Available expression data suggest that the condition of duplicated Hox modules may have played a role in promoting body plan disparity in the posterior tagma of some orders, such as spiders and scorpions, but functional data substantiating this postulate are critically missing. Spatiotemporal dynamics of duplicated transcription factors in spiders may represent cases of developmental system drift, rather than neofunctionalization. Developmental system drift may represent an important, but overlooked, null hypothesis for studies of paralogs in chelicerate developmental biology. To distinguish between subfunctionalization, neofunctionalization, and developmental system drift, concomitant establishment of comparative functional datasets from taxa exhibiting the genome duplication, as well as those that lack the paralogy, is sorely needed.

Evolution and phylogenetic diversity of the aquaporin gene family in arachnids.

Water flux across cells predominantly occurs through the pore formed by the aquaporin channels. Since water balance is one of the most important challenges to terrestrial animals, aquaporin evolution and diversity is known to play roles in animal terrestrialisation. Arachnids (Arthropoda: Chelicerata: Arachnida) are the second most diverse group and represent the pioneer land colonists in animals; however, there remains no thorough investigation on aquaporin evolution and diversity in this evolutionarily important lineage. Here we reported a phylogenetic study of aquaporin evolution and diversity using genomic data from 116 arachnid species covering almost all (15/16) extant orders. A previously unrecognised subfamily related to aquaporin-4 (i.e. Aqp4-like subfamily) via phylogenetic analysis was identified, suggesting certain underestimate of the arachnid aquaporin diversity in earlier studies probably due to limited taxonomic sampling. Further analysis indicates that this subfamily emerged deep within the life tree of arthropods. Gene tree of another Aqp4-like subfamily (PripL) shows an unexpected basal split between acariform mites (Acariformes) and other arachnids. A closer inspection demonstrated that the PripL evolved quickly and has been under differential selection pressure in acariform mites. Evidence is provided that the evolutionarily ancient Glp subfamily (i.e. aquaglyceroporin) is significantly expanded in terrestrial arachnids compared with their marine relatives. Finally, in spite of the phylogenetic diversity, there exists conservation of some exons in size, functional domain, and intron-insertion phase: an 81-bp and a 218-bp exon, respectively, in apq4-like and glp genes across Eumetazoa lineages including arachnids and human beings. Both exons encode the carboxyl-terminal NPA motif, implying the coding and splicing pressure during hundreds of million years of animal evolution. Hypotheses were tested to explore the possible link between these findings and arachnid terrestrialisation.

Comparative analysis of mitochondrial genomes reveals family-specific architectures and molecular features in scorpions (Arthropoda: Arachnida: Scorpiones).

Scorpions are a group of arachnids with great evolutionary success that comprise more than 2,000 described species. Mitochondrial genomes have been little studied in this clade. We describe and compare different scorpion mitochondrial genomes and analyze their architecture and molecular characteristics. We assembled eight new scorpion mitochondrial genomes from transcriptomic datasets, annotated them, predicted the secondary structures of tRNAs, and compared the nucleotide composition, codon usage, and relative synonymous codon usage of 16 complete scorpion mitochondrial genomes. Lastly, we provided a phylogeny based on all mitochondrial protein coding genes. We characterized the mitogenomes in detail and reported particularities such as dissimilar synteny in the family Buthidae compared to other scorpions, unusual tRNA secondary structures, and unconventional start and stop codons in all scorpions. Our comparative analysis revealed that scorpion mitochondrial genomes exhibit different architectures and features depending on taxonomic identity. We highlight the parvorder Buthida, particularly the family Buthidae, as it invariably exhibited different mitogenome features such as synteny, codon usage, and AT-skew compared to the parvorder Iurida that included the rest of the scorpion families we analyzed in this study. Our results provide a better understanding of the evolution of mitogenome features and phylogenetic relationships in scorpions.

A novel probe set for the phylogenomics and evolution of RTA spiders.

