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PubMed2002

Identification of beta-type phospholipase A(2) inhibitor in a nonvenomous snake, Elaphe quadrivirgata.

A novel serum protein inhibiting specifically the enzymatic activity of the basic phospholipase A(2) (PLA(2)) from the venom of the Chinese mamushi snake (Agkistrodon blomhoffii siniticus) was purified from a nonvenomous Colubridae snake, Elaphe quadrivirgata. The purified inhibitor was a 150-kDa glycoprotein having a trimeric structure, composed of two homologous 50-kDa subunits. Their amino acid sequences, containing leucine-rich repeats, were typical of the beta-type PLA(2) inhibitor (PLIbeta), previously identified from the serum of A. blomhoffii siniticus. The inhibitor inhibited exclusively group II basic PLA(2)s and did not inhibit other kinds of PLA(2)s. This is the first paper reporting the existence of PLIbeta in a nonvenomous snake. The existence of PLIbeta in the nonvenomous snake reflects that PLIbetas are widely distributed over the snake species and participate commonly in regulating the physiological activities of the unidentified target PLA(2)s.

Amino Acid Sequence↗
PubMed2001

Cloning of cDNAs encoding C-type lectins from Elapidae snakes Bungarus fasciatus and Bungarus multicinctus.

A number of C-type lectins with various biological activities have been purified and characterized from Viperidae snake venoms. In contrast, only a few reports could be found in literature concerning the C-type lectins in Elapidae snake venoms. Based on the published cDNA sequences of C-type lectins from Viperidae snake venoms, oligonucleotide primers were designed and used to screen the cDNA libraries made from the venom glands of Bungarus fasciatus and Bungarus multicinctus. This allowed the cloning of three full length cDNAs encoding C-type lectins. The encoded proteins, named BFL-1, BFL-2 and BML, exhibit high degrees of sequence identities with Viperidae snake venom saccharide-binding lectins (around 60% with Trimeresurus stejnegeri venom lectin, Crotalus atrox venom lectin and Agkistrodon piscivorus venom lectin). They show much less identities with other venom C-type lectin-like proteins (around 30% with the platelet glycoprotein Ib-binding protein from Agkistrodon blomhoffi venom and the factor IX/X-binding protein from Trimeresurus flavoviridis venom). The cDNAs revealed that the precursors contain potential signal peptides characterized by a hydrophobic core. To our knowledge, these are the first cDNA cloning of group VII C-type lectins (Drickamer K. 1993. Prog. Nucleic Acid Res. Mol. Biol. 45, 207-232) from Elapidae snake venom glands.

Animals↗
PubMed1997

Prothrombin and factor X activator activities in the venoms of Viperidae snakes.

A Ca(2+)-dependent prothrombin activator, carinactivase-1 (CA-1), was previously found in the venom of Echis carinatus leucogaster. In the present study, the activities of CA-1-like enzymes were screened in the venoms of various Viperidae snakes. The addition of 1 mM Ca2+ ions to the venoms of only Echis snakes in Viperidae produced considerably high prothrombin activator activity, indicating that only the Echis snake venoms contain not only the Ca(2+)-independent prothrombin activator, ecarin, but also Ca(2+)-dependent activator(s). CA-1-like activators and ecarin in the venom of each Echis snake were efficiently separated by Blue Sepharose column chromatography. The venoms of the various Viperidae snakes were also examined for factor X activator activity. The venoms of genera Daboia, Vipera, Cerastes, Echis, Calloselasma and Bothrops contained factor X activator activity in the presence of Ca2+ ions. Cerastes cerastes and Calloselasma rhodostoma venoms also had Ca(2+)-independent factor X activator activity.

Animals↗
PubMed1998

The Bov-B lines found in Vipera ammodytes toxic PLA2 genes are widespread in snake genomes.

In the fourth intron of two toxic Vipera ammodytes PLA2 genes a Ruminantia specific 5'-truncated Bov-B LINE element was identified. Southern blot analysis of Bov-B LINE distribution in vertebrates shows that, apart from the Ruminantia, it is limited to Viperidae snakes (V. ammodytes, Vipera palaestinae, Echis coloratus, Bothrops alternatus, Trimeresurus flavoviridis and Trimeresurus gramineus). The copy number of the 3' end of Bov-B LINE in the V. ammodytes genome is between 62,000 and 75,000. At orthologous positions in other snake PLA2 genes the Bov-B LINE element is absent, indicating that its retrotransposition in the V. ammodytes PLA2 gene locus has occurred quite recently, about 5 Myr ago. The amplification of Bov-B LINEs in snakes may have occurred before the divergence of the Viperinae and Crotalinae subfamilies. Due to its wide distribution in Viperidae snakes it should be a valuable phylogenetic marker. The neighbour-joining phylogenetic tree shows two clusters of truncated Bov-B LINE, a Bovidae and a snake cluster, indicating an early horizontal transfer of this transposable element.

