The kinematics of amblypygid (Arachnida) pedipalps during predation: extreme elongation in raptorial appendages does not result in a proportionate increase in reach and closing speed.

The link between form and function is key to understanding the evolution of unique and/or extreme morphologies. Amblypygids, or whip spiders, are arachnids that often have highly elongated spined pedipalps. These limbs are used to strike at, and secure, prey before processing by the chelicerae. Amblypygi pedipalps are multifunctional, however, being used in courtship and contest, and vary greatly in form between species. Increased pedipalp length may improve performance during prey capture, but length could also be influenced by factors including territorial contest and sexual selection. Here, for the first time, we used high-speed videography and manual tracking to investigate kinematic differences in prey capture between amblypygid species. Across six morphologically diverse species, spanning four genera and two families, we created a total dataset of 86 trials (9-20 per species). Prey capture kinematics varied considerably between species, with differences being expressed in pedipalp joint angle ranges. In particular, maximum reach ratio did not remain constant with total pedipalp length, as geometric scaling would predict, but decreased with longer pedipalps. This suggests that taxa with the most elongated pedipalps do not deploy their potential length advantage to proportionally increase reach. Therefore, a simple mechanical explanation of increased reach does not sufficiently explain pedipalp elongation. We propose other factors to help explain this phenomenon, such as social interactions or sexual selection, which would produce an evolutionary trade-off in pedipalp length between prey capture performance and other behavioural and/or anatomical pressures.

The male reproductive system in whip spiders (Arachnida: Amblypygi).

Whip spiders (Arachnida, Amblypygi), like many other soil arthropods, transfer their spermatozoa indirectly via a stalked spermatophore. While the complex courtship behavior as well as the morphological differences of spermatophores and corresponding female genitalia between taxa have received great attention in the past, comparative research on the internal reproductive system is lacking so far. In this study, the morphology of the male whip spiders of four neoamblypygid taxa has been comparatively studied via computer tomography and subsequent 3D reconstruction. We investigated four species belonging to the Neoamblypygi, that is, the phrynichid species Damon medius (Damoninae), and Euphrynichus bacillifer (Phrynichidae), the phrynid species Phrynus hispaniolae (Phrynidae), and the charontid species Charon grayi (Charontidae). The male reproductive organs consist of paired testes and two pairs of accessory glands, the ventral and lateral glands, which project their ducts anteriorly into the ventrally located unpaired spermatophore producing organ where the respective seminal and secretory reservoirs are located. While this general organization of the male reproductive system is similar among all investigated taxa, there are some notable differences in some structures. The most surprising findings include the complete absence of ventral glands in D. medius, the presence of unique spherical ventral gland reservoirs in C. grayi as well as differences in the organization of the seminal and secretory reservoirs and their connections to the inner genital slit. In addition, the secretory products of both, ventral and lateral glands, are stored in combined secretory reservoirs in E. bacillifer and P. hispaniolae. This study is the first to show that there is some morphological variation in the male reproductive system in Neoamblypygi. These results are the basis for the reconstruction of the Bauplan for the reproductive organs of the whip spiders.

Cerotegument microstructure of whip spiders (Amblypygi: Euamblypygi Weygoldt, 1996) reveals characters for systematics from family to species level.

Like other arthropods, whip spiders of the arachnid order Amblypygi Thorell, 1883 protect themselves against external environmental influences. In this taxon, in addition to the epicuticle, the outermost layer of the exoskeleton, a cement layer (cerotegument) with superhydrophobic properties is deposited over certain body parts. Due to the high level of interspecific variation, the cerotegument structure and the morphology of its associated gland openings have been suggested to be informative for whip spider systematics. The first comparative study of the cerotegument is presented herein, based on a survey across 4 families, 16 genera, and 62 species of Euamblypygi Weygoldt, 1996, the suborder comprising all extant whip spiders except the rare monotypic family Paracharontidae Weygoldt, 1996. Results confirmed that the morphology of the colloidal particles and their assembly on cement globules differ considerably among taxa, but that the level of variation differs among lineages. Interspecific variation in cerotegument ultrastructure was highest among species of Neoamblypygi Weygoldt, 1996, making it an informative character in this clade. Evolutionary trends and intraspecific variation in the structure of the amblypygid cerotegument are briefly discussed.