Spiders are important models for evolutionary studies of web building, sexual selection and adaptive radiation. The recent development of probes for UCE (ultra-conserved element)-based phylogenomic studies has shed light on the phylogeny and evolution of spiders. However, the two available UCE probe sets for spider phylogenomics (Spider and Arachnida probe sets) have relatively low capture efficiency within spiders, and are not optimized for the retrolateral tibial apophysis (RTA) clade, a hyperdiverse lineage that is key to understanding the evolution and diversification of spiders. In this study, we sequenced 15 genomes of species in the RTA clade, and using eight reference genomes, we developed a new UCE probe set (41 845 probes targeting 3802 loci, labelled as the RTA probe set). The performance of the RTA probes in resolving the phylogeny of the RTA clade was compared with the Spider and Arachnida probes through an in-silico test on 19 genomes. We also tested the new probe set empirically on 28 spider species of major spider lineages. The results showed that the RTA probes recovered twice and four times as many loci as the other two probe sets, and the phylogeny from the RTA UCEs provided higher support for certain relationships. This newly developed UCE probe set shows higher capture efficiency empirically and is particularly advantageous for phylogenomic and evolutionary studies of RTA clade and jumping spiders.

Too early for the ferry: The biogeographic history of the Assamiidae of southeast Asia (Chelicerata: Opiliones, Laniatores).

Opiliones (harvestmen) have come to be regarded as an abundant source of model groups for study of historical biogeography, due to their ancient age, poor dispersal capability, and high fidelity to biogeographic terranes. One of the least understood harvestman groups is the Paleotropical Assamiidae, one of the more diverse families of Opiliones. Due to a labyrinthine taxonomy, poorly established generic and subfamilial boundaries, and the lack of taxonomic keys for the group, few efforts have been undertaken to decipher relationships within this arachnid lineage. Neither the monophyly of the family, nor its exact placement in the harvestman phylogeny, have been established. Here, we assessed the internal phylogeny of Assamiidae using a ten-locus Sanger dataset, sampling key lineages putatively ascribed to this family for five of the ten markers. Our analyses recovered Assamiidae as a monophyletic group, in a clade with the primarily Afrotropical Pyramidopidae and the southeast Asian Beloniscidae. Internal relationships of assamiids disfavored the systematic validity of subfamilies, with biogeography reflecting much better phylogenetic structure than the existing higher-level taxonomy. To assess whether the Asian assamiids came to occupy Indo-Pacific terranes via rafting on the Indian subcontinent, we performed divergence dating to infer the age of the family. Our results show that Indo-Pacific clades are ancient, originating well before the Cretaceous and therefore predate a vicariant mechanism commonly encountered for Paleotropical taxa.

Highly diversified mitochondrial genomes provide new evidence for interordinal relationships in the Arachnida.

Arachnida is an exceptionally diverse class in the Arthropoda, consisting of 20 orders and playing crucial roles in the terrestrial ecosystems. However, their interordinal relationships have been debated for over a century. Rearranged or highly rearranged mitochondrial genomes (mitogenomes) were consistently found in this class, but their various extent in different lineages and efficiency for resolving arachnid phylogenies are unclear. Here, we reconstructed phylogenetic trees using mitogenome sequences of 290 arachnid species to decipher interordinal relationships as well as diversification through time. Our results recovered monophyly of ten orders (i.e. Amblypygi, Araneae, Ixodida, Mesostigmata, Opiliones, Pseudoscorpiones, Ricinulei, Sarcoptiformes, Scorpiones and Solifugae), while rejecting monophyly of the Trombidiformes due to the unstable position of the Eriophyoidea. The monophyly of Acari (subclass) was rejected, possibly due to the long-branch attraction of the Pseudoscorpiones. The monophyly of Arachnida was further rejected because the Xiphosura nested within arachnid orders with unstable positions. Mitogenomes that are highly rearranged in mites but less rearranged or conserved in the remaining lineages point to their exceptional diversification in mite orders; however, shared derived mitochondrial (mt) gene clusters were found within superfamilies rather than interorders, confusing phylogenetic signals in arachnid interordinal relationships. Molecular dating results show that arachnid orders have ancient origins, ranging from the Ordovician to the Carboniferous, yet have significantly diversified since the Cretaceous in orders Araneae, Mesostigmata, Sarcoptiformes, and Trombidiformes. By summarizing previously resolved key positions of some orders, we propose a plausible arachnid tree of life. Our results underline a more precise framework for interordinal phylogeny in the Arachnida and provide new insights into their ancient evolution.

