Crotalus Adamanteus Research Articles

Neurotropic effects of venoms and other factors that promote prey acquisition

Mammals envenomed by either the Eastern diamondback rattlesnake (Crotalus adamanteus) or the cottonmouth (Agkistrodon piscivorus piscivorus) exhibit an immediate but transitory pupillar contraction, a parasympathomimetic effect mediated through the ciliary ganglion that can be prevented by a retrobulbar injection of anesthetic. The venom of the cottonmouth injected into the lymph spaces of the frog (Rana pipiens) produces an immediate and total collapse of the lung sacs. Applied locally to the surface, it produces a constriction that eventually collapses the entire sac. Tests of venoms and toxins from both anterior and posterior parts of the venom apparatus indicate that the lung-collapsing moiety originates in the accessory, not the main portion of the venom gland. This is the first example of a functional specialization within the whole structure. It seems that this factor is elaborated primarily in snakes that prey upon frogs, although insufficient data are available from this study to confirm this. In both reptile species, the predatory strike is accompanied by an immediate effect, perhaps mediated by the parasympathetic nervous system, designed to incapacitate the prey and facilitate capture. These effects cannot now be attributed to neurotoxins because the effect of the former is transitory (and not lethal) and neither has been purified sufficiently to determine potency or structure. Both take part in securing, but not killing, the prey, and both directly oppose the sympathetic nervous system "fright-fight/flight" response. Evidence is presented to support the possibility that known epigenetic mechanisms are capable of effecting heritable changes in gene expression that could allow for the development of factors that facilitate prey acquisition and promote rapid adaptation to environmental change.

Proximate developmental mediators of sexual dimorphism in size: case studies from squamate reptiles

Sexual dimorphism in size (sexual size dimorphism; SSD) is nearly ubiquitous, but the relative importance of genetic versus environmental control of SSD is not known for most species. We investigated proximate determinants of SSD in several species of squamate reptiles, including three species of Sceloporus lizards and the diamond-backed rattlesnake (Crotalus atrox). In natural populations of these species, SSD is caused by sexual differences in age-specific growth. Males and females, however, may often share similar potentials for growth: growth is strongly responsive to the availability of food, and sexual differences in growth can be greatly suppressed or completely absent under common environmental conditions in the laboratory. Sexually divergent growth is expressed in natural environments because of inherent ecological differences between males and females and because of potential epigenetic effects of sex-specific growth regulators. In field-active Sceloporus, sexual differences in growth rate are associated with sexual divergence in plasma testosterone. Experiments confirm that testosterone inhibits growth in species in which females are larger (for example, S. undulatus and S. virgatus) and stimulates growth in those in which males are larger (for example, S. jarrovii). Interestingly, however, sexual divergence in plasma testosterone is not accompanied by divergence in growth in S. jarrovii or in male-larger C. atrox in the laboratory. Furthermore, experimental effects of castration and testosterone replacement on growth are not evident in captive S. jarrovii, possibly because growth effects of testosterone are superseded by an abundant, high-quality diet. In female-larger S. undulatus, growth may be traded-off against testosterone-induced reproductive costs of activity. In male-larger species, costs of reproduction in terms of growth are suggested by supplemental feeding of reproductive female C. atrox in their natural environment and by experimental manipulation of reproductive cost in female S. jarrovii. Growth costs of reproduction, however, do not contribute substantially to the development of SSD in male-larger S. jarrovii. We conclude that the energetic costs of testosterone-induced, male reproductive behavior may contribute substantially to the development of SSD in some female-larger species. However, despite strong evidence that reproductive investment exacts a substantial cost in growth, we do not support the reproductive cost hypothesis as a general explanation of SSD in male-larger species.

Habitat Use of Sympatric Rattlesnake Species Within the Gulf Coastal Plain

Abstract: The eastern diamondback rattlesnake (Crotalus adamanteus) and timber rattlesnake (Crotalus horridus) are sympatric throughout most of southern Georgia, USA. We used rattlesnake sightings to quantify and compare habitat use by these 2 species in the Gulf Coastal Plain. At the largest scale examined, univariate statistics and logistic regression models indicated that eastern diamondback rattlesnakes were associated with roads but not with any of the specific habitat types we examined. In contrast, timber rattlesnakes were closely associated with hardwood habitat and riverine systems but not with roads and edges. To effectively conserve and manage both species in the Southeast, a habitat matrix of large intact patches of both hardwood and pine (Pinus spp.) forest may be necessary.