Animals↗
PubMed2001

A dual-snake model of high penetrability for ultrasound image boundary extraction.

Most deformable models require the initial contour to be placed close to the boundary of the object of interest for boundary extraction of ultrasound (US) images, which is impractical in many clinical applications. To allow a distant initial contour, a new dual-snake model promising high penetrability through the interference of the noises is proposed in this paper. The proposed dual-snake model features a new far-reaching external force, called the discrete gradient flow, a connected component-weighted image force, and an effective stability evaluation of two underlying snakes. The experimental results show that, with a distant initial contour, the mean distance from the derived boundary to the desired boundary is less than 1.4 pixels, and most snake elements are within 2.7 pixels of the desired boundaries for the synthetic images with CNR > or =1. For the clinical US images, the mean distance is less than 1.9 pixels, and most snake elements are within 3 pixels of the desired boundaries.

Algorithms↗
PubMed2000

An early vision-based snake model for ultrasound image segmentation.

Due to the speckles and the ill-defined edges of the object of interest, the classic image-segmentation techniques are usually ineffective in segmenting ultrasound (US) images. In this paper, we present a new algorithm for segmenting general US images that is composed of two major techniques; namely, the early-vision model and the discrete-snake model. By simulating human early vision, the early-vision model can capture both grey-scale and textural edges while the speckle noise is suppressed. By performing deformation only on the peaks of the distance map, the discrete-snake model promises better noise immunity and more accurate convergence. Moreover, the constraint for most conventional snake models that the initial contour needs to be located very close to the actual boundary has been relaxed substantially. The performance of the proposed snake model has been shown to be comparable to manual delineation and superior to that of the gradient vector flow (GVF) snake model.

Algorithms↗
PubMed1998

Bothrops sp. snake venoms: comparison of some biochemical and physicochemical properties and interference in platelet functions.

Procoagulant, proteolytic, phospholipase and platelet pro-aggregating and inhibiting activities were screened for pooled venoms of seven Bothrops species as well as Crotalus durissus terrificus and Lachesis muta snakes typical of the Brazilian territory. As reported by other authors, we also found that examination of the electrophoretic and gel filtration patterns of Bothrops snakes venoms could not be used for identification of the species of a given venom because of the lack of marked interspecific differences within the same genus. Our data indicated that B. cotiara, B. alternatus and B. atrox possess no platelet pro-aggregating activity, low inhibitory effect on platelet aggregation and very low or intermediate levels for the other activities. B. moojeni, B. neuwiedi and B. jararacussu whose venoms possess high procoagulant, platelet pro-aggregating and phospholipase activities are low in both proteolytic and platelet inhibitory activities. B. jararaca venom showed the highest inhibitory effect on platelet aggregation and very low platelet pro-aggregating activity. Compared with the Bothrops venoms studied, L. muta venom showed that highest proteolytic activity while C. d. terrificus venom presented remarkable high platelet pro-aggregating and phospholipase activities. In all venoms, proteolytic activity could be completely inhibited by EDTA (2 mM) alone. In contrast, the presence of phenylmethylsulfonyl fluoride (5 mM) inhibited partially the caseinolytic activity of all venoms, except that L. muta venom, which was almost completely blocked by this reagent. Altogether, these data confirm the presence of high levels of metalloproteinases in the venoms of Crotalinae snakes. Most of these enzymes are dependent of the availability of Ca2+, being much less the same concerning the presence of serine residues in their active sites. The data indicated that the presence and levels of procoagulant, azocaseinolytic and phospholipase A2 activities alone could not differentiated the species of the Bothrops venoms studied, particularly in the cases of B. jararaca, B. moojeni and B. atrox. However, the platelet inhibiting property of low doses of B. jararaca venom can be useful to differentiate it from B. moojeni venom. In the same way, the platelet pro-aggregating activity of high doses of B. jararaca venom may be used to distinguish it from B. atrox crude venom, otherwise very similar but incapable to activate platelets. In conclusion, our comparative screening of biological properties has indicated that platelet studies may serve as a tool to distinguish among venoms that otherwise behave biochemically in a very similar way. Although promising, the general applicability of platelet activation studies by snake venoms for classification or taxanomical purposes has yet to be extended to other family of snakes to be proven useful.

