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Evaluating leukocytes provides insight in the overall health of animals by characterizing pathologies related to infection, inflammation, and stress. In reptile species, many leukocyte quantification methods are used to evaluate white blood cell (WBC) parameters; however, agreement and analytical variability (repeatability) between the various methods have yet to be reported and critically assessed in many species, including the prairie rattlesnake (Crotalus viridis). The objectives of this study were to 1) test agreement between hematology parameters determined by phloxine-based stain (Eosinophilic leukopet, EO), Natt–Herrick's (NH), and estimate (EST) methods using fresh whole blood and lithium-heparinized blood; 2) determine the agreement between estimates from a veterinary student and from a board-certified clinical pathologist; and 3) determine the analytical variability of EO, NH, and EST methods. Venipuncture was performed in 27 adult snakes, and agreement was then assessed using Bland–Altman plots and Passing–Bablock regression between all methods. There was a variable degree of proportional and constant error between many of the quantification methods, but descriptive statistics for all methods fell within biologically relevant values. There was good agreement in the EST method between the clinical pathologist and the veterinary student, but the student had a significantly higher mean WBC count in fresh whole blood. The lowest analytical variability was observed with the NH method (15.4%), whereas the highest was observed with the EST method (58.6% for the clinical pathologist, 59.4% for the veterinary student). These results identify the intrinsic variability in reptile hematological methods currently used and emphasize the need to develop a singular diagnostic standard for leukocyte quantification in reptile species. [ABSTRACT FROM AUTHOR]

An individual's migration path shape should affect its fitness, because patchily distributed features (e.g., prey) are encountered more often on straight than tortuous paths. We hypothesized that Prairie Rattlesnakes ( Crotalus viridis viridis (Rafinesque, 1818)) with straighter migration paths should have better body condition, because they encounter prey patches more frequently, and higher migration mortality, because they also encounter predators and hazardous human land uses more frequently, than individuals with tortuous paths. If true, then a straighter path should be favoured when the benefit (resource acquisition) outweighs the cost (mortality risk). Humans pose a significant mortality risk for migrants; thus, the cost of straight-line movement should increase relative to the benefit in more human-dominated landscapes, favouring more tortuous movements. We tested these hypotheses using data on the body condition, mortality, and migration movements of 25 female Prairie Rattlesnakes in one human-dominated and one seminatural landscape. As hypothesized, we found better body condition and higher migration mortality for snakes with straighter migration paths, and that snakes followed more tortuous paths in the human-dominated landscape. Although selection for tortuous movements may reduce rates of migration mortality in human-dominated landscapes, this may ultimately contribute to population declines if poorer body condition reduces overwinter survival or reproductive success. [ABSTRACT FROM AUTHOR]

Morphological data are a conduit for the recognition and description of species, and their acquisition has recently been broadened by geometric morphometric (GM) approaches that co-join the collection of digital data with exploratory ‘big data’ analytics. We employed this approach to dissect the Western Rattlesnake (Crotalus viridis) species-complex in North America, currently partitioned by mitochondrial (mt)DNA analyses into eastern and western lineages (two and seven subspecies, respectively). The GM data (i.e., 33 dorsal and 50 lateral head landmarks) were gleaned from 2,824 individuals located in 10 museum collections. We also downloaded and concatenated sequences for six mtDNA genes from the NCBI GenBank database. GM analyses revealed significant head shape differences attributable to size and subspecies-designation (but not their interactions). Pairwise shape distances among subspecies were significantly greater than those derived from ancestral character states via squared-change parsimony, with the greatest differences separating those most closely related. This, in turn, suggests the potential for historic character displacement as a diversifying force in the complex. All subspecies, save one, were significantly differentiated in a Bayesian discriminant function analysis (DFA), regardless of whether our priors were uniform or informative (i.e., mtDNA data). Finally, shape differences among sister-clades were significantly greater than expected by chance alone under a Brownian model of evolution, promoting the hypothesis that selection rather than drift was the driving force in the evolution of the complex. Lastly, we combine head shape and mtDNA data so as to derived an integrative taxonomy that produced robust boundaries for six OTUs (operational taxonomic units) of the C. viridis complex. We suggest these boundaries are concomitant with species-status and subsequently provide a relevant nomenclature for its recognition and representation. [ABSTRACT FROM AUTHOR]