Widespread retention of ohnologs in key developmental gene families following whole-genome duplication in arachnopulmonates.

Whole-genome duplications (WGDs) have occurred multiple times during animal evolution, including in lineages leading to vertebrates, teleosts, horseshoe crabs, and arachnopulmonates. These dramatic events initially produce a wealth of new genetic material, generally followed by extensive gene loss. It appears, however, that developmental genes such as homeobox genes, signaling pathway components and microRNAs are frequently retained as duplicates (so-called ohnologs) following WGD. These not only provide the best evidence for WGD, but an opportunity to study its evolutionary consequences. Although these genes are well studied in the context of vertebrate WGD, similar comparisons across the extant arachnopulmonate orders are patchy. We sequenced embryonic transcriptomes from two spider species and two amblypygid species and surveyed three important gene families, Hox, Wnt, and frizzled, across these and 12 existing transcriptomic and genomic resources for chelicerates. We report extensive retention of putative ohnologs, further supporting the ancestral arachnopulmonate WGD. We also found evidence of consistent evolutionary trajectories in Hox and Wnt gene repertoires across three of the six arachnopulmonate orders, with interorder variation in the retention of specific paralogs. We identified variation between major clades in spiders and are better able to reconstruct the chronology of gene duplications and losses in spiders, amblypygids, and scorpions. These insights shed light on the evolution of the developmental toolkit in arachnopulmonates, highlight the importance of the comparative approach within lineages, and provide substantial new transcriptomic data for future study.

Insights into the Karyotype Evolution of Charinidae, the Early-Diverging Clade of Whip Spiders (Arachnida: Amblypygi).

Whip spiders (Amblypygi) represent an ancient order of tetrapulmonate arachnids with a low diversity. Their cytogenetic data are confined to only a few reports. Here, we analyzed the family Charinidae, a lineage almost at the base of the amblypygids, providing an insight into the ancestral traits and basic trajectories of amblypygid karyotype evolution. We performed Giemsa staining, selected banding techniques, and detected 18S ribosomal DNA and telomeric repeats by fluorescence in situ hybridization in four Charinus and five Sarax species. Both genera exhibit a wide range of diploid chromosome numbers (2n = 42-76 and 22-74 for Charinus and Sarax, respectively). The 2n reduction was accompanied by an increase of proportion of biarmed elements. We further revealed a single NOR site (probably an ancestral condition for charinids), the presence of a (TTAGG)n telomeric motif localized mostly at the chromosome ends, and an absence of heteromorphic sex chromosomes. Our data collectively suggest a high pace of karyotype repatterning in amblypygids, with probably a high ancestral 2n and its subsequent gradual reduction by fusions, and the action of pericentric inversions, similarly to what has been proposed for neoamblypygids. The possible contribution of fissions to charinid karyotype repatterning, however, cannot be fully ruled out.

Fine structure of the epicuticular secretion coat and associated glands of Pedipalpi and Palpigradi (Arachnida).

