Variation in Morphology, Diet, and Venom Composition in Crotalus pyrrhus (Cope 1867)

Because rattlesnakes rely heavily on their venom for predation and defense, we can expect selection pressures from several sources to act on its composition, including, ontogeny, sex, and environmental variation. In this dissertation, I summarize the results of four studies of the southwestern speckled rattlesnake (Crotalus pyrrhus). First, I describe two cases of envenomation at separate Arizona localities (Tinajas Altas Mountains, Yuma County, and Phoenix Mountains, Maricopa County). Both patients experienced swelling, but neither demonstrated coagulopathy, thrombocytopenia, or hypofibrinogenemia. The latter patient required amputation of the distal portion of his middle finger. I also investigated variation in morphology, diet, and venom protein composition from 23 populations within six biogeographic regions across the species’ United States range of Arizona, California, and Nevada. For morphological variation, snakes varied in in size among biogeographic regions, with the largest snakes occurring in the Mojave Desert, Transverse Ranges, and Peninsular Ranges of California and Nevada; the smallest snakes restricted to the Lower Colorado River subdivision of the Sonoran Desert; and snakes of intermediate size inhabiting the Colorado Desert (California) and Arizona Uplands subdivision of the Sonoran Desert. I also documented sexual body component dimorphism, with females possessing larger heads and longer trunks than males relative to overall body size. The diet of C. pyrrhus consisted predominately of mammals (80.8%), in particular terrestrial squirrels (39.4%) and the heteromyid rodent genus Chaetodipus (26.9%). An ontogenetic shift occurred from primarily lizards to rodents, but neither biogeographic region nor sex significantly influenced prey class consumed. However, the Tinajas Altas Mountains population apparently relies on birds to a greater extent than other populations. For venom composition, I used reverse-phase liquid chromatography and mass spectrometry to define eight elution regions, each dominated by one to three protein families. Seven elution regions varied among the biogeographic regions and three by sex. The biggest distinction was between venoms of eastern (Arizona) and western (California/Nevada) snakes. We failed to detect Mojave Toxin (or a homolog) in any population. Collectively, these findings document differences in phenotypic and behavioral traits of C. pyrrhus and suggest that variable fitness landscapes provide different fitness optima

Enter the Dragon: The Dynamic and Multifunctional Evolution of Anguimorpha Lizard Venoms

While snake venoms have been the subject of intense study, comparatively little work has been done on lizard venoms. In this study, we have examined the structural and functional diversification of anguimorph lizard venoms and associated toxins, and related these results to dentition and predatory ecology. Venom composition was shown to be highly variable across the 20 species of Heloderma, Lanthanotus, and Varanus included in our study. While kallikrein enzymes were ubiquitous, they were also a particularly multifunctional toxin type, with differential activities on enzyme substrates and also ability to degrade alpha or beta chains of fibrinogen that reflects structural variability. Examination of other toxin types also revealed similar variability in their presence and activity levels. The high level of venom chemistry variation in varanid lizards compared to that of helodermatid lizards suggests that venom may be subject to different selection pressures in these two families. These results not only contribute to our understanding of venom evolution but also reveal anguimorph lizard venoms to be rich sources of novel bioactive molecules with potential as drug design and development lead compounds

Varanid lizard venoms disrupt the clotting ability of human fibrinogen through destructive cleavage

The functional activities of Anguimorpha lizard venoms have received less attention compared to serpent lineages. Bite victims of varanid lizards often report persistent bleeding exceeding that expected for the mechanical damage of the bite. Research to date has identified the blockage of platelet aggregation as one bleeding-inducing activity, and destructive cleavage of fibrinogen as another. However, the ability of the venoms to prevent clot formation has not been directly investigated. Using a thromboelastograph (TEG5000), clot strength was measured after incubating human fibrinogen with and lizard venoms. Clot strengths were found to be highly variable, with the most potent effects produced by incubation with venoms from the and clades. The most fibrinogenolytically active venoms belonged to arboreal species and therefore prey escape potential is likely a strong evolutionary selection pressure. The results are also consistent with reports of profusive bleeding from bites from other notably fibrinogenolytic species, such as . Our results provide evidence in favour of the predatory role of venom in varanid lizards, thus shedding light on the evolution of venom in reptiles and revealing potential new sources of bioactive molecules useful as lead compounds in drug design and development

