Article

Venom Ontogeny in the Pacific Rattlesnakes Crotalus viridis helleri and C. v. oreganus

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Abstract

Ontogenetic variation in venom composition was examined in the Pacific rattlesnakes Crotalus viridis helleri and C. v. oreganus. Venoms were analyzed for protease, phospholipase A2, L-amino acid oxidase, exonuclease and elastinolytic activities, and toxicity toward a native prey (Sceloporus graciosus). Protease activity increased significantly with size; L-amino acid oxidase and exonuclease activities also tended to increase. Phospholipase A2 activity decreased significantly with size, as did venom toxicity. These factors produce a highly toxic venom with low protease activity in juvenile snakes, which facilitates efficient handling of lizards and young rodents. Analysis of gut contents of museum specimens showed that lizards constitute a major fraction of prey taken by juvenile rattlesnakes. Lizards continue to be taken with high frequency until snakes reach approx. 500 mm in total length; above this size, mammals are taken exclusively. As snakes increase in size, they feed on larger mammalian prey, and a functionally different venom is produced. Venom from adult Pacific rattlesnakes is less toxic but has high protease activity, aiding in the digestion of prey in a thermally variable environment.

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... The increase of venom proteolytic activity as age advances has already been observed in generalist species present in this study and in other species of snakes (Andrade; Abe, 1999; [23,36]). Some authors suggest this differentiation may be due to the shift in animals' diet (Andrade; Abe, 1999; [23,36]). ...
... The increase of venom proteolytic activity as age advances has already been observed in generalist species present in this study and in other species of snakes (Andrade; Abe, 1999; [23,36]). Some authors suggest this differentiation may be due to the shift in animals' diet (Andrade; Abe, 1999; [23,36]). However, a change in this activity relative to sex may perhaps be related to the fact that females generate offspring and might need greater defense mechanisms, since the main symptomatology caused by proteases (SVMP and SVSP) involves tissue degradation, coagulant, hemorrhagic and fibrinogenolytic action ( [5]; Secretaria Da Saúde Do Estado De São Paulo, 1993 [7]; Andrade; Abe, 1999 [36]; Gutiérrez et al., 2006;[6]). ...
... Some authors suggest this differentiation may be due to the shift in animals' diet (Andrade; Abe, 1999; [23,36]). However, a change in this activity relative to sex may perhaps be related to the fact that females generate offspring and might need greater defense mechanisms, since the main symptomatology caused by proteases (SVMP and SVSP) involves tissue degradation, coagulant, hemorrhagic and fibrinogenolytic action ( [5]; Secretaria Da Saúde Do Estado De São Paulo, 1993 [7]; Andrade; Abe, 1999 [36]; Gutiérrez et al., 2006;[6]). ...
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The genus Bothrops are considered Category 1 of medical importance by the World Health Organization, responsible for approximately 85 % of snakebites occurring throughout Brazil. Main factors determining snake venom variations can be genetics, diet, gender, geographic distribution, age, or even seasonality. In this study, we compared the composition of protein profile, biochemical activities, and immunorecognition of toxins present in the venom of eight adults of Bothrops species (B. alternatus, B. atrox, B. jararaca, B. jararacussu, B. leucurus, B. moojeni, B. neuwiedi and B. pauloensis). The following methods were used to analyze the venoms: protein dosage; electrophoresis in polyacrylamide gel containing SDS; High Performance Liquid Chromatography – Reverse Phase; enzymatic activities, western blotting and Enzyme Linked Immuno Sorbent Assay. The results show inter and intraspecific differences in the electrophoretic profile. LAAO and PLA2 activities, in general, were higher in males than females and proteolytic activity was higher in females than males. The bothropic antivenom produced by Instituto Butantan recognized most of the protein bands in all Bothrops species analyzed, with only the regions between 37 and 25 kDa presenting lower intensity. A notable variability in the chromatograms was observed. Bothrops venom demonstrated inter-intraspecific disparities in protein composition and biochemical activity.
... Overall, IC 50 estimates also highlighted that the Premium Serums antivenom exhibited better recognition potential against the N. naja and D. russelii venoms (Additional file 1: Fig S11). Therefore, based on the titre values and the IC 50 estimates, the best binding antivenom (Premium Serums) was further downselected for animal protection studies. ...
... For instance, given the gradual increase in gape size associated with the developmental stage of snakes, variations in diet has been reported [48,49]. In turn, to facilitate such shifts in diet, ontogenetic shifts in venoms have also been reported [50,51]. For example, in the case of certain Crotalus spp. ...
... Eventually, as adults, with an increased gape size, they could shift to feeding on larger mammals. Field observations in crotaline snakes have also identified a diet shift from ectothermic (arthropod, reptilian or amphibian) to endothermic (mammalian) prey with age, which is underpinned by a corresponding shift in venom profiles [50,55,68,69]. Behavioural observations in this study point to a similar shift in diet from ectotherms (lizards and frogs) to endotherms (rodents) in D. russelii. ...
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Background Snake venoms can exhibit remarkable inter- and intraspecific variation. While diverse ecological and environmental factors are theorised to explain this variation, only a handful of studies have attempted to unravel their precise roles. This knowledge gap not only impedes our understanding of venom evolution but may also have dire consequences on snakebite treatment. To address this shortcoming, we investigated the evolutionary ecology of venoms of Russell’s viper (Daboia russelii) and spectacled cobra (Naja naja), India’s two clinically most important snakes responsible for an alarming number of human deaths and disabilities. Methodology Several individuals (n = 226) of D. russelii and N. naja belonging to multiple clutches (n = 9) and their mothers were maintained in captivity to source ontogenetic stage-specific venoms. Using various in vitro and in vivo assays, we assessed the significance of prey, ontogeny and sex in driving venom composition, function, and potency. Results Considerable ontogenetic shifts in venom profiles were observed in D. russelii, with the venoms of newborns being many times as potent as juveniles and adults against mammalian (2.3–2.5 ×) and reptilian (2–10 ×) prey. This is the first documentation of the ontogenetic shift in viperine snakes. In stark contrast, N. naja, which shares a biogeographic distribution similar to D. russelii, deployed identical biochemical cocktails across development. Furthermore, the binding kinetics of cobra venom toxins against synthetic target receptors from various prey and predators shed light on the evolutionary arms race. Conclusions Our findings, therefore, provide fascinating insights into the roles of ecology and life history traits in shaping snake venoms.
... The geographic variation of snake venom is well known in several rattlesnake species [20,30,31] as well as ontogenetic changes [30,[32][33][34][35], yet the evolutionary cause of these patterns remains unclear. One of the most supported hypotheses to explain venom variation is the coevolution of venom and prey [31,36,37]. ...
... On the other hand, the evaluation of antivenom neutralization needs to be documented even when a species causes few bites [39]. While no clinical cases have been reported for Crotalus helleri caliginis [33], it is important to know the composition of its venom and understand the changes in the venom in a species that has been isolated for years. There is a permanent Mexican Navy presence, the island is remote, and the snake densities are high, therefore it is important to document if the Mexican antivenoms are effective in neutralizing the venom in case of an ophidian accident. ...
... As the composition of the diet of C. helleri caliginis on the island differs from the diet of C. helleri helleri on the mainland, our hypothesis was that their venom would show some differences among the two subspecies [42]. Previous studies showed that C. helleri helleri and other closely related species of rattlesnakes show ontogenetic changes in the venom [33,43], therefore, we also expected to see ontogenetic changes in the venom of C. helleri caliginis. This knowledge will be important to understand the evolutionary changes in this insular species in a geographically isolated region and their possible clinical implications. ...
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The Baja California Peninsula has over 250 islands and islets with many endemic species. Among them, rattlesnakes are the most numerous but also one of the least studied groups. The study of island rattlesnake venom could guide us to a better understanding of evolutionary processes and the description of novel toxins. Crotalus helleri caliginis venom samples were analyzed to determine possible ontogenetic variation with SDS-PAGE in one and two dimensions and with RP-HPLC. Western Blot, ELISA, and amino-terminal sequencing were used to determine the main components of the venom. The biological and biochemical activities demonstrate the similarity of C. helleri caliginis venom to the continental species C. helleri helleri, with both having low proteolytic and phospholipase A2 (PLA2) activity but differing due to the absence of neurotoxin (crotoxin-like) in the insular species. The main components of the snake venom were metalloproteases, serine proteases, and crotamine, which was the most abundant toxin group (30–35% of full venom). The crotamine was isolated using size-exclusion chromatography where its functional effects were tested on mouse phrenic nerve–hemidiaphragm preparations in which a significant reduction in muscle twitch contractions were observed. The two Mexican antivenoms could neutralize the lethality of C. helleri caliginis venom but not the crotamine effects.
... The fact that rattlesnakes are more likely to hold onto smaller prey compared with larger prey indicates that the snakes recognize, to some extent, the retaliatory capacity of prey based on body size (Allon and Kochva 1974;Kardong 1986a). Mackessy (1988) discovered an ontogenetic shift in both prey handling and venom chemistry in C. oreganus. He found that juveniles more often struck-and-held lizards (Uta stansburiana) and neonate mice (species not given, but presumably M. musculus), and began to strike-and-release comparatively more dangerous rodent prey as the snakes attained a larger body size. ...
... In Hayes (1991), C. viridis never held struck prey (deer mice, Peromyscus maniculatus), irrespective of snake or prey size, although small snakes gripped deer mice for slightly longer durations before releasing them. Hayes (1991) explained the disparity between the prey-handling behavior of C. viridis in his study and C. oreganus in Mackessy (1988) with reference to the different dietary habits of these populations. The C. oreganus from Mackessy (1988) primarily feed on lizards, whereas the C. viridis from Hayes (1991) primarily feed on mice. ...
... Hayes (1991) explained the disparity between the prey-handling behavior of C. viridis in his study and C. oreganus in Mackessy (1988) with reference to the different dietary habits of these populations. The C. oreganus from Mackessy (1988) primarily feed on lizards, whereas the C. viridis from Hayes (1991) primarily feed on mice. Lizards are generally not immobilized as quickly by venom, so the cost of releasing them might be greater because they could presumably travel further distances once released, making relocation more difficult. ...
Article
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Chemosensory searching in squamates with specialized tongue–vomeronasal systems is well-documented. By tongue-flicking, these reptiles gather important chemical cues from their environment to guide their feeding behavior. Strike-induced chemosensory searching (SICS) is a specific expression of chemosensory reception that is central to the predatory strategy used by venomous reptiles that strike, release, and relocate their prey. This complex behavioral process consists of multiple sequential steps and has been mainly studied in viperids, particularly rattlesnakes. Although this phenomenon has been extensively researched, there is no comprehensive review of the SICS literature. Here, we provide such a review, centered on the idea that SICS is a result of suppression, and then enhancement, of chemosensory searching that serves as a key element of the ambush hunting strategy that most viperid snakes use to consume large and well-defended prey. SICS is also present in other venomous and nonvenomous taxa, and we include a taxonomic categorization of SICS studies in our review. We summarize the key findings discovered during decades of research into this remarkable feeding behavior and highlight areas where our knowledge remains incomplete in an effort to foster further research that will increase our understanding of reptilian feeding ecology.
... From an animal biology perspective, the link between venom variations and shifts in dietary habits between species and populations in snakes is well established (Daltry et al., 1996;Barlow et al., 2009;Cipriani et al., 2017;Lyons et al., 2020), with venom and resistance to toxins in prey animals continuously driving each other's evolution in what has often been described as an arms race (Casewell et al., 2013;Fry et al., 2008;Jackson et al., 2016). Examples of such a marked influence of diet on venom abound in rattlesnakes (Mackessy, 1988;Sanz et al., 2006;Mackessy, 2010;Sunagar et al., 2014;Saviola et al., 2017;Holding et al., 2018). However, prey specificity is unlikely to be the only driver of venom diversification in this clade, since significant differences in venom composition can be observed in populations and species characterized by similar prey preferences (Mackessy, 2008;Zancolli et al., 2019). ...
... Furthermore, venom variation in several species of rattlesnakes is known to be ontogenetic, with juveniles generally possessing a Type II venom phenotype that gradually transitions to Type I in adults (Mackessy et al., 2003;Calvete et al., 2010). While this process often reflects ontogenetic dietary shifts (Mackessy, 1988;Gibbs et al., 2011), age-related modifications in venom composition have been documented in species that likely feed on the same prey type throughout their life (Calvete et al., 2010;Seneci et al., 2021). Consequently, a concatenation of multiple factors, from environmental variables ranging from local climate and elevation (Zancolli et al., 2019) to interspecific and even intergeneric hybridization (Dowell et al., 2016, Dowell et al., 2018, but see Zancolli et al., 2016), is likely at the root of inter-and intraspecific variation in venom composition in rattlesnakes. ...
