Measurement of voluntary bite forces in large carnivores using a semi-automated reward-driven system.

Bite force is a key performance trait of the feeding system, but maximal in vivo bite force has been measured in few large mammals. The alternative, modelling of bite force from anatomy, cannot be validated without in vivo measurements. To overcome existing limitations of ethics, safety and animal well-being, we propose a semi-automated method to obtain voluntary maximum bite forces from large mammals using bite plates that automatically dispense a food reward if an incrementally increasing threshold force value is reached. We validated our method using two Malayan sun bears, two Andean spectacled bears and a lioness. We show that voluntary bite force measurement using positive reinforcement is a non-invasive and reliable method to record maximum voluntary bite force performance in large mammals. Our results further show that in vivo data are critical as modeling efforts from osteology have greatly underestimated bite forces in Andean spectacled bears.

The effect of change in mass distribution due to defensive posture on gait in fat-tailed scorpions.

In terrestrial legged locomotion, the distribution of mass can influence the gait characteristics. This can be due to a change in the magnitude or distribution of the load. The latter occurs in scorpions when they lift their large metasoma from a trailing position in ambulatory posture to the well-known arched forward position in the defensive posture. We measured how locomotion changes between these two postures by recording scorpions walking using high-speed video. We found that the metasoma in the fat-tailed scorpion (Androctonus australis) represents about a quarter of the total mass. Moving this mass anteriorly over the body changes the position of the center of mass forward 8.15 ± 1.86 mm. We found this increases the overall duty factor, and particularly that of the second leg pair, even when taking the reduced speed in defensive posture into account. In the five scorpions we recorded, also the ipsilateral phase of leg pairs 3 and 4 differed in defensive posture. We found that the trajectory the 4th foot describes during a single stride also differed significantly between postures, showing this to be a sensitive measure of changes in gait. The change from an ambulatory to a defensive posture places different demands on the gait of scorpions, possibly largely due to the forward displacement of the center of mass.

Unexpected lack of specialisation in the flow properties of spitting cobra venom.

Venom spitting is a defence mechanism based on airborne venom delivery used by a number of different African and Asian elapid snake species ('spitting cobras'; Naja spp. and Hemachatus spp.). Adaptations underpinning venom spitting have been studied extensively at both behavioural and morphological level in cobras, but the role of the physical properties of venom itself in its effective projection remains largely unstudied. We hereby provide the first comparative study of the physical properties of venom in spitting and non-spitting cobras. We measured the viscosity, protein concentration and pH of the venom of 13 cobra species of the genus Naja from Africa and Asia, alongside the spitting elapid Hemachatus haemachatus and the non-spitting viper Bitis arietans By using published microCT scans, we calculated the pressure required to eject venom through the fangs of a spitting and a non-spitting cobra. Despite the differences in the modes of venom delivery, we found no significant differences between spitters and non-spitters in the rheological and physical properties of the studied venoms. Furthermore, all analysed venoms showed a Newtonian flow behaviour, in contrast to previous reports. Although our results imply that the evolution of venom spitting did not significantly affect venom viscosity, our models of fang pressure suggests that the pressure requirements to eject venom are lower in spitting cobras than in non-spitting cobras.

Unexpected lack of specialisation in the flow properties of spitting cobra venom.

Venom spitting is a defence mechanism based on airborne venom delivery used by a number of different African and Asian elapid snake species ('spitting cobras'; Naja spp. and Hemachatus spp.). Adaptations underpinning venom spitting have been studied extensively at both behavioural and morphological level in cobras, but the role of the physical properties of venom itself in its effective projection remains largely unstudied. We hereby provide the first comparative study of the physical properties of venom in spitting and non-spitting cobras. We measured the viscosity, protein concentration and pH of the venom of 13 cobra species of the genus Naja from Africa and Asia, alongside the spitting elapid Hemachatus haemachatus and the non-spitting viper Bitis arietans. By using published microCT scans, we calculated the pressure required to eject venom through the fangs of a spitting and a non-spitting cobra. Despite the differences in the modes of venom delivery, we found no significant differences between spitters and non-spitters in the rheological and physical properties of the studied venoms. Furthermore, all analysed venoms showed a Newtonian flow behaviour, in contrast to previous reports. Although our results imply that the evolution of venom spitting did not significantly affect venom viscosity, our models of fang pressure suggests that the pressure requirements to eject venom are lower in spitting cobras than in non-spitting cobras.

