This study presents the first total evidence dated phylogenetic analysis of the scorpion genus Urophonius, integrating 115 morphological characters and five molecular markers (28S, 18S, H3, 16S, COI). Our comprehensive phylogenetic framework provides novel insights into the genus' diversification timeline and evolutionary processes. Additionally, we described Urophonius andinus n. sp. from the central Chilean Andes, a high-altitude species found at 2400 m.s.a.l., representing the highest elevational record for the genus. This new species is placed within the granulatus species group, characterized by a spring-summer activity period. Este estudio representa el primer an & aacute;lisis filogen & eacute;tico de evidencia total del g & eacute;nero de escorpiones Urophonius, integrando 115 caracteres morfol & oacute;gicos y cinco marcadores moleculares (28S, 18S, H3, 16S, COI). Nuestro amplio marco filogen & eacute;tico brinda nuevas perspectivas sobre los tiempos y los procesos de diversificaci & oacute;n del g & eacute;nero. Adicionalmente, describimos a Urophonius andinus n. sp. de los Andes centrales de Chile, una especie de altura encontrada a unos 2400 m sobre el nivel del mar, lo que representa el r & eacute;cord altitudinal para el g & eacute;nero. Esta nueva especie forma parte del grupo de especies granulatus, que presenta un ciclo de actividad en primavera-verano.

The paradigm of 'selective females' and 'competing males' has evolved into a multifactorial interpretation of reproductive decisions, which can lead to prudence in reproductive efforts and adjustability in mate selectivity patterns. In mutual mate choice scenarios, sexual selection may act on the same or different male and female traits, with both sexes potentially assessing the quality of their partner. Meanwhile, during courtship, there is an exchange of signals that may influence mate choice patterns, establishing a sexual dialogue between behaviours and responses of the opposite sex. Scorpions are a fascinating group to study mate choice and behaviour adjustability because both sexes undergo substantial costs during mating and exhibit a wide range of elaborate behaviours during courtship. Here, we investigated the occurrence of mutual mate choice in a scorpion species for the first time, Urophonius achalensis, which is characterized by monandrous females (with genital plugs) and polygynous males. We expected a mutual mate choice scenario in this scorpion species, where both sexes are selective according to the quality of potential mates, preferring individuals of higher body condition and females without plugs in the case of males. In addition, we expected a sexual dialogue during courtship involving adjustability between male behaviours and female responses, rather than a coerciveeaggressive scenario. During this dialogue, female resistance response operates as a screening mechanism for male stimulation/ appeasement behaviours. Our results indicate the presence of mutual mate choice in U. achalensis, where female mating status influenced the initial sexual selectivity of both sexes and where male body condition affected female selectivity. We observed behavioural adjustability in both sexes influenced by their own quality and that of their partner, which could be considered a form of sexual dialogue during courtship. This research highlights the value of detailed studies that examine the interplay between behaviours and responses in courtship and contributes to the field of sexual communication and mate choice in scorpions. (c) 2023 The Association for the Study of Animal Behaviour. Published by Elsevier Ltd. All rights reserved.

Sexual dimorphism (SD), the divergence of secondary sexual traits between males and females within a species, can arise from diverse evolutionary forces, such as natural selection, mate choice, and intrasexual competition. Allometric scaling patterns of dimorphic traits are related to their functional roles and the different selective pressures that affect each sex. Generally, traits that threaten rivals involved in intrasexual competition tend to exhibit the highest allometric slopes. Conversely, non-sexual traits often display isometric scaling, while genitalia and traits in direct contact between the sexes during courtship and copulation typically show hypoallometry. A good approach to study patterns of SD and allometry is to complement interspecific studies with analyzes of case studies, where the functional aspect is known in detail. Here, we review the occurrence of SD and evaluation of allometry in the Order Scorpiones, allowing us to compare general trends in a broader comparative framework within the group. In addition, we examined SD and allometric slopes of multiple traits (including somatic traits used in sexual and non-sexual interactions, as well as genitalia) in adult individuals of the scorpion Timogenes elegans (Scorpiones, Bothriuridae). We found that at an interspecific level there was a variation in SD between species and morphological traits, with most traits showing a male-biased SD, except for the chelicerae, which were found to be wider in females. Regarding SD studies, we found relatively few reports of functional allometry showing differences in allometric patterns between species. The results in T. elegans follow some of the general patterns found in other scorpions. We found hypoallometry in genital traits and hyperallometry in the pedipalps of both sexes, with steeper allometric slopes observed for pedipalp height in males. These results suggest that genital traits are under stabilizing selective pressure, while pedipalps in both sexes may be under natural and sexual selective pressure. Understanding allometric patterns and their relationship to function in scorpions provides significant insights into the evolutionary pressures driving the divergence of morphological traits used in both sexual and non-sexual contexts.