Animals↗
PubMed2003

Non-invasive measure of body composition of snakes using dual-energy X-ray absorptiometry.

Non-invasive techniques to measure body composition are critical for longitudinal studies of energetics and life histories and for investigating the link between body condition and physiology. Previous attempts to determine, non-invasively, the body composition of snakes have proven problematic. Therefore, we explored whether dual-energy X-ray absorptiometry (DXA) could be used to determine the body composition of snakes. We analyzed 20 adult diamondback water snakes (Nerodia rhombifer) with a DXA instrument and subsequently quantified their body composition by gravimetric and chemical extraction methods. Body composition components scaled with body mass with mass exponents between 0.88 and 1.53. DXA values for lean tissue mass, fat mass and total-body bone mineral mass were significantly correlated with observed masses of lean tissue, fat and ash from chemical analysis. Using regression models incorporating DXA values we predicted the fat-free tissue mass, lean tissue mass, fat mass, ash mass and total body water content for this sample of water snakes. A cross-validation procedure demonstrated that these models estimated fat-free tissue mass, lean tissue mass, fat mass, ash mass and total-body water content with respective errors of 2.2%, 2.3%, 16.0%, 6.6% and 3.5%. Compared to other non-invasive techniques, include body condition indices, total body electrical conductivity (TOBEC) and cyclopropane absorption, DXA can more easily and accurately be used to determine the body composition of snakes.

Absorptiometry, Photon↗
PubMed2002

Computerized detection of pulmonary nodules in chest radiographs based on morphological features and wavelet snake model.

We have developed a new computer-aided diagnosis scheme for automated detection of lung nodules in digital chest radiographs based on a combination of morphological features and the wavelet snake. In our scheme, two processes were applied in parallel to reduce the false-positive detections after initial nodule candidates were selected. One process consisted of adaptive filtering for enhancement of nodules and suppression of normal lung structures, followed by extraction of conventional morphological features. The other process consisted of a novel approach for elimination of false positives called the edge-guided wavelet snake model. In the latter process, multiscale edges of the candidate nodules were extracted to yield parts of the nodule boundaries. A wavelet snake was then used for fitting of these multiscale edges for approximation of the true boundaries of nodules. A boundary feature called the weighted overlap between the snake and the multiscale edges was calculated and used for elimination of false positives. Finally, the weighted overlap and the morphological features were combined by use of an artificial neural network for efficient reduction of false positives. Our scheme was applied to a publicly available database of digital chest images for pulmonary nodules. Receiver operating characteristic analysis was employed for evaluation of the performance of each process in the scheme. The combined features yielded a large reduction of false positives, and thus achieved a high performance in discriminating between true and false positives. These results show that our new method, in particular the false-positive reduction method based on the wavelet snake, is effective in improving the performance of a computerized scheme for detection of pulmonary nodules in chest radiographs.

Adult↗
PubMed2004

A resistant predator and its toxic prey: persistence of newt toxin leads to poisonous (not venomous) snakes.

The common garter snake (Thamnophis sirtalis) preys upon the rough-skinned newt (Taricha granulosa), which contains the neurotoxin tetrodotoxin (TTX) in the skin. TTX is toxic, large quantities are present in a newt, and highly resistant snakes have the ability to ingest multiple newts; subsequently snakes harbor significant amounts of active toxin in their own tissues after consuming a newt. Snakes harbor TTX in the liver for 1 mo or more after consuming just one newt, and at least 7 wk after consuming a diet of newts. Three weeks after eating one newt, snakes contained an average of 42 microg of TTX in the liver. This amount could severely incapacitate or kill avian predators, and mammalian predators may be negatively affected as well.

Animals↗
PubMed2001

Molecular cloning and expression of a functional snake venom vascular endothelium growth factor (VEGF) from the Bothrops insularis pit viper. A new member of the VEGF family of proteins.