Transcriptome annotation and characterization of novel toxins in six scorpion species.

BACKGROUND: Venom has evolved in parallel in multiple animals for the purpose of self-defense, prey capture or both. These venoms typically consist of highly complex mixtures of toxins: diverse bioactive peptides and/or proteins each with a specific pharmacological activity. Because of their specificity, they can be used as experimental tools to study cell mechanisms and develop novel medicines and drugs. It is therefore potentially valuable to explore the venoms of various animals to characterize their toxins and identify novel toxin-families. This study focuses on the annotation and exploration of the transcriptomes of six scorpion species from three different families. The transcriptomes were annotated with a custom-built automated pipeline, primarily consisting of Basic Local Alignment Search Tool searches against UniProt databases and filter steps based on transcript coverage. RESULTS: We annotated the transcriptomes of four scorpions from the family Buthidae, one from Iuridae and one from Diplocentridae using our annotation pipeline. We found that the four buthid scorpions primarily produce disulfide-bridged ion-channel targeting toxins, while the non-buthid scorpions have a higher abundance of non-disulfide-bridged toxins. Furthermore, analysis of the "unidentified" transcripts resulted in the discovery of six novel putative toxin families containing a total of 37 novel putative toxins. Additionally, 33 novel toxins in existing toxin-families were found. Lastly, 19 novel putative secreted proteins without toxin-like disulfide bonds were found. CONCLUSIONS: We were able to assign most transcripts to a toxin family and classify the venom composition for all six scorpions. In addition to advancing our fundamental knowledge of scorpion venomics, this study may serve as a starting point for future research by facilitating the identification of the venom composition of scorpions and identifying novel putative toxin families.

Distinct spinning patterns gain differentiated loading tolerance of silk thread anchorages in spiders with different ecology.

Building behaviour in animals extends biological functions beyond bodies. Many studies have emphasized the role of behavioural programmes, physiology and extrinsic factors for the structure and function of buildings. Structure attachments associated with animal constructions offer yet unrealized research opportunities. Spiders build a variety of one- to three-dimensional structures from silk fibres. The evolution of economic web shapes as a key for ecological success in spiders has been related to the emergence of high performance silks and thread coating glues. However, the role of thread anchorages has been widely neglected in those models. Here, we show that orb-web (Araneidae) and hunting spiders (Sparassidae) use different silk application patterns that determine the structure and robustness of the joint in silk thread anchorages. Silk anchorages of orb-web spiders show a greater robustness against different loading situations, whereas the silk anchorages of hunting spiders have their highest pull-off resistance when loaded parallel to the substrate along the direction of dragline spinning. This suggests that the behavioural 'printing' of silk into attachment discs along with spinneret morphology was a prerequisite for the evolution of extended silk use in a three-dimensional space. This highlights the ecological role of attachments in the evolution of animal architectures.

A biomechanical view on stinger diversity in scorpions.