Components of the same structure or characters of the same individual might respond differently to natural and sexual selective pressures, showing complex morphological patterns. Besides, studying interactions between species plays a crucial role in understanding the diversification of sex-linked phenotypes. Specifically, when two closely related species coexist and exhibit interspecific sexual interactions (reproductive interference—IR), key traits for mating can diverge in sympatric areas to prevent interbreeding and ensure reproductive isolation (reproductive character displacement—RCD). RCD is primarily driven by natural selection, although sexual selection pressures can alter the pattern of phenotypic variation. Additionally, to gain a comprehensive understanding of the patterns of morphological diversification, it is essential to consider changes related to phenotypic plasticity across environmental gradients. To date, there are no studies evaluating this topic in scorpions, and two sympatric species ( Urophonius brachycentrus and U. achalensis ) with RI, provide an ideal model for evaluating phenotypic variation across environmental gradients and the presence of RCD. In this study, we compared intra-specific variation, as well as the size and shape of multiple characters involved in courtship and sperm transfer, between individuals from sympatric and allopatric populations using geometric morphometrics. Our findings revealed an increase in the size of various characters at lower temperatures (higher altitudes) for U. brachycentrus , making them more similar to heterospecifics in sympatric areas, resulting in a pattern of morphological convergence between these species. Increased similarity between species combined with a scramble competition mating system could intensify sexual selection pressures on particular characters. Furthermore, we identified asymmetric RCD in the shape of several sexual characters crucial for mating success (grasping structures) and sperm transfer (genital characters), which could potentially be significant for mechanical isolation during interspecific interactions. Our results highlight significant morphological variability in the size and shape of somatic and genital characters in two scorpion species. This variability may reflect different evolutionary responses, driven in part by natural selection pressures associated with geographic and environmental variations and species recognition mechanisms, and in part by sexual selection pressures at both the intra- and interspecific levels. This comprehensive study reveals the complexity of evolving multifunctional traits in an understudied model and offers valuable insights into traits subject to multiple selective pressures in animal systems experiencing RI.

We describe Bothriurus mistral n. sp. (Scorpiones, Bothriuridae) from the Chilean north-central Andes of the Coquimbo Region. This is the highest elevational discovery for Bothriurus in the western slopes of the Andes. This species was collected in the Estero Derecho Private Protected Area and Natural Sanctuary as part of the First National Biodiversity Inventory of Chile of the Integrated System for Monitoring and Evaluation of Native Forest Ecosystems (SIMEF). Bothriurus mistral n. sp. is closely related to Bothriurus coriaceus Pocock, 1893, from the lowlands of central Chile. This integrative research includes a combination of traditional morphometrics and geometric morphometric analyses to support the taxonomic delimitation of the species.

In this contribution we describe Brachistosternus diaguita n. sp. from the Argentinean central Andes, which is closely related to Brachistosternus montanus Roig-Alsina 1977, also from the same region. To corroborate reciprocal monophyly of the species, Bayesian inference was carried out on a selection of taxa, for which three mitochondrial (16S rDNA, 12SrDNA and COI) and two nuclear (28S rDNA and 18S rDNA) gene fragments had previously been sequenced for previous contributions on this genus by our group. Finally, we explored the po-tential distribution of Argentinean high Andean species of this genus using Maxent.

