Corallus hortulanus testes histology: morphological and reproductive aspects.

In general snakes show differentiate anatomical, biological and behavioral particularities compared to other species. Basic information about the snakes anatomy, physiology and reproductive biology is scarce in several species, making the reproduction a challenge. Thus, the present work aims to evaluate morphological aspects of the Corallus hortulanus testes, correlating these findings with environmental factors and reproductive aspects. The testes of three specimens of Corallus hortulanus were cut to a thickness of 3µm in microtome, stained with 1% toluidine blue, photo documented and described. Seasonality was observed in the sperm production of Corallus hortulanus, with the presence of mature spermatozoa in the wettest and hottest periods of the year, as well as the largest testicular volume in these periods.

Synergistic effects of musking and autohaemorrhaging on the duration of death feigning in dice snakes (Natrix tessellata).

Predation exerts a significant selection pressure on prey, shaping a multitude of traits that serve as antipredator defences. In turn, natural selection could favour combinations of antipredator defences with synergistic effects that enhance prey survival. An especially interesting antipredator defence is death feigning (DF), present in a wide variety of taxa and usually characterized by the prey lying motionless often along with defaecation, musking and autohaemorrhaging (AH). All these aspects of the DF display should work in conjunction with one another, intensifying the overall effect of the display and in turn facilitating quicker escape. To confirm this hypothesis, we tested 263 dice snakes (Natrix tessellata) directly in the field. We noted the occurrence of smearing faeces, musk and AH, and we measured the duration of DF, expecting to see a negative association between the occurrence of these behaviours and the duration of DF. Our results affirm our hypothesis: dice snakes that smeared themselves in musk and faeces prior to DF and had AH during DF spent significantly less time in DF. Our results highlight the functional integration of antipredator behaviours across different phases of predator-prey interactions, emphasizing the need for future research to prioritize studying the sequential display of behaviours.

Natrix natrix after dark: citizen science sheds light on the common grass snake's nightlife.

Activity patterns in animals are often species-specific, and can be generally categorized as diurnal, crepuscular, or nocturnal. Understanding these patterns provides insight into ecological adaptations and behaviors. The common grass snake (Natrix natrix), one of the most common and widespread European snake species, is traditionally considered diurnal, with scarce evidence of its crepuscular and nocturnal activity. We aimed to document the distribution, environmental conditions, and potential phenotype associations of nighttime activity in N. natrix. We used citizen science data from iNaturalist (1992-2022), Observation.org (2012-2022), together with personal field observations (2010-2023) to collect 127 crepuscular and nocturnal activity records. Most observations occurred between May and August, coinciding with the peak activity period of grass snakes across their distribution range. Statistical analyses revealed no significant difference in mean daily temperatures between crepuscular and nocturnal observations. However, striped individuals displayed nocturnal activity at higher temperatures, consistent with their distribution in warmer regions, but failed to register any difference when tested on a geographic subsample, that accounted for sympatry of the phenotypes. Surprisingly, we found no significant impact of moon presence or moonlight on nighttime activity or age class, contrary to expectations based on other snake species' responses. While our study reveals that nocturnal activity in the common grass snake is geographically widespread, further research is warranted to understand its drivers and ecological implications. This study highlights the value of citizen science platforms for biological and ecological research, offering unparalleled spatial and temporal coverage by their users. In conclusion, our work extends the knowledge of nocturnal behavior in N. natrix and underlines the critical role of citizen science in discovering behavioral aspects of common and widespread species.

Dynamic gap crossing in Dendrelaphis, the sister taxon of flying snakes.

Arboreal animals commonly use dynamic gap-crossing behaviors such as jumping. In snakes, however, most species studied to date only employ the quasi-static cantilever crawl, which involves a whole-body reach. One exception is the paradise tree snake (Chrysopelea paradisi), which exhibits kinematic changes as gap distance increases, culminating in dynamic behaviors that are kinematically indistinguishable from those used to launch glides. Because Chrysopelea uses dynamic behaviors when bridging gaps without gliding, we hypothesized that such dynamic behaviors evolved ancestrally to Chrysopelea. To test this predicted occurrence of dynamic behaviors in closely related taxa, we studied gap bridging locomotion in the genus Dendrelaphis, which is the sister lineage of Chysopelea. We recorded 20 snakes from two species (D. punctulatus and D. calligastra) crossing gaps of increasing size, and analyzed their 3D kinematics. We found that, like C. paradisi, both species of Dendrelaphis modulate their use of dynamic behaviors in response to gap distance, but Dendrelaphis exhibit greater inter-individual variation. Although all three species displayed the use of looped movements, the highly stereotyped J-loop movement of Chrysopelea was not observed in Dendrelaphis. These results support the hypothesis that Chrysopelea may have co-opted and refined an ancestral behavior for crossing gaps for the novel function of launching a glide. Overall, these data demonstrate the importance of gap distance in governing behavior and kinematics during arboreal gap crossing.