Scorpions have elongated metasomas that bear a telson, which is used as a stinger for venom injection. There is a remarkable diversity in the use of the stinger among scorpions, comprising defensive behavior, prey subjugation and mating. This diversity could be reflected by the shape of the telson, as different stinging behaviors will result in very different functional demands. Here we explored the diversity of telson shapes in scorpions by providing morphological measurements, such as curvature and tip angle, as well as by testing stingers under load using finite element analysis (FEA). FEA models were loaded with forces scaled to the surface area of the models, to allow comparison of the relative strain energy based on shape alone. Load force angle was rotated to identify the optimal stinging angle based on the lowest strain energy. Aculeus length and mean aculeus height correlated with minimal strain energy. Optimal stinging angle correlated with tip angle, and differed from the tip angle by about 28.4 ±â€…6.22 °. We found that species that are more venomous have long aculei (stinger barbs) with a larger radius of curvature. FEA models of these longer aculei showed basal stress concentrations, indicating a potential greater risk of basal breakage due to shape alone. Telsons with shorter and thicker aculeus shapes showed stress concentrations at the tip only. Despite these marked differences in shape, we found no difference in the scaled strain energy between groups of species that are more venomous and less venomous groups of species. These results show that scorpion stingers may be biomechanically optimized, and this may indicate different usage of the stinger in different species.

Intersexual Differences in the Gene Expression of Phoneutria depilata (Araneae, Ctenidae) Toxins Revealed by Venom Gland Transcriptome Analyses.

The wandering spider, Phoneutria depilata, is one of Colombia's most active nocturnal arthropod predators of vertebrates and invertebrates. Its venom has been a relevant subject of study in the last two decades. However, the scarcity of transcriptomic data for the species limits our knowledge of the distinct components present in its venom for linking the mainly neurotoxic effects of the spider venom to a particular molecular target. The transcriptome of the P. depilata venom gland was analyzed to understand the effect of different diets or sex and the impact of these variables on the composition of the venom. We sequenced venom glands obtained from ten males and ten females from three diet treatments: (i) invertebrate: Tenebrio molitor, (ii) vertebrate: Hemidactylus frenatus, and (iii) mixed (T. molitor + H. frenatus). Of 17,354 assembled transcripts from all samples, 65 transcripts relating to venom production differed between males and females. Among them, 36 were classified as neurotoxins, 14 as serine endopeptidases, 11 as other proteins related to venom production, three as metalloprotease toxins, and one as a venom potentiator. There were no differences in transcripts across the analyzed diets, but when considering the effect of diets on differences between the sexes, 59 transcripts were differentially expressed. Our findings provide essential information on toxins differentially expressed that can be related to sex and the plasticity of the diet of P. depilata and thus can be used as a reference for venomics of other wandering spider species.

Orchids reduce attachment of herbivorous snails with leaf trichomes.

Protective structures in the epidermis are essential for land plants to defend themselves against herbivores. In this study, we investigated the effect of different types of trichomes of three orchids, Calanthe triplicata, Dendrochilum pallidiflavens and Trichotosia ferox, on attachment of herbivorous land snails, using histochemistry and centrifuge experiments. Size, ornamentation and histochemistry of epicuticular trichomes on the orchid leaves were assessed with light microscopy, scanning electron microscopy and transmission electron microscopy. Total forces needed to detach two differently shaped snail species, Subulina octona and Pleurodonte isabella, were measured using a turntable equipped with a synchronized strobe. Snails were placed in two positions, either perpendicular or parallel to the main veins on the orchid leaves, both on the adaxial (= upper) or abaxial (= lower) side. The results obtained provided three new insights. First, a perpendicular or parallel position of the snails to the main veins did not significantly affect the attachment performance of either species tested. Secondly, snails detached significantly easier on leaf sides covered with a high density of lignin filled epicuticular trichomes. Thirdly, the removal of glandular trichomes did not affect the attachment forces; however, the absence of lignified trichomes increased the attachment of the snails. Our study highlights the importance of studying micro-ornamentation in combination with performance for obtaining a better understanding of the defense mechanisms employed by different species of orchids to deter herbivorous snails.

Packing a pinch: functional implications of chela shapes in scorpions using finite element analysis.