... Rattlesnakes are a prime example of this, as they display remarkably high genetic and species diversity and have successfully colonized a variety of ecosystems throughout their vast range. Specialization of venom activity towards a specific prey item is documented for numerous species such as C. pricei (Grabowsky and Mackessy, 2019), C. helleri (Mackessy, 1988;Holding et al., 2016;Holding et al., 2018;Gibbs et al., 2020), C. lepidus, C. willardi (Holycross et al., 2012;Martínez-Romero et al., 2013;Saviola et al., 2017) and Sistrurus as a whole (Sanz et al., 2006;Gibbs and Mackessy, 2009;Gibbs et al., 2011;Gibbs et al., 2013). Our findings did reveal widespread intraspecific and/or intra-lineage variation in venom activity in many clades (Figs. 1 & 2), although it must be emphasized that we performed our analysis entirely on human plasma and fibrinogen when several rattlesnakes specialize on non-mammalian prey whose physiology may be radically different from ours. ...
Article
What factors influence the evolution of a heavily selected functional trait in a diverse clade? This study adopts rattlesnakes as a model group to investigate the evolutionary history of venom coagulotoxicity in the wider context of phylogenetics, natural history, and biology. Venom-induced clotting of human plasma and fibrinogen was determined and mapped onto the rattlesnake phylogenetic tree to reconstruct the evolution of coagulotoxicity across the group. Our results indicate that venom phenotype is often independent of phylogenetic relationships in rattlesnakes, suggesting the importance of diet and/or other environmental variables in driving venom evolution. Moreover, the striking inter- and intraspecific variability in venom activity on human blood highlights the considerable variability faced by physicians treating envenomation. This study is the most comprehensive effort to date to describe and characterize the evolutionary and biological aspects of coagulotoxins in rattlesnake venom. Further research at finer taxonomic levels is recommended to elucidate patterns of variation within species and lineages.
... In multiple snake species, variations in venom composition have been associated with ontogeny [30]. This was first documented in detail by Mackessy in 1988 [31]. In this seminal work, the ontogenetic variation in venom composition was examined in rattlesnakes of various lengths [31]. ...
... This was first documented in detail by Mackessy in 1988 [31]. In this seminal work, the ontogenetic variation in venom composition was examined in rattlesnakes of various lengths [31]. In both Crotalus helleri and Crotalus oreganus, increased protease activity is positively correlated with size, and toxicity is more pronounced in juveniles [31]. ...
... In this seminal work, the ontogenetic variation in venom composition was examined in rattlesnakes of various lengths [31]. In both Crotalus helleri and Crotalus oreganus, increased protease activity is positively correlated with size, and toxicity is more pronounced in juveniles [31]. The separation of venom pharmacology among ontogenetic changes may have evolved due to changes in diet requirements. ...
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Venomous animals are a striking example of the convergent evolution of a complex trait. These animals have independently evolved an apparatus that synthesizes, stores, and secretes a mixture of toxic compounds to the target animal through the infliction of a wound. Among these distantly related animals, some can modulate and compartmentalize functionally distinct venoms related to predation and defense. A process to separate distinct venoms can occur within and across complex life cycles as well as more streamlined ontogenies, depending on their life-history requirements. Moreover, the morphological and cellular complexity of the venom apparatus likely facilitates the functional diversity of venom deployed within a given life stage. Intersexual variation of venoms has also evolved further contributing to the massive diversity of toxic compounds characterized in these animals. These changes in the biochemical phenotype of venom can directly affect the fitness of these animals, having important implications in their diet, behavior, and mating biology. In this review, we explore the current literature that is unraveling the temporal dynamics of the venom system that are required by these animals to meet their ecological functions. These recent findings have important consequences in understanding the evolution and development of a convergent complex trait and its organismal and ecological implications.
... Animal venoms are composed of an assortment of proteins and peptides that are produced in a gland and injected into prey or in defense against predators [21]. Ontogenetic shifts in composition can yield measurable variation in venom function as stark as shifts from primarily neurotoxic and myotoxic to primarily hemotoxic and hemorrhagic venoms in snakes [22][23][24]. Venom ontogeny has medical relevance, as snakebite outcomes and antivenom efficacy can depend upon venom composition [25][26][27][28]. It also has biological relevance, as ontogenetic shifts in venom composition can impact the ability of snakes to incapacitate certain species of prey [22]. ...
... Venom ontogeny has medical relevance, as snakebite outcomes and antivenom efficacy can depend upon venom composition [25][26][27][28]. It also has biological relevance, as ontogenetic shifts in venom composition can impact the ability of snakes to incapacitate certain species of prey [22]. Although the presence of shifts has been demonstrated across many species, less is known about their tempo, either as gradual or sudden processes, and the proximate physiological signals that might initiate shifts in venom composition remain largely unexplored [10,29]. ...
... If the trajectory of the shift were more continuous, this would confound studies of venom variation by introducing variation from mid-sized animals that blurs the lines between the juvenile and adult study groups. Previous work in Pacific rattlesnakes (Crotalus oreganus ssp.) suggested that protease activity in venoms begins to increase between 50 and 60 cm in snout-vent length (SVL) [22], while in Crotalus adamanteus there is evidence for a shift in venom composition at a length of 102 cm, the size that indicates sexual maturity in the species [30]. Prior works on ontogenetic shifts in venom across developmental stages tend to employ a "venom census", surveying populations by collecting single samples from many individuals of different sizes. ...
Article
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Ontogenetic shifts in venom occur in many snakes but establishing their nature as gradual or discrete processes required additional study. We profiled shifts in venom expression from the neonate to adult sizes of two rattlesnake species, the eastern diamondback and the timber rattlesnake. We used serial sampling and venom chromatographic profiling to test if ontogenetic change occurs gradually or discretely. We found evidence for gradual shifts in overall venom composition in six of eight snakes, which sometimes spanned more than two years. Most chromatographic peaks shift gradually, but one quarter shift in a discrete fashion. Analysis of published diet data showed gradual shifts in overall diet composition across the range of body sizes attained by our eight study animals, while the shifts in abundance of different prey classes varied in form from gradual to discrete. Testosterone concentrations were correlated with the change in venom protein composition, but the relationship is not strong enough to suggest causation. Venom research employing simple juvenile versus adult size thresholds may be failing to account for continuous variation in venom composition lifespan. Our results imply that venom shifts represent adaptive matches to dietary shifts and highlight venom for studies of alternative gene regulatory mechanisms. Key Contribution: We found that differences in venom composition, including the phospholipase A2 and snake venom metalloproteinase enzymes that contribute significantly to venom function, were correlated with increases in body size. These changes tended to occur gradually rather than discretely, although several clear examples of discrete shifts in individual peaks occurred, suggesting regulatory mechanisms may differ among venom proteins. Both gradual and discrete shifts in venom protein expression may be adaptive for the similar modes of shift in diet composition.
... Venom compositional variation is a ubiquitous phenomenon that has been observed in a variety of contexts throughout venomous snake taxa. Previously, venom composition has been shown to vary taxonomically, ontogenetically, geographically, and based on dietary preference (Mackessy, 1988(Mackessy, , 2010Chippaux et al., 1991;Mackessy et al., 2003;Massey et al., 2012). Understanding these types of venom variation has significant evolutionary implications and can inform snakebite treatment (Massey et al., 2012). ...
... This has been previously assumed in a variety of studies that either pool venom from both glands (e.g. Smith and Mackessy, 2016;Mackessy, 1988;2003;2010;Chippaux et al., 1982) or assume comparable transcript expression between left and right glands (Aird et al., 2015;Rokyta et al., 2012). ...
... The snake was an adult female, with snout to vent/tail lengths of 640/37 (2014) and 715/ 41 (2017); therefore, ontogenetic effects (Saviola et al., 2015) were not a concern. Venom was manually extracted as previously described (Mackessy, 1988); however, left and right gland capillary tube volumes were not pooled as usual, because of their differing appearance upon extraction. While the physiological cause of the gland abnormality was unknown, differences in venom quality between the left and right glands was initially evaluated based on thousands of previous extractions of this species. ...
Article
It is assumed that toxin expression is equivalent between left and right glands of a single snake. In the current study, we report venoms that differ in enzyme functionality and overall composition between the left and right gland of a single snake. The right gland produced venom of comparable composition to venom previously extracted from the same individual; however, the left gland produced venom with overall lower protein content and considerably less enzyme activity. Snake venoms are complex toxic mixtures composed primarily of potent bioactive proteins used for prey incapacitation or defense. Venom compositional variation is a ubiquitous phenomenon that has been observed in a variety of contexts throughout venomous snake taxa. Previously, venom composition has been shown to vary taxonomically, ontogenetically, geographically, and based on dietary preference (Mackessy, 1988, 2010; Chippaux et al., 1991; Mackessy et al., 2003; Massey et al., 2012). Understanding these types of venom variation has significant evolutionary implications and can inform snakebite treatment (Massey et al., 2012).
... Neutral evolution (Aird et al., 2017;Mebs, 2001;Sasa, 1999), positive selection (Aird et al., 2015;Rokyta et al., 2011) and purifying selection (Sunagar and Moran, 2015;Sunagar et al., 2014) have all been suggested to be main drivers of venom protein gene evolution. Although it is well accepted that the key role of venom is to immobilize and digest prey, the direct and indirect processes resulting in venom variation (Aird et al., 2017;Casewell et al., 2014;Durban et al., 2017;Mackessy, 1988;Margres et al., 2017;Rokyta et al., 2015a;Zancolli et al., 2019) and the evolutionary origin of snake venom (Fry, 2005;Hargreaves et al., 2014;Reyes-Velasco et al., 2015) are still debated. ...
... The biological significance behind venom variation is being actively explored, and more data is needed documenting and verifying factors contributing to the distribution, diversity and the genetic regulatory processes resulting in this variation. Currently, phylogenetic signal (Lomonte et al., 2014;Mackessy, 2010a;Sanz et al., 2006;Smith and Mackessy, 2016), age (Alape-Giron et al., 2008;Mackessy, 1988;Mackessy et al., 2006;Modahl et al., 2016), geographic locality (Massey et al., 2012;Rokyta et al., 2015b;Sunagar et al., 2014;Tan et al., 2015b) and diet (da Silva and Aird, 2001;Daltry et al., 1996;Li et al., 2005;Mackessy, 1988;Pawlak et al., 2006Pawlak et al., , 2009 are all known contributing biological factors. ...
... The biological significance behind venom variation is being actively explored, and more data is needed documenting and verifying factors contributing to the distribution, diversity and the genetic regulatory processes resulting in this variation. Currently, phylogenetic signal (Lomonte et al., 2014;Mackessy, 2010a;Sanz et al., 2006;Smith and Mackessy, 2016), age (Alape-Giron et al., 2008;Mackessy, 1988;Mackessy et al., 2006;Modahl et al., 2016), geographic locality (Massey et al., 2012;Rokyta et al., 2015b;Sunagar et al., 2014;Tan et al., 2015b) and diet (da Silva and Aird, 2001;Daltry et al., 1996;Li et al., 2005;Mackessy, 1988;Pawlak et al., 2006Pawlak et al., , 2009 are all known contributing biological factors. ...
Article
The genera Ophiophagus and Naja comprise part of a clade of snakes referred to as cobras, dangerously venomous front-fanged snakes in the family Elapidae responsible for significant human mortality and morbidity throughout Asia and Africa. We evaluated venom enzyme variation for eleven cobra species and three N. kaouthia populations using SDS-PAGE venom fingerprinting and numerous enzyme assays. Acetylcholinesterase and PLA2 activities were the most variable between species, and PLA2 activity was significantly different between Malaysian and Thailand N. kaouthia populations. Venom metalloproteinase activity was low and significantly different among most species, but levels were identical for N. kaouthia populations; minor variation in venom L-amino acid oxidase and phosphodiesterase activities were seen between cobra species. Naja siamensis venom lacked the α-fibrinogenolytic activity common to other cobra venoms. In addition, venom from N. siamensis had no detectable metalloproteinase activity and exhibited an SDS-PAGE profile with reduced abundance of higher mass proteins. Venom profiles from spitting cobras (N. siamensis, N. pallida, and N. mossambica) exhibited similar reductions in higher mass proteins, suggesting the evolution of venoms of reduced complexity and decreased enzymatic activity among spitting cobras. General, the venom proteomes of cobras show highly abundant three-finger toxin diversity, followed by large quantities of PLA2s. However, PLA2 bands and activity were very reduced for N. haje, N. annulifera and N. nivea. Venom compositional analysis provides insight into the evolution, diversification and distribution of different venom phenotypes that complements venomic data, and this information is critical for the development of effective antivenoms and snakebite treatment.
... Adaptive differences may be produced by changes in prey preference at different life-history stages (Mushinsky et al. 1982) or optimal foraging strategy that promotes faster growth rates and reduces time spent in more vulnerable size classes (Werner and Gilliam 1984;Klauber 1997). For example, the production of large toxin enzymes such as SVMPs may be more metabolically costly (Mackessy 1988), leading to limited expression in juveniles. Although the precise mechanism remains unknown, the venom phenotype was significantly variable across age classes with only a limited number of toxins exhibiting differential expression across populations, suggesting that changes in venom expression due to maturity may have greater ecological implications (i.e. ...