Hyrtiazepine, an Azepino-indole-Type Alkaloid from the Red Sea Marine Sponge Hyrtios erectus

Biological and chemical investigations of the methanolic crude extract of the Red Sea marine sponge Hyrtios erectus led to the isolation of a novel azepino-indole-type alkaloid named hyrtiazepine (2) and 5-hydroxy-1H-indole-3-carboxylic acid methyl ester (3), together with the known metabolites hyrtiosulawesine (1), 5-hydroxyindole-3-carbaldehyde (4), hyrtiosin A (5), and hyrtiosin B (6). Their structures were elucidated on the basis of mass spectrometry and detailed 2D NMR spectroscopic data. Hyrtiosulawesine (1) displayed a significant antiphospholipase A2 activity with an IC50 value of 14 microM in a fluorometric assay using Crotalus adamanteus venom phospholipase A2.

Antibacterial activity of snake, scorpion and bee venoms: a comparison with purified venom phospholipase A2enzymes

Venoms of snakes, scorpions, bees and purified venom phospholipase A(2) (PLA(2)) enzymes were examined to evaluate the antibacterial activity of purified venom enzymes as compared with that of the crude venoms. Thirty-four crude venoms, nine purified PLA(2)s and two L-amino acid oxidases (LAAO) were studied for antibacterial activity by disc-diffusion assay (100 microg ml(-1)). Several snake venoms (Daboia russelli russelli, Crotalus adamanteus, Naja sumatrana, Pseudechis guttata, Agkistrodon halys, Acanthophis praelongus and Daboia russelli siamensis) showed activity against two to four different pathogenic bacteria. Daboia russelli russelli and Pseudechis australis venoms exhibited the most potent activity against Staphylococcus aureus, while the rest showed only a moderate activity against one or more bacteria. The order of susceptibility of the bacteria against viperidae venoms was -S. aureus > Proteus mirabilis > Proteus vulgaris > Enterobacter aerogenes > Pseudomonas aeruginosa and Escherichia coli. The minimum inhibitory concentrations (MIC) against S. aureus was studied by dilution method (160-1.25 microg ml(-1)). A stronger effect was noted with the viperidae venoms (20 microg ml(-11)) as compared with elapidae venoms (40 microg ml(-1)). The MIC were comparable with those of the standard drugs (chloramphenicol, streptomycin and penicillin). The present findings indicate that viperidae (D. russelli russelli) and elapidae (P. australis) venoms have significant antibacterial effects against gram (+) and gram (-) bacteria, which may be the result of the primary antibacterial components of laao, and in particular, the PLA(2) enzymes. The results would be useful for further purification and characterization of antibacterial agents from snake venoms. The activity of LAAO and PLA(2) enzymes may be associated with the antibacterial activity of snake venoms.

Habitat specificity and home‐range size as attributes of species vulnerability to extinction: a case study using sympatric rattlesnakes

AbstractLarge home‐range size and habitat specificity are two commonly cited ecological attributes that are believed to contribute to species vulnerability. The eastern diamondback rattlesnake Crotalus adamanteus is a declining species that occurs sympatrically with the more abundant canebrake rattlesnake Crotalus horridus in a portion of the south‐eastern Coastal Plain, USA. In this study, we use the ecological similarities of the two species as experimental controls to test the role of home‐range size and habitat specificity in the imperilment of the eastern diamondback rattlesnake. We used analysis of variance to investigate differences in home‐range size between the two species, and home‐range selection was modeled as habitat use versus availability with a case control sampling design using logistic regression. We failed to detect differences in home‐range size between the two species; therefore, we could not identify home‐range size as an attribute contributing to the imperilment of eastern diamondback rattlesnakes. Eastern diamondback rattlesnakes selected pine savannas to a degree that suggests that the species is a habitat specialist. Of the two factors examined, habitat specificity to the imperiled longleaf pine ecosystem may be a significant contributor to the decline of the eastern diamondback rattlesnake.