Intraspecific venom variation in the medically significant Southern Pacific Rattlesnake (Crotalus oreganus helleri): biodiscovery, clinical and evolutionary implications

Due to the extreme variation of venom, which consequently results in drastically variable degrees of neutralization by CroFab antivenom, the management and treatment of envenoming by Crotalus oreganus helleri (the Southern Pacific Rattlesnake), one of the most medically significant snake species in all of North America, has been a clinician's nightmare. This snake has also been the subject of sensational news stories regarding supposed rapid (within the last few decades) evolution of its venom. This research demonstrates for the first time that variable evolutionary selection pressures sculpt the intraspecific molecular diversity of venom components in C. o. helleri. We show that myotoxic beta-defensin peptides (aka: crotamines/small basic myotoxic peptides) are secreted in large amounts by all populations. However, the mature toxin-encoding nucleotide regions evolve under the constraints of negative selection, likely as a result of their non-specific mode of action which doesn't enforce them to follow the regime of the classic predator prey chemical arms race. The hemorrhagic and tissue destroying snake venom metalloproteinases (SVMPs) were secreted in larger amounts by the Catalina Island and Phelan rattlesnake populations, in moderate amounts in the Loma Linda population and in only trace levels by the Idyllwild population. Only the Idyllwild population in the San Jacinto Mountains contained potent presynaptic neurotoxic phospholipase A(2) complex characteristic of Mohave Rattlesnake (Crotalus scutulatus) and Neotropical Rattlesnake (Crotalus durissus terrificus). The derived heterodimeric lectin toxins characteristic of viper venoms, which exhibit a diversity of biological activities, including anticoagulation, agonism/antagonism of platelet activation, or procoagulation, appear to have evolved under extremely variable selection pressures. While most lectin alpha- and beta-chains evolved rapidly under the influence of positive Darwinian selection, the beta-chain lectin of the Catalina Island population appears to have evolved under the constraint of negative selection. Both lectin chains were conspicuously absent in both the proteomics and transcriptomics of the Idyllwild population. Thus, we not only highlight the tremendous biochemical diversity in C. o. venom-arsenal, but we also show that they experience remarkably variable strengths of evolutionary selection pressures, within each toxin class among populations and among toxin classes within each population. The mapping of geographical venom variation not only provides additional information regarding venom evolution, but also has direct medical implications by allowing prediction of the clinical effects of rattlesnake bites from different regions. Such information, however, also points to these highly variable venoms as being a rich source of novel toxins which may ultimately prove to be useful in drug design and development

Canopy venom: proteomic comparison among new world arboreal pit-viper venoms

Central and South American pitvipers, belonging to the genera Bothrops and Bothriechis, have independently evolved arboreal tendencies. Little is known regarding the composition and activity of their venoms. In order to close this knowledge gap, venom proteomics and toxin activity of species of Bothriechis, and Bothrops (including Bothriopsis) were investigated through established analytical methods. A combination of proteomics and bioactivity techniques was used to demonstrate a similar diversification of venom composition between large and small species within Bothriechis and Bothriopsis. Increasing our understanding of the evolution of complex venom cocktails may facilitate future biodiscoveries

The Bold and the Beautiful: a Neurotoxicity Comparison of New World Coral Snakes in the Micruroides and Micrurus Genera and Relative Neutralization by Antivenom

Coral snake envenomations are well characterized to be lethally neurotoxic. Despite this, few multispecies, neurotoxicity and antivenom efficacy comparisons have been undertaken and only for the Micrurus genus; Micruroides has remained entirely uninvestigated. As the USA's supplier of antivenom has currently stopped production, alternative sources need to be explored. The Mexican manufacturer Bioclon uses species genetically related to USA species, thus we investigated the efficacy against Micrurus fulvius (eastern coral snake), the main species responsible for lethal envenomations in the USA as well as additional species from the Americas. The use of Coralmyn® coral snake antivenom was effective in neutralizing the neurotoxic effects exhibited by the venom of M. fulvius but was ineffective against the venoms of Micrurus tener, Micrurus spixii, Micrurus pyrrhocryptus, and Micruroides euryxanthus. Our results suggest that the Mexican antivenom may be clinically useful for the treatment of M. fulvius in the USA but may be of only limited efficacy against the other species studied