... Snakes, as gape limited predators, may select prey at different life-history stages (Shine 1991); therefore, the venom phenotype may adaptively shift as size increases to more effectively subdue and/or digest different, larger prey species (Margres et al. 2015b). Variable efficacy of adult and juvenile venom in differing prey items is observed in multiple snake species (Mackessy 1988;Andrade and Abe 1999;Margres et al. 2016b;Cipriani et al. 2017;Borja et al. 2018), suggesting that ontogenetic venom Where the "ruber" meets the road: Using the genome GBE Genome Biol. Evol. ...
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Understanding the proximate and ultimate causes of phenotypic variation is fundamental in evolutionary research, as such variation provides the substrate for selection to act upon. Although trait variation can arise due to selection, the importance of neutral processes is sometimes understudied. We presented the first reference-quality genome of the Red Diamond Rattlesnake (Crotalus ruber) and used range-wide ‘omic data to estimate the degree to which neutral and adaptive evolutionary processes shaped venom evolution. We characterized population structure and found substantial genetic differentiation across two populations, each with distinct demographic histories. We identified significant differentiation in venom expression across age classes with substantially reduced but discernible differentiation across populations. We then used conditional redundancy analysis to test whether venom expression variation was best predicted by neutral divergence patterns or geographically-variable (a)biotic factors. Snake size was the most significant predictor of venom variation, with environment, prey availability, and neutral sequence variation also identified as significant factors, though to a lesser degree. By directly including neutrality in the model, our results confidently highlight the predominant, yet not singular, role of life history in shaping venom evolution.
... Together, these families account for about 77% of the venom proteome, and the remaining 22% is made up of other protein families: L-amino acid oxidases (LAAO), cysteine-rich secretory proteins (CRISP), C-type lectins (CTL), disintegrins (DIS) and natriuretic peptides (NP) [10,11]. In addition to the inherent intergeneric and interspecific diversity in venom composition, reports have correlated variation in the venom of viperids with diet [12][13][14][15], age of the organism [15][16][17][18][19][20], and sex [21]. These factors multiply the number of venoms with different characteristics and make the challenge for public health response even more complex. ...
... Together, these families account for about 77% of the venom proteome, and the remaining 22% is made up of other protein families: L-amino acid oxidases (LAAO), cysteine-rich secretory proteins (CRISP), C-type lectins (CTL), disintegrins (DIS) and natriuretic peptides (NP) [10,11]. In addition to the inherent intergeneric and interspecific diversity in venom composition, reports have correlated variation in the venom of viperids with diet [12][13][14][15], age of the organism [15][16][17][18][19][20], and sex [21]. These factors multiply the number of venoms with different characteristics and make the challenge for public health response even more complex. ...
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Background Each year, 3,800 cases of snakebite envenomation are reported in Mexico, resulting in 35 fatalities. The only scientifically validated treatment for snakebites in Mexico is the use of antivenoms. Currently, two antivenoms are available in the market, with one in the developmental phase. These antivenoms, produced in horses, consist of F(ab’)2 fragments generated using venoms from various species as immunogens. While previous studies primarily focused on neutralizing the venom of the Crotalus species, our study aims to assess the neutralization capacity of different antivenom batches against pit vipers from various genera in Mexico. Methodology We conducted various biological and biochemical tests to characterize the venoms. Additionally, we performed neutralization tests using all three antivenoms to evaluate their effectiveness against lethal activity and their ability to neutralize proteolytic and fibrinogenolytic activities. Results Our results reveal significant differences in protein content and neutralizing capacity among different antivenoms and even between different batches of the same product. Notably, the venom of Crotalus atrox is poorly neutralized by all evaluated batches despite being the primary cause of envenomation in the country’s northern region. Furthermore, even at the highest tested concentrations, no antivenom could neutralize the lethality of Metlapilcoatlus nummifer and Porthidium yucatanicum venoms. These findings highlight crucial areas for improving existing antivenoms and developing new products. Conclusion Our research reveals variations in protein content and neutralizing potency among antivenoms, emphasizing the need for consistency in venom characteristics as immunogens. While Birmex neutralizes more LD50 per vial, Antivipmyn excels in specific neutralization. The inability of antivenoms to neutralize certain venoms, especially M. nummifer and P. yucatanicum, highlights crucial improvement opportunities, given the medical significance of these species.
... This results in considerable differences in size between age-classes within many populations, raising the potential for younger, smaller animals to utilize different sets of resources for a significant portion of their life. An ontogenetic shift in diet is one such phenomenon that has received considerable attention: smaller prey, for example, may be required for juvenile animals that lack the capability to subdue and swallow larger prey (Hampton, 2011;Patterson et al., 2022); a notable example is neonate rattlesnakes in some populations that rely heavily on ectothermic prey (e.g., lizards) while adults use larger prey such as birds and mammals (Mackessy, 1988;Saviola et al., 2012). Of obvious importance is knowing whether ontogenetic patterns of habitat use exist, whether driven by diet, thermoregulatory opportunities, or other factors associated with body size (Shine et al., 2002;Blouin-Demers et al., 2007;Eskew et al., 2009). ...
... For adult snakes, the selection of ambush sites is related to the presence of chemical cues left by prey (Theodoratus and Chiszar, 2000). With a paucity of small reptilian prey in the grasslands (e.g., lizardsa common feature in the ecology of the species further south; Mackessy, 1988), the diet of Western Rattlesnakes in British Columbia consists primarily of small mammals such as mice, voles, and shrews (McAllister et al., 2016). The primary prey species at our study site (Maida et al., 2020), the deer mouse (Peromyscus maniculatus) and Great Basin pocket mouse (Perognathus parvus), are also known to select for shrub cover, which can likely be attributed to food availability (Harris, 1986;Melaschenko and Hodges, 2020). ...
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Studies on habitat selection are central to our understanding of animal ecology and play an important role in the conservation and management of species. However, habitat selection is not necessarily consistent across sectors of populations and failing to understand both similarities and differences between age classes, for instance, may be problematic. Knowledge on habitat use by juvenile vertebrates in general is sorely lacking, particularly for species with precocial offspring, and here we address this gap by describing habitat use by juvenile Western Rattlesnakes ( Crotalus oreganus ). We measured habitat at two scales for radio-telemetered juvenile snakes along active-season movement paths at a site in southern British Columbia, Canada, where seasonal migrations and habitat associations of adult animals have been particularly well studied. Despite the relatively short distances these animals travelled and their diminutive size, we found there was selection for structurally stable cover (e.g., woody debris, shrub, and rock cover) similar to that documented for adult snakes in the same region. Additionally, we tested for differences in microhabitat features at sites used for short-duration (fewer than seven days) and long-duration (seven days or longer) stopovers: we detected negative selection for leaf litter at long-duration stopover sites, but otherwise identified no difference in the microhabitat features associated with these two categories of locations. Overall, this study contributes rare data to our growing understanding of the complexity of habitat requirements for migratory snake species, including northern crotalid vipers, while underscoring the crucial role of habitat selection research across all segments of populations. Comparative data on habitat selection by early-age classes within reptile populations informs conservation planning for the long-term survival of wild populations.
... Ontogenetic changes in the composition and activities of the venom have been observed in different species of snakes in many works in the field of proteomics and biochemistry (Andrade and Abe, 1999;Calvete, 2017;Gibbs et al., 2011;Gutiérrez et al., 1990Gutiérrez et al., , 1991Lomonte et al., 1983;Mackessy, 1988;Pla et al., 2017a;Rokyta et al., 2017;Zelanis et al., 2009). The venom of young snakes of other Bothrops species exhibited higher coagulant activity than that of adult individuals, while the latter generally have greater proteolytic activity (Andrade and Abe, 1999;Furtado et al., 1991;Guércio et al., 2006;Kamiguti, 1988;Pereira, 2006). ...
... The shift to a more proteolytic venom as the animal grows has already been observed in other snakes' species described by other authors, who have suggested that this variation in venom could be linked to dietary change (Andrade and Abe, 1999;Mackessy, 1988). Despite the fact that B. pauloensis presents diet shift, an increase in the proteolytic activity performed in adult venom in vitro was not observed. ...
Article
Considerable heterogeneity and ontogenetic changes in venom composition have already been observed in different species of snakes within the Viperidae family. Since the venom of young and adult can cause distinct pathological effects and because the antivenom may be less effective in neutralizing envenoming by young snakes compared to adults, it is of paramount importance to understand the ontogenetic variation of snake venom. Thus, the present study aimed to analyze and compare the venom of Bothrops pauloensis snakes, searching for possible influences of ontogeny and sex in their biochemical and biological aspects. The venom of younger individuals was more complex in relation to high molecular mass proteins, with a greater abundance of metalloproteinases, while adults showed a greater abundance of medium and low molecular mass proteins, such as phospholipases A2 (PLA2), C-type lectins and serine proteases. The antivenom showed better immunorecognition towards the venom of adult snakes than younger ones, in addition to a deficiency in the recognition of medium molecular mass proteins, suggesting the need for an improvement in the antivenom. Younger snakes showed higher coagulant, caseinolytic, and hemorrhagic activity, while adult snakes showed higher L-amino acid oxidase (LAAO) activity and acted faster in lethality. Differences between males and females were observed mainly in the rate of loss of coagulant activity, change in PLA2 activity and lethality action time. Furthermore, considering only the adult groups, males showed a higher LAAO and thrombin-like activity, while females showed a higher caseinolytic and hyaluronidase activity. With the results obtained in this work, it was possible to conclude that there is an ontogenetic variation in the composition and some activities of the B. pauloensis snake venom, in addition to differences between the venom of males and females, reinforcing that there is an intraspecific variation that may result in different symptoms in their envenoming and, consequently, differences in the response to treatment with the antivenom.
... However, the rationale for the observed ontogenetic changes remains obscure, as little is known about the different pathophysiological changes in renal functions after envenomation with D. siamensis and the differences in the protein abundance of its venom. The mechanisms of venom action within the body to induce AKI during envenomation from D. siamensis have not been determined, although several studies in other snake species have reported ontogenetic differences among venoms of the same species, including variations in the biological and biochemical features [4,5,6] differences in venom compositions [7,8,9,10,11], toxicity [4,12,13,14] and enzymatic activity [8,14]. The abundance of different toxic and non-toxic proteins in snake venom are influenced by many factors, including variations in taxonomy, age, sex, geography, diet and seasons [15,16,17]. ...
... However, the rationale for the observed ontogenetic changes remains obscure, as little is known about the different pathophysiological changes in renal functions after envenomation with D. siamensis and the differences in the protein abundance of its venom. The mechanisms of venom action within the body to induce AKI during envenomation from D. siamensis have not been determined, although several studies in other snake species have reported ontogenetic differences among venoms of the same species, including variations in the biological and biochemical features [4,5,6] differences in venom compositions [7,8,9,10,11], toxicity [4,12,13,14] and enzymatic activity [8,14]. The abundance of different toxic and non-toxic proteins in snake venom are influenced by many factors, including variations in taxonomy, age, sex, geography, diet and seasons [15,16,17]. ...
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Background: Eastern Russell’s viper (Daboia siamensis) is one of the most medically significant snakes responsible for the development of acute renal failure. However, variation of the clinical picture and renal pathophysiology following bites by young and adult D. siamensis have not been elucidated. Methods: In this study, we analyzed the venomic profiles of D. siamensis at different maturation stages of juvenile, subadult and adult groups. The same pooled venom from each group was subjected to enzymatic, electrophoretic and proteomic analysis, including sublethal toxicity (0.1 mg/kg iv.) examined on bodily functions by comparing the venom compositional and functional profiles among venom specimens from juvenile, subadult and adult D. siamensis by correlating them with the renal pathophysiology in experimental rabbits. Results: The comparative studies revealed that juvenile venom possessed higher phospholipase A2, metalloproteinase and serine proteinase levels, while subadult and adult venoms contained more L-amino acid oxidase, phosphodiesterase, the Kunitz-type serine protease inhibitor, disintegrin families and endothelial growth factor. An in vivo study revealed that the adult and subadult venoms caused persistent hypotension and bradycardia, while thrombocytopenia was a more characteristic effect of juvenile venom. All venom age groups showed significant reductions in renal hemodynamics and electrolyte excretions. The juvenile venom caused a higher tubulonephrosis lesion score than adult and subadult venoms. Conclusions: The D. siamensis venom shows an ontogenetic shift in its compositions and activities. Renal function alterations after envenomation depend on either the synergistic actions of different venom components or the disproportionate expression between the concentrations of enzymatic and non-enzymatic proteins in each age venom group. The high proportion of enzymatic toxin proteins in the juvenile venom results in greater nephrotoxicity.
