Machine learning approaches delimit cryptic taxa in a previously intractable species complex.

Cryptic species are not diagnosable via morphological criteria, but can be detected through analysis of DNA sequences. A number of methods have been developed for identifying species based on genetic data; however, these methods are prone to over-splitting taxa with extreme population structure, such as dispersal-limited organisms. Machine learning methodologies have the potential to overcome this challenge. Here, we apply such approaches, using a large dataset generated through hybrid target enrichment of ultraconserved elements (UCEs). Our study taxon is the Aoraki denticulata species complex, a lineage of extremely low-dispersal arachnids endemic to the South Island of Aotearoa New Zealand. This group of mite harvesters has been the subject of previous species delimitation studies using smaller datasets generated through Sanger sequencing and analytical approaches that rely on multispecies coalescent models and barcoding gap discovery. Those analyses yielded a number of putative cryptic species that seems unrealistic and extreme, based on what we know about species' geographic ranges and genetic diversity in non-cryptic mite harvesters. We find that machine learning approaches, on the other hand, identify cryptic species with geographic ranges that are similar to those seen in other morphologically diagnosable mite harvesters in Aotearoa New Zealand's South Island. We performed both unsupervised and supervised machine learning analyses, the latter with training data drawn either from animals broadly (vagile and non-vagile) or from a custom training dataset from dispersal-limited harvesters. We conclude that applying machine learning approaches to the analysis of UCE-derived genetic data is an effective method for delimiting species in complexes of low-vagility cryptic species, and that the incorporation of training data from biologically relevant analogues can be critically informative.

Habituation to a predatory stimulus in a harvester (Arachnida, Opiliones).

Several studies have investigated habituation in a defensive context, but few have addressed responses to dangerous stimuli. In such cases, animals should not habituate since this could cost their lives. Here we have stimulated individuals of the harvester Mischonyx squalidus with a predatory stimulus (squeezing with tweezers) in repeated trials within and between days, and measured the occurrence and magnitude of nipping, a defensive behavior. Contrary to our expectations, they did habituate to this stimulus. The probability and magnitude of response declined over trials during each of three days of testing in a typical habituation pattern. During the trials we also observed other defensive behaviors. We discuss our results mainly considering alternative defensive responses. Our data show that we lack information on (1) the role played by the ambiguity of stimuli, (2) the role played by subsequent stimuli and (3) the importance of the array of defensive behaviors of a species in understanding habituation. Although ubiquitous across animals and therefore expected, habituation is described for the first time in the order Opiliones.

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.

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.

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.

A new cave-dwelling species of Charinus (Amblypygi: Charinidae) from northern Brazil.

We present the description and illustration of a new species within the genus Charinus Simon, 1892, discovered in a cave located in the state of Tocantins, situated in the northern region of Brazil. Charinus tocantinensis sp. nov., can be distinguished from its congeners by the following combination of distinctive characters: pedipalp femur with four dorsal spines and four ventral spines; well-developed median eyes and median ocular tubercle; female gonopods sucker-like, slightly longer than wide, opening rounded and margins with fold with each a small posterior slit; cheliceral claw with ten denticles. In addition to the species description, we have included a key for identifying Charinus species in Brazil, along with a distribution map that highlights the known species in the country. This newly described species marks the first formal documentation of a Charinus species within the state of Tocantins. With the addition of this species, the global tally of recognized Charinus species now stands at 98, and Brazil, in particular, is home to a total of 44 formally described species within this genus.

Description of a new fossil Thelyphonida (Arachnida, Uropygi) and further record of Cratosolpuga wunderlichi Selden, in Selden and Shear, 1996 (Arachnida, Solifugae) from Crato Formation (Aptian/Albian), Araripe Basin, Brazil.

