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Alpha- and beta-keratins of the snake epidermis.

Snake scales contain specialized hard keratins (beta-keratins) and alpha- or cyto-keratins in their epidermis. The number, isoelectric point, and the evolution of these proteins in snakes and their similarity with those of other vertebrates are not known. In the present study, alpha- and beta-keratins of snake molts and of the whole epidermis have been studied by using two-dimensional electrophoresis and immunocytochemistry. Specific keratins in snake epidermis have been identified by using antibodies that recognize acidic and basic cytokeratins and avian or lizard scale beta-keratin. Alpha keratins of 40-70 kDa and isoelectric point (pI) at 4.5-7.0 are present in molts. The study suggests that cytokeratins in snakes are acidic or neutral, in contrast to mammals and birds where basic keratins are also present. Beta keratins of 10-15 kDa and a pI of 6.5-8.5 are found in molts. Some beta-keratins appear as basic proteins (pI 8.2) comparable to those present in the epidermis of other reptiles. Some basic "beta-keratins" associate with cytokeratins as matrix proteins and replace cytokeratins forming the corneous material of the mature beta-layer of snake scales, as in other reptiles. The study also suggests that more forms of beta-keratins (more than three different types) are present in the epidermis of snakes.

Animals↗

Altered visual experience and acute visual deprivation affect predatory targeting by infrared-imaging Boid snakes.

Boid and Crotaline snakes use both their eyes and infrared-imaging facial pit organs to target homeothermic prey. These snakes can target in complete darkness, but the eyes can also effectively direct predatory strikes. We investigated the behavioral correlates of boid snakes' simultaneous use of two imaging systems by testing whether congenital unilateral visual deprivation affects targeting performance. Normally sighted Burmese pythons exhibited average targeting angle of zero (on the midline axis of the head), but three unilaterally anophthalmic Burmese pythons targeted preferentially on the sighted side. A unilaterally anophthalmic amethystine python also targeted on the sighted side, and a unilaterally anophthalmic Brazilian rainbow boa tended to target on the sighted side, though its mean targeting angle was not significantly different from zero. When unilaterally anophthalmic Burmese pythons were temporarily blinded, mean strike angle changed to that of normally sighted snakes. These results show that while infrared-imaging snakes can shift between visual and infrared information under acute experimental conditions, loss of part of the visual field during development results in abnormal predatory targeting behavior. In contrast, normally sighted snakes subjected to temporary unilateral blinding do not target preferentially on the sighted side. Therefore, while loss of part of the visual field may be compensated for by infrared input in normal snakes, partial absence of visual input during development may alter central organization of visual information. Conversely, absence of half the visual field during development does not alter targeting performance based upon infrared input alone, suggesting that organization of the central infrared map does not depend upon normal organization of visual input.

Animals↗

Sublingual plicae (anterior processes) are not necessary for garter snake vomeronasal function.

The anterior processes of snakes may transfer odorants from the tongue to the vomeronasal (VN) organ. To test whether the anterior processes are required for a vomeronasally mediated behavior, the authors tested garter snakes (Thamnophis sirtalis) preoperatively and after cauterization of the anterior processes or control cauterization with artificial earthworms covered with earthworm wash (EWW) or distilled water. Snakes in both groups attacked EWW-covered artificial worms but not controls both pre- and postoperatively. In addition, snakes with anterior processes cauterization or control cauterization tongue flicked 3H-proline. Radioautographs of the VN organs of snakes with and without anterior processes were indistinguishable: Snakes in both groups had reduced silver grains over the VN sensory epithelium as had been reported previously with intact snakes. These findings indicate that the anterior processes are unnecessary both for a behavior known to require a functional VN system and for delivery of odorants to the VN organ.

Animals↗

Dynamic directional gradient vector flow for snakes.

Snakes, or active contour models, have been widely used in image segmentation. However, most present snake models do not discern between positive and negative step edges. In this paper, a new type of dynamic external force for snakes named dynamic directional gradient vector flow (DDGVF) is proposed that uses this information for better performance. It makes use of the gradients in both x and y directions and deals with the external force field for the two directions separately. In snake deformation, the DDGVF field is utilized dynamically according to the orientation of snake in each iteration. Experimental results demonstrate that the DDGVF snake provides a much better segmentation than GVF snake in situations when edges of different directions are present which pose confusion for segmentation.

Algorithms↗

An adaptive snake model for ultrasound image segmentation: modified trimmed mean filter, ramp integration and adaptive weighting parameters.