... This is corroborated by our thromboelastography and fluorometry results in terms of time to clot formation and FX zymogen activation. Ontogenetic shifts in venom composition and/or activity have been extensively documented in a variety of rattlesnake species and lineages (60,112,(121)(122)(123)(124), particularly with respect to a pattern of loss of crotoxin-like neurotoxic PLA 2 s (Type II phenotype) in favor of hemorrhagic SVMPs (Type I phenotype) as the snake ages (38,45,46). This phenomenon is recurrent in the C. durissus complex (35,43,45). ...
... This phenomenon is recurrent in the C. durissus complex (35,43,45). Such agedriven changes in venom composition are generally thought to stem from shifts in prey preference between juvenile and adult snakes (10,121,122), as seen in a variety of snakes ranging from Australian elapids (110) to lancehead pit vipers of the genus Bothrops (113,125). However, our current knowledge-albeit fragmentary-points to C. durissus, C. simus, and C. tzabcan being rodent specialists throughout their life (10,24,(126)(127)(128)(129). ...
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Rattlesnakes are a diverse clade of pit vipers (snake family Viperidae, subfamily Crotalinae) that consists of numerous medically significant species. We used validated in vitro assays measuring venom-induced clotting time and strength of any clots formed in human plasma and fibrinogen to assess the coagulotoxic activity of the four medically relevant Mexican rattlesnake species Crotalus culminatus, C. mictlantecuhtli, C. molossus, and C. tzabcan. We report the first evidence of true procoagulant activity by Neotropical rattlesnake venom in Crotalus culminatus. This species presented a strong ontogenetic coagulotoxicity dichotomy: neonates were strongly procoagulant via Factor X activation, whereas adults were pseudo-procoagulant in that they converted fibrinogen into weak, unstable fibrin clots that rapidly broke down, thereby likely contributing to net anticoagulation through fibrinogen depletion. The other species did not activate clotting factors or display an ontogenetic dichotomy, but depleted fibrinogen levels by cleaving fibrinogen either in a destructive (non-clotting) manner or via a pseudo-procoagulant mechanism. We also assessed the neutralization of these venoms by available antivenom and enzyme-inhibitors to provide knowledge for the design of evidence-based treatment strategies for envenomated patients. One of the most frequently used Mexican antivenoms (Bioclon Antivipmyn®) failed to neutralize the potent procoagulant toxic action of neonate C. culminatus venom, highlighting limitations in snakebite treatment for this species. However, the metalloprotease inhibitor Prinomastat substantially thwarted the procoagulant venom activity, while 2,3-dimercapto-1-propanesulfonic acid (DMPS) was much less effective. These results confirm that venom-induced Factor X activation (a procoagulant action) is driven by metalloproteases, while also suggesting Prinomastat as a more promising potential adjunct treatment than DMPS for this species (with the caveat that in vivo studies are necessary to confirm this potential clinical use). Conversely, the serine protease inhibitor 4-(2-aminoethyl)benzenesulfonyl fluoride hydrochloride (AEBSF) inhibited the direct fibrinogen cleaving actions of C. mictlantecuhtli venom, thereby revealing that the pseudo-procoagulant action is driven by kallikrein-type serine proteases. Thus, this differential ontogenetic variation in coagulotoxicity patterns poses intriguing questions. Our results underscore the need for further research into Mexican rattlesnake venom activity, and also highlights potential limitations of current antivenom treatments.
... Свойства ядов некоторых видов гадюковых змей изменяются в течение онтогенеза (Mackessy, 1988Wray et al., 2015). Ферментативная активность яда восточной степной гадюки (Christoph, 1861) до недавнего времени изучалась по образцам, полученным только от взрослых особей (Бакиев и др., 2015). ...
... При этом в период с июля по октябрь 2015 г. увеличивалось как число особей с желтоокрашенным ядом, так и активность самого фермента. Схожие различия в активности оксидазы L-аминокислот характерны для некоторых видов гремучников, у которых с возрастом и увеличением размеров змей активность оксидазы L-аминокислот возрастала (Mackessy, 1988). ...
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The paper presents the results of our long-term (August 2014–October 2016) observations of changes in some properties of the poisonous secretion of eastern steppe vipers of the nominative subspecies Vipera renardi renardi (Christoph, 1861) during their postembryonic ontogenesis. The poisonous secretion of newborn vipers differed from the venom of adult snakes by an increased protease activity and the absence of any L-amino acid oxidase activity; all newborns had colorless venom. Adults produce venom either colorless, where no L-amino acid oxidase activity is detected, or yellow, where it is detected. It was found that the enzymatic activity of the venom of young vipers between their first and second winterings corresponded to the level of adults. After the second wintering, young vipers showed statistically insignificant seasonal changes in the activity of proteases and L-amino acid oxidase.
... The results of these studies indicate that individual (intra-population) venom variation is genetically inherited and is not altered ontogenetically by environmental factors, seasonal variation, or diet [14,15,[20][21][22]. An exception to this is ontogenetic venom changes in species with a bi-phasic juvenile to adult shift in dietary ecology [23][24][25]. Inter-population venom variation can be significant [13,26], and as venom is a trophic adaptation, these divergences are assumed to be the result of selection pressure due to local differences in diet, although this level of dietary data is usually lacking. Taipans are a dietary specialist, so all populations have a broadly similar diet, and it can reasonably be assumed that their venom composition would be fairly uniform throughout their distribution. ...
... All the other bands on the gel were higher molecular weights-ranging from approximately 25 to 110 kDa. MS analysis of the bands at 25,30,37,70,90, and 100 kDa gave the highest confidence matches for all of these bands to the serine protease prothrombin activator Oscutarin C (9% to 22% sequence coverage and 5 to 65 peptides). The strongest match for this protein was the 30kDa band (20% sequence coverage with 65 peptides matched). ...
Article
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Intra-specific venom variation has the potential to provide important insights into the evolution of snake venom, but remains a relatively neglected aspect of snake venom studies. We investigated the venom from 13 individual coastal taipans Oxyuranus scutellatus from four localities on the north-east coast of Australia, spanning a distance of 2000km. The intra-specific variation in taipan venom was considerably less than the inter-specific variation between it and the other Australian elapids to which it was compared. The electrophoretic venom profile of O. scutellatus was visually different to six other genera of Australian elapids, but not to its congener inland taipan O. microlepidotus. There was minimal geographical variation in taipan venom, as the intra-population variation exceeded the inter-population variation for enzymatic activity, procoagulant activity, and the abundance of neurotoxins. The pre-synaptic neurotoxin (taipoxin) was more abundant than the post-synaptic neurotoxins (3FTx), with a median of 11.0% (interquartile range (IQR): 9.7% to 18.3%; range: 6.7% to 23.6%) vs. a median of 3.4% (IQR: 0.4% to 6.7%; range: 0% to 8.1%). Three taipan individuals almost completely lacked post-synaptic neurotoxins, which was not associated with geography and occurred within two populations. We found no evidence of sexual dimorphism in taipan venom. Our study provides a basis for evaluating the significance of intra-specific venom variation within a phylogenetic context by comparing it to the inter-specific and inter-generic variation. The considerable intra-population variation we observed supports the use of several unpooled individuals from each population when making inter-specific comparisons.
... Age-related changes in venom composition, as it happens in L. stenophrys, have been reported in a number of New World pit viper species from different genera, i.e. Crotalus, Bothrops, Sistrurus, and Bothiechis (Alape-Girón et al., 2008;Calvete et al., 2011Calvete et al., , 2010Gibbs et al., 2011;Gonçalves-Machado et al., 2016;Guércio et al., 2006;Mackessy, 1988;Pla et al., 2017;Saldarriaga et al., 2003;Zelanis et al., 2010Zelanis et al., , 2008Zelanis et al., , 2011. A similar pattern was noted for the Brown Treesnake (Boiga irregularis), with juvenile venoms more toxic than adult venoms towards geckos (Mackessy et al., 2006), suggesting that this trend has not exclusively evolved in pit vipers. ...
... A shift in the feeding habits of juvenile versus adult snakes, from cold-blooded (arthropods, frogs and lizards) to warm-blooded (mammals) prey, has been invoked to explain the ontogenetic venom change in some pitviper species (Andrade and Abe, 1999). In concordance with this view, it has been documented that venoms from neonate snakes are more toxic to lizards and inbred mice than adult venoms (Mackessy, 1988). Young Bothrops snakes preferentially eat amphibians, lizards, birds, and shift to mammals when they become adults (Campbell and Lamar, 2004). ...
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Snakes of the genus Lachesis, commonly known as bushmasters, are the largest venomous snakes in the Americas. Because these snakes have their habitats in areas of remote forests they are difficult to find, and consequently there are few studies of Lachesis taxa in their natural ecosystems. Bushmasters are distributed in tropical forest areas of South and Central America. In Brazil they can be found in the Amazon Rainforest and the Atlantic Forest. Despite the low incidence of cases, laquetic envenoming causes severe permanent sequelae due to the high amount of inoculated venom. These accidents are characterized by local pain, hemorrhage and myonecrosis that can be confused with bothropic envenomings. However, victims of Lachesis bites develop symptoms characteristic of Lachesis envenoming, known as vagal syndrome. An important message of this bibliographic synthesis exercise is that, despite having the proteomic profiles of all the taxa of the genus available, very few structure-function correlation studies have been carried out. Therefore the motivation for this review was to fill a gap in the literature on the genus Lachesis, about which there is no recent review. Here we discuss data scattered in a number of original articles published in specialized journals, spanning the evolutionary history and extant phylogeographic distribution of the bushmasters, their venom composition and diet, as well as the pathophysiology of their bites to humans and the biological activities and possible biotechnological applicability of their venom toxins.
... Changing biotic conditions are assumed to drive snake venom diversity, e.g., variation in prey composition can promote the expression of specific toxins [14]. Diet-dependent feeding success can be a strong selective force in venom composition [8,15,16], particularly in relation to ontogenetic changes in diet [17,18]. Variation in snake venom has also been observed between individuals from the same population (e.g., sex differences [19,20]) and between populations occurring in distinct ecological conditions [21][22][23][24][25][26]. ...
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Allopatric populations living under distinct ecological conditions are excellent systems to infer factors underlying intraspecific venom variation. The venom composition of two populations of Vipera ammodytes, insular with a diet based on ectotherms and mainland with a diet based on ectotherms and endotherms, was compared considering the sex and age of individuals. Ten toxin families, dominated by PLA2, svMP, svSP, and DI, were identified through a bottom-up approach. The venom profiles of adult females and males were similar. Results from 58 individual SDS-PAGE profiles and venom pool analysis revealed significant differences between juveniles compared to subadults and adults. Two venom phenotypes were identified: a juvenile svMP-dominated and KUN-lacking phenotype and an adult PLA2/svMP-balanced and KUN-containing phenotype. Despite differences in prey availability (and, therefore, diet) between populations, no significant differences in venom composition were found. As the populations are geographically isolated, the lack of venom diversification could be explained by insufficient time for natural selection and/or genetic drift to act on the venom composition of island vipers. However, substantial differences in proteomes were observed when compared to venoms from geographically distant populations inhabiting different conditions. These findings highlight the need to consider ecological and evolutionary processes when studying venom variability.
... At the same time, the existence and importance of analyzing intra-specific variations in the venoms of individual snakes are noted by many researchers [6,26,27]. There are multiple factors responsible for intra-specific venom variations, such as geographic origin [8,28], etc. Identification of such factors in many cases is carried out when using pooled venoms of snakes of the same group (from the same territory, the same age, etc.), but it is important as well to analyze the differences between individuals within the same groups. ...
Article
The knowledge of variations in the composition of venoms from different snakes is important from both theoretical and practical points of view, in particular, at developing and selecting an antivenom. Many studies on this topic are conducted with pooled venoms, while the existence and significance of variations in the composition of venoms between individual snakes of the same species are emphasized by many authors. It is important to study both inter- and intra-specific, including intra-population, venom variations, because intra-specific variations in the venom composition may affect the effectiveness of antivenoms as strongly as inter-specific. In this work, based on venom Raman spectroscopy with principal component analysis, we assessed the variations in venoms of individual snakes of the Vipera nikolskii species from two populations and compared these intra-specific variations with inter-specific variations (with regard to the other related species). We demonstrated intra-specific (inter- and intra-population) differences in venom compositions which are smaller than inter-specific variations. We also assessed the compositions of V. nikolskii venoms from two populations to explain inter-population differences. The method used is rapid and requires virtually no preparation of samples, used in extremely small quantities, allowing the venoms of individual snakes to be analyzed. In addition, the method is informative and capable of detecting fairly subtle differences in the composition of venoms.
... Venom composition varies between species depending on several factors, e.g. phylogenetic affinities [32], geographic localities [33,34], snake age [35][36][37][38] and diet [39,40]. ...