Background: A new fossil species of whipscorpion, Mesoproctus rayoli n. sp., is described. The specimen originates from the Crato Formation, dating to the Lower Cretaceous (Aptian/Albian) period within the Araripe Sedimentary Basin. This species has been provisionally assigned to Mesoproctus Dunlop, 1998, as it represents the sole known Thelyphonida fossil genus discovered in South America and within Araripe Lagerstätte. Methods: The material underwent detailed description and illustration processes. Key diagnostic characters, such as body length, pedipalpal coxae apophysis, the form of the opisthosoma, and the length of leg IV, were meticulously examined. SEM methods were applied in this study. Results: Through the detailed analysis, comparisons and differences to Mesoproctus rowlandi Dunlop, 1998 were made possible. Additionally, a well-preserved specimen of the rare camel spider, Cratosolpuga wunderlichi Selden, in Selden and Shear, 1996, was identified from the limestones of the Crato Formation. The newly discovered fossil specimen of Cratosolpuga wunderlichi suggests two characters not previously described: (i) a segmented tarsomere on leg IV; and (ii) a leg I with one tarsal claw.

Phylogenomic data reveal three new families of poorly studied Solifugae (camel spiders).

The systematics of the arachnid order Solifugae have been an enigma, owing to challenges in interpreting morphology, a paucity of molecular phylogenetic studies sampling across the group, and a dearth of taxonomic attention for many lineages. Recent work has suggested that solifuge families largely exhibit contiguous distributions and reflect patterns of vicariance, with the exception of three families: Melanoblossidae, Daesiidae and Gylippidae. Morphological studies have cast doubt on their existing circumscriptions and the present composition of these taxa renders their distributions as disjunct. We leveraged ultraconserved elements (UCEs) to test the phylogenetic placement of three key lineages of Solifugae that cause these anomalous distributions: Dinorhax rostrumpsittaci (putative melanoblossid), Namibesia (putative daesiid), and Trichotoma (putative gylippid). Phylogenetic placement of these three genera based on UCEs rendered the families that harbor them as para- or polyphyletic, recovering instead relationships that better accord with a biogeographic history driven by vicariance. Toward a stable and phylogenetically informed classification of Solifugae, we establish three new families, Dinorhaxidae new rank, Namibesiidae new rank and Lipophagidae new rank.

Opiliofauna (ARACHNIDA: OPILIONES) of the Atlantic Forest in the state of Paraná, Brazil.

Opiliones are arachnids that provide different services in terrestrial ecosystems, especially in tropical forests, which justifies inventory studies to better understand the occurrence and distribution of these arthropods in Brazil, since little is known about the harvestmen fauna in Conservation Units such as in the state of Paraná. In this context, the objective of the present study was to survey the harvestmen fauna at the Iguaçu National Park (PARNA) and in the Bela Vista Biological Refuge (BVBR), from September 2021 to May 2022, using a sampling effort of 40 hours in the BVBR, and 66 hours at the PARNA Iguaçu. Ten species/morphospecies were recorded, highlighting a new occurrence for Brazil, the species Opisthoplatus vegetus. The present study reduces the information gap about Opiliones in the state, which reaffirms the importance of Conservation Units for the protection of the biota in the state of Paraná.

A new pseudoscorpion genus (Garypinoidea: Garypinidae) from the Eocene supports extinction and range contraction in the European paleobiota.

During the Paleogene, the Holarctic experienced drastic climatic oscillations, including periods of extensive glaciation. These changes had a severe impact on both the flora and fauna causing widespread extinction and range shifts with some taxa retreating to refugia in the Mediterranean Basin. Here we provide evidence for this hypothesis using fossils from the pseudoscorpion family Garypinidae Daday, 1889 (Arachnida: Pseudoscorpiones). This family comprises 21 extant genera from all continents except Antarctica but is restricted to low mid-latitudes (<44°N) in the Northern Hemisphere. We provide the second record of garypinids from the European succinite ambers of the Eocene by describing the first extinct genus in Garypinidae, Baltamblyolpium gen. nov., which includes two species: Baltamblyolpium gizmotum sp. nov. from Baltic amber and Baltamblyolpium grabenhorsti sp. nov. from Bitterfeld amber. The new genus exhibits a morphology that closely resembles Neoamblyolpium Hoff, 1956 from western North America and the genus Amblyolpium Simon, 1898, which is widespread but includes taxa restricted to Mediterranean refugia in Europe. The discovery of a new fossil genus of Garypinidae from Europe confirms that the family was found at more northerly latitudes during the Eocene, however, extinction and range contraction resulted in their present-day relictual distribution in southern Europe like many other lineages that once thrived in the European "Baltic amber forest" of the Eocene.

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.