Crotalines (pitvipers) in the Americas are distributed from southern Canada to southern Argentina, and are represented by 13 genera and 163 species that constitute a monophyletic group. Their phylogenetic relationships have been assessed mostly based on DNA sequences, while morphological data have scarcely been used for phylogenetic inquiry. We present a total-evidence phylogeny of New World pitvipers, the most taxon/character comprehensive phylogeny to date. Our analysis includes all genera, morphological data from external morphology, cranial osteology and hemipenial morphology, and DNA sequences from mitochondrial and nuclear genes. We performed analyses with parsimony as an optimality criterion, using different schemes for character weighting. We evaluated the contribution of the different sources of characters to the phylogeny through analyses of reduced datasets and calculation of weighted homoplasy and retention indexes. We performed a morphological character analysis to identify synapomorphies for the main clades. In terms of biogeography, our results support a single colonization event of the Americas by pitvipers, and a cladogenetic event into a Neotropical clade and a North American/Neotropical clade. The results also shed light on the previously unstable position of some taxa, although they could not sufficiently resolve the position of Bothrops lojanus, which may lead to the paraphyly of either Bothrops or Bothrocophias. The morphological character analyses demonstrated that an important phylogenetic signal is contained in characters related to head scalation, the jaws and the dorsum of the skull, and allowed us to detect morphological convergences in external morphology associated with arboreality.

Despite having >1200 described species and despite their nearly worldwide distribution and prevalence in many xeric ecosystems, relative to many other arachnid groups, we know little about the natural history and behaviour of animals in the order Solifugae (camel spiders, sun spiders, sun scorpions, etc.). Here, we review the current solifuge literature through the lens of conceptual research areas in ecology, evolution, and behaviour and propose ways in which solifuges can contribute to research in specific subfields, as follows: (i) ecology: community and trophic dynamics; connecting food webs; habitat specialization; and biodiversity and conservation; (ii) evolution: speciation and diversification; activity cycles and associated traits; adaptations for speed; and living in extreme environments; and (iii) behaviour and sensory systems: sleep, quiescence, and diapause; sensory systems and sensory ecology; learning and cognition; and mating systems, sexual selection, and sexual conflict. This resource can provide a starting point for identifying research programmes that will simultaneously contribute basic natural history information about this under-studied group and provide a broader understanding of fundamental concepts and theories across the life sciences. We hope that scientists will take this review as a challenge to develop creative ways of leveraging the unique features of solifuges to advance scientific knowledge and understanding.

There are few works on the reproductive behavior of species of cribellate spiders. Even more scarce are studies of the reproductive behavior of representatives of the Desidae, such as the genus Metaltella Mello-Leitao, 1931. In this paper, we describe for the first time the reproductive behavior of Metaltella iheringi (Keyserling, 1891), a species that is characterized by complex genitalia in both sexes. We determined frequencies and durations of the behaviors in the different phases. Thirty virgin males were exposed to virgin females and the behaviors performed by each sex, as well as their frequencies, were recorded. Three phases were identified: pm-copulatory, copulatory, and post-copulatory. The most frequent behaviors were chelicera-palp rubbing, abdomen vibration and web-stretching by the male, and body-shaking and leg-tapping on the web by the female. When the male grabbed the female, she typically fell into a state of quiescence (the female remained motionless with legs in a semi-flexed position) and she remained so even after the male ended the copulation. Mating was characterized by low aggression by the female and a low incidence of cannibalism. In the post-copulatory phase, the male performed frequent behaviors such as abdomen vibration, sperm induction, and post-copulatory cohabitation. We discuss the possible implications of these behaviors in a pre- and post-copulatory sexual selection context. We also provide information that serves as a basis for future studies to understand the mechanisms involved in these behaviors.