Stabilized Morphological Evolution of Spiders Despite Mosaic Changes in Foraging Ecology.

A prominent question in animal research is how the evolution of morphology and ecology interacts in the generation of phenotypic diversity. Spiders are some of the most abundant arthropod predators in terrestrial ecosystems and exhibit a diversity of foraging styles. It remains unclear how spider body size and proportions relate to foraging style, and if the use of webs as prey capture devices correlates with changes in body characteristics. Here, we present the most extensive data set to date of morphometric and ecological traits in spiders. We used this data set to estimate the change in spider body sizes and shapes over deep time and to test if and how spider phenotypes are correlated with their behavioral ecology. We found that phylogenetic variation of most traits best fitted an Ornstein-Uhlenbeck model, which is a model of stabilizing selection. A prominent exception was body length, whose evolutionary dynamics were best explained with a Brownian Motion (free trait diffusion) model. This was most expressed in the araneoid clade (ecribellate orb-weaving spiders and allies) that showed bimodal trends toward either miniaturization or gigantism. Only few traits differed significantly between ecological guilds, most prominently leg length and thickness, and although a multivariate framework found general differences in traits among ecological guilds, it was not possible to unequivocally associate a set of morphometric traits with the relative ecological mode. Long, thin legs have often evolved with aerial webs and a hanging (suspended) locomotion style, but this trend is not general. Eye size and fang length did not differ between ecological guilds, rejecting the hypothesis that webs reduce the need for visual cue recognition and prey immobilization. For the inference of the ecology of species with unknown behaviors, we propose not to use morphometric traits, but rather consult (micro-)morphological characters, such as the presence of certain podal structures. These results suggest that, in contrast to insects, the evolution of body proportions in spiders is unusually stabilized and ecological adaptations are dominantly realized by behavioral traits and extended phenotypes in this group of predators. This work demonstrates the power of combining recent advances in phylogenomics with trait-based approaches to better understand global functional diversity patterns through space and time. [Animal architecture; Arachnida; Araneae; extended phenotype; functional traits; macroevolution; stabilizing selection.].

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.

Evidence of positive selection on six spider developmental genes.

Spiders constitute more than 49,000 described species distributed all over the world, and all ecological environments. Their order, Araneae, is defined by a set of characteristics with no parallel among their arachnid counterparts (e.g., spinnerets, silk glands, chelicerae that inoculate venom, among others). Changes in developmental pathways often underlie the evolution of morphological synapomorphies, and as such spiders are a promising model to study the role of developmental genes in the origin of evolutionary novelties. With that in mind, we investigated changes in the evolutionary regime of a set of six developmental genes, using spiders as our model. The genes were mainly chosen for their roles in spinneret ontogeny, yet they are pleiotropic, and it is likely that the origins of other unique morphological phenotypes are also linked to changes in their sequences. Our results indicate no great differences in the selective pressures on those genes when comparing spiders to other arachnids, but a few site-specific positive selection evidence were found in the Araneae lineage. These findings lead us to new insights on spider evolution that are to be further tested.

The genome of a daddy-long-legs (Opiliones) illuminates the evolution of arachnid appendages.