In vitro antimicrobial activity of natural toxins and animal venoms tested against Burkholderia pseudomallei

BackgroundBurkholderia pseudomallei are the causative agent of melioidosis. Increasing resistance of the disease to antibiotics is a severe problem in treatment regime and has led to intensification of the search for new drugs. Antimicrobial peptides are the most ubiquitous in nature as part of the innate immune system and host defense mechanism.MethodsHere, we investigated a group of venoms (snakes, scorpions and honey bee venoms) for antimicrobial properties against two strains of Gram-negative bacteria Burkholderia pseudomallei by using disc-diffusion assay for in vitro susceptibility testing. The antibacterial activities of the venoms were compared with that of the isolated L-amino acid oxidase (LAAO) and phospholipase A2 (PLA2s) enzymes. MICs were determined using broth dilution method. Bacterial growth was assessed by measurement of optical density at the lowest dilutions (MIC 0.25 mg/ml). The cell viability was measured using tetrazolium salts (XTT) based cytotoxic assay.ResultsThe studied venoms showed high antimicrobial activity. The venoms of C. adamanteus, Daboia russelli russelli, A. halys, P. australis, B. candidus and P. guttata were equally as effective as Chloramphenicol and Ceftazidime (30 μg/disc). Among those tested, phospholipase A2 enzymes (crotoxin B and daboiatoxin) showed the most potent antibacterial activity against Gram-negative (TES) bacteria. Naturally occurring venom peptides and phospholipase A2 proved to possess highly potent antimicrobial activity against Burkholderia pseudomallei. The XTT-assay results showed that the cell survival decreased with increasing concentrations (0.05–10 mg/mL) of Crotalus adamanteus venom, with no effect on the cell viability evident at 0.5 mg/mL.ConclusionThis antibacterial profile of snake venoms reported herein will be useful in the search for potential antibacterial agents against drug resistant microorganisms like B. pseudomallei.

Heating and cooling rates of eastern diamondback rattlesnakes, Crotalus adamanteus

1. Establishing if and how organisms modulate temperature changes is an important component of understanding their thermal biology. 2. We used temperature-sensitive radio-transmitters to monitor heating and cooling rates between 5 and 35 °C of four Crotalus adamanteus in the laboratory. 3. We found no difference between heating and cooling rates in C. adamanteus. Additionally, rates of temperature change mirrored those of a biophysical model, further suggesting a lack of physiological thermoregulation. 4. Our findings contrast previously published studies that demonstrate active temperature control of similarly sized reptiles and demonstrate a need for more investigations of physiological thermoregulation in reptiles.

Removal or Maintenance of Inositol-linked Acyl Chain in Glycosylphosphatidylinositol Is Critical in Trypanosome Life Cycle

The protozoan parasite Trypanosoma brucei is coated by glycosylphosphatidylinositol (GPI)-anchored proteins. During GPI biosynthesis, inositol in phosphatidylinositol becomes acylated. Inositol is deacylated prior to attachment to variant surface glycoproteins in the bloodstream form, whereas it remains acylated in procyclins in the procyclic form. We have cloned a T. brucei GPI inositol deacylase (GPIdeAc2). In accordance with the acylation/deacylation profile, the level of GPIdeAc2 mRNA was 6-fold higher in the bloodstream form than in the procyclic form. Knockdown of GPIdeAc2 in the bloodstream form caused accumulation of an inositol-acylated GPI, a decreased VSG expression on the cell surface and slower growth, indicating that inositol-deacylation is essential for the growth of the bloodstream form. Overexpression of GPIdeAc2 in the procyclic form caused an accumulation of GPI biosynthetic intermediates lacking inositol-linked acyl chain and decreased cell surface procyclins because of release into the culture medium, indicating that overexpression of GPIdeAc2 is deleterious to the surface coat of the procyclic form. Therefore, the GPI inositol deacylase activity must be tightly regulated in trypanosome life cycle.