The spermatozoa of scorpions are often bundled together, forming a type of sperm conjugation known as a sperm package. Sperm packages may be found inside the testes and seminal vesicles but vanish in the female atrium, leaving free spermatozoa. Previous studies, based on a limited number of taxa, suggested a diversity of sperm package morphology across the order Scorpiones C.L. Koch, 1850. However, the sperm packages of most scorpion taxa remained unknown. The present study provides the first systematic survey of sperm package morphology across the order, covering 89 exemplar species in 66 genera and 19 families representing all suprafamilial ranks, with a more detailed investigation of the family Bothriuridae Simon, 1880. Whereas all exemplar species of scorpions exhibit sperm packages, Buthida Soleglad and Fet, 2003, including Chaerilidae Pocock, 1893, and most Buthidae C.L. Koch, 1837, present unorganized sperm or loosely organized bundles. Although the details vary, three main types of sperm packages may be recognized in all other families: single folded; straight; and multiple folded. Subtypes may be identified according to general shape and folding patterns, mainly among sperm packages that are folded multiple times. Single-folded sperm packages are the most common type observed in the order. Although most sperm packages lack a covering, a conspicuous secretion sheath may be evident, e.g., in some Chactidae Pocock, 1893. Sperm packages vary in length from 112-354 mu m and bent sperm packages are not necessarily longer than straight sperm packages. Four exemplar species of Bothriuridae reveal that variation in sperm count within a single sperm package is consistent with the count derived in spermatogenesis. The diversity of sperm packages suggests a path from free spermatozoa, via bent sperm packages, to other forms. Sperm packages may aid in the transport, cooperation, competition, and survival of spermatozoa. The diverse morphology, function, and evolution of sperm packages merit further investigation.

Different organisms (mainly poikilotherms) are subject to environmental fluctuations that could affect their normal physiological functioning (e.g., by destabilization of biomembranes and rupture of biomolecules). As a result, animals regulate their body temperature and adapt to different environmental conditions through various physiological strategies. These adaptations are crucial in all organisms, although they are more relevant in those that have reached a great adaptive diversity such as scorpions. Within scorpions, the genus Urophonius presents species with winter activity, being this a peculiarity within the Order and an opportunity to study the strategies deployed by these organisms when facing different temperatures. Here, we explore three basic issues of lipid remodeling under high and low temperatures, using adults and juveniles of Urophonius achalensis and U. brachycentrus. First, as an indicator of metabolic state, we analyzed the lipidic changes in different tissues observing that low temperatures generate higher quantities of triacylglycerols and fewer amount of structural lipids and sphyngomielin. Furthermore, we studied the participation of fatty acids in adaptive homeoviscosity, showing that there are changes in the quantity of saturated and unsaturated fatty acids at low temperature (mainly 16:0, 18:0, 18:1 and 18:2). Finally, we observe that there are quantitative and qualitative variations in the cuticular hydrocarbons (with possible water barrier and chemical recognition function). These fluctuations are in some cases species-specific, metabolic-specific, tissue-specific and in others depend on the ontogenetic state.

Urbanization causes the loss of large amounts of habitat and produces significant changes in environmental conditions with consequences in the individual’s behavior, morphology, and physiology in natural populations. The urbanization can impact in the individual’s development instability (DI) that may be measured with the fluctuating asymmetry (FA) level making it an environmental quality bioindicator. In addition, the FA of each trait should be interpreted considering the trait functional importance, morphogenesis and history of selection pressures. In this paper, we evaluated the impact of two decades of increasing urbanization levels over a population of the pampean scorpion Bothriurus bonariensis. We analyze the FA level in somatic and genital traits in adult individuals of both sexes, collected every ~ 10 years (1997, 2008, 2018). Interestingly, in this study the somatic traits (pedipalp chelae) would be under directional selection and are shaped throughout the individual’s growth. In contrast, the genital traits (male’s hemispermatophore) are involved in the sperm transfer process and could be under stabilizing selection. Also, hemispermatophores are affected for less time by environmental stressors since they are formed once the individual has reached sexual maturity. Our results showed a significant increase in the FA levels in somatic traits of males and females, according to the increase of urbanization along years. In contrast, genital traits showed no changes in the level of FA in females, and only a few male genital traits presented an increase in the FA level. This work shows evidence that the measured scorpion traits (under different selective pressures and morphogenesis) might present differences in their susceptibility to environmental stress. We discuss these results in a context of DI of individuals caused by urbanization. We support the idea that the level of FA can be useful as an index to evaluate the effects of environmental stress on invertebrate populations. Furthermore, our paper presents the scorpions as a good model that could be used as an indicator group for studying the impact of the anthropogenic disturbance factors on natural ecosystems.