Material properties of skin in the flying snake Chrysopelea ornata.

In snakes, the skin serves for protection, camouflage, visual signaling, locomotion, and its ability to stretch facilitates large prey ingestion. The flying snakes of the genus Chrysopelea are capable of jumping and gliding through the air, requiring additional functional demands: its skin must accommodate stretch in multiple directions during gliding and, perhaps more importantly, during high-speed, direct-impact landing. Is the skin of flying snakes specialized for gliding? Here, we characterized the material properties of the skin of Chrysopelea ornata and compared them with two nongliding species of colubrid snakes, Thamnophis sirtalis and Pantherophis guttatus, as well as with previously published values. The skin was examined using uniaxial tensile testing to measure stresses, and digital image correlation methods to determine strains, yielding metrics of strength, elastic modulus, strain energy, and extensibility. To test for loading orientation effects, specimens were tested from three orientations relative to the snake's long axis: lateral, circumferential, and ventral. Specimens were taken from two regions of the body, pre- and pos-tpyloric, to test for regional effects related to the ingestion of large prey. In comparison with T. sirtalis and P. guttatus, C. ornata exhibited higher post-pyloric and lower pre-pyloric extensibility in circumferential specimens. However, overall there were few differences in skin material properties of C. ornata compared to other species, both within and across studies, suggesting that the skin of flying snakes is not specialized for gliding locomotion. Surprisingly, circumferential specimens demonstrated lower strength and extensibility in pre-pyloric skin, suggesting less regional specialization related to large prey.

Perils of ingesting harmful prey by advanced snakes.

The advanced snakes (Alethinophidia) include the extant snakes with a highly evolved head morphology providing increased gape and jaw flexibility. Along with other physiological and morphological adaptations, this allows them to immobilize, ingest, and transport prey that may be disproportionately large or presents danger to the predator from bites, teeth, horns, or spines. Reported incidents of snakes failing to consume prey and being injured or killed during feeding mostly reflect information in the form of natural-history notes. Here we provide the first extensive review of such incidents, including 101 publications describing at least 143 cases of mortality (including six of 'multiple individuals') caused by ingestion or attempted consumption of injurious prey. We also report on 15 previously unpublished injurious feeding incidents from the USA, Austria, and Bulgaria, including mortality of five juvenile piscivorous dice snakes (Natrix tessellata) from a single location. Occurrences are spread across taxa, with mortality documented for at least 73 species from eight families and 45 genera. Incidents were generally well represented within each of three major categories: oversized prey (40.6%), potentially harmful prey (40.6%), and predator's behavioural/mechanical errors (18.9%). Reptile (33%) and fish (26%) prey caused disproportionately high mortality compared to mammals (16%). Feeding can be dangerous throughout a snake's life, with the later stages of feeding likely being more perilous. The number of reports has increased over time, and the data seem biased towards localities with a higher number of field-working herpetologists. We propose a standardized framework, comprising a set of basic information that should ideally be collected and published, and which could be useful as a template for future data collection, reporting, and analyses. We conclude that incidents of mortality during feeding are likely to be more common than previously assumed, and this hypothesis has implications for the ecology of persistence where populations are impacted by changing trophic environments.

Invasive brown treesnakes (Boiga irregularis) move short distances and have small activity areas in a high prey environment.