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Breast and hepatic cancers are the leading incidences in the globe occurring of the human sufferings from various cancers. Snake venoms have been reported to provide effective therapeutic agents. The current study investigates the anticancer potency of Egyptian venoms snakes on two cells: breast cancer cells (MCF-7) and hepato-cancer cells (HepG2) (In vitro assay). The examined venoms were more potent on MCF-7 than HepG2 cells. Their inhibition % on MCF-7 ranged from 71.47 to 99.02% with medium inhibition concentrations (IC50s): 3.48, 3.60, 3.70, 4.33, and 4.49 μg/ml for venoms: Echis pyramid (E.H), Cerastes vipera (C.V), Naja haje (N.H), Echis coloratus (E.C), and Cerastes cerastes (C.C), respectively. The values of IC50s on HepG2 were 4.32, 17.77, 59.72, 63.75, and 217.90 μg/ml for toxins: E.C, E.P, C.V, C.C, and N.H, respectively. Some biomarkers were conducted to investigate the apoptotic effects of toxins into the cells. Increasing profiles of lactate dehydrogenase (LDH) activity and levels of glutathione content (GSH) and malodialdhyde (MDA) as well as repairment of DNA indicated such these actions. So, more reliable investigations on these venoms were needed to provide intelligent therapeutic agent for cancer treatment.
... All snake collections and field-collected samples were conducted according to permits obtained from Colorado Parks and Wildlife (scientific collecting permits #14HP974, 17HP0974, 20HP0974, to SPM), Wyoming Game and Fish Department (scientific collecting permit #1544, SPM), New Mexico Game and Fish Department (permit #3418, SPM), Utah Division of Wildlife Resources (permit #1COLL10567, SPM), Texas Parks and Wildlife Department (permit #SPR-0604-391, SPM), and Montana Fish, Wildlife and Parks (permits 2019-075-W and 2020-076-W, TAC). Venom was manually extracted as described [81]; samples were either dried in the field over calcium carbonate and placed on ice until storage at − 20 °C or were placed in liquid nitrogen until lyophilization and storage at − 20 °C. Due to the potential for ontogenetic venom variation, only venoms from adult snakes were utilized in this study. ...
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Background Snake venoms are trophic adaptations that represent an ideal model to examine the evolutionary factors that shape polymorphic traits under strong natural selection. Venom compositional variation is substantial within and among venomous snake species. However, the forces shaping this phenotypic complexity, as well as the potential integrated roles of biotic and abiotic factors, have received little attention. Here, we investigate geographic variation in venom composition in a wide-ranging rattlesnake (Crotalus viridis viridis) and contextualize this variation by investigating dietary, phylogenetic, and environmental variables that covary with venom. Results Using shotgun proteomics, venom biochemical profiling, and lethality assays, we identify 2 distinct divergent phenotypes that characterize major axes of venom variation in this species: a myotoxin-rich phenotype and a snake venom metalloprotease (SVMP)-rich phenotype. We find that dietary availability and temperature-related abiotic factors are correlated with geographic trends in venom composition. Conclusions Our findings highlight the potential for snake venoms to vary extensively within species, for this variation to be driven by biotic and abiotic factors, and for the importance of integrating biotic and abiotic variation for understanding complex trait evolution. Links between venom variation and variation in biotic and abiotic factors indicate that venom variation likely results from substantial geographic variation in selection regimes that determine the efficacy of venom phenotypes across populations and snake species. Our results highlight the cascading influence of abiotic factors on biotic factors that ultimately shape venom phenotype, providing evidence for a central role of local selection as a key driver of venom variation.
... While overall enzymatic activity in this study is based on combined averages of adult and neonate C. tancitarensis venom, ontogenetic shifts in venom composition are common and should be taken into consideration when evaluating venom compositional trends [16,[30][31][32]. Neonate C. tancitarensis venoms have lower metalloproteinase activity based on faint PI and PIII bands, and the presence of PI, PI-II, and P-III SVMPs in both adult C. tancitarensis venoms was confirmed by mass spectrometry. ...
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The Crotalus intermedius group is a clade of rattlesnakes consisting of several species adapted to a high elevation habitat, primarily in México. Crotalus tancitarensis was previously classified as C. intermedius, until individuals occurring on Cerro Tancítaro in Michoacán, México, were reevaluated and classified as a new species (C. tancitarensis) based on scale pattern and geographic location. This study aimed to characterize the venom of C. tancitarensis and compare the venom profile to those of other species within the Crotalus intermedius group using gel electrophoresis, biochemical assays, reverse-phase high performance liquid chromatography, mass spectrometry, and lethal toxicity (LD50)assays. Results show that the venom profiles of species within the Crotalus intermedius group are similar, but with distinct differences in phospholipase A2 (PLA2), metalloproteinase PI (SVMP PI), and kallikrein-like serine proteinase (SVSP) activity and relative abundance. Proteomic analysis indicated that the highland forms produce venoms with 50–60 protein isoforms and a composition typical of type I rattlesnake venoms (abundant SVMPs, lack of presynaptic PLA2-based neurotoxins), as well as a diversity of typical Crotalus venom components such as serine proteinases, PLA2s, C-type lectins, and less abundant toxins (LAAOs, CRiSPs, etc.). The overall venom profile of C. tancitarensis appears most similar to C. transversus, which is consistent with a previous mitochondrial DNA analysis of the Crotalus intermedius group. These rattlesnakes of the Mexican highlands represent a radiation of high elevation specialists, and in spite of divergence of species in these Sky Island habitats, venom composition of species analyzed here has remained relatively conserved. The majority of protein family isoforms are conserved in all members of the clade, and as seen in other more broadly distributed rattlesnake species, differences in their venoms are largely due to relative concentrations of specific components.
... Venom variation is observable on the interspecific level (between species) [7,10] and intraspecific level (within species) [11][12][13][14]. Whilst intraspecific variation is often identified between age classes (i.e., "ontogenetic shifts") or populations (regional variation), the degree to which variation exists within populations is poorly understood. ...
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Snake venoms are complex mixtures of toxins that differ on interspecific (between species) and intraspecific (within species) levels. Whether venom variation within a group of closely related species is explained by the presence, absence and/or relative abundances of venom toxins remains largely unknown. Taipans (Oxyuranus spp.) and brown snakes (Pseudonaja spp.) represent medically relevant species of snakes across the Australasian region and provide an excellent model clade for studying interspecific and intraspecific venom variation. Using liquid chromatography with ultraviolet and mass spectrometry detection, we analyzed a total of 31 venoms covering all species of this monophyletic clade, including widespread localities. Our results reveal major interspecific and intraspecific venom variation in Oxyuranus and Pseudonaja species, partially corresponding with their geographical regions and phylogenetic relationships. This extensive venom variability is generated by a combination of the absence/presence and differential abundance of venom toxins. Our study highlights that venom systems can be highly dynamical on the interspecific and intraspecific levels and underscores that the rapid toxin evolvability potentially causes major impacts on neglected tropical snakebites.
... Thus, the difference observed in M. obscurus may be due to an ontogenic shift from a more "complex" venom that is expressed by juveniles to a "simple" venom that is expressed by adults. Ontogenetic shifts in venom composition have been observed in many snake species, and this change is potentially due to differences in the diet as individuals age [17,22,53,54]. However, more intensive sampling is needed to determine if this is indeed the case in M. obscurus. ...
Article
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Many venomous animals express toxins that show extraordinary levels of variation both within and among species. In snakes, most studies of venom variation focus on front-fanged species in the families Viperidae and Elapidae, even though rear-fanged snakes in other families vary along the same ecological axes important to venom evolution. Here we characterized venom gland transcriptomes from 19 snakes across two dipsadine rear-fanged genera (Leptodeira and Helicops, Colubridae) and two front-fanged genera (Bothrops, Viperidae; Micrurus, Elapidae). We compared patterns of composition, variation, and diversity in venom transcripts within and among all four genera. Venom gland transcriptomes of rear-fanged Helicops and Leptodeira and front-fanged Micrurus are each dominated by expression of single toxin families (C-type lectins, snake venom metalloproteinase, and phospholipase A2, respectively), unlike highly diverse front-fanged Bothrops venoms. In addition, expression patterns of congeners are much more similar to each other than they are to species from other genera. These results illustrate the repeatability of simple venom profiles in rear-fanged snakes and the potential for relatively constrained venom composition within genera.
... These toxins were particularly abundant in the venoms of juvenile Lataste's vipers (Table 1; Fig. 3). Given the direct relationship existing between a snake's size and the venom volume it can produce [87], it is possible that high levels of PIII-SVMPs might aid young V. latastei specimens in subduing their prey despite the low amount of venom they can deploy. PIII-SVMPs might also be at play in aiding prey digestion, but evidence supporting this role is controversial (see [88]). ...
Article
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Lataste's viper (Vipera latastei) is a venomous European viper endemic to the Iberian Peninsula, recognised as medically important by the World Health Organization. To date, no comprehensive characterisation of this species' venom has been reported. Here, we analysed the venoms of juvenile and adult specimens of V. latastei from two environmentally different populations from northern Portugal. Using bottom-up venomics, we produced six venom proteomes (three per population) from vipers belonging to both age classes (i.e., two juveniles and four adults), and RP-HPLC profiles of 54 venoms collected from wild specimens. Venoms from juveniles and adults differed in their chromatographic profiles and relative abundances of their toxins, suggesting the occurrence of ontogenetic changes in venom composition. Specifically, snake venom metalloproteinase (SVMP) was the most abundant toxin family in juvenile venoms, while snake venom serine proteinases (SVSPs), phospholipases A2 (PLA2s), and C-type lectin-like (CTLs) proteins were the main toxins comprising adult venoms. The RP-HPLC venom profiles were found to vary significantly between the two sampled localities, indicating geographic variability. Furthermore, the presence/absence of certain peaks in the venom chromatographic profiles appeared to be significantly correlated also to factors like body size and sex of the vipers. Our findings show that V. latastei venom is a variable phenotype. The intraspecific differences we detected in its composition likely mirror changes in the feeding ecology of this species, taking place during different life stages and under different environmental pressures. Significance Lataste's viper (Vipera latastei) is a medically important viper endemic to the Iberian Peninsula, inhabiting different habitats and undergoing a marked ontogenetic dietary shift. In the current study, we report the first proteomic analysis of V. latastei venom from two environmentally different localities in northern Portugal. Our bottom-up venomic analyses show that snake venom serine proteinases (SVSPs), phospholipases A2 (PLA2s), and C-type lectin-like (CTLs) proteins are the major components of adult V. latastei venom. The comparative analysis of young and adult venoms suggests the occurrence of ontogenetic shift in toxin abundances, with snake venom metalloproteinases (SVMPs) being the predominant toxins in juvenile venoms. Moreover, geographic venom variation between the two studied populations is also detected, with our statistical analyses suggesting that factors like body size and sex of the vipers are possibly at play in its determination. Our work represents the first assessment of the composition of V. latastei venom, and the first step towards a better understanding of the drivers behind its variability.
... In addition, the proteome of N. kaouthia venom (NkV) from two different locations in Assam (Guwahati and Jamurighat) was revealed by ESI-LC-MS/MS analysis of fractions from reversedphase high-performance liquid chromatography (RP-HPLC), though the proteome was represented only in terms of several identified proteins (qualitative) rather than percent abundance of individual protein families [9]. Variations in venom composition are well known to occur in the same species of snake due to different geographic locations, dietary habits, genders [10][11][12][13][14][15][16][17][18][19]. Moreover, the variations in venoms may dictate different severities in the clinical symptoms post N. kaouthia envenomation [20,21]. ...
Article
The Indian monocled cobra (Naja kaouthia) is one of the most prevalent venomous snakes in northeast India (NEI) and is the cause of many fatalities. The composition of NEI N. kaouthia venom (NkV) was deciphered using two different proteomic approaches: (i) 1D SDS-PAGE coupled to label-free quantification of protein bands using stringent identification criteria and (ii) reversed-phase high-performance liquid chromatography (RP-HPLC) followed by quantification based on area under the RP-HPLC peaks. The proteomic data from both strategies were compared. Proteomic analyses from both workflows identified 32 proteins (toxins) distributed over 10–14 snake venom protein families in NEI NkV. The relative abundances of the venom proteins determined from the analytical workflows coincided with the densitometry band intensities of the NEI NkV. Phospholipase A2 (13.1–16.0%) and three-finger toxins (58.5–64.2%) represented the most abundant enzymatic and non-enzymatic proteins in NEI NkV, respectively. Immuno-cross-reactivity studies by enzyme-linked immunoassay and immunoblot analyses pointed to the poor efficacy of commercial PAVs in recognizing the low molecular mass (<15 kDa) toxins of NEI NkV. Spectrofluorometric titration determined the presence of NEI NkV-specific antibodies in commercial PAV, at a level that was higher than that previously reported for eastern India NkV-specific antibodies in commercial antivenom.