During the generation of abundant expressed sequence tags from the Viperidae snake Bothrops insularis venom glands, we identified for the first time a cDNA coding for a putative vascular endothelial growth factor-like (VEGF-like) protein. The deduced primary sequence, after complete sequencing of the longest snake venom VEGF (svVEGF) cDNA, displayed similarity with vertebrate VEGFs and with the hypotensive factor from Vipera aspis venom. Its cDNA was subcloned, expressed in Escherichia coli with a His(6) tag as an insoluble monomer, and purified by Ni(2+)-affinity chromatography after 8 m urea extraction. Antiserum against svVEGF was generated and tested in Western blot against proteins from snake venoms and cellular extracts. The mature svVEGF appears to be ubiquitously distributed throughout snake venoms and was also confirmed by Northern blot studies of other related Viperidae species and by cDNA cloning of svVEGF from Bothrops jararaca pit viper. The produced recombinant protein dimerizes after refolding processes and was biologically characterized, showing ability to increase vascular permeability. These results established that svVEGF is a novel and important active toxin during the early stages of bothropic snake bite envenoming and represents a new member of the VEGF family of proteins.

Amino Acid Sequence↗
PubMedDate not supplied

Oxygen consumption and the energetics of island-dwelling Florida cottonmouth snakes.

We measured oxygen consumption (Vo(2)) to estimate standard metabolic rates (SMR) in cottonmouth snakes (Agkistrodon piscivorus conanti) from Seahorse Key and the adjacent peninsula of northern Florida. The island population is unusual because adult snakes feed on fish that are dropped by colonial nesting birds, and food resources are temporally limited relative to that of mainland populations. We found no differences in SMR between island and mainland snakes at any of four experimental temperatures (15 degrees -30 degrees C), suggesting that any adjustments to energy limitations involve other aspects of physiology or behavior. As with other viperid species, the SMR of cottonmouths is about one-half of that expected from interspecific allometric regressions previously reported for snakes generally. Allometric mass exponents of SMR averaged 0.76 and were not affected by temperature. We found that Vo(2) increased with temperature (Q(10) = 2.4-2.8) and was elevated 29% during scotophase compared with photophase. Neonates exhibited elevated Vo(2)compared with older juveniles of similar size, apparently due to assimilation of yolk that is present in the neonatal gut. In adult snakes, specific dynamic action (SDA) following feeding resulted in four- to eightfold increases in Vo(2), with magnitude and duration related positively to relative meal size. The total energy devoted to SDA increased with meal size and averaged 32.8%+/-4.4% of total ingested energy. We estimate that a nonreproductive 500-g adult cottonmouth at Seahorse Key uses 3,656 kJ of assimilated energy annually for maintenance and activity, which requires ingestion of approximately 1 kg of fish.

Animals↗
PubMed2006

Experimental infection of common garter snakes (Thamnophis sirtalis) with West Nile virus.

The role of various reptilian species in the infectious cycle of several arboviruses is documented, but their role in that of West Nile virus (WNV) is uncertain. Common garter snakes (Thamnophis sirtalis) were infected subcutaneously with 10(5) plaque forming units (PFU) WNV-Isr 98, five of nine snakes became viremic, and five exhibited persistent low levels of neutralizing antibodies. Four of the parentally infected snakes died and high titers of virus were found in multiple organ samples. In contrast, orally infected garter snakes did not become viremic, but viral RNA was detected in cloacal swabs. Since oral infection of predator birds by WNV is known, their ingestion of infected snakes may also result in their becoming infected.

Administration, Oral↗
PubMed2005

Snake Venom: Any Clue for Antibiotics and CAM?

Lately several naturally occurring peptides presenting antimicrobial activity have been described in the literature. However, snake venoms, which are an enormous source of peptides, have not been fully explored for searching such molecules. The aim of this work is to review the basis of antimicrobial mechanisms revealing snake venom as a feasible source for searching an antibiotic prototype. Therefore, it includes (i) a description of the constituents of the snake venoms involved in their main biological effects during the envenomation process; (ii) examples of snake venom molecules of commercial use; (iii) mechanisms of action of known antibiotics; and (iv) how the microorganisms can be resistant to antibiotics. This review also shows that snake venoms are not totally unexplored sources for antibiotics and complementary and alternative medicine (CAM).

Journal Article↗
PubMedDate not supplied

Heterologous amplification of microsatellite markers from colubroid snakes in European natricines (serpentes: natricinae).

Eighteen microsatellite loci developed for a range of snake species (New World natricines, elapids, crotalids) were tested against European natricines (Natrix natrix, N. maura, and N. tessellata) in cross-species amplification experiments. Five loci were polymorphic (average expected heterozygosity 0.749 for a population of N. natrix in Amsterdam, mean sample size 47.8) and three loci were monomorphic. The remainder could not be consistently scored or failed to amplify. Further tests on single individuals of a diverse set of eight species of colubroid snakes showed that 15 of the 18 loci could be cross-amplified in at least one of these species. We conclude that our results show promise for the utilization of these markers for experimental assessments of genetic variation in the phylogenetically closely related group of European natricine snakes with emphasis on N. natrix. The full suite of microsatellite markers now available for snakes may show additional potential for subsequent investigation across a broader range of colubroid snakes.