Isolation of a new l-amino acid oxidase from Crotalus durissus cascavella venom

A novel l-amino acid oxidase (LAO) (Casca LAO) from Crotalus durissus cascavella venom was purified to a high degree of molecular homogeneity using a combination of molecular exclusion and ion-exchange chromatography system. The purified monomer of LAO presented a molecular mass of 68 kDa and pI estimated in 5.43, which were determined by two-dimensional electrophoresis. The 71st N-terminal amino acid sequence of the LAO from Crotalus durissus cascavella presented a high amino acid sequence similarities with other LAOs from Colloselasma rhosostoma, Crotalus adamanteus, Agkistrodon h. blomhoffi, Agkistrodon h. halys and Trimeresurus stejnegeri. LAO displayed a Michaelis-Menten behavior with a kilometer of 46.7 μM and an optimum pH for enzymatic activity of 6.5. Casca LAO induced a dose-dependent platelet aggregation, which was abolished by catalase and inhibited by indomethacin and aspirin. These results suggest that the production of H 2O 2 is involved in subsequent activation of inflammatory enzymes, such as thromboxane. Casca LAO also inhibited the bacterial growth of Gram-negative ( Xanthomonas axonopodis pv passiflorae) and Gram-positive ( S. mutans) strains. Electron microscopy assessments of both bacterial strains suggest that the hydrogen peroxide produced by LAO induce bacterial membrane rupture and consequently loss of cytoplasmatic content. This LAO exhibited a high antileishmanic activity against the promastigote of Leishmania amazonensis in vitro, its activity was dependent on the production of hydrogen peroxide, and the 50% inhibitory concentration was estimated in 2.39 μg/ml.

Modelling of l-DOPA enzymatic oxidation catalyzed by l-amino acid oxidases from Crotalus adamanteus and Rhodococcus opacus

l-amino acid oxidases ( l-AAO) are well known for their broad substrate specificity. l-amino acid oxidases from Crotalus adamanteus and Rhodococcus opacus were applied for biotransformation of 3,4-dihydroxyphenyl- l-alanine (L-DOPA) as a substrate to its corresponding α-keto acid. In this reaction, hydrogen peroxide formed as a by-product causes chemical decarboxylation of α-keto acids and acts as competitive product inhibitor. Beef liver catalase was used to decompose it. It was shown that both enzymes were able to oxidize l-DOPA to corresponding products. l-AAO from R. opacus was more specific (lower K m l − DOPA value) and more active towards l-DOPA substrate than l-AAO from C. adamanteus. Its catalytic constant, k 3, estimated by Levenspiel's method, was found to be 10-fold higher than the one for l-AAO from C. adamanteus. l-AAO from R. opacus exhibits slightly l-DOPA inhibition, which is not the case for l-AAO from C. adamanteus. The biotransformations of l-DOPA were carried out in batch enzyme membrane reactor (EMR), as well as in the repetitive batch EMR. The reactor and kinetics were modelled. Parameters were estimated by differential and integral method and presented in this article.

Rationalisation of First-Aid Measures for Elapid Snakebite

Summary The plasma of monkeys envenomated with tiger snake (Notechis scutatus) venom was monitored by radioimmunoassay for both crude venom and a neurotoxin. When the injected limb was immobilised and a pressure of 55 mm Hg applied to the injection site, only very low levels of circulating venom or neurotoxin were detectable. In practical terms, venom movement can be effectively delayed for long periods by the application of a firm crepe bandage to the length of the bitten limb combined with immobilisation by a splint. Pressure alone or immobilisation alone did not delay venom movement.

Serum sickness in a dog associated with antivenin therapy for snake bite caused by Crotalus adamanteus

AbstractObjective: This case report describes antivenin‐associated acute and delayed hypersensitivity reactions in a dog envenomated by an Eastern diamondback rattlesnake (Crotalus adamanteus), specifically reviewing the syndrome of antivenin‐associated serum sickness. Clinician awareness of this syndrome is important in order to allow for its recognition and appropriate treatment.Case summary: A Boxer dog was envenomated by an Eastern diamondback rattlesnake. Shock, echinocytosis, and coagulopathy were manifested, and the dog was given antivenin crotalidae polyvalent therapy and supportive care. The early onset of an anaphylactoid reaction was attributed to antivenin therapy and was managed with diphenhydramine and subcutaneous epinephrine therapy. Fever, chemosis, and limb edema occurred during the 3rd through 6th hospital days following antivenin therapy and were consistent with serum sickness syndrome as described in humans. Further immunoassay support reflecting complement activation and response to treatment were characteristic.New information provided: To our knowledge, this describes the first reported case of antivenin‐associated serum sickness in a dog.