Objective To compare the effect of antivenom administration on mortality, hospitalization duration, and cost of hospitalization for dogs envenomated by Crotalus viridis. Design Retrospective study (January 2004-December 2012). Setting Private veterinary emergency and referral center. Animals One hundred thirteen dogs with confirmed C. viridis envenomation. Interventions None. Measurements and Main Results Dogs were divided into groups treated with either supportive care only (group 1) or supportive care plus antivenin crotalidae polyvalent (group 2). A modified snakebite severity score was used to compare patients from group 1 and group 2. Patients in group 2 received one 10 mL vial of antivenin crotalidae polyvalent over 4-6 hours. The mortality rate was 1.8% (2/113). Group 1 had a median duration of hospitalization of 20 hours (range, 8-50 hours), while group 2 had a median duration of hospitalization of 24 hours (range, 1.5-74 hours). Group 1 had a median cost of hospitalization of 1050.00 USD (range, 423.52-2266.09 USD) while group 2 had a median cost of hospitalization of 2002.19 USD (range, 1139.91-6908.01 USD). Both the duration of hospitalization ( P < 0.01) and the cost of hospitalization ( P < 0.01) were significantly greater in the group of dogs receiving antivenom. Conclusions Both the cost and the duration of hospitalization were significantly greater in the group of dogs that received antivenom. The difference in cost between the groups was approximately the cost of the antivenom vial and administration. Amongst the parameters evaluated, no significant benefit was associated with antivenom administered to dogs envenomated by C. viridis. [ABSTRACT FROM AUTHOR]

Many snake species make lengthy linear migrations between overwintering sites and summer foraging or mating habitats. Although mountainous topography may restrict migratory movements, most previous studies on migratory snake populations have occurred in areas with low to moderate topographic relief. The objectives of this study were to describe the movement patterns of Prairie Rattlesnakes ( Crotalus v. viridis) in a mountainous landscape, compare those patterns to those of migratory snake populations from areas with lower topographic relief, and test for variation in movement patterns between sexes and among years. We used radiotelemetry to monitor the movements of 21 male and 6 nonpregnant female Prairie Rattlesnakes in the Frank Church Wilderness in central Idaho during the summers of 2006-2008. Mean total distance moved during the entire activity season in 2008 was 4.46 km (range 1.38-7.67); mean maximum distance moved from the hibernaculum was 1.46 km (range 0.69-2.71). Although the movement distances reported here are intermediate to those reported for other migratory snake populations, they are similar to some distances reported from areas with low to moderate topographic relief. This suggests that rattlesnakes are capable of making considerable movements in a mountainous landscape, although factors such as prey availability could also contribute to differences in reported movement distances. Rattlesnakes displayed moderate fidelity to summer activity areas but had similar mean bearings during outbound migration across multiple years. We hypothesize that linear migrations reported from rattlesnakes in many populations actually represent the most-direct movement to annual foraging areas rather than true searching movements. [ABSTRACT FROM AUTHOR]