Sexual communication in mate choice scenarios involves nonmutually exclusive functions: species recognition, mate location and mate assessment. Most studies on sexual communication in mate choice have focused on female assessment of conspicuous male traits, such as visual and acoustic signals. Only a few studies have addressed male assessment of female traits, especially those that are inconspicuous to the human sensory system, such as chemical signals. Although chemical signals are important in sexual communication of many arachnids, there is a gap in our knowledge about how male scorpions use these signals to evaluate potential partners. Here we studied sexual communication functions in two scorpion species (Urophonius brachycentrus and Urophonius achalensis) with sympatric populations and synchronic reproduction, where males are under a scramble competition mating system. We addressed the volatile pheromones role in each of the sexual communication functions. We used Y-mazes to test volatile pheromones, exposing males to different female stimulus arrays in single-signal (conspecific only) and mixed-signal (conspecific and heterospecific) contexts. Males located conspecific females through volatile pheromones, and the time spent in proximity to the female was proportional to males' perception of the female's quality, reinforcing the pheromones' function in mate location and assessment in single-signal environments. However, these sexually selected functions were overridden in mixed-signal contexts, in which males could not differentiate conspecific from heterospecific females, suggesting that pheromones do not allow species recognition. This interaction between sexual communication functions can lead to reproductive interference in this sympatric zone. Male indiscriminateness mediated by scramble competition could attenuate pheromones' species recognition function during mate choice. We provide a comprehensive study of the pheromones' role for mate and species recognition in different social contexts in scorpions. We found support for nonindependent and interacting functions of sexual communication, indicating that mate choice is a complex process in scorpions.

The subfamily Nothopuginae is revised. Titanopuga salinarum gen. et sp. nov., Cuyanopuga bilobata gen. et sp. nov. and Nothopuga telteca sp. nov. are described. We performed a cladistic analysis based on 144 morphological characters and 24 terminals, including representatives of all ammotrechid subfamilies. Nothopuginae is recovered monophyletic with moderate support, but its sister group relation with Oltacolinae is well supported. The genus Mortola is considered a junior synonym of Pseudocleobis, and Pseudocleobis mortola comb. nov. is considered nomen dubium. The subfamily Mortolinae is represented by a monophyletic clade including Chinchippus, Dasycleobis and Pseudocleobis. A dichotomous key for the genera and species of Nothopuginae is provided.

Mating plugs have been proposed as a mechanism that has evolved to avoid sperm competition. Their structure and composition vary across taxa and are related to the effectiveness of its function. This effectiveness could be related to different evolutionary interests of the sexes. Urophonius brachycentrus and Urophonius achalensis (Scorpiones, Bothriuridae) are highly suitable species to study mating plugs because both are monandrous species with specific morphological and physiological responses in the female's genitalia. Here, we analyze (a) the morphology and fine structure of the mating plugs of both species, (b) the site of production in males and the formation process of the mating plug, and (c) the changes that it undergoes over time in the female's reproductive tract. In both species, a complex mating plug obliterates the female's genital aperture and fills the genital atrium. We observed considerable interspecific variation in the mating plug morphology. A mating hemi‐plug was found surrounding the capsular lobes of the hemispermatophore, which could have a mixed composition (involving portions of the hemispermatophore and glandular products). The glandular portion was transferred in a semi‐solid state filling the female's genital atrium and then hardening. Changes that the plug undergoes in the female's genitalia (darkening and increase of the “distal” area of the plug) indicate a participation of the female to the formation of this type of plug. Our study provides new insights into the plugging phenomenon in scorpions, and we discussed the adaptive significance as a post‐copulatory mechanism to avoid sperm competition.