Eastern Diamondback Rattlesnake (Crotalus adamanteus) Envenomation of Dogs: 31 Cases (1982–2002)

The medical records of 31 dogs treated for envenomation by the Eastern Diamondback Rattlesnake (Crotalus adamanteus) were reviewed. Twenty-four of 25 dogs that survived were hospitalized for an average of 4.3 days. The most common presenting signs were tachycardia, swelling/edema, depressed mentation, tachypnea, and bleeding puncture wounds. Thirteen (42%) of the 31 dogs were presented with or developed cardiac arrhythmias, predominantly ventricular premature contractions. Hematological disorders, including defibrination, elevated fibrin split products, hemolytic anemia, thrombocytopenia, and prolonged clotting times, were recorded in 81% of the dogs. Polyvalent crotalid antivenin was administered (mean of 4.0 vials per dog) to 88% of the surviving dogs and 50% of the nonsurviving dogs.

Enzymatic activity of L-amino acid oxidase from snake venom Crotalus adamanteus in supercritical CO2

L-amino acid oxidase (L-AAO) from snake venom Crotalus adamanteus was successfully tested as a catalyst in supercritical CO2 (SC-CO2). The enzyme activity was measured before and after exposure to supercritical conditions (40°C, 110 bar). It was found that L-AAO activity slightly increased after SC-CO2 exposure by up to 15%. L-AAO was more stable in supercritical CO2 than in phosphate buffer under atmospheric pressure, as well as in the enzyme membrane reactor (EMR) experiment. 3,4-Dihydroxyphenyl-L-alanine (L-DOPA) oxidation was performed in a batch reactor made of stainless steel that could withstand the pressures of SC-CO2, in which L-amino acid oxidase from C. adamanteus was able to catalyze the reaction of oxidative deamination of L-DOPA in SC-CO2. For the comparison L-DOPA oxidation was performed in the EMR at 40°C and pressure of 2.5 bar. Productivity expressed as mmol-s of converted L-DOPA after 3 h per change of enzyme activity after 3 h was the highest in SC-CO2 (1.474 mmol U−1), where catalase was present, and the lowest in the EMR (0.457 mmol U−1).

Effects of Body Mass and Temperature on Standard Metabolic Rate in the Eastern Diamondback Rattlesnake (Crotalus adamanteus)

Abstract Determining the consequences of body size and body temperature (Tb) variation is critical to understanding many aspects of snake ecology, because size and temperature play such important roles in the biology of ectotherms. Here, we investigate the effects of body size and temperature variation on the energetics of the largest species of rattlesnake, the Eastern Diamondback Rattlesnake (Crotalus adamanteus). Specifically, we measured oxygen consumption to estimate the standard metabolic rate (SMR) of five C. adamanteus (mass range 800–4980 g) at 5-degree increments from 5–35 C. A multiple regression model indicated that SMR increased with body size and temperature. Q10s were generally high (range 1.82–4.20) compared to other squamates but were similar to the high values calculated for other large rattlesnakes. An energy balance model for C. adamanteus predicted that as Tb increases, so must prey consumption to meet annual SMR energy demands. Thus, Tb variation likely affects patterns of energy acq...

Electrochemical measurement of the interaction of Crotalus adamanteus venom with DMPC vesicles.

Electrochemical measurements of large unilamellar vesicles encapsulating potassium ferrocyanide showed that the lysis of vesicles by Crotalus adamanteus venom could be measured showing both time and concentration dependent responses.

An Illustrated Guide to Trunk Vertebrae of Cottonmouth (Agkistrodon piscivorus) and Diamondback Rattlesnake (Crotalus adamanteus) in Florida

The cottonmouth, Or water moccasin (Agkistrodon piscivorus), and the diamondback rattlesnake (Crotalus adamanteus) are distributed throughout Florida and their skeletal remains, usually vertebrae, are often present in zooarchaeological assemblages. Although the two viperid snakes exhibit different habitat preferences—one a semi-aquatic snake, the other terrestrial-their vertebrae are very similar. This illustrated guide helps to distinguish between the vertebrae of the two taxa. A strategy of limiting species identifications to the middle trunk series of mature adults and employing multiple characteristics is recommended to overcome the intracolumnar and individual variability that occurs in the vertebrae of these snakes.