Animal movements reflect temporal and spatial availability of resources as well as when, where, and how individuals access such resources. To test these relationships for a predatory reptile, we quantified the effects of prey abundance on the spatial ecology of invasive brown treesnakes (Boiga irregularis) on Guam. Five months after toxicant-mediated suppression of a brown treesnake population, we simultaneously used visual encounter surveys to generate relative rodent abundance and radiotelemetry of snakes to document movements of surviving snakes. After snake suppression, encounter rates for small mammals increased 22-fold and brown treesnakes had smaller mean daily movement distances (24 ± 13 m/day, [Formula: see text] ± SD) and activity areas (5.47 ± 5 ha) than all previous observations. Additionally, snakes frequenting forest edges, where our small mammal encounters were the highest, had smaller mean daily movement distances and three-dimensional activity volumes compared to those within the forest interior. Collectively, these results suggest that reduced movements by snakes were in part a response to increased prey availability. The impact of prey availability on snake movement may be a management consideration when attempting to control cryptic invasive species using tools that rely on movement of the target species to be effective.

THE EFFECT OF VARIED ENRICHMENT TYPES ON SNAKE BEHAVIOR.

Environmental enrichment is a strategy used to improve the welfare of animals under human care. While enrichment techniques for mammals and birds have been studied extensively, reptilian enrichment has received less attention. There has been an increase in enrichment programs for reptiles in zoological institutions, however many are not accompanied by behavioral studies. Detailed recording of behavioral responses to enrichment is necessary to assess the efficacy of the enrichment type and to determine its utility in various settings. In this study, 18 snakes of multiple species, from two Families (Colubridae, Pythonidae), were exposed to four enrichment types (Humid Hide, Olfactory, Climbing, Suspended Hide). Baseline recordings were conducted prior to the introduction of enrichment. Snakes were recorded for two hours after introduction of each item. Five behavior types were identified based on baseline videos: tongue flicking, climbing, hiding, interacting with transparent boundaries, and utilizing non-enrichment items. Interacting with transparent boundaries was classified as an undesirable behavior, while the other four behaviors were classified as desirable. Changes in climbing and tongue flicking behaviors were noted with introduction of each item- these changes were not statistically significant. The increase in these behaviors may indicate clinical importance, and shows that snakes under human care respond to environmental enrichment. As some snakes showed a reduction in undesirable behaviors when compared to baseline conditions, this may suggest increased welfare during times when enrichment is offered. The extent to which these results can be applied to other species merits further study.

Chemical Ecology of the North American Newt Genera Taricha and Notophthalmus.

The North American newt genera Taricha and Notophthalmus (order Caudata) are well known for the combination of potent toxicity, aposematic coloration, and striking defense postures that protects these animals from predation. This suite of traits is centered around the neurotoxin tetrodotoxin, which causes paralysis and death in metazoans by disrupting the initiation and propagation of electrical signals in the nerves and muscles. Tetrodotoxin defends newts from predation across multiple life history stages and its role in generating arms-race coevolution between Taricha newts and garter snake (genus Thamnophis) predators is well studied. However, understanding the broader picture of chemical defenses in Taricha and Notophthalmus requires an expanded comprehension of the defensive chemical ecology of tetrodotoxin that includes possible coevolutionary interactions with insect egg predators, protection against parasites, as well as mimicry complexes associated with tetrodotoxin and aposematic coloration in both genera. Herein the authors review what is known about the structure, function, and pharmacology of tetrodotoxin to explore its evolution and chemical ecology in the North American newt. Focus is made specifically on the origin and possible biosynthesis of tetrodotoxin in these taxa as well as providing an expanded picture of the web of interactions that contribute to landscape level patterns of toxicity and defense in Taricha and Notophthalmus.

The molecular basis of venom resistance in the non-venomous snake Sinonatrix annularis.

Various snake species and snake predators have natural neutralization against snake toxins, which their antidotal abilities are commonly attributed to the intrinsic inhibitors produced by the liver, e.g., phospholipase A2 inhibitor (PLI) and metalloproteinase inhibitor (SVMPI). Sinonatrix annularis was found to possess broad-spectrum neutralization to different snake venoms in our lab. Although the anti-venom compound PLIγ has been previously characterized in our laboratory, the mechanism of resistance of S. annularis to snake venoms remains obscure. In this research, a venom affinity chromatography was constructed by immobilizing D. acutus venom to NHS-agarose beads and applied for antitoxins mining from S. annularis. The binding capacity of the venom column was validated using a self-prepared rabbit antivenom against D. acutus. Serum and liver homogenate of S. annularis were then applied to the column, the bound components were profiled using SDS-PAGE and mass spectrometry. PLIs, snake venom metalloproteins inhibitor (SVMPI), small serum protein (SSP), heat shock proteins, etc were identified. To identify their toxin targets in D. acutus venom, a reverse separation was conducted by coupling the fractionated S. annularis serum proteins to NHS-agarose beads. Fifteen toxins of five families were captured and identified as follows: PLA2s, metalloproteinases, cysteine-rich secretory proteins, snake venom serine proteinases, and C-type lectins. These discoveries increased our understanding of the capacity and mechanism of the natural neutralization of S. annularis to snake venom. These natural inhibitors are medically significant due to their powerful and broad antidotal activities, which may provide alternative and promising drug candidates for snakebite treatment.