Miniaturization does not change conserved spider anatomy, a case study on spider Rayforstia (Araneae: Anapidae).

Miniaturization is an evolutionary trend observed in many animals. Some arachnid groups, such as spiders and mites, demonstrate a strong tendency toward miniaturization. Some of the most miniaturized spiders belong to the family Anapidae. In this study, using light and confocal microscopy and 3D modelling, we provide the first detailed description of the anatomy of a spider of the genus Rayforstia, which is only 900 µm long. In comparison with larger spiders, Rayforstia has no branching of the midgut in the prosoma and an increased relative brain volume. In contrast to many miniature insects and mites, the spider shows no reduction of whole organ systems, no allometry of the digestive and reproductive systems, and also no reduction of the set of muscles. Thus, miniature spider shows a more conserved anatomy than insects of a similar size. These findings expand our knowledge of miniaturization in terrestrial arthropods.

Morphological and Molecular Identification of Hard Ticks in Hainan Island, China.

Ticks are small, blood-sucking arachnids, known vectors of various diseases, and found throughout the world. They are distributed basically in almost all regions of China. At present, there is not much information regarding tick species on Hainan Island. They were subjected to morphological identification and imaging on an individual basis. Molecular phylogenetic analyses, based on cox1 and 16S rRNA genes, were utilized to identify the species and determine their approximate phylogenetic origin and genetic diversity. The genomic DNA of tick species was extracted, and cytochrome oxidase subunit 1 (cox1) and 16S ribosomal RNA (rRNA) genes were amplified and sequenced. The identification of five tick species, namely Rhipicephalus microplus, Rhipicephalus sanguineus, Rhipicephalus haemaphysaloides, Haemaphysalis cornigera and Haemaphysalis mageshimaensis, was carried out by morphological analysis. When employing the cox1 and 16S rRNA phylogenetic tree, all isolates of R. microplus from Hainan Island were classified as clade A and B, respectively. R. sanguineus was recognized as a member of the tropical lineage by phylogenetic analysis on the cox1 and 16S rRNA genes. Three phylogenetic groups of R. haemaphysaloides were recognized and found to be related closely to strains from China. H. cornigera and H. mageshimaensis formed one phylogenetic group, presumably from tick strains prevalent in Japan and China. The haplotype network analysis indicated that R. microplus is classed into 26 and 6 haplotypes, which correspond to cox1 and 16S rRNA gene assemblages, respectively. In addition, four cox1 haplotypes were detected in R. sanguineus. This is the first evidence that suggests genetic diversity, host range and geographical distribution of hard ticks in Hainan Island, China.

First description of the male genitalia in a short-tailed whipscorpion (Arachnida: Schizomida), description of the female, and comments on pygidial glands and cuticular ultrastructure of Surazomus algodoal Ruiz & Valente, 2017.

The male genitalia of Surazomus algodoal Ruiz & Valente, 2017 are described. Since this is the first attempt to describe male genitalia in the schizomids, we propose new terminology to describe the sclerites of the male genital chamber in the order. We believe that the male genitalia may provide a large set of characters for taxonomic research in schizomids, as the female genitalia have proven to do. The setae and other cuticular structures were investigated using light and scanning electron microscopy (SEM). The female of S. algodoal is described for the first time, including the genitalia, and we highlight the sexual dimorphism present in the species. We also present descriptions of: 1) microtrichia, with respect to their presence/length/distribution, 2) cuticular sensilla, glandular openings and other cuticular structures, proposing new terminology, 3) setae of the pedipalpal trochanter and tarsus. Some changes on the terminology of setae of pedipalpal tibia are also given. These advances may be useful for description in other schizomids. The pygidial glands, mostly ignored in modern papers, were assessed and commented upon.

Palpigrades from Cuba (Arachnida: Palpigradi: Eukoeneniidae).

The knowledge of palpigrades in Cuba is limited to the species Eukoenenia orghidani, discovered and described from Cueva de Bellamar. In this work, a survey for palpigrades in the suburbs of La Habana, Cuba revealed three species, Eukoenenia berlesei, Eukoenenia florenciae and a new species described here as Eukoenenia glandulosa sp. nov. Interestingly, the three species coexist in the microspaces of wet soil. A total of 16 arthropod species were identified living in the same microhabitat as the palpigrades.