Here we describe and compare the venomic and antivenomic characteristics of both neonate and adult Prairie Rattlesnake ( Crotalus viridis viridis ) venoms. Although both neonate and adult venoms contain unique components, similarities among protein family content were seen. Both neonate and adult venoms consisted of myotoxin, bradykinin-potentiating peptide (BPP), phospholipase A 2 (PLA 2 ), Zn 2 + -dependent metalloproteinase (SVMP), serine proteinase, L-amino acid oxidase (LAAO), cysteine-rich secretory protein (CRISP) and disintegrin families. Quantitative differences, however, were observed, with venoms of adults containing significantly higher concentrations of the non-enzymatic toxic compounds and venoms of neonates containing higher concentrations of pre-digestive enzymatic proteins such as SVMPs. To assess the relevance of this venom variation in the context of snakebite and snakebite treatment, we tested the efficacy of the common antivenom CroFab® for recognition of both adult and neonate venoms in vitro . This comparison revealed that many of the major protein families (SVMPs, CRISP, PLA 2 , serine proteases, and LAAO) in both neonate and adult venoms were immunodepleted by the antivenom, whereas myotoxins, one of the major toxic components of C . v . viridis venom, in addition to many of the small peptides, were not efficiently depleted by CroFab®. These results therefore provide a comprehensive catalog of the venom compounds present in C . v . viridis venom and new molecular insight into the potential efficacy of CroFab® against human envenomations by one of the most widely distributed rattlesnake species in North America. Biological significance Comparative proteomic analysis of venoms of neonate and adult Prairie Rattlesnake ( Crotalus viridis viridis ) from a discrete population in Colorado revealed a novel pattern of ontogenetic shifts in toxin composition for viperid snakes. The observed stage-dependent decrease of the relative content of disintegrins, catalytically active D49-PLA 2 s, L-amino acid oxidase, and SVMPs, and the concomitant increase of the relative abundance of paralytic small basic myotoxins and ohanin-like toxin, and hemostasis-disrupting serine proteinases, may represent an age-dependent strategy for securing prey and avoiding injury as the snake switches from small ectothermic prey and newborn rodents to larger endothermic prey. Such age-dependent shifts in venom composition may be relevant for antivenom efficacy and treatment of snakebite. However, applying a second-generation antivenomics approach, we show that CroFab®, developed against venom of three Crotalus and one Agkistrodon species, efficiently immunodepleted many, but not all, of the major compounds present in neonate and adult C . v . viridis venoms. [ABSTRACT FROM AUTHOR]

Prairie Rattlesnakes ( Crotalus viridis) are a species of conservation concern in Canada because of suspected population declines. Data on the habitat selection of Prairie Rattlesnakes at northern range limits are essential to inform management strategies and implement recovery plans in Canada and other northern regions. We used radio-telemetry and generalized linear mixed models to create resource selection functions for Prairie Rattlesnakes ( N = 23) in and around Grasslands National Park, Saskatchewan, Canada. Eight habitat variables were compared between used and available sites to quantify microhabitat selection. We found that Prairie Rattlesnakes select for specific sites (i.e., within 1 m) with shrub cover and burrows and avoid bare ground. Shrub cover was two-times higher at used sites compared to available ones, while bare ground showed the opposite trend. In addition, two-thirds of all snake location points were within 1 m of a burrow, which suggests that retreat sites are important. None of the habitat variables measured at 10 m from used points were significant predictors of rattlesnake habitat selection, indicating that rattlesnakes select habitat at a very fine level. Northern range limits present a thermally challenging environment in which burrows and shrubs are likely crucial for thermoregulation. In addition, these retreat sites likely provide a prey source, protection from predators, and suitable areas for reproduction. Retreat sites such as burrows and shrubs are therefore critical components of habitat for Prairie Rattlesnakes and should be included in management strategies for the species. [ABSTRACT FROM AUTHOR]

Background: The neglected human diseases caused by trypanosomatids are currently treated with toxic therapy with limited efficacy. In search for novel anti-trypanosomatid agents, we showed previously that the Crotalus viridis viridis (Cvv) snake venom was active against infective forms of Trypanosoma cruzi. Here, we describe the purification of crovirin, a cysteine-rich secretory protein (CRISP) from Cvv venom with promising activity against trypanosomes and Leishmania. Methodology/Principal Findings: Crude venom extract was loaded onto a reverse phase analytical (C8) column using a high performance liquid chromatographer. A linear gradient of water/acetonitrile with 0.1% trifluoroacetic acid was used. The peak containing the isolated protein (confirmed by SDS-PAGE and mass spectrometry) was collected and its protein content was measured. T. cruzi trypomastigotes and amastigotes, L. amazonensis promastigotes and amastigotes and T. brucei rhodesiense procyclic and bloodstream trypomastigotes were challenged with crovirin, whose toxicity was tested against LLC-MK2 cells, peritoneal macrophages and isolated murine extensor digitorum longus muscle. We purified a single protein from Cvv venom corresponding, according to Nano-LC MS/MS sequencing, to a CRISP of 24,893.64 Da, henceforth referred to as crovirin. Human infective trypanosomatid forms, including intracellular amastigotes, were sensitive to crovirin, with low IC50 or LD50 values (1.10–2.38 µg/ml). A considerably higher concentration (20 µg/ml) of crovirin was required to elicit only limited toxicity on mammalian cells. Conclusions: This is the first report of CRISP anti-protozoal activity, and suggests that other members of this family might have potential as drugs or drug leads for the development of novel agents against trypanosomatid-borne neglected diseases. [ABSTRACT FROM AUTHOR]