... The authors speculate that the differences could have ecological consequences. Although these ontogenetic diet shifts often are associated with observable alterations on venom-related adaptations (28)(29)(30)(31)(32), that is not always the case. In rattlesnakes (Crotalus oreganus), the ontogenetic diet shift from small lizards in juvenile snakes to rodents and small mammals in mature snakes is not linked to significant changes in venom toxicity (33). ...
Article
Toxin evolution in animals is one of the most fascinating and complex subjects of scientific inquiry today. Gaining an understanding of toxins poses a multifaceted challenge given the diverse modes of acquisition, evolutionary adaptations, and abiotic components that affect toxin phenotypes. Here, we highlight some of the main genetic and ecological factors that influence toxin evolution and discuss the role of antagonistic interactions and coevolutionary dynamics in shaping the direction and extent of toxicity and resistance in animals. We focus on toxic Pacific newts (family Salamandridae, genus Taricha) as a system to investigate and better evaluate the widely distributed toxin they possess, tetrodotoxin (TTX), and the hypothesized model of arms-race coevolution with snake predators that is used to explain phenotypic patterns of newt toxicity. Finally, we propose an alternative coevolutionary model that incorporates TTX-producing bacteria and draws from an elicitor–receptor concept to explain TTX evolution and ecology.
... These observations are in concordance with the PLA 2 s activities in the development of the hemolytic effects (Vishwanath et al., 1988;Stoykova et al., 2013) and are consistent with the findings of Lomonte et al. (2012), where the procoagulant activity of venom of the genus Bothriechis was directly proportional to the concentration of PLA 2 s in the total venom. PLA 2 may be related to the pre-digestion process complementing the paralysis and death caused by other toxins (Mackessy, 1988;Queiroz et al., 2008). ...
Article
Bothriechis schlegelii is a venomous snake found in Central and South America, mainly sighted in regions devoted to agriculture. However, in Colombia, little is known about its contribution to the total envenoming cases. Furthermore, there are no reports of the biochemical and functional activities of venoms from the southwest populations, and the differences respecting other populations are unknown. This study analyzed the protein profiles of venom samples obtained from three specimens originating from this region of Colombia using electrophoresis and chromatography. The lethality, edema-induction, hemorrhagic, defibrinating, coagulant, and indirect hemolytic activities were also evaluated. As a result, venoms were composed of proteins with a wide range of molecular weights, most of them below <37 kDa, with differences between male and female electrophoretic and chromatographic profiles. These variations were also observed in the evaluation of venom functional activities such as pro-coagulant, indirect hemolytic, and edema-inducing activities, whereas neither hemorrhagic nor defibrinating activities were detected. These results are also different considering reports with venom samples from other geographical locations, restating the existence of high intraspecific variability in B. schlegelii venoms, which could have relevant pathophysiological and therapeutic implications.
... As observed herein, higher proteolytic activity presented by adult venoms was reported by other authors [26,87], including Furtado et al. studying B. moojeni [28]. Interestingly, caseinolytic activity among females increased until 2 years-old and decreased afterwards. ...
Article
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The Brazilian lancehead (Bothrops moojeni) has a wide distribution in Brazil and represents a serious public health hazard. Previous works reported that the symptoms of snakebites caused by B. moojeni juveniles’ bites were mainly related to coagulation, while those caused by adults’ bites had a more prominent local damage. In this work, we analyzed the venoms of B. moojeni at different life stages to better understand the ontogeny shift in this species. Snakes were grouped by age and sex, and venom pools were formed accordingly. Compositional analyses by one-dimensional electrophoresis (1-DE), chromatography, and mass spectrometry revealed that ontogenetic changes might be mostly related to phospholipase A2 (PLA2) and metalloproteases. Regarding the venoms functional aspect, proteolytic, L-amino acid oxidase, PLA2, and coagulant in vitro activities were assayed, but only the first and the last ones showed age-related changes, with the venom of snakes up to 1 year-old displaying lower proteolytic and higher coagulant activities, while those from 2 years-old onward presented the opposite relation. The venoms of 3 years-old snakes were exceptions to the compositional and functional pattern of adults as both venoms presented profiles similar to neonates. Sex-related differences were observed in specific groups and were not age-related. In vivo experiments (median lethal dose and hemorrhagic activity) were statistically similar between neonates and adults, however we verified that the adult venom killed mice faster comparing to the neonates. All venoms were mostly recognized by the antibothropic serum and displayed similar profiles to 1-DE in western blotting. In conclusion, the Brazilian lancehead venom showed ontogenetic shift in its composition and activities. Furthermore, this change occurred in snakes from 1 to 2 years-old, and interestingly the venom pools from 3 years-old snakes had particular characteristics, which highlights the importance of comprehensive studies to better understand venom variability.
... Most research on snake venom has been done on adult snakes, mostly because the bites of young snakes cause less pathological effects than adult snakes in humans (ZELANISE et al., 2011), but still numerous reports exist detailing differences in the venoms of snake with age (LOMONTE et al., 1983;MACKESSY, 1988;CHIPPAUX et al., 1991) MINTON and WEINSTEIN (1985) confirmed that venom form young C. atrox was much more lethal than that of adults from same locality but had only about one fourth the proteolytic activity. They tested same snakes fifteen months later, and lethality had declined almost five-fold, and protease activity had approached adult levels. ...
Conference Paper
At the time of birth, newborn llamas and alpacas present in the anterior longitudinal presentation and dorso-sacral position with the head resting on the metacarpal bones of the extended forelegs. The newborn (called a cria) stands very soon to suckle colostrum within the first 2 hours, and may suckle up to 10 times per day during the first seven days of life. Colostrum is only available for 3-4 days, but immunoglobulin absorption occurs only during the first 24 hours of life. Milk production reaches its peak by 2-3 weeks of lactation and then declines by the first month of lactation. Preweaning mortality is 20-80 % in South America, as opposed to 2-5 % in developed countries. The most common diagnoses of neonatal llamas and alpacas are systemic inflammations, congenital defects, sepsis, gastrointestinal diseases, and others.
... Most research on snake venom has been done on adult snakes, mostly because the bites of young snakes cause less pathological effects than adult snakes in humans (ZELANISE et al., 2011), but still numerous reports exist detailing differences in the venoms of snake with age (LOMONTE et al., 1983;MACKESSY, 1988;CHIPPAUX et al., 1991) MINTON and WEINSTEIN (1985) confirmed that venom form young C. atrox was much more lethal than that of adults from same locality but had only about one fourth the proteolytic activity. They tested same snakes fifteen months later, and lethality had declined almost five-fold, and protease activity had approached adult levels. ...
Conference Paper
At the time of birth, newborn llamas and alpacas present in the anterior longitudinal presentation and dorso-sacral position with the head resting on the metacarpal bones of the extended forelegs. The newborn (called a cria) stands very soon to suckle colostrum within the first 2 hours, and may suckle up to 10 times per day during the first seven days of life. Colostrum is only available for 3-4 days, but immunoglobulin absorption occurs only during the first 24 hours of life. Milk production reaches its peak by 2-3 weeks of lactation and then declines by the first month of lactation. Preweaning mortality is 20-80 % in South America, as opposed to 2-5 % in developed countries. The most common diagnoses of neonatal llamas and alpacas are systemic inflammations, congenital defects, sepsis, gastrointestinal diseases, and others.
... This may be due to differences in species and variation in venom compositions [25]. Overall, changes (increase or decrease (in the intensity of toxic activities of venom from "captive vs. wild" and "adult vs. neonate" snakes may be related to the type of prey consumed and habitat shifts for adaptive venom evolution [49,50,51,52]. A study conducted by Gibbs et al. [53], illustrated that adult snakes are likely influenced by different diets, e.g. ...
Article
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Gloydius caucasicus (NIKOLSKY, 1916) is a member of the Viperidae family in Iran. Comprehensive understanding of the toxigenic characteristics of snake venom is important for clinical monitoring of snakebite patients and effective therapy. We compared the toxic activities of venoms and the neutralization capacity of antivenoms produced with venoms from wild adult (WA) with long-term captive adult (LCA) of G. caucasicus in order to obtain more effective antivenom from LCA in therapy, and subsequently protect G. caucasicus from overharvesting for its venom, which poses a real threat of extinction for the species. Our results showed that LD50 of WA and LCA were 16.8 μg/dose and 17.7 μg/dose, respectively. Lower hemorrhagic and necrotic (p ≥ 0.05), and higher coagulative and edematogenic activities (p ≤ 0.05) were observed in WA compared with LCA venom. Also, captive-born neonates exhibited weaker toxic activities compared with captive adult snakes, which could be an age-related difference. Study data illustrated that effective capacity of LCA antivenom to neutralize the toxic activities of WA viper venom. According to the results, about 0.4–4 μl of LCA antivenom is required to neutralize the toxic activities of 1 μg of WA venom, indicating its efficacy in treatment of snakebites in humans. On this basis, it is recommended that capture of wild snakes for their venom be discontinued to reduce their future extinction risk.
... The finding that, compared to adults, newborn specimens from both sides of Costa Rica have more proteolytic, hemorrhagic, edema-forming and lethal venoms, whereas venom from adult specimens are more hemolytic and myotoxic [75,76], supports the view that the age-dependent transition in the composition of the venom may be functionally relevant. In accordance with this evidence, optimal foraging theory predicts that predators should either maximize energy gains or minimize time spent obtaining a fixed amount of energy [77,78], and ontogenetic shift in diet from ectothermic prey (arthropods, lizards, and amphibians) to endothermic prey (mammals) has been associated with a shift in snake venom toxicity, with juvenile venom being more toxic to ectotherms and adult venom more toxic towards mammals [4,[79][80][81]. ...
Article
Bothrops asper is a venomous pitviper that is widely distributed and of clinical importance in Mesoamerica and northern South America, where it is responsible for 50–80% of all envenomations by Viperidae species. Previous work suggests that B. asper has a complex phylogeographic structure, with the existence of multiple evolutionarily distinct lineages, particularly in the inter-Andean valleys of north South America. To explore the impact of the evolutionary history of B. asper on venom composition, we have investigated geographic variation in the venom proteome of this species from the populations from the Pacific side of Ecuador and south-western Colombia. Among the 21 classes of venom components identified, proteins from mainly four major toxin families, snake venom metalloproteases (PI- and PII-SVMP), phospholipases A2 (K49- and D49-PLA2s), serine proteinases (SVSP), and C-type lectins-like (CTL) proteins are major contributors to the geographic variability in venom. Principal component analyses demonstrate significant differences in venom composition between B. asper lineages previously identified through combination of molecular, morphological and geographical data, and provide additional insights into the selection pressures modulating venom phenotypes on a geographic scale. In particular, altitudinal zonation within the Andean mountain range stands out as a key ecological factor promoting diversification in venom. In addition, the pattern of distribution of PLA2 molecules among B. asper venoms complements phylogenetic analysis in the reconstruction of the dispersal events that account for the current biogeographic distribution of the present-day species' phylogroups. Ontogenic variation was also evident among venoms from some Ecuadorian lineages, although this age-related variation was less extreme than reported in B. asper venoms from Costa Rica. The results of our study demonstrate a significant impact of phylogenetic history on venom composition in a pitviper and show how analyses of this variation can illuminate the timing of the cladogenesis and ecological events that shaped the current distribution of B. asper lineages. Biological significance Bothrops asper, called “the ultimate pitviper” due to its defensive behavior, large body size, and medical importance, represents a species complex that is widely distributed from southern México southwards across north‐western South America to north-western Perú. This work reports the characterization of the venom proteomes of B. asper lineages from the Pacific sides of Ecuador and south-western Colombia. Multivariate analyses indicate that variability in venom composition among the B. asper lineages is driven by proteins from four major toxin families, presumably in response to selection pressures created by recent and historical ecological conditions created by geological and climatic events from the Pliocene-Pleistocene to the present along the Central and South American Continental Divide. The emerging biogeographic pattern of venom variation, interpreted in the context of the current phylogenetic hypotheses, support and complement previously proposed evolutionary Plio-Pleistocene dispersal events that shaped the present-day distribution range of B. asper lineages. In addition, our venomics data indicate the occurrence of genetic exchange between Colombian and Pacific Costa Rican populations, which may have occurred during the second wave of B. asper migration into Mesoamerica. Our work represents a foundation for a future broader sampling and more complete “-omics” analyses to deepen our understanding of the patterns and causes of venom variation in this medically important pitviper.
... The results of these studies indicate that individual (intra-population) venom variation is genetically inherited and is not altered ontogenetically by environmental factors, seasonal variation, or diet [14,15,[20][21][22]. An exception to this is ontogenetic venom changes in species with a bi-phasic juvenile to adult shift in dietary ecology [23][24][25]. Interpopulation venom variation can be significant [13,26], and as venom is a trophic adaptation, these divergences are assumed to be the result of selection pressure due to local differences in diet, although this level of dietary data is usually lacking. Taipans are a dietary specialist, so all populations have a broadly similar diet, and it can reasonably be assumed that their venom composition would be fairly uniform throughout their distribution. ...