Pedipalpi Latreille, 1810 is a poorly studied clade of arachnids comprising the whip spiders (Amblypygi Thorell, 1883), short-tailed whip scorpions (Schizomida Petrunkevitch, 1945) and whip scorpions (Thelyphonida Cambridge, 1872). It has recently been shown that whip spiders coat their exoskeleton with a solid cement layer (cerotegument) that forms elaborate microstructures and turns the cuticle into a super-hydrophobic state. The amblypygid cerotegument provides taxonomic information due to its fine structural diversity, but its presence and variation in the sister groups was previously unknown. The present contribution reports the surface structure of the cuticle in species of Palpigradi, Thelyphonida, and Schizomida to determine if these taxa possess a solid epicuticular secretion coat. Scanning electron microscopy revealed that in addition to Amblypygi only species of Thelyphonida possess solid epicuticular secretion layers. Unlike in Amblypygi, in the Thelyphonida this layer does not usually form microstructures and is less rigidly attached to the underlying cuticle. A species of Typopeltis Pocock, 1894, which exhibited globular structures analogous to the amblypygid cerotegument, was an exception. Glandular structures associated with cement secretions in Amblypygi and Thelyphonida were considered homologous due to similar structure. Solid epicuticular secretion coats were absent from Schizomida, which is interpreted as a secondary loss despite the presence of slit-like glandular openings that appear to produce such epicuticular secretions. The micro-whip scorpion order Palpigradi Thorell, 1900 exhibited markedly different cuticular surface structures and lacked solid epicuticular secretions, consistent with the hypothesis that this order is not closely related to Pedipalpi. These results enhance the knowledge of the small, enigmatic orders of Arachnida.

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.

Amblypygid-fungal interactions: The whip spider exoskeleton as a substrate for fungal growth.

Fungi and arthropods represent some of the most diverse organisms on our planet, yet the ecological relationships between them remain largely unknown. In animals, fungal growth on body surfaces is often hazardous and is known to cause mortality. In contrast, here we report the presence of an apparently non-harmful mycobiome on the cuticle of whip spiders (Arachnida: Amblypygi). The associations are not species-specific and involve a diversity of fungal species, including cosmopolitan and local decomposers as well as entomopathogens. We discuss the ecology of the detected fungal species and hypothesize that the thick epicuticular secretion coat of whip spiders (the cerotegument) promotes fungal growth. It is possible that this relationship is beneficial towards the host if it leads to parasite control or chemical camouflage. Our findings, which are the first from this arthropod lineage, indicate that non-pathogenic interactions between arthropods and fungi may be much more widespread than predicted and call for more studies in this area.

Description of a new Charinus species (Amblypygi: Charinidae) from the Monseñor Nouel province, Dominican Republic.

A new species of Charinus Simon, 1892 from the Dominican Republic is described. With the addition of Charinus magua sp. nov. from the Monseñor Nouel province, the number of known members of the genus Charinus occurring on the island Hispaniola is increased to three. The frontal process, the trichobothria on leg IV, the number and shape of the articles of leg I and the shape of the tarsomers of legs II-IV are often included in taxonomical descriptions of Charinus species. We present these characters in the detailed description of C. magua sp. nov. and illustrate these for the first time for the two other known Charinus from Hispaniola, Charinus dominicanus and Charinus bahoruco. Furthermore, we present and discuss the cerotegument ultrastructure of all three species.

Numerical simulation of colloidal self-assembly of super-hydrophobic arachnid cerotegument structures.

Certain arachnids exhibit complex coatings of their exoskeleton, consisting of globular structures with complex surface features. This, so-called, cerotegument is formed by a multi-component colloidal secretion that self-assembles and cures on the body surface, and leads to high water repellency. Previous ultrastructural studies revealed the involvement of different glandular cells that contribute different components to the secretion mixture, but the overall process of self-assembly into the complex regular structures observed remained highly unclear. Here we study this process from a theoretical point of view, starting from the so-called Tammes-problem. We show that slight changes of simple parameters lead to a variety of morphologies that are highly similar to the ones observed in the species specific cerotegument structures of whip-spiders. These results are not only important for our understanding of the formation of globular hierarchical structures in nature, but also for the fabrication of novel surface coatings by colloidal lithography.

Transgenerational loss and recovery of early learning ability in foraging predatory mites.