Over a decade of field physiology reveals life-history specific strategies to drought in garter snakes (Thamnophis legans).

Changing climates and severe weather events can affect population viability. Individuals need to buffer such negative fitness consequences through physiological plasticity. Whether certain life-history strategies are more conducive to surviving changing climates is unknown, but theory predicts that strategies prioritizing maintenance and survival over current reproduction should be better able to withstand such change. We tested this hypothesis in a meta-population of garter snakes having naturally occurring variation in life-history strategies. We tested whether slow pace-of-life (POL) animals, that prioritize survival over reproduction, are more resilient than fast POL animals as measured by several physiological biomarkers. From 2006 to 2019, which included two multi-year droughts, baseline and stress-induced reactivity of plasma corticosterone and glucose varied annually with directionalities consistent with life-history theory. Slow POL animals exhibited higher baseline corticosterone and lower baseline glucose, relative to fast POL animals. These patterns were also observed in stress-induced measures; thus, reactivity was equivalent between ecotypes. However, in drought years, measures of corticosterone did not differ between different life histories. Immune cell distribution showed annual variation independent of drought or life history. These persistent physiological patterns form a backdrop to several extirpations of fast POL populations, suggesting a limited physiological toolkit to surviving periods of extreme drought.

Decreased attractivity in female garter snakes treated with an aromatase inhibitor.

Most experimental studies on sexual signal regulation via hormone manipulation have focused on male signals, yet female signals demonstrate substantial phenotypic variation and hormone-dependent expression. Female red-sided garter snakes (Thamnophis sirtalis parietalis) produce a skin-based sex pheromone used by males in mate selection. The principle female sex steroid, 17 ß-estradiol, controls pheromone production in snakes, but studies manipulating female garter snakes have produced conflicting results, relied on behavioral tests with males in the laboratory, and did not quantify pheromone expression. Because aromatase is the terminal enzyme in estradiol biosynthesis, we hypothesized that female garter snakes rely on aromatase to ultimately control pheromone production during the annual cycle of this species. To test this, we used a known pharmacological inhibitor of aromatase, fadrozole (FAD). Wild-caught female garter snakes were chronically treated via subcutaneous injections of either FAD (1.0 mg kg-1 ) or saline (control) for six months in the laboratory during the active period of the annual cycle then hibernated. In two separate field bioassays the next spring at the den site, FAD females received approximately 50% less courtship from wild, sexually active male garter snakes compared to SHAM females. Pheromone analysis revealed that four of the largest, unsaturated methyl ketones were specifically downregulated in FAD females, indicating that aromatase action is a crucial, permissive step in the maintenance of female attractivity.

Testing the febrile response of snakes inoculated with Ophidiomyces ophidiicola, the causative agent of snake fungal disease.