Preclinical assessment of the ability of polyvalent (Crotalinae) and anticoral (Elapidae) antivenoms produced in Costa Rica to neutralize the venoms of North American snakes

Polyvalent (Crotalinae) and anticoral (Elapidae) antivenoms produced by Instituto Clodomiro Picado, Costa Rica, were assessed for their ability to neutralize various toxic activities of the venoms of North American snakes of the genera Crotalus, Agkistrodon and Micrurus, in assays involving preincubation of venom and antivenom. When the intraperitoneal route of injection was utilized, polyvalent (Crotalinae) antivenom was effective in the neutralization of the venoms of Crotalus atrox, Crotalus adamanteus, Crotalus viridis viridis, Crotalus horridus atricaudatus, Agkistrodon contortrix contortrix and Agkistrodon piscivorus piscivorus, whereas the venom of Crotalus scutulatus was not neutralized. When the intravenous route was used, results differed depending on the ‘challenge dose’ of venom employed. Polyvalent antivenom neutralized all venoms when mice were challenged with 2 LD 50s of venom. When 5 LD 50s were used, antivenom neutralized the venoms of C. atrox, C. adamanteus, C. v. viridis and C. h. atricaudatus, being ineffective in the neutralization of C. scutulatus, A. c. contortrix and A. p. piscivorus. Polyvalent antivenom was effective in the neutralization of hemorrhagic and myotoxic activities of all venoms studied. It also neutralized coagulant activity of C. adamanteus venom, whereas most of the venoms were devoid of clotting activity on plasma in vitro. Moreover, it neutralized defibrinating activity of the only three venoms that induced this effect (i.e. C. adamanteus, A. c. contortrix and A. p. piscivorus). Anticoral (Elapidae) antivenom neutralized lethality induced by the venom of Micrurus fulvius, using either the intravenous or the intraperitoneal routes of injection. Moreover, it neutralized myotoxic effect of this venom as well. It is concluded that polyvalent antivenom neutralizes lethality and other activities of most of the crotaline venoms tested. However, since it is ineffective in neutralizing the lethal effect of C. scutulatus venom, it is suggested that a venom containing presynaptically-active neurotoxic phospholipases A 2 related to ‘mojave toxin’ needs to be introduced in the immunizing mixture in order to increase the neutralizing scope of this product in North America. Anticoral antivenom is highly effective in the neutralization of the venom of M. fulvius.

Kallikrein-like proteinase from bushmaster snake venom

A kallikrein-like proteinase of Lachesis muta muta (bushmaster) venom, designated LV-Ka, was purified by gel filtration and anion exchange chromatographies. Physicochemical studies indicated that the purified enzyme is a 33 kDa monomeric glycoprotein, the Mr of which fell to 28 kDa after deglycosylation with PNGase F. Approximately 77% of the protein sequence was determined by sequencing the various fragments derived from digestions with endoproteases. The partial sequence obtained suggests that LV-Ka is of a similar size to other serine proteinases (i.e., approximately 234 amino acid residues). Sequence studies on the NH 2-terminal region of the protein indicate that LV-Ka shares a high degree of sequence homology with the kallikrein-like enzymes EI and EII from Crotalus atrox, with crotalase from Crotalus adamanteus and significant homology with other serine proteinases from snake venoms and vertebrate serum enzymes. LV-Ka showed kallikrein-like activity, releasing bradikinin from kininogen as evidenced by guinea pig bioassay. In addition, intravenous injection of the proteinase (0.8 μg/g) was shown to lower blood pressure in experimental rats. In vitro, the isolated proteinase was shown to have neither fibrin(ogeno)lytic activity nor coagulant effect. LV-Ka was active upon the kallikrein substrates S-2266 and S-2302 (specific activity=13.0 and 31.5 U/mg, respectively; crude venom=0.25 and 6.0 U/mg) but had no proteolytic effect on dimethylcasein and insulin B chain. Its enzymatic activity was inhibited by NPGB and PMSF, indicating that the enzyme is a serine proteinase. Interestingly, one of the other reactions catalyzed by plasma kallikrein, the activation of plasminogen was one of the activities exhibited by LV-Ka.