The ability to learn is ubiquitous in animals but highly variable within and between species, populations and individuals. Diet-related circumstances, such as diet quantity and quality can influence both long-term constitutive (genetic; by selection) and short-term operational (non-genetic; by the immediate circumstances) learning performance. Here, we scrutinized the causes of loss of learning ability, following multi-generational feeding on pollen, in a line of the predatory mite Amblyseius swirskii, which was previously well able to learn prey during early life, enhancing foraging later in life. We investigated whether, and, if so, how quickly, a transgenerational diet switch to live prey restores the early learning ability of foraging predatory mites. The first experiment shows that the early learning ability was restored after switching the diet of the pollen-fed predator line to live spider mites for two generations before conducting the behavioral assay. The second experiment reveals that offspring regained their learning ability if the diet of their mothers was switched from pollen to spider mites for 3 or 10 days before offspring production. Both experiments in concert suggest transgenerational, pollen-induced operational loss of learning ability in the predatory mite A. swirskii. Maternally-transmitted nutrient deficiency and/or maternally-induced epigenetic changes are the most plausible explanations for the pollen diet-induced loss of learning ability. Our study represents a key example for maternal diet-induced variation in learning ability.

Stygophrynus orientalis sp. nov. (Amblypygi: Charontidae) from Indonesia with the description of a remarkable spermatophore.

Here Stygophrynus orientalis sp. nov. is described, a new charontid whip spider from Banggai Island, Indonesia, representing the most eastern record of the genus exceeding the formerly postulated restriction of its distribution western of the Wallace Line. In addition, the third known spermatophore of a Stygophrynus species is presented, which differs remarkably from those previously described from the genus. Finally, the cerotegument structure of the new species is depicted and compared with that of other Charontidae and Charinidae.

Correction: Constitutive and Operational Variation of Learning in Foraging Predatory Mites.

[This corrects the article DOI: 10.1371/journal.pone.0166334.].

The water-repellent cerotegument of whip-spiders (Arachnida: Amblypygi).

The cuticle of arthropods is usually composed of layers of a chitin-protein-microcomposite, a proteinaceous epicuticle and a thin lipid coating. However, in some instances a thick cement layer (cerotegument) covers the cuticle and may produce elaborate microstructures. This has previously been described for millipedes and mites. Here we report the previously unknown presence of a superhydrophobic cerotegument in whip-spiders (Ambypygi) and reveal its variation in ultrastructure and water-repellence between species. We discuss the relevance of found micro-morphological and physical characters for taxonomy and phylogenetics of this group, and the potential biological functions.

Whip spiders (Amblypygi) become water-repellent by a colloidal secretion that self-assembles into hierarchical microstructures.

BACKGROUND: Among both plants and arthropods, super-hydrophobic surfaces have evolved that enable self-cleaning, locomotion on water surfaces, or plastron respiration. Super-hydrophobicity is achieved by a combination of non-polar substances and complex micro- and nano-structures, usually acquired by growing processes or the deposition of powder-like materials. RESULTS: Here we report on a multi-phasic secretion in whip spiders (Arachnida, Amblypygi), which externally forms durable, hierarchical microstructures on the basically smooth cuticle. The solidified secretion crust makes the previously highly wettable cuticle super-hydrophobic. We describe the ultrastructure of secretory cells, and the maturation and secretion of the different products involved. CONCLUSION: Whip spiders represent intriguing objects of study for revealing the mechanisms of the formation of complex microstructures in non-living systems. Understanding the physical and chemical processes involved may, further, be of interest for bio-inspired design of functional surface coatings.

Constitutive and Operational Variation of Learning in Foraging Predatory Mites.

Learning is widely documented across animal taxa but studies stringently scrutinizing the causes of constitutive or operational variation of learning among populations and individuals are scarce. The ability to learn is genetically determined and subject to constitutive variation while the performance in learning depends on the immediate circumstances and is subject to operational variation. We assessed variation in learning ability and performance of plant-inhabiting predatory mites, Amblyseius swirskii, caused by population origin, rearing diet, and type of experience. Using an early learning foraging paradigm, we determined that homogeneous single prey environments did not select for reduced learning ability, as compared to natural prey-diverse environments, whereas a multi-generational pollen diet resulted in loss of learning, as compared to a diet of live prey. Associative learning produced stronger effects than non-associative learning but both types of experience produced persistent memory. Our study represents a key example of environmentally caused variation in learning ability and performance.