An unexpected diversity of Cyphophthalmi (Arachnida: Opiliones) in Upper Cretaceous Burmese amber.

Ten new Cyphophthalmi specimens (Arachnida: Opiliones) from the Upper Cretaceous (Lower Cenomanian) Burmese amber of northern Myanmar are described. Seven of these are placed in Stylocellidae, the predominant extant family found today in Southeast Asia. Sirocellus iunctus gen. et sp. nov. represents the first fossil with a combination of sironid and stylocellid characters, suggesting a still ongoing transition in some lineages during the Upper Cretaceous. Mesopsalis oblongus gen. et sp. nov. represents a second fossil with elongated ozophores, a character not known from modern species. Leptopsalis breyeri sp. nov. is the first Cretaceous cyphophthalmid assignable to an extant genus. The species Foveacorpus cretaceus gen. et sp. nov. and F. parvus gen. et sp. nov., which cannot be placed in an extant family, show morphological novelties for Cyphophthalmi such as numerous pits covering the whole body. The possible function of these pits is discussed. Three more adult males with unique adenostyles and two juveniles are not formally named but further indicate an already highly diverse cyphophthalmid fauna during the Cretaceous. The total number of named Burmese amber Cyphophthalmi species is raised from one to six, and the total fossil record for this suborder now stands at eight.

An additional species of the genus Nemaspela Silhavý, 1966 from a cave in Georgia, with a note on Nemaspela melouri (Opiliones: Nemastomatidae).

A cavernicolous harvestman species, Nemaspela kotia sp. nov., is described from the Kotia Cave (=Kotiasklde Cave) in Zemo Imereti plateau, Imereti Region, West Georgia. Its males lack a cheliceral apophysis, a feature shared with the Georgian cave-dwelling Nemaspela femorecurvata Martens, 2006 and N. prometheus Martens, Maghradze & Barjadze, 2021. Besides, Nemaspela melouri Martens, Maghradze & Barjadze, 2021, known only from its type locality, is found in Solkota Cave (Tskaltubo Municipality, Sataplia-Tskaltubo karst massif, Imereti region, West Georgia). A key for the identification of the known species of Nemaspela is provided.

Two new cave-dwelling pseudoscorpion species (Arachnida: Pseudoscorpiones) from Northeastern Brazil.

In a study of pseudoscorpions in caves from the northeastern Brazil, eight species of Pseudoscorpiones were identified. The known distribution of the species Pseudochthonius biseriatus Mahnert, 2001, Cryptoditha aff. elegans, Parachernes (P.) nitidimanus (Ellingsen, 1905), Petterchernes brasiliensis Heurtault, 1986, Ceriochernes foliaceosetosus Beier, 1974, and Progarypus setifer Mahnert, 2001 is extended to include the State of Pernambuco. Additionally, Geogarypus gollumi sp. nov. (Geogarypidae) and Progarypus smaugi sp. nov. (Hesperolpiidae) are described as new species.

New cave-dwelling pseudoscorpions of the genus Lagynochthonius (Pseudoscorpiones, Chthoniidae) from Guizhou in China.

Eleven new Lagynochthonius species from thirteen karst caves of Guizhou Province are described, including detailed diagnosis and illustrations: L. biyunensis sp. nov. from Biyun Cave (Panzhou City), L. guanniuensis sp. nov. from Guanniu Cave (Xifeng County), L. latipectus sp. nov. from Xianren Cave and Shenxian Cave (Jiangkou County), L. longedentatus sp. nov. from Hongbaituo Cave (Ceheng County), L. longyanensis sp. nov. from Longyan Cave (Bozhou District), L. maanensis sp. nov. from Pianxiao Cave (Zheng'an County), L. mawangensis sp. nov. from Mawang Cave (Wuchuan County), L. nigriculus sp. nov. from Yuqiyan Cave and unnamed cave (Weining County), L. tuoluoensis sp. nov. from Tuoluodadong Cave (Shuicheng District), L. xiaoensis sp. nov. from Xiao Cave (Suiyang County) and L. zhijinensis sp. nov. from Long Cave (Zhijin County). All species are endemic to Guizhou. An identification key to all known Lagynochthonius species from China is also provided.