Numerous studies have reported genetic fragmentation of species whose habitat has been modified by roads and other anthropogenic features, but it is still not clear how most species respond to roads and whether genetic effects can be detected over a limited number of generations. We used road-crossing models and population genetic analysis (based on microsatellite loci) to make inferences about functional connectivity between populations of the Prairie Rattlesnake ( Crotalus viridis (Rafinesque, 1818)) on opposite sides of the Trans-Canada Highway near Medicine Hat (Alberta, Canada). The road-crossing model predicted a high probability of mortality while crossing the Trans-Canada Highway. However, model-based genetic clustering methods (STRUCTURE and BAPS) did not detect structure; a nonmodel-based clustering method (DAPC) found structure, but most groups consisted of individuals captured throughout the study area. Estimates of effective population size were immeasurably large and power to detect genic differentiation was diminished if the effective size exceeded 500; this reduction in power was intensified when the number of loci was reduced (from eight to five to account for null alleles). Our results corroborate accounts of long-distance migration by this species and indicate that genetic fragmentation may not be easily detectable over this spatial and temporal scale. [ABSTRACT FROM AUTHOR]

Highlights: [•] We infer a dated phylogeny of the longtailed rattlesnakes from nuclear and mtDNA. [•] BI, ML and species tree analysis supported the monophyly of the longtailed species. [•] Longtailed rattlesnakes split form the C. atrox and C. viridis groups 5.1–8.9mya. [•] Longtailed rattlesnakes diverged from each other between 1.5 and 5.6mya. [•] The three longtailed rattlesnakes should be considered separate species. [ABSTRACT FROM AUTHOR]

On account of their unique anatomy, physiology, natural history, ecology, and behavior, rattlesnakes make ideal subjects for a variety of different scientific disciplines. The prairie rattlesnake (Crotalus viridis) in Colorado was selected for investigation of its relationship to colonies of black-tailed prairie dogs (Cynomys ludovicianus) with regard to spatial ecology. A total of 31 snakes were anesthetized and had radiotransmitters surgically implanted. In addition, at the time of their capture, all snakes underwent the following: (1) they had bacterial culture taken from their mouths for potential isolation of pathogenic bacteria; (2) similarly, they had cloacal bacterial cultures taken to assess potentially harmful bacteria passed in the feces; and (3) they had blood samples drawn to investigate the presence of any zoonotic agents in the serum of the snakes. The results of the study and their implications are discussed here. Traditionally, a low incidence of bacterial wound infection has been reported following snakebite. Nevertheless, the oral cavity of snakes has long been known to house a wide variety of bacterial flora. In our study, 10 different bacterial species were isolated from the mouths of the rattlesnakes, 6 of which are capable of being zoonotic pathogens and inducing human disease. More studies are necessary to see why more rattlesnake bites do not become infected despite the presence of such pathogenic bacteria. The results of fecal bacteria isolated revealed 13 bacterial species, 12 of which can cause disease in humans. Of the snakes whose samples were cultured, 26% were positive for the presence of the pathogen Salmonella arizonae, one of the causative agents of reptile-related salmonel-losis in humans. It has long been reported that captive reptiles have a much higher incidence than wild, free-ranging species. This study shows the incidence of Salmonella in a wild, free-ranging population of rattlesnakes. In addition, Stenotrophomonas maltophilia was isolated. This bacterium is associated with wound and soft tissue infections that can lead to sepsis, endocarditis, meningitis, and peritonitis. In addition, this bacterium has been increasingly implicated as an opportunistic pathogen to humans during pregnancies, hospitalizations, malignancies and chemotherapy, chronic respiratory diseases, and presurgical endotracheal intubation. Furthermore, S. maltophilia has an intense resistance to broad-spectrum antibiotics, the results of our study showed the bacterium was resistant to multiple antibiotics. Our results indicate that anyone working with snake feces, dead skin, or their carcasses must follow reasonable hygiene protocols. Rattlesnakes tested for West Nile antibodies had positive results but these were invalidated owing to possible cross-reactivity with other unknown viruses, interference with snake serum proteins, and the fact that the test was not calibrated for rattlesnake serum. Still, the interesting implication remains, should we be regularly testing these animals as sentinels against potentially zoonotic diseases. The results of this study clearly show the value of veterinarians in a multidisciplinary study of this sort and the particular skill set they can offer. Veterinarians must get involved in conservation studies if the biodiversity of the planet is to be preserved. [ABSTRACT FROM AUTHOR]