Whip spiders (Amblypygi, Arachnida) of the Western Palaearctic-a review.

All records of the two amblypygid species occurring in the Western Palaearctic are mapped and both species (Charinus ioanniticus and Musicodamon atlanteus) are discussed. Charinus ioanniticus is known from the Eastern Mediterranean (Greece, Turkey, Israel and Egypt) from 10 localities and Musicodamon atlanteus is known from the Maghreb (Morocco, Algeria) from three localities. All records are mapped.

A new species of the South East Asian genus Sarax Simon, 1892 (Arachnida: Amblypygi: Charinidae) and synonymization of Sarax mediterraneus Delle Cave, 1986.

A new species of the whip spider genus Sarax Simon, 1892 from Cebu Island in the Philippines is described: Sarax huberi sp. nov. With the description of this species, the diversity of the genus is increased to three species in the Philippines. Some additional data on their natural environment and their specific habitat are presented and compared with sibling species. The synonymization of Sarax mediterraneus Delle Cave, 1986 with Sarax buxtoni (Gravely, 1915) is carried out.

Maternal intraguild predation risk affects offspring anti-predator behavior and learning in mites.

Predation risk is a strong selective force shaping prey morphology, life history and behavior. Anti-predator behaviors may be innate, learned or both but little is known about the transgenerational behavioral effects of maternally experienced predation risk. We examined intraguild predation (IGP) risk-induced maternal effects on offspring anti-predator behavior, including learning, in the predatory mite Phytoseiulus persimilis. We exposed predatory mite mothers during egg production to presence or absence of the IG predator Amblyseius andersoni and assessed whether maternal stress affects the anti-predator behavior, including larval learning ability, of their offspring as protonymphs. Protonymphs emerging from stressed or unstressed mothers, and having experienced IGP risk as larvae or not, were subjected to choice situations with and without IG predator traces. Predator-experienced protonymphs from stressed mothers were the least active and acted the boldest in site choice towards predator cues. We argue that the attenuated response of the protonymphs to predator traces alone represents optimized risk management because no immediate risk existed. Such behavioral adjustment could reduce the inherent fitness costs of anti-predator behaviors. Overall, our study suggests that P. persimilis mothers experiencing IGP risk may prime their offspring to behave more optimally in IGP environments.

Functional anatomy of the pretarsus in whip spiders (Arachnida, Amblypygi).

Whip spiders (Amblypygi) are a small, cryptic order of arachnids mainly distributed in the tropics. Some basal lineages (families Charinidae and Charontidae) have adhesive pads on the tips of their six walking legs. The present study describes the macro- and ultrastructure of these pads and investigates their contact mechanics and adhesive strength on smooth and rough substrates. Furthermore, the structure of the pretarsus and its kinematics are compared in Charon cf. grayi (with an adhesive pad) and Phrynus longipes (without an adhesive pad). The adhesive pads exhibit an elaborate structure with a unique combination of structural features of smooth and hairy foot pads including a long transversal contact zone performing lateral detachment, a thick internally-branched cuticle with longitudinal ribs and hexagonal surface microstructures with spatulate keels. The contact area of the pads on smooth glass is discontinuous due to the spatulate microstructures with a discontinuous detachment, which could be observed in vivo by high speed videography at a rate of up to 10,000 fps. Adhesive strength was measured with vertical whole animal pull-off tests, obtaining mean values between 55 and 200 kPa. The occurrence of viscous lipid secretions between microstructures was occasionally observed, which, however, seems not to be a necessity for good foothold. The results are discussed in relation to the whip spider's ecology and evolution. Structure-function relationships of the adhesive pads are compared to those of insects and vertebrates.