Chelicerate arthropods exhibit dynamic genome evolution, with ancient whole-genome duplication (WGD) events affecting several orders. Yet, genomes remain unavailable for a number of poorly studied orders, such as Opiliones (daddy-long-legs), which has hindered comparative study. We assembled the first harvestman draft genome for the species Phalangium opilio, which bears elongate, prehensile appendages, made possible by numerous distal articles called tarsomeres. Here, we show that the genome of P. opilio exhibits a single Hox cluster and no evidence of WGD. To investigate the developmental genetic basis for the quintessential trait of this group-the elongate legs-we interrogated the function of the Hox genes Deformed (Dfd) and Sex combs reduced (Scr), and a homologue of Epidermal growth factor receptor (Egfr). Knockdown of Dfd incurred homeotic transformation of two pairs of legs into pedipalps, with dramatic shortening of leg segments in the longest leg pair, whereas homeosis in L3 is only achieved upon double Dfd + Scr knockdown. Knockdown of Egfr incurred shortened appendages and the loss of tarsomeres. The similarity of Egfr loss-of-function phenotypic spectra in insects and this arachnid suggest that repeated cooption of EGFR signalling underlies the independent gains of supernumerary tarsomeres across the arthropod tree of life.

Implications of a cheliceral axial duplication in Tetragnatha versicolor (Araneae: Tetragnathidae) for arachnid deuterocerebral appendage development.

The homology of the arachnid chelicera with respect to other head appendages in Panarthropoda has long been debated. Gene expression data and the re-interpretation of early transitional fossils have supported the homology of the deutocerebrum and its associated appendages, implying a homology between primary antennae (mandibulates), chelicerae (euchelicerates), and chelifores (sea spiders). Nevertheless, comparatively little is known about the mechanistic basis of proximo-distal (PD) axis induction in chelicerates, much less the basis for cheliceral fate specification. Here, we describe a new cheliceral teratology in the spider Tetragnatha versicolor Walckenaer, 1841, which consists on a duplication of the PD axis of the left chelicera associated with a terminal secondary schistomely on the fang of the lower axis. This duplication offers clues as to potential shared mechanisms of PD axis formation in the chelicera. We review the state of knowledge on PD axis induction mechanisms in arthropods and identify elements of gene regulatory networks that are key for future functional experiments of appendage development in non-insect model systems. Such investigations would allow a better understanding of PD axis induction of modified and poorly studied arthropod limbs (e.g., chelicerae, chelifores, and ovigers).

DNA screening of Drosophila suzukii predators in berry field orchards shows new predatory taxonomical groups.

Drosophila suzukii (spotted wing drosophila, SWD) is a pandemic quarantine pest that attacks mostly red fruits. The high number of life cycles per year, its ability to rapidly invade and spread across new habitats, and highly polyphagous nature, makes this a particularly aggressive invasive species, for which efficient control methods are currently lacking. The use of native natural predators is particularly promising to anchor sustainable and efficient measures to control SWD. While several field studies have suggested the presence of potential predatory species in infested orchards, only a few confirmed the presence of SWD DNA in predators' gut content. Here, we use a DNA-based approach to identify SWD predators among the arthropod diversity in South Europe, by examining the gut content of potential predator specimens collected in SWD-infested berry fields in North Portugal. These specimens were morphologically identified to the family/order, and their gut content was screened for the presence of SWD DNA using PCR. New SWD predatory taxonomical groups were identified, as Opiliones and Hemerobiidae, in addition to known SWD predators, such as Hemerobiidae, Chrysopidae, Miridae, Carabidae, Formicidae and Araneae. Additionally, the presence of a spider family, Uloboridae, in the orchards was recorded for the first time, posing this family as another SWD-candidate predator. This study sets important bases to further investigate the potential large-scale use of some of these confirmed predator taxa for SWD control in South Europe.

The Evolution of Sox Gene Repertoires and Regulation of Segmentation in Arachnids.