ABSTRACT The grasslands of southwestern Saskatchewan, Canada are home to several snake species of conservation concern at the northern extreme of their geographic range. To aid conservation assessment and management planning for these snakes, we used radio-telemetry, a geographic information system, and multivariate modeling to identify and compare macrohabitat selection by eastern yellow-bellied racers ( n = 33; Coluber constrictor flaviventris), bullsnakes ( n = 16; Pituophis catenifer sayi), and prairie rattlesnakes ( n = 23; Crotalus viridis). All 3 species shared communal hibernacula in the inactive winter season, but dispersed into different macrohabitats across the landscape during the spring and summer. Their home ranges varied in size by species and were dumbbell-shaped with activity centers near hibernacula and in well-defined summer grounds; activity centers were connected by narrow movement corridors. Racers strongly selected for riparian areas, bullsnakes selected for valley grassland habitats, and rattlesnakes selected for areas associated with prairie dog colonies. Some rattlesnakes traveled great distances (over 11 km) from the dens compared to the other species (bullsnake max. = 4 km; racer max. = 5 km), which may be a result of their selected macrohabitat being more patchily distributed in the landscape. Our results indicate that management plans for these snakes must consider the den area, corridors, and separate summering grounds, as well as differences in home range size and movement patterns for each species. © 2013 The Wildlife Society. [ABSTRACT FROM AUTHOR]

Rattlesnakes use chemical stimuli in ambush site selection and for relocation of envenomated prey through strike-induced chemosensory searching. Shifts in responsiveness to prey chemicals have been documented in many snakes, and often correlate with prey commonly taken as snakes increase in age and size as well as geographical locations of the species. For instance, neonate rattlesnakes that prey primarily on ectotherms responded most strongly to chemical cues of commonly taken lizard prey, whereas adult rattlesnakes that prey primarily on small mammals responded significantly to chemical cues of commonly taken rodents. In the current study, 11 Prairie Rattlesnakes Crotalus viridis viridis which were classified as large neonates based on measures of snout-vent length (SVL) and body mass, yet chronologically were at or near adulthood, were tested for their responsiveness to chemical extracts of natural and non-natural prey items. Although the snakes had eaten only neonate lab mice (Mus musculus), they responded significantly more to chemical cues of natural prey items and particularly to chemical cues of prey normally taken by subadults (Peromyscus mice and Sceloporus lizard). These results suggest that ontogenetic shifts in responsiveness to natural prey chemical cues are innately programmed and are not based on body size or feeding experience in C. v. viridis. This does not imply, however, that growth and experience are without effects, especially with novel prey or rare prey that have experienced recent population expansion [ABSTRACT FROM AUTHOR]

ABSTRACT.-Ten Prairie Rattlesnakes (Crotalus viridis viridis) and 10 Western Diamondback Rattlesnakes (Cro talus atrox) were offered choices between envenomated (E) and nonenvenomated (NE) mice. Both species preferred E over NE mice when envenomation was accomplished by a conspecific but not when envenomation was accomplished by the other species. However, when envenomation was accomplished by Crotalus oreganus oreganus, E mice were preferred by C. v. viridis but not by C. atrox. Accordingly, we conclude that rattlesnakes of closely related species recognize each other's E prey, whereas rattlesnakes of distantly related species do not. [ABSTRACT FROM AUTHOR]