Scorpions depend on their pedipalps for prey capture, defense, mating and sensing their environment. Some species additionally use their pedipalps for burrowing or climbing. Because the pincers or chelae at the end of the pedipalps vary widely in shape, they have been used as part of a suite of characters to delimit ecomorphotypes. We here evaluate the influence of the different chela cuticular shapes on their performance under natural loading conditions. Chelae of 20 species, representing seven families and spanning most of the range of chela morphologies, were assigned to clusters based on chela shape parameters using hierarchical cluster analysis. Several clusters were identified corresponding approximately to described scorpion ecomorphotypes. Finite element models of the chela cuticulae were constructed from CT scans and loaded with estimated pinch forces based on in vivo force measurements. Chela shape clusters differed significantly in mean Von Mises stress and strain energy. Normalized FEA showed that chela shape significantly influenced Von Mises stress and strain energy in the chela cuticula, with Von Mises stress varying up to an order of magnitude and strain energy up to two orders of magnitude. More elongate, high-aspect ratio chela forms showed significantly higher mean stress compared with more robust low-aspect ratio forms. This suggests that elongate chelae are at a higher risk of failure when operating near the maximum pinch force. Phylogenetic independent contrasts (PIC) were calculated based on a partly resolved phylogram with branch lengths based on an alignment of the 12S, 16S and CO1 mitochondrial genes. PIC showed that cuticular stress and strain in the chela were correlated with several shape parameters, such as aspect ratio, movable finger length, and chela height, independently of phylogenetic history. Our results indicate that slender chela morphologies may be less suitable for high-force functions such as burrowing and defense. Further implications of these findings for the ecology and evolution of the different chela morphologies are discussed.

Functional morphology and bite performance of raptorial chelicerae of camel spiders (Solifugae).

Solifugae are an understudied group of relatively large arachnids with well over 1000 species distributed on almost all major continents. These highly active predators utilize their large chelicerae for feeding, defense, burrowing and mating. We investigated the differences in cheliceral morphology and performance of two ecologically divergent species from North Africa; the cursorial Galeodes sp. and the burrowing Rhagodes melanus. Morphological data show differences in aspect ratio between the two species. Bite force measurements show Rhagodes (N=11) to be a much stronger biter than Galeodes (N=8), in terms of both absolute maximum force (Rhagodes 5.63 N, Galeodes 2.12 N) and force relative to cheliceral size. Synchrotron microtomographs of one specimen for each species reveal large differences in physiological cross-sectional area (PCSA) and estimated muscle stress, resulting in a much higher muscle stress in Rhagodes. This species also showed a longer muscle fiber length. Muscle volume and PCSA were found to differ between the two chelicerae in the two scanned specimens. Whereas Rhagodes reflects this morphological asymmetry in having a higher bite force in the right chelicera, Galeodes shows no such bias.

Reevaluating scorpion ecomorphs using a naïve approach.

BACKGROUND: Ecomorphs create the opportunity to investigate ecological adaptation because they encompass organisms that evolved characteristic morphologies under similar ecological demands. For over 50 years, scorpions have been empirically assigned to ecomorphs based on the characteristic morphologies that rock, sand, vegetation, underground, and surface dwellers assume. This study aims to independently test the existence of scorpion ecomorphs by quantifying the association between their morphology and ecology across 61 species, representing 14 families of the Scorpiones order. RESULTS: Without a priori categorization of species into ecomorphs, we identified four groups based on microhabitat descriptors, which reflect how scorpion ecospace is clustered. Moreover, these microhabitat groups, i.e., ecotypes, have significantly divergent morphologies; therefore, they represent ecomorphs. These ecomorphs largely correspond with the ones previously described in the literature. Therefore, we retained the names Lithophilous, Psammophilous, and Pelophilous, and proposed the name Phytophilous for vegetation dwellers. Finally, we sought to map the morphology-ecology association in scorpions and found that the morphological regions most tightly associated with ecology are at the extremities. Moreover, the major trend in ecomorphological covariation is that longer walking legs and relatively slender pedipalps (pincers) are associated with sandy microhabitats, while the inverse morphological proportions are associated with rocky microhabitats. CONCLUSIONS: Scorpion ecomorphs are validated in a naïve approach, from ecological descriptors and whole body anatomy. This places them on a more solid quantitative footing for future studies of ecological adaptation in scorpions. Our results verify most of the previously defined ecomorphotypes and could be used as a current practice to understand the adaptive significance of ecological morphology.