In most animal species, body condition has a fundamental role in fitness. In males, sexual selection generally favors larger body size or greater weight. This may result in males with better condition performing more vigorous courtships, and biasing female preferences. The effects of body condition on mating performance have been extensively studied in different animal groups. Among arachnids, scorpions are an interesting group for evaluating the effects of these sexual traits on mating performance, since they exhibit an ancient mode of indirect sperm transfer. Scorpion males deposit a single spermatophore on the soil to transfer the sperm to the females, and therefore, the production of spermatophores involves a high cost for them. In this study, we use the scorpion Bothriurus bonariensis as a model to evaluate different patterns of sexual behavior as a function of the body condition of both males and females. We found that males with a better body condition performed the mating dance stage more quickly than males with a lower condition. In addition, males performed the sexual sting behavior for a longer time with females in a better condition. Our results suggest that a better condition provides a mating advantage to males and represents an indicator of courtship performance. Given that female quality is usually correlated with fecundity, males mating with females with a better body condition probably have higher reproductive success.

Two new species in the South American sun-spider family Mummuciidae are herein described. Gaucha ramirezi sp. nov. is known from the Chancaní Provincial Park and Forest Reserve, Córdoba province, Argentina, and further reported for a single locality to the northeast, in Santiago del Estero province. The systematic position of this species is uncertain and it is not assigned to any species-group of Gaucha Mello-Leitão, 1924. The other species, Gaucha santana sp. nov., is only known from the Ibirapuitã Environmental Protection Area, in the southern Brazilian state of Rio Grande do Sul, and is a member of the fasciata species-group. With these descriptions, the number of known species of Gaucha is raised to eleven.

Immune defense is a key feature in the life history of organisms, expensive to maintain, highly regulated by individuals and exposed to physiological and evolutionary trade-offs. In chelicerates, relatively scarce are the studies that relate postcopulatory mechanisms and immune response parameters. This work makes an approximation to the female's immunological consequences produced after the placement of a foreign body in the genitalia of three scorpions species, two species that normally receive genital plugs during mating (Urophonius brachycentrus and U. achalensis) and one that does not (Zabius fuscus). Here we performed the first morphological description of the natural plugs of the two Urophonius species. We described complex three zoned structure anchored to the female genital atrium and based on this information we placed implants in the genitalia (for eliciting the local immune response) of virgin females of the three species and measured the immune encapsulation response to this foreign body. We found a greater and heterogeneous response in different zones of the implants in the plug producing species. To corroborate the specificity of this immune response, we compared the local genital reaction with the triggered response at a systemic level by inserting implants into the female body cavity of U. brachycentrus and Zabius fuscus. We found that the systemic response did not differ between species and that only in the plug producing species the local response in the genitalia was higher than the systemic one. We also compared the total hemocyte load before and after the genital implantation to see if this parameter was compromised by the immunological challenge. We confirmed that in Urophonius species the presence of a strange body in the genitalia caused a decrease in the hemocyte load. Besides, we find correlations between the body weight and the immunological parameters, as well as between different immunological parameters with each other. Complementarily, we characterized the hemocytes of the three scorpion species for the first time. This comparative study can help to provide a wider framework of the immunological characteristics of the species, their differences and their relationship with the particular postcopulatory mechanism such as the genital plugs.

Abstract We studied the male holotype of Cercophonius himalayensis Lourenço, 1996, the sole member of the scorpion family Bothriuridae from India, and concluded that it belongs to a species of the genus Phoniocercus Pocock, 1893, which is endemic to the temperate forests of Patagonia. The presence of a Patagonian genus in India is extremely unlikely; therefore, we consider this to be a case of mislabeling of the specimen, and consequently exclude the scorpion family Bothriuridae from Indian fauna. Cercophonius himalayensis is transferred to the genus Phoniocercus, and formally synonymized with Phoniocercus sanmartini Cekalovic, 1968. A brief illustrated description of the type specimen is made, emphasizing important diagnostic characters and some body parts not previously described, such as the hemispermatophore. We also present a probable explanation for the origin of the material.