The collection of an Agkistrodon piscivorus (Cottonmouth) on a barge in Winona, MN lends credence to the belief that this species may occasionally be carried up the Mississippi River via commercial boat traffic. Consideration is given to reports of other crotaline snakes not expected to occur in the upper Mississippi River drainage, including Agkistrodon contortrix (Copperhead). Crotalus viridis (Prairie Rattlesnake), and Crotalus atrox (Western Diamondback Rattlesnake). [ABSTRACT FROM AUTHOR]

The broad and imprecise type localities of Crotalinus catenatus Rafinesque, 1818 (now Sistrurus catenatus) and Crotalinus viridis Rafinesque, 1818 (now Crotalus viridis) are re-identified based on a published account of the collector's travels as well as an examination of the relationship and contacts between the collector and the author of the type description. Both type specimens are lost. Unfortunately, enhanced resolution of the type locality of S. catenatus places it within the range of S. c. tergeminus Say, 1823 (a subsequently described subspecies). Thus, the name S. c. catenatus incontrovertibly applies to the form currently recognized as S. c. tergeminus, which in turn renders the latter as a subjective junior synonym of the former, leaving the eastern form with an unfamiliar name (Crotalus messasaugus Kirtland, 1838). To conserve prevailing usage, we are preparing an appeal to the International Commission of Zoological Nomenclature (ICZN) to allow a neotype of Sistrurus catenatus to be designated from the type locality of Crotalus massassaugus. In the interim, existing use of the names should be continued. [ABSTRACT FROM AUTHOR]

Geographic venom samples of Crotalus viridis viridis were obtained from South Dakota, Wyoming, Colorado, Oklahoma, Texas, New Mexico, and Arizona. From these samples, the phospholipases A2 (PLA2s) were purified and their N-terminal sequences, precise masses, and in vitro enzymatic activities were determined. We purified two to four distinct acidic PLA2s from each sample; some of them displayed different inhibition specificities toward mammalian platelets. One of the acidic PLA2s induced edema, but had no anti-platelet activity. There was also a common basic PLA2 myotoxin in all the samples. We have cloned five acidic PLA2s and several hybrid-like nonexpressing PLA2s. Molecular masses and N-terminal sequences of the purified PLA2s were matched with those deduced from the cDNA sequences, and the complete amino acid sequences of five novel acidic PLA2s were thus solved. They share 78% or greater sequence identity, and a cladogram based on the sequences of many venom acidic PLA2s of New World pit vipers revealed at least two subtypes. The results contribute to a better understanding of the ecogenetic adaptation of rattlesnakes and the structure–activity relationships and evolution of the acidic PLA2s in pit viper venom. [Copyright &y& Elsevier]

Presents a study which investigated the diet of the Prairie Rattlesnake Crotalus viridis viridis in a multiple land use area in southeastern Alberta by examining the gut contents of road-killed individuals and the composition of scats recovered from sequestered individuals. Materials and methods used; Results of the study; Discussion.

We quantified sexual dimorphism and allometric changes in the lung morphology of 160 juvenile and adult specimens of prairie rattlesnake, Crotalus viridis viridis, from a single population. In virtually all lung components, those of males are located more posteriorly than are those of females of the same body size. Males display a longer vascular component than females but there is no sexual dimorphism in size of the avascular component. Thus, males generally have longer lungs than do females at all body sizes. With increasing body size, the lung components are found more anteriorly, relative length of the vascular lung decreases, and relative length of the avascular lung increases in both sexes. However, total lung length increases isometrically with body size. These sexual and allometric differences suggest that intraspecific variation should be taken into account when lung size characters are used in snake systematic studies. [ABSTRACT FROM AUTHOR]

Predatory behavior of captive prairie rattlesnakes ( Crotalus viridis) was observed when artificial rock burrows were and were not present in terraria. Euthanized mice were suspended from tongs in front of the snakes, each of which immediately struck and released these prey. The rodent was placed 30-35 cm from the snake. We measured latency to retrieve the envenomated carcass and to swallow the carcass once it was retrieved. Retrieval was faster in snakes living in terraria containing cover than in snakes without cover. Swallowing time did not differ between the conditions. Burrows were used not only as réfugia during inactive periods, but also as centers of activity during predatory episodes. [ABSTRACT FROM AUTHOR]