Animals↗
PubMed2003

Laser capture microdissection and real-time PCR for analysis of glomerular endothelin-1 gene expression in mesangiolysis of rat anti-Thy 1.1 and murine Habu Snake Venom glomerulonephritis.

Molecular analysis of pathologic changes in glomeruli requires methods allowing rapid and exact detection of alterations in gene expression. Here, we analyzed endothelin-1 (ET-1) mRNA expression in mesangiolytic glomeruli during the course of a rat and murine model of mesangioproliferative glomerulonephritis (GN). A novel method combining laser capture microdissection (LCM), which permits the precise removal of selected mesangiolytic glomeruli, with a highly sensitive real-time RT-PCR technique was used. Anti-Thy 1.1. GN was introduced in male Sprague-Dawley rats (1.0 mg/kg body weight of OX-7 IV) and Habu Snake Venom GN was introduced in C57BL6 mice (habu snake venom toxin 6 mg/kg body weight IV). The degree of mesangiolysis during both GNs was analyzed using a semiquantitative scoring system. Mesangiolytic glomeruli were microdissected at different days of the diseases (day 2, 6, and 12 in anti-Thy 1.1 GN and days 1, 3, 7, and 14 in Habu Snake Venom GN) and from normal control animals. After RNA extraction and cDNA synthesis, ET-1 gene expression was measured by real-time RT-PCR. In parallel, in anti-Thy 1.1. GN ET-1 mRNA expression was analyzed using semiquantitative nonradioactive in situ hybridization; ET-1 protein expression was investigated by immunohistochemistry. Mesangiolysis peaked at day 6 in anti-Thy1.1 GN and at day 1 in Habu Snake Venom GN. Mesangiolytic glomeruli were easily microdissected on cryostat sections in both models; quantification of mRNA with RT-PCR was reliable and reproducible. Glomerular ET-1 mRNA expression increased during the course of anti-Thy 1.1 GN and Habu Snake Venom GN peaked when mesangiolysis was most pronounced. This was seen by RT-PCR after glomerular LCM and by in situ hybridization; in parallel, glomerular ET-1 protein expression was increased. Combination of LCM and RT-PCR is a reliable method for quantification of localized gene expression in isolated renal structures. The above data argue for an important role of ET-1 in pathogenesis and/or repair of mesangiolysis in experimental mesangioproliferative GN.

Animals↗
PubMed2006

An invasive species induces rapid adaptive change in a native predator: cane toads and black snakes in Australia.

Rapid environmental change due to human activities has increased rates of extinction, but some species may be able to adapt rapidly enough to deal with such changes. Our studies of feeding behaviour and physiological resistance to toxins reveal surprisingly rapid adaptive responses in Australian black snakes (Pseudechis porphyriacus) following the invasion of a lethally toxic prey item, the cane toad (Bufo marinus). Snakes from toad-exposed localities showed increased resistance to toad toxin and a decreased preference for toads as prey. Separate laboratory experiments suggest that these changes are not attributable to learning (we were unable to teach naive snakes to avoid toxic prey) or to acquired resistance (repeated sub-lethal doses did not enhance resistance). These results strongly suggest that black snake behaviour and physiology have evolved in response to the presence of toads, and have done so rapidly. Toads were brought to Australia in 1935, so these evolved responses have occurred in fewer than 23 snake generations.

Adaptation, Physiological↗
PubMed1998

Detection of the carpal bone contours from 3-D MR images of the wrist using a planar radial scale-space snake.

In this paper we consider the problems encountered when applying snake models to detect the contours of the carpal bones in 3-D MR images of the wrist. In order to improve the performance of the original snake model introduced by Kass [1], we propose a new image force based on one-dimensional (1-D) second-order Gaussian filtering and contrast equalization. The improved snake is less sensitive to model initialization and has no tendency to cut off contour sections of high curvature, because 1-D radial scale-space relaxation is used. Contour orientation is used to minimize the influence of neighboring image structures. Due to 1-D contrast equalization an intensity insensitive measure of external energy is obtained. As a consequence a good balance between internal and external energetic contributions of the snake is established, which also improves convergence. By incorporating this new image force into the snake model, we succeed in accurate contour detection, even when relatively high noise levels are present and when the contrast varies along the contours of the bones.

Algorithms↗