Linking toxicity and predation in a venomous arthropod: the case of Tityus fuhrmanni (Scorpiones: Buthidae), a generalist predator scorpion.

BACKGROUND: Scorpions are arachnids that have a generalist diet, which use venom to subdue their prey. The study of their trophic ecology and capture behavior is still limited compared to other organisms, and aspects such as trophic specialization in this group have been little explored. METHODS: In order to determine the relationship between feeding behavior and venom toxicity in the scorpion species Tityus fuhrmanni, 33 specimens were offered prey with different morphologies and defense mechanisms: spiders, cockroaches and crickets. In each of the experiments we recorded the following aspects: acceptance rate, immobilization time and the number of capture attempts. The median lethal dose of T. fuhrmanni venom against the three different types of prey was also evaluated. RESULTS: We found that this species does not have a marked difference in acceptance for any of the evaluated prey, but the number of capture attempts of spiders is higher when compared to the other types of prey. The immobilization time is shorter in spiders compared to other prey and the LD50 was higher for cockroaches. CONCLUSIONS: These results indicate that T. fuhrmanni is a scorpion with a generalist diet, has a venom with a different potency among prey and is capable of discriminating between prey types and employing distinct strategies to subdue them.

Pinching or stinging? Comparing prey capture among scorpions with contrasting morphologies.

Background: Scorpions can use their pincers and/or stingers to subdue and immobilize their prey. A scorpion can thus choose between strategies involving force or venom, or both, depending on what is required to subdue its prey. Scorpions vary greatly in the size and strength of their pincers, and in the efficacy of their venom. Whether this variability is driven by their defensive or prey incapacitation functionis unknown. In this study, we test if scorpion species with different pincer morphologies and venom efficacies use these weapons differently during prey subjugation. To that end, we observed Opisthacanthus elatus and Chactas sp. with large pincers and Centruroides edwardsii and Tityus sp. with slender pincers. Methods: The scorpion pinch force was measured, and behavioral experiments were performed with hard and soft prey (Blaptica dubia and Acheta domesticus). Stinger use, sting frequency and immobilization time were measured. Results: We found that scorpions with large pincers such as O. elatus produce more force and use the stinger less, mostly subjugating prey by crushing them with the pincers. In C. edwardsii and Tityus sp. we found they use their slender and relatively weak pincers for holding the prey, but seem to predominantly use the stinger to subjugate them. On the other hand, Chactas sp. uses both strategies although it has a high pinch force. Conclusions: Our results show that scorpionspecies with massive pincers and high pinch force as O. elatus use the stinger less for prey subjugation than scorpionspecies with slenderpincers.

Biomechanical analyses of pterygotid sea scorpion chelicerae uncover predatory specialisation within eurypterids.