Snake Fungal Disease (SFD) negatively impacts wild snake populations in the eastern United States and Europe. Ophidiomyces ophidiicola causes SFD and manifests clinically by the formation of heterophilic granulomas around the mouth and eyes, weight loss, impaired vision, and sometimes death. Field observations have documented early seasonal basking behaviors in severely infected snakes, potentially suggesting induction of a behavioral febrile response to combat the mycosis. This study tested the hypothesis that snakes inoculated with Ophidiomyces ophidiicola would seek elevated basking temperatures to control body temperature and behaviorally induce a febrile response. Eastern ribbon snakes (Thamnophis saurita, n = 29) were experimentally or sham inoculated with O. ophidiicola. Seven days after inoculation, snakes were tested on a thermal gradient and the internal body temperature and substrate temperature of each snake was recorded over time. Quantitative PCR was used when snakes arrived, during pre-inoculation, and post-inoculation to test snakes for the presence of O. ophidiicola. Some snakes arrived with O. ophidiicola and were subsequently inoculated, allowing for an assessment of secondary exposure effects. Snake thermoregulatory behavior was compared between 1) O. ophidiicola inoculated vs. sham inoculated treatments, 2) infected vs. disease negative groups, and 3) disease naïve vs. pre-exposed immune response categories. Neither internal nor substrate temperatures differed among initially prescribed, and qPCR recovered disease states, although infected snakes tended to reach a preferred body temperature faster than disease negative snakes. Snakes experiencing their first exposure (disease naïve) sought higher substrate temperatures than snakes experiencing their second exposure (pre-exposed). Here, we recover no evidence for behaviorally induced fever in snakes with SFD but do elucidate a febrile immune response associated with secondary exposure.

Lasso locomotion expands the climbing repertoire of snakes.

The diverse ways and environments in which animals move are correlated with morphology1, but morphology is not sufficient to predict how animals move because behavioral innovations can create new capacities. We document a new mode of snake locomotion - 'lasso locomotion' - that allows the brown treesnake (Boiga irregularis) to ascend much larger smooth cylinders than any previously known behavior. This lasso locomotion may facilitate exploiting resources that might otherwise be unobtainable and contribute to the success and impact of this highly invasive species. VIDEO ABSTRACT.

Embryonic heart rate correlates with maternal temperature and developmental stage in viviparous snakes.

Interactions between the environment and maternal and embryonic physiology can have critical ramifications for early-life phenotypes and survival in a range of species. A major component of the environment-maternal-embryonic nexus is the regulation of embryonic heart rate, which can have important ramifications for developmental phenology, but remains relatively unexplored in viviparous reptiles. The goal of this study was to test for a relationship between embryonic heart rate and maternal body temperature in two species of viviparous garter snakes. The embryonic heart rates of Thamnophis elegans and T. sirtalis were assessed using a field-portable ultrasound. For both T. elegans and T. sirtalis, embryonic heart rate was strongly correlated to maternal temperature. Interestingly, there was also a strong correlation between embryonic and maternal heart rate that was most likely mediated by a common response to maternal body temperature, in spite of the effects of handling during ultrasound on maternal heart rate. Furthermore, embryos at earlier developmental stages had lower heart rates. To our knowledge, this study is the first to explore embryonic heart rate in viviparous reptiles, providing a foundation for future work using ultrasonography to test ecological and evolutionary hypotheses related to developmental dynamics in free-ranging viviparous species.

Effect of Capture, Phenotype, and Physiological Status on Blood Glucose and Plasma Corticosterone Levels in Free-Ranging Dice Snakes.

AbstractTheoretically, animals integrate intrinsic and extrinsic factors to respond appropriately to the wide range of stressors they encounter during their life span. We examined how stress response varies between sexes and among morphotypes in wild dice snakes (Natrix tessellata). We also considered reproductive and feeding status and antipredator behavior. We used two indicators of stress (glucose [GLUC] and corticosterone [CORT] levels) at eight sampling time intervals (immediately after capture, up to 17 h after) and a large sample size (N=113 snakes). Concentrations of both markers increased sharply after capture (an equivalent of predation). This acute phase occurred earlier for GLUC (30 min) compared to CORT (60 min). Then the values plateaued to very high levels without decline over time, indicating prolonged saturation of the hypothalamus-pituitary-adrenal axis. In contrast to our expectations, we found no effect of sex, morphotype, or reproductive status. Yet the CORT stress response of those individuals displaying death-feigning (DF) antipredator behavior was attenuated compared to those that did not. Low stress hormones levels may facilitate the expression of DF (high levels supporting fleeing behavior). The presence of partially digested material in the stomach was associated with higher blood GLUC during the plateau. Assaying blood GLUC requires very little blood but was as good as CORT at gauging acute stress response. The prolonged plateau suggests that captivity should be minimized during field studies.

High-elevation hypoxia impacts perinatal physiology and performance in a potential montane colonizer.