... Rattlesnake envenomation typically manifests as pain, edema, hemorrhage, coagulopathy, and necrosis, sometimes resulting in permanent disfigurement and/or kidney failure (Arnold, 1982;Ownby, 1982). Systemic neurotoxicity resulting in respiratory paralysis is seldom seen in rattlesnake envenomations in the United States (Bush and Siedenburg, 1999;Jansen et al., 1992;Massey et al., 2012), even though neurotoxins have evolved in some species, or populations within species (Glenn and Straight, 1978;Mackessy, 1988;Massey et al., 2012;Sunagar et al., 2014;Wilkinson et al., 1991). The only effective treatment for rattlesnake envenomation is antivenom administration (Arnold, 1982;Boyer et al., 2015). ...
Article
Envenomations by the Southwestern Speckled Rattlesnake (Crotalus pyrrhus) are fairly rare. Previous descriptions in the literature do not include locality, an important factor in the clinical symptoms or syndromes of snakebites resulting from geographic variation in venom composition. Here, we describe two cases of envenoming by C. pyrrhus from two Arizona localities (Tinajas Altas Mountains, Yuma County, and Phoenix Mountains, Maricopa County). Both patients experienced swelling, but neither demonstrated coagulopathy, thrombocytopenia, or hypofibrinogenemia. The Phoenix Mountains patient developed hemorrhagic bullae and tissue damage in his bitten extremity, necessitating the amputation of the distal portion of his middle finger. Treatment for both consisted of medication for pain, isotonic crystalloid, and antivenom therapy with recovery in each case. Based on visual inspection of 1D-gels and RP-HPLC chromatograms, venom samples were largely similar but appeared to differ quantitatively for several toxin families between and within populations.
Article
Snake venoms are complex biochemical secretions under strong selection for prey subjugation, and venoms are tightly linked to the biotic communities that snakes inhabit. Physiological adaptations for venom resistance have been identified in various snake prey species, but fewer snake predators, with research in this area largely biased towards mammalian species. Fewer investigations have assayed for the presence of resistance mechanisms in avian systems. Birds of prey (hereafter “raptors”; orders Accipitriformes, Falconiformes, and Strigiformes) represent major sources of predation for snakes. Raptor dietary habits range from snake specialists to non-snake feeders, and this continuum of snake predation frequency among species creates the ideal system in which to explore the presence and strength of venom resistance. We assayed sera from a suite of Great Plains raptors against snake venom metalloproteinases (SVMPs) of the Prairie Rattlesnake (Crotalus v. viridis) to test the general hypotheses that 1) raptor sera will display elevated SVMP inhibition compared to a naïve avian model (domestic chicken; Gallus gallus) and 2) raptor species with high levels of rattlesnake predation will more effectively inhibit SVMP activity than those that are not known to feed on rattlesnakes. We found that raptors do possess elevated SVMP inhibition in comparison to a naïve avian model, but this level of inhibition remains low and is unlikely to be biologically significant in detoxifying venoms. We found no evidence suggesting that inhibitory potential of different raptor sera corresponds to the level of rattlesnake predation associated with each species. The widespread lack of SVMP inhibition in diverse raptors underscores the complexity of venom resistance dynamics in natural systems and further suggests that physiological venom resistance mechanisms may be poorly developed in birds more broadly.
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Predation has the potential to impart strong selective pressures on organisms within their environments, resulting in adaptive changes in prey that minimize risk of predation. Pressures from venomous snakes present an exceptional challenge to prey, as venom represents a unique chemical arsenal evolutionarily tailored to incapacitate prey. In response, venom resistance has been detected in various snake prey species, and to varying degrees. This study analyzes venom resistance in an eastern Colorado grassland habitat, where the Prairie Rattlesnake (Crotalus viridis) and Desert Massasauga Rattlesnake (Sistrurus tergeminus edwardsii) co-occur with a suite of grassland rodents. We test for venom resistance across rodent and snake pairings using two geographically distant field sites to determine the role of 1) predation pressure and trophic ecology, and 2) sympatric and allopatric patterns of venom resistance. Resistance was measured using serum-based metalloproteinase inhibition assays to determine potential inhibition of proteolytic activity, augmented by median lethal dose (LD50) assays on rodent species to assess toxicity of crude venoms. Resistance is present in several rodent species, with strong resistance present in populations of Eastern Woodrat (Neotoma floridana), Ord’s Kangaroo Rat (Dipodomysordii), and Northern Grasshopper Mouse (Onychomys leucogaster). Resistance is less developed in other species, including the House Mouse (Mus musculus) and Plains Pocket Mouse (Perognathus flavescens). An unexpected differential is present, where Lincoln County Kangaroo Rats are highly resistant to venom of co-occurring Prairie Rattlesnakes yet are sensitive to an allopatric population of Prairie Rattlesnakes in Weld County. Lincoln Co. Northern Grasshopper Mice also demonstrate extremely elevated resistance to Weld Co. Prairie Rattlesnake venoms, and they may possess resistance mechanisms for myotoxin a, an abundant component of Weld Co. C. v. viridis venoms. This study illustrates the complexity of venom resistance in biological communities that can exist when incorporating multiple species interactions. Future studies aimed at characterizing resistance mechanisms at the molecular level will provide a more detailed physiological context for understanding mechanisms by which resistance to venoms occurs.
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Studying the consequences of hybridization between closely related species with divergent traits can reveal patterns of evolution that shape and maintain extreme trophic adaptations. Snake venoms are an excellent model system for examining the evolutionary and ecological patterns that underlie highly selected polymorphic traits. Here we investigate hybrid venom phenotypes that result from natural introgression between two rattlesnake species that express highly divergent venom phenotypes: Crotalus o. concolor and C. v. viridis. Though not yet documented, interbreeding between these species may lead to novel venom phenotypes with unique activities that break the typical trends of venom composition in rattlesnakes. The characteristics of these unusual phenotypes could unveil the roles of introgression in maintaining patterns of venom composition and variation, including the near ubiquitous dichotomy between neurotoxic or degradative venoms observed across rattlesnakes. We use RADseq data to infer patterns of gene flow and hybrid ancestry between these diverged lineages and link these genetic data with analyses of venom composition, biological activity, and whole animal model toxicity tests to understand the impacts of introgression on venom composition. We find that introgressed populations express admixed venom phenotypes that do not sacrifice biological activity (lethal toxicity) or overall abundance of dominant toxins compared to parental venoms. These hybridized venoms therefore do not represent a trade-off in functionality between the typical phenotypic extremes but instead represent a unique combination of characters whose expression appears limited to the hybrid zone.
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In many species, the spatial ecology of early age classes can differ significantly from adults. Adult Western Rattlesnakes (Crotalus oreganus Holbrook, 1840) at the northern extent of their range undertake small-scale but important seasonal migrations between communal hibernacula and summer foraging and mating grounds. Mature snakes also show annual fidelity to their migratory paths, providing a useful system to examine the development of migratory behaviour. We examined and compared spring outbound migratory movements of juveniles and adults at a site in southern British Columbia, Canada, using radiotelemetry data collected between 2011 and 2016 (adult snakes) and in 2021 (juvenile snakes). We found that compared with adult rattlesnakes, juveniles displayed similar directional orientation, direction of vertical migration, and path sinuosity, but initiated spring migrations later and exhibited shorter movements in terms of distances and rates. For example, juvenile straight-line migration distance (262 ± 90 m) was significantly shorter than that for adults (1069 ± 134 m; P < 0.001). We provide a starting point in attempting to understand an important question in migration—how individuals early on in their lives adopt different tactics—while contributing to our growing understanding of the complexity of patterns and variation in the movement ecology of a far-ranging snake.
Article
Understanding the molecular basis of adaptations in coevolving species requires identifying the genes that underlie reciprocally selected phenotypes, such as those involved in venom in snakes and resistance to the venom in their prey. In this regard, California ground squirrels (CGS; Otospermophilus beecheyi) are eaten by northern Pacific rattlesnakes (Crotalus oreganus oreganus), but individual squirrels may still show substantial resistance to venom and survive bites. A recent study using proteomics identified venom interactive proteins (VIPs) in the blood serum of CGS. These VIPs represent possible resistance proteins, but the sequences of genes encoding them are unknown despite the value of such data to molecular studies of coevolution. To address this issue, we analyzed a de novo assembled transcriptome from CGS liver tissue-where many plasma proteins are synthesized-and other tissues from this species. We then examined VIP sequences in terms of three characteristics that identify them as possible resistance proteins: evidence for positive selection, high liver expression, and nonsynonymous variation across CGS populations. Based on these characteristics, we identified five VIPs (i.e., α-2-macroglobulin, α-1-antitrypsin-like protein GS55-LT, apolipoprotein A-II, hibernation-associated plasma protein HP-20, and hibernation-associated plasma protein HP-27) as the most likely candidates for resistance proteins among VIPs identified to date. Four of these proteins have been previously implicated in conferring resistance to the venom in mammals, validating our approach. When combined with the detailed information available for rattlesnake venom proteins, these results set the stage for future work focused on understanding coevolutionary interactions at the molecular level between these species.
Article
Venom is an integral feeding trait in many animal species. Although venom often varies ontogenetically, little is known about the proximate physiological mediators of venom variation within individuals. The glucocorticoid hormone corticosterone can alter transcription and activation of proteins, including homologues of snake venom components such as snake venom metalloproteinases (SVMP) and phospholipase A2 (PLA2). Corticosterone is endogenously produced by snakes, varying seasonally and also in response to stress, and is a candidate endogenous mediator of changes in venom composition and functional activity. Here, we tested the hypothesis that corticosterone induces changes in snake venom by sampling the venom of wild adult rattlesnakes before and after being treated with either empty (control) or corticosterone-filled (treatment) silastic implants. We measured longitudinal changes in whole-venom composition, whole-venom total protein content, and enzymatic activity of SVMP and PLA2 components of venom. We also assessed within-individual repeatability of venom components. Despite successfully elevating plasma corticosterone in the treatment group, we found no effect of corticosterone treatment or average plasma corticosterone level on any venom variables measured. Except for total protein content, venom components were highly repeatable within individuals (r > 0.9). Our results indicate that the effects of corticosterone, a hormone commonly associated with stress and metabolic functions, in adult rattlesnake venom are negligible. Our findings bode well for venom researchers and biomedical applications that rely on consistency of venoms produced from potentially stressed individuals, and provide an experimental framework for future studies on proximate mediators of venom variation across an individual’s lifespan.
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Venomous snakes are among the world's most specialised predators. During feeding, they use fangs to penetrate the body tissues of their prey, but the success of this penetration depends on the shape of these highly specialised teeth. Here, we examined the evolution of fang shape in a wide range of snakes using 3D geometric morphometrics (3DGM) and cross‐sectional tooth sharpness measurements. We investigated the relationship of these variables with six diet categories based on the prey's biomechanical properties, and tested for evolutionary convergence using two methods. Our results show that slender elongate fangs with sharp tips are used by snakes that target soft‐skinned prey (e.g. mammals), while fangs become more robust and blunter as the target's skin becomes scaly (e.g. fish, reptiles) and eventually hard‐shelled (e.g. crustaceans), both with and without correction for evolutionary allometry. Convergence in fang shape is present, indicating that fangs of snakes with the same diet are more similar than those of closely related species with different diets. Establishing the relationship between fang morphology and diet helps to explain how snakes became adapted to different lifestyles, while also providing a proxy to infer diet in lesser‐known species or extinct snakes from the fossil record. This article is protected by copyright. All rights reserved
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The Tzabcan rattlesnake (Crotalus tzabcan) is endemic to the Yucatán Peninsula and is part of the C. durissus complex. Although relatively widespread, it is rather an uncommon species; therefore, little is known about its natural history. Herein, we describe the diet of C. tzabcan on the basis of data from field encounters, museum specimens, and published data. Dietary samples were collected from Campeche, Quintana Roo, and Yucatán, Mexico, and literature records from Belize, representing most of the species’ distribution range. Examination of 50 individuals resulted in 28 prey items obtained from 27 snakes. The diet of C. tzabcan consisted exclusively of mammals, including the orders Rodentia (86% of the prey items) and Soricomorpha (7%), and no ontogenetic shift in prey type was detected. However, an ontogenetic telescope is evident, where adults consume larger prey than juveniles but continue feeding on small prey. No sexual dimorphism in snout–vent length and total length was detected in C. tzabcan. No sexual differences in prey mass were found when controlling for snake body length, nor when comparing between sexes in adults and juveniles. The presence of prey was not related to snake collection date, suggesting year-round feeding. There was no difference in prey class and size among snakes from Yucatán dry forest and moist forest. These results suggest a homogenous diet among sexes, seasons, and populations. This is the first detailed study on the diet and feeding ecology of C. tzabcan, and it adds five new prey species: Cryptotys mayensis, Heteromys desmarestianus, H. gaumeri, Oryzomys couesi, and Rattus rattus, as well as two previously reported ones: Mus musculus and Sigmodon toltecus. These findings contrast with anecdotal reports of C. tzabcan consuming reptiles and birds and show many similarities with the related species C. durissus. Additional studies on the natural history of C. tzabcan and related species would help to better understand how the feeding ecology of Neotropical rattlesnakes differs from those species of temperate zones.