The Sox family of transcription factors regulates many processes during metazoan development, including stem cell maintenance and nervous system specification. Characterizing the repertoires and roles of these genes can therefore provide important insights into animal evolution and development. We further characterized the Sox repertoires of several arachnid species with and without an ancestral whole-genome duplication and compared their expression between the spider Parasteatoda tepidariorum and the harvestman Phalangium opilio. We found that most Sox families have been retained as ohnologs after whole-genome duplication and evidence for potential subfunctionalization and/or neofunctionalization events. Our results also suggest that Sox21b-1 likely regulated segmentation ancestrally in arachnids, playing a similar role to the closely related SoxB gene, Dichaete, in insects. We previously showed that Sox21b-1 is required for the simultaneous formation of prosomal segments and sequential addition of opisthosomal segments in P. tepidariorum. We studied the expression and function of Sox21b-1 further in this spider and found that although this gene regulates the generation of both prosomal and opisthosomal segments, it plays different roles in the formation of these tagmata reflecting their contrasting modes of segmentation and deployment of gene regulatory networks with different architectures.

Genomic Determination of Reproductive Mode in Facultatively Parthenogenetic Opiliones.

Sexual reproduction may pose myriad short-term costs to females. Despite these costs, sexual reproduction is near ubiquitous. Facultative parthenogenesis is theorized to mitigate some of the costs of sex, as individuals can participate in occasional sex to limit costs while obtaining many benefits. However, most theoretical models assume sexual reproduction is fixed following mating, with no possibility of clutches of mixed reproductive ontogeny. Therefore, we asked: if coercive males are present at high frequency in a population of facultative parthenogens, will their clutches be solely sexually produced, or will there be evidence of sexually and asexually-produced offspring? How will their offspring production compare to conspecifics in low-frequency male populations? We addressed our questions by collecting females and egg clutches of the facultatively parthenogenetic Opiliones species Leiobunum manubriatum and L. globosum. In L. manubriatum, females from populations with few males were not significantly more fecund than females from populations with higher male relative frequency, despite the potential release of the former from sexual conflict. We used 3 genotyping methods along with a custom set of DNA capture probes to reveal that offspring of L. manubriatum from these high male populations were primarily produced via asexual reproduction. This is surprising because sex ratios in these southern populations approach equality, increasing the probability for females to encounter mates and produce offspring sexually. We additionally found evidence for reproductive polymorphisms within populations. Rapid and accurate SNP genotyping data will continue to allow us to address broader evolutionary questions regarding the role of facultative reproductive modes in the maintenance of sex.

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.].

Systemic paralogy and function of retinal determination network homologs in arachnids.

BACKGROUND: Arachnids are important components of cave ecosystems and display many examples of troglomorphisms, such as blindness, depigmentation, and elongate appendages. Little is known about how the eyes of arachnids are specified genetically, let alone the mechanisms for eye reduction and loss in troglomorphic arachnids. Additionally, duplication of Retinal Determination Gene Network (RDGN) homologs in spiders has convoluted functional inferences extrapolated from single-copy homologs in pancrustacean models. RESULTS: We investigated a sister species pair of Israeli cave whip spiders, Charinus ioanniticus and C. israelensis (Arachnopulmonata, Amblypygi), of which one species has reduced eyes. We generated embryonic transcriptomes for both Amblypygi species, and discovered that several RDGN homologs exhibit duplications. We show that duplication of RDGN homologs is systemic across arachnopulmonates (arachnid orders that bear book lungs), rather than being a spider-specific phenomenon. A differential gene expression (DGE) analysis comparing the expression of RDGN genes in field-collected embryos of both species identified candidate RDGN genes involved in the formation and reduction of eyes in whip spiders. To ground bioinformatic inference of expression patterns with functional experiments, we interrogated the function of three candidate RDGN genes identified from DGE using RNAi in the spider Parasteatoda tepidariorum. We provide functional evidence that one of these paralogs, sine oculis/Six1 A (soA), is necessary for the development of all arachnid eye types. CONCLUSIONS: Our work establishes a foundation to investigate the genetics of troglomorphic adaptations in cave arachnids, and links differential gene expression to an arthropod eye phenotype for the first time outside of Pancrustacea. Our results support the conservation of at least one RDGN component across Arthropoda and provide a framework for identifying the role of gene duplications in generating arachnid eye diversity.