Climate change is generating range shifts in many organisms, notably along the elevational gradient in mountainous environments. However, moving up in elevation exposes organisms to lower oxygen availability, which may reduce the successful reproduction and development of oviparous organisms. To test this possibility in an upward-colonizing species, we artificially incubated developing embryos of the viperine snake (Natrix maura) using a split-clutch design, in conditions of extreme high elevation (hypoxia at 2877 m above sea level; 72% sea-level equivalent O2 availability) or low elevation (control group; i.e. normoxia at 436 m above sea level). Hatching success did not differ between the two treatments. Embryos developing at extreme high elevation had higher heart rates and hatched earlier, resulting in hatchlings that were smaller in body size and slower swimmers compared to their siblings incubated at lower elevation. Furthermore, post-hatching reciprocal transplant of juveniles showed that snakes which developed at extreme high elevation, when transferred back to low elevation, did not recover full performance compared to their siblings from the low elevation incubation treatment. These results suggest that incubation at extreme high elevation, including the effects of hypoxia, will not prevent oviparous ectotherms from producing viable young, but may pose significant physiological challenges on developing offspring in ovo. These early-life performance limitations imposed by extreme high elevation could have negative consequences on adult phenotypes, including on fitness-related traits.

Mandibular sawing in a snail-eating snake.

The jaws of vertebrates display a striking diversity in form and function, but they typically open and close like a trapdoor rather than sliding like a saw. Here, we report unique feeding behaviour in the blunt-headed snail-eating snake, Aplopeltura boa (family Pareidae), where the snake cuts off and circumvents the indigestible part (the operculum) of its prey in the mouth using long sliding excursions of one side of the mandible, while the upper jaws and the mandible on the other side maintain a stable grasp on the prey. This behaviour, which we call 'mandibular sawing', is made possible by extraordinarily independent movements of the jaw elements and is a surprising departure from usual feeding behaviour in vertebrates.

Energy Metrics of Red-Sided Garter Snakes (Thamnophis sirtalis parietalis) Vary with Sex but Not Life-History Stage.

Because reproduction is energetically expensive, an organism's energy stores are likely involved in mediating transitions between reproductive and self-maintenance activities. We investigated whether body condition index, adipocyte follicle size, and liver glycogen differ with the life-history transition from reproduction to migration and foraging in red-sided garter snakes (Thamnophis sirtalis parietalis). Females primarily investing in mating behavior located at the den had a significantly higher body condition index than females migrating to summer feeding grounds. The body condition index of male snakes did not differ between snakes located at the den and those migrating to summer feeding grounds. Neither adipocyte follicle area nor liver glycogen stores differed significantly between snakes performing mating activities at the den and those migrating to summer feeding grounds. We did find a sexual dimorphism in that female red-sided garter snakes had significantly larger adipocyte follicles and higher liver glycogen compared with males. Our findings support the across-species phenomenon of females and males displaying a sexual dimorphism in stored energy substrates. Conversely, we did not find evidence to suggest that red-sided garter snakes primarily utilize fatty acids to fuel the initiation of migration, a finding that is not consistent with other long-distance migrators, such as birds. Because we did not find evidence to suggest that stored energy metrics influence the decision to migrate, a physiological mechanism that induces migration in red-sided garter snakes remains elusive.

Melanism, body size, and sex ratio in snakes-new data on the grass snake (Natrix natrix) and synthesis.

It is postulated that melanism in ectotherms is adaptive by enhancing thermoregulation, subsequent resource acquisition, and growth. Such effects may differ between the sexes as a result of the differential costs of self-maintenance and reproduction, but empirical support for the sex-specific consequences of melanism remains inconsistent. We studied the effects of melanism on body size and sex ratio in a population of the European grass snake (Natrix natrix) in SE Poland and also carried out a systematic review of the literature on the consequences of melanism in terrestrial snakes. Melanistic grass snakes of both sexes appeared to be smaller than the typical phenotype, which indicates higher predation pressure and minimal thermal benefits for black individuals. A female-biased sex ratio was observed in the typical phenotype, but not in melanistic snakes, suggesting that the costs for females and/or benefits for males are higher in melanistic individuals. In conjunction with earlier studies, our data indicate that the consequences of melanism may be related to the reproductive mode of species. In viviparous species, melanism tends to improve growth and/or body size and is more frequent in females, whereas the opposite holds for oviparous snakes. Further studies on melanism should examine a wider array of species with different reproductive strategies and traits beyond the usual thermal benefits.