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Identified, in 2 experiments, how proximate factors of the rapid rattlesnake strike affect predatory behavior and the resulting envenomation and capture of the prey. Results indicate that the larger the snake, the more likely it was to hold, rather than quickly release, prey. Snakes possessed a ready reserve of venom sufficient to envenomate up to 4 mice in close succession without loss of killing effectiveness. The head/thorax site on mice was the region most frequently struck and also the site of venom injection that led to the fastest prey death. Small mice were more often retained in the jaws rather than released and occasionally evoked no envenomation at all. A retaliatory bite to the head or body of an attacking snake encouraged a quick release of the prey. Retention of mice following the strike enhanced the severity of envenomation. Poor envenomation on the 1st strike led to a 2nd or even a 3rd follow-up strike. Unlike defensive strikes, offensive (predatory) strikes resulted in no dry bites. This suggests that jaw mechanics may be disrupted during defensive strikes or that the snake can actually control its expenditure of venom. Artificial reduction of venom reserved by milking the venom glands resulted in poor envenomation but stimulated no change in the basic hold/release behavior. (49 ref) (PsycINFO Database Record (c) 2012 APA, all rights reserved)
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Rattlesnake venom injected into live mice before they were swallowed by non-venomous snakes speeded the process of digestion. This effect of venom was more pronounced when digestion was tested at 15°C than at 25°C. The proteolytic activity of viperid venom appears to facilitate the entry of a snake's stomach secretions into the prey's body cavity, inhibiting bacterial activity and thereby reducing the risk of prey putrefying before it can be digested. This effect would be particularly valuable for snakes that eat very large prey in relation to their own body size or that normally experience low body temperatures after feeding. Viperid snakes comprise a disproportionately large fraction of the total snake fauna at high altitudes in North America and Africa. Proteolytic venom appears to be one of a group of characters that permit viperid snakes to swallow and digest large prey. A digestive function of venom probably appeared at an early stage in the evolution of vipers.
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Viperid snakes have stouter bodies, larger heads, and longer jaws than snakes in other families; there are no major differences between the two subfamilies of vipers in these features. A suite of morphological characters that facilitates swallowing large prey finds its greatest expression among vipers, but certain elapid and colubrid snakes have converged upon the same body form. The number of jaw movements required to swallow prey is linearly related to the size of a prey item when shape is held constant. Very small and very large prey are not disproportionately difficult for a snake to ingest. Vipers swallow their prey with fewer jaw movements than do colubrids or boids and can swallow prey that is nearly three times larger in relation to their own size. Proteolytic venom assists in digestion of prey, and melanin deposits shield the venom glands from light that would degrade the venom stores. Ancillary effects of the morphological features of vipers, plus the ability to ingest a very large quantity of food in one meal, should produce quantitative and qualitative differences in the ecology and behavior of vipers and other snakes.
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L-amino acid oxidase prepared from Crotalus adamanteus venom after the method of Wellner and Meister does not contribute to the production of the profound fall in systemic arterial pressure usually seen following injection of crude Crotalus venom, nor does it provoke any early deleterious changes in the dependent variables of the cardiovascular system.The enzyme has no effect on neuromuscular transmission in the mammal. Its LD50 is 9·13 (7·83–10·35) mg per kg test animal body weight.
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Rattlesnakes of three taxa (Crotalus durissus, C. enyo, and C. viridis) struck and released large rodent prey but held smaller rodents in their jaws after striking. The specimens of C. enyo were clearest in this regard; almost all large prey were released, and all small prey were held. This finding is consistent with the fact that large rodents are more dangerous to rattlesnakes than are small rodents. Accordingly, it appears that rattlesnakes have evolved differential predatory strategies for dealing with prey of varying size. Speculation is offered about the manner in which these respective strategies are activated or selected during predatory episodes.
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B. Lomonte, J. A. Gené, J. M. Gutiérrez and L. Cerdas. Estudio comparativo de los venenos de serpiente Cascabel (Crotalus durissus durissus) de ejemplares adultos y recién nacidos. Toxicon21, 379 – 384, 1983. — Venoms from adult and newborn Central American rattlesnakes (Crotalus durissus durissus) were compared for lethal, proteolytic, hemorrhagic, myonecrotic, edematigenous and in vitro hemolytic activities. Electrophoretic and immunoelectrophoretic patterns showed some differences between these venoms. Venom from newborn snakes was devoid of hemorrhagic and edematigenous activities, whereas the venom from adult specimens induced these effects. On the other hand, newborn snake venom showed higher lethality and indirect hemolytic activity, and lower proteolytic activity, than venom from adult specimens. Both types of venoms induced only slight myonecrosis in mice, as judged by histological observation. The ed50 of an antivenom, in terms of absolute weight neutralized per ml of serum, was lower for the newborn specimens venom than for adult's venom, however, for each venom the number of ld50 neutralized was similar.
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Venom samples separately collected at monthly intervals from three Crotalus atrox specimens as they aged from 2 to 22 months showed many quantitative changes of biological activities. But more important were qualitative changes of coagulation activity. Up to 8 months the venoms clotted fibrinogen solutions directly. At 9 to 10 months, plasma was clotted but not fibrinogen. Subsequently the venoms no longer clotted plasma. Qualitative venom changes with growth of snakes could explain some of the conflicting reports both on clinical aspects of snake bite in man and on experimental venom work. The findings emphasize the importance for clinicians dealing with snake bite to monitor the clot-quality of their patient's blood--a simple bedside test for defibrinogenation, no-clot indicating zero fibrinogen and speck-clot representing fibrinogen concentrations under 50 mg/100 ml. With strongly defibrinating venoms, non-clotting blood is a very sensitive sign of systemic envenoming. In contrast, the relatively feeble defibrinating activity of juvenile C. atrox venom suggests that, if the blood is non-clotting in C. atrox bites, this indicates the victim has received a potentially lethal or near-lethal dose of venom urgently requiring effective antivenom therapy.
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Crotalus ruber ruber venom contains several different proteases, and the proteolytic activity of the crude venom is 6-15 times greater in adult than in juvenile venom. Venom samples were assayed for proteolytic, phosphodiesterase, L-amino acid oxidase and elastinase-like activities and were subjected to gel filtration on BioGel P-100. Two major size classes of proteases were resolved (mol. wt 67,000 and 20,500). EDTA, N-ethylmaleimide (N-EM) and 1,10-phenanthroline inhibited proteolytic activity of crude venom, and EDTA, Zn2+ and Cu2+ inhibited proteolytic activity of the fractionated venom.
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Venom samples from western diamondback rattlesnakes (Crotalus atrox) from 13 localities in the United States were tested for i.v. and s.c. lethality for mice, protease activity, hemorrhagic activity, and the presence of Mojave toxin. Electrophoresis on polyacrylamide gel was used to compare protein composition. The neutralizing effect of two commercial antivenoms was evaluated against selected samples of venom. Venom of young snakes from north Texas was compared with that of adults from the same locality. Venom samples from the southwest portion of the range showed highest lethality, those from the northeast portion lowest. This trend was reversed with respect to protease activity. Hemorrhagic activity showed little geographic variation, but northern samples tended to be slightly higher. Differences in venom protein composition were evident between snakes from the eastern and western portions of the range. Mojave toxin in small to trace amounts was detected in two Arizona venom samples and one from west Texas. Antivenoms were relatively ineffective in neutralizing lethality. Venom of young snakes from north Texas was much more lethal by s.c. injection than that of adult snakes from any part of the range, but very low in protease activity. Hemorrhagic activity was about equal to that of adult snakes from the same region. Fifteen months later, lethality had declined almost five-fold, and protease activity had approached adult levels. There was a distinct change in protein composition. Mojave toxin was not detected in venoms of the young snakes.
Article
Hybridomas secreting monoclonal antibodies against Mojave toxin were established. The antibodies were used for identifying cross-reacting proteins in individual C. s. scutulatus and other Crotalus venoms and to isolate Mojave toxin. The antibodies recognized five bands with a pI range from 5.1 to 6.1 in immunoblots of electrofocused crude venom and Mojave toxin purified by immunoaffinity chromatography. The specificity of the antibodies was for the basic subunit of the toxin, which resolved into four bands of pI between 9.3 and 9.6. Individual C. s. scutulatus venoms of snakes from Texas and southern Arizona had multiple bands with pI's ranging from 4.9 to 6.3. Cross-reacting proteins were also recognized by the antibodies in the electrophoresed venoms of C. basiliscus, C. d. durissus, C. d. terrificus, C. h. horridus and C. v. concolor, and may be isolated by immunoaffinity chromatography with the monoclonal antibodies.
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Purified phospholipase A2 from Bothrops alternatus venom is one single protein species with a molecular weight of 15,000 and isoelectric point 5.08. When injected i.p. or i.v. at a dose of 0.7 microgram/g body weight it is lethal to mice, eliciting a typical syndrome of dyspnea, tachycardia, arrhythmia and irreversible shock. Post mortem and histopathologic studies have demonstrated that the lungs (massive pulmonary hemorrhage), heart (foci of myocardial and endocardial necrosis with interfibrillar hemorrhage), liver (congestion, hepatocytic microvacuolization with zones of massive necrosis) and kidneys (foci of tubular and glomerular necrosis) were severely injured. Except for the less extensive hemorrhages and the significantly longer survival time, the observed lesions are similar to those observed after the injection of lethal doses of whole venom. The lethal potency of the purified enzyme (LD50 i.p. 0.14 microgram/g body weight) is 46-fold greater than that of the whole venom (LD50 i.p. 6.4 micrograms/g body weight). The contribution of phospholipase A2 to the overall lethal effect of B. alternatus venom is suggested by the decreased lethal potency of a venom sample in which a significant amount of phospholipase A2 has been removed and the full restoration of the lethal potency upon supplementation of the depleted sample with purified enzyme. It is concluded that phospholipase A2 is a major component responsible for lethality of the whole B. alternatus venom, while the contribution of other venom components appears to be significant mainly in reducing the time of survival.
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In vitro incubation of rat aorta with rattlesnake venom led to the destruction of the elastic fibers of the aorta. Correlated with this action, venom contained enzyme activity which solubilized Congo red-elastin and catalyzed the release of p-nitrophenol from the synthetic elastase substrate t-BOC-L-alanine p-nitrophenol. Therefore, rattlesnake venom contains an elastase-like enzyme.
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Caudoxin, a single-chain phospholipase A2 isolated from the venom of Bitis caudalis is a toxic polypeptide with a formula weight of 13,332 dalton. The LD50 in mice (i.p.) was 0.18 (0.15-0.22) mg/kg. In the chick biventer cervicis muscle preparation the toxin (1-10 micrograms per ml) caused complete neuromuscular blockade without affecting the response of the muscle to acetylcholine. In the mouse phrenic nerve-diaphragm preparation, the toxin abolished the indirectly elicited contraction without inhibiting that evoked directly. When this preparation was bathed in a low calcium (0.6 mM) medium, the toxin induced a triphasic change in the indirectly evoked contractions: an immediate initial inhibition followed by augmentation and then a second phase of inhibition leading to irreversible neuromuscular blockade. Electrophysiological studies in the same preparation showed a similar triphasic change in the quantal content of endplate potentials. The frequency of miniature endplate potentials first increased and then decreased, while the resting membrane potential was not significantly decreased by the toxin. Histological study showed that the toxin caused local myonecrosis only at a higher dose (2 mg/kg mouse). It is concluded that caudoxin produced a neuromuscular block by acting selectively on a presynaptic site. However, the site of binding appears to be different from that of beta-bungarotoxin since combination of the toxin with beta-bungarotoxin caused potentiation of its neuromuscular blocking action rather than addition.
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Spectrophotometric assays for snake venom l-amino acid oxidase have been developed based on the conversion of l-kynurenine (I) to kynurenic acid (III) and of 3,4-dehydro-l-proline (IV) to pyrrole-2-carboxylic acid (VI).
Enzymes in snake venoms In: Snake venoms: handbook of experimental pharmacology Intraspecific variation in composition of venom of the jumping viper, Bo-throps nummifera
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Rattlesnake venom enzymes that interact with components of the hemostatic system Observations on toxicity and antigenic makeup of venoms from juvenile snakes
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A field guide to western rep-tiles and amphibians The deter-mination of proteolytic activity of snake venoms by means of a chromogenic substrate
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Venoms: chemistry and molecular biology Chemistry of rattlesnake venoms In: Rattlesnake venoms: their action and treatment Proteolytic activity of snake venoms
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Uber eine neue L-aminosaure-oxydase
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