Eurypterids (sea scorpions) are extinct aquatic chelicerates. Within this group, members of Pterygotidae represent some of the largest known marine arthropods. Representatives of this family all have hypertrophied, anteriorly-directed chelicerae and are commonly considered Silurian and Devonian apex predators. Despite a long history of research interest in these appendages, pterygotids have been subject to limited biomechanical investigation. Here, we present finite element analysis (FEA) models of four different pterygotid chelicerae-those of Acutiramus bohemicus, Erettopterus bilobus, Jaekelopterus rhenaniae, and Pterygotus anglicus-informed through muscle data and finite element models (FEMs) of chelae from 16 extant scorpion taxa. We find that Er. bilobus and Pt. anglicus have comparable stress patterns to modern scorpions, suggesting a generalised diet that probably included other eurypterids and, in the Devonian species, armoured fishes, as indicated by co-occurring fauna. Acutiramus bohemicus is markedly different, with the stress being concentrated in the proximal free ramus and the serrated denticles. This indicates a morphology better suited for targeting softer prey. Jaekelopterus rhenaniae exhibits much lower stress across the entire model. This, combined with an extremely large body size, suggests that the species likely fed on larger and harder prey, including heavily armoured fishes. The range of cheliceral morphologies and stress patterns within Pterygotidae demonstrate that members of this family had variable diets, with only the most derived species likely to feed on armoured prey, such as placoderms. Indeed, increased sizes of these forms throughout the mid-Palaeozoic may represent an 'arms race' between eurypterids and armoured fishes, with Devonian pterygotids adapting to the rapid diversification of placoderms.

The Terrific Skink bite force suggests insularity as a likely driver to exceptional resource use.

Natural history museum collections hold extremely rare, extinct species often described from a single known specimen. On occasions, rediscoveries open new opportunities to understand selective forces acting on phenotypic traits. Recent rediscovery of few individuals of Bocourt´s Terrific Skink Phoboscincus bocourti, from a small and remote islet in New Caledonia allowed to genetically identify a species of land crab in its diet. To explore this further, we CT- and MRI-scanned the head of the holotype, the only preserved specimen dated to about 1870, segmented the adductor muscles of the jaw and bones, and estimated bite force through biomechanical models. These data were compared with those gathered for 332 specimens belonging to 44 other skink species. Thereafter we recorded the maximum force needed to generate mechanical failure of the exoskeleton of a crab specimen. The bite force is greater than the prey hardness, suggesting that predation on hard-shelled crabs may be an important driver of performance. The high bite force seems crucial to overcome low or seasonal variations in resource availability in these extreme insular environments. Phoboscincus bocourti appears to be an apex predator in a remote and harsh environment and the only skink known to predate on hard-shelled land crabs.

Armed stem to stinger: a review of the ecological roles of scorpion weapons.

Scorpions possess two systems of weapons: the pincers (chelae) and the stinger (telson). These are placed on anatomically and developmentally well separated parts of the body, that is, the oral appendages and at the end of the body axis. The otherwise conserved body plan of scorpions varies most in the shape and relative dimensions of these two weapon systems, both across species and in some cases between the sexes. We review the literature on the ecological function of these two weapon systems in each of three contexts of usage: (i) predation, (ii) defense and (iii) sexual contests. In the latter context, we will also discuss their usage in mating. We first provide a comparative background for each of these contexts of usage by giving examples of other weapon systems from across the animal kingdom. Then, we discuss the pertinent aspects of the anatomy of the weapon systems, particularly those aspects relevant to their functioning in their ecological roles. The literature on the functioning and ecological role of both the chelae and the telson is discussed in detail, again organized by context of usage. Particular emphasis is given on the differences in morphology or usage between species or higher taxonomic groups, or between genders, as such cases are most insightful to understand the roles of each of the two distinct weapon systems of the scorpions and their evolutionary interactions. We aimed to synthesize the literature while minimizing conjecture, but also to point out gaps in the literature and potential future research opportunities.

MS/MS analysis of four scorpion venoms from Colombia: a descriptive approach.

BACKGROUND: Scorpions are widely known for the neurotoxic effects of their venoms, which contain peptides affecting ionic channels. Although Colombia is recognized for its scorpion diversity, only a few studies are available describing the venom content. METHODS: In this descriptive study, we analyzed the MS/MS sequence, electrophoretic and chromatographic profile linked to a bioinformatics analysis of the scorpions Chactas reticulatus (Chactidae), Opisthacanthus elatus (Hormuridae), Centruroides edwardsii (Buthidae) and Tityus asthenes (Buthidae) from Colombia. RESULTS: Each scorpion showed a specific electrophoretic and chromatographic profile. The electrophoretic profiles indicate the presence of high molecular mass compounds in all venoms, with a predominance of low molecular mass compounds in the Buthidae species. Chromatographic profiles showed a similar pattern as the electrophoretic profiles. From the MS/MS analysis of the chromatographic collected fractions, we obtained internal peptide sequences corresponding to proteins reported in scorpions from the respective family of the analyzed samples. Some of these proteins correspond to neurotoxins affecting ionic channels, antimicrobial peptides and metalloproteinase-like fragments. In the venom of Tityus asthenes, the MSn analysis allowed the detection of two toxins affecting sodium channels covering 50% and 84% of the sequence respectively, showing 100% sequence similarity. Two sequences from Tityus asthenes showed sequence similarity with a phospholipase from Opisthacanthus cayaporum indicating the presence of this type of toxin in this species for the first time. One sequence matching a hypothetical secreted protein from Hottentotta judaicus was found in three of the studied venoms. We found that this protein is common in the Buthidae family whereas it has been reported in other families - such as Scorpionidae - and may be part of the evolutionary puzzle of venoms in these arachnids. CONCLUSION: Buthidae venoms from Colombia can be considered an important source of peptides similar to toxins affecting ionic channels. An interesting predicted antimicrobial peptide was detected in three of the analyzed venoms.

Deciphering the diet of a wandering spider (Phoneutria boliviensis; Araneae: Ctenidae) by DNA metabarcoding of gut contents.

Arachnids are the most abundant land predators. Despite the importance of their functional roles as predators and the necessity to understand their diet for conservation, the trophic ecology of many arachnid species has not been sufficiently studied. In the case of the wandering spider, Phoneutria boliviensis F. O. Pickard-Cambridge, 1897, only field and laboratory observational studies on their diet exist. By using a DNA metabarcoding approach, we compared the prey found in the gut content of males and females from three distant Colombian populations of P. boliviensis. By DNA metabarcoding of the cytochrome c oxidase subunit I (COI), we detected and identified 234 prey items (individual captured by the spider) belonging to 96 operational taxonomic units (OTUs), as prey for this wandering predator. Our results broaden the known diet of P. boliviensis with at least 75 prey taxa not previously registered in fieldwork or laboratory experimental trials. These results suggest that P. boliviensis feeds predominantly on invertebrates (Diptera, Lepidoptera, Coleoptera, and Orthoptera) and opportunistically on small squamates. Intersex and interpopulation differences were also observed. Assuming that prey preference does not vary between populations, these differences are likely associated with a higher local prey availability. Finally, we suggest that DNA metabarcoding can be used for evaluating subtle differences in the diet of distinct populations of P. boliviensis, particularly when predation records in the field cannot be established or quantified using direct observation.

A non-lethal method for studying scorpion venom gland transcriptomes, with a review of potentially suitable taxa to which it can be applied.

Scorpion venoms are mixtures of proteins, peptides and small molecular compounds with high specificity for ion channels and are therefore considered to be promising candidates in the venoms-to-drugs pipeline. Transcriptomes are important tools for studying the composition and expression of scorpion venom. Unfortunately, studying the venom gland transcriptome traditionally requires sacrificing the animal and therefore is always a single snapshot in time. This paper describes a new way of generating a scorpion venom gland transcriptome without sacrificing the animal, thereby allowing the study of the transcriptome at various time points within a single individual. By comparing these venom-derived transcriptomes to the traditional whole-telson transcriptomes we show that the relative expression levels of the major toxin classes are similar. We further performed a multi-day extraction using our proposed method to show the possibility of doing a multiple time point transcriptome analysis. This allows for the study of patterns of toxin gene activation over time a single individual, and allows assessment of the effects of diet, season and other factors that are known or likely to influence intraindividual venom composition. We discuss the gland characteristics that may allow this method to be successful in scorpions and provide a review of other venomous taxa to which this method may potentially be successfully applied.