The snake model is a widely-used approach to finding the boundary of the object of interest in an ultrasound image. However, due to the speckles, the weak edges and the tissue-related textures in an ultrasound image, conventional snake models usually cannot obtain the desired boundary satisfactorily. In this paper, we propose a new adaptive snake model for ultrasound image segmentation. The proposed snake model is composed of three major techniques, namely, the modified trimmed mean (MTM) filtering, ramp integration and adaptive weighting parameters. With the advantages of the mean and median filters, the MTM filter is employed to alleviate the speckle interference in the segmentation process. The weak edge enhancement by ramp integration attempts to capture the slowly varying edges, which are hard to capture by conventional snake models. The adaptive weighting parameter allows weighting of each energy term to change adaptively during the deformation process. The proposed snake model has been verified on the phantom and clinical ultrasound images. The experimental results showed that the proposed snake model achieves a reasonable performance with an initial contour placed 10 to 20 pixels away from the desired boundary. The mean minimal distances from the derived boundary to the desired boundary have been shown to be less than 3.5 (for CNR > or = 0.5) and 2.5 pixels, respectively, for the phantom and ultrasound images.

Algorithms↗

Natural phospholipase A(2) myotoxin inhibitor proteins from snakes, mammals and plants.

A renewed interest in the phenomenon of inter- and intra-species resistance towards the toxicity of snake venoms, coupled with the search for new strategies for treatment of snake envenomations, has prompted the discovery of proteins which neutralize the major toxic components of these venoms. Among these emerging groups of proteins are inhibitors of toxic phospholipases A2 (PLA2s), many of which exhibit a wide range of toxic effects including muscle-tissue damage, neurotoxicity, and inflammation. These proteins have been isolated from both venomous and non-venomous snakes, mammals, and most recently from medicinal plant extracts. The snake blood-derived inhibitors have been grouped into three major classes, alpha, beta, and gamma, based on common structural motifs found in other proteins with diverse physiological properties. In mammals, DM64, an anti-myotoxic protein isolated from opossum serum, belongs to the immunoglobulin super gene family and is homologous to human alpha1B-glycoprotein and DM43, a metalloproteinase inhibitor from the same organism. In plants, a short note is made of WSG, a newly described anti-toxic-PLA2 glycoprotein isolated from Withania somnifera (Ashwaganda), a medicinal plant whose aqueous extracts neutralize the PLA2 activity of the Naja naja venom. The implications of these new groups of PLA2 toxin inhibitors in the context of our current understanding of snake biology as well as in the development of novel therapeutic reagents in the treatment of snake envenomations worldwide are discussed.

Animals↗

Characterization of venom (Duvernoy's secretion) from twelve species of colubrid snakes and partial sequence of four venom proteins.

R.E. Hill and S.P. Mackessy. Characterization of venom (Duvernoy's secretion) from twelve species of colubrid snakes and partial sequence of four venom proteins. Toxicon XX, xx-yy, 2000. - Venomous colubrids, which include more than 700 snake species worldwide, represent a vast potential source of novel biological compounds. The present study characterized venom (Duvernoy's gland secretion) collected from twelve species of opisthoglyphous (rear-fanged) colubrid snakes, an extremely diverse assemblage of non-venomous to highly venomous snakes. Most venoms displayed proteolytic activity (casein), though activity levels varied considerably. Low phosphodiesterase activity was detected in several venoms (Amphiesma stolata, Diadophis punctatus, Heterodon nasicus kennerlyi, H. n. nasicus and Thamnophis elegans vagrans), and acetylcholinesterase was found in Boiga irregularis saliva and venom, but no venoms displayed hyaluronidase, thrombin-like or kallikrein-like activities. High phospholipase A(2) (PLA(2)) activity was found in Trimorphodon biscutatus lambda venom, and moderate levels were detected in Boiga dendrophila and D. p. regalis venoms as well as B. dendrophila and H. n. nasicus salivas. Non-reducing SDS-PAGE revealed 7-20 protein bands (3.5 to over 200 kD, depending on species) for all venoms analyzed, and electrophoretic profiles of venoms were typically quite distinct from saliva profiles. Components from A. stolata, Hydrodynastes gigas, Tantilla nigriceps and T. e. vagrans venoms showed protease activity when run on gelatin zymogram gels. N-terminal protein sequences for three 26 kD venom components of three species (H. gigas, H. torquata, T. biscutatus) and one 3.5 kD component (T. nigriceps) were also obtained, and the 3.5 kD peptide showed apparent sequence homology with human vascular endothelial growth factor; these data represent the first sequences of colubrid venom components. Protease, phosphodiesterase and PLA(2) activities are also common to elapid and viperid snake venoms, but it is apparent that numerous other (as yet undescribed) components make up the majority of colubrid venom proteins. The complex nature of venoms produced by most species surveyed, and the high levels of protease or phospholipase A(2) activity of some venoms, suggest that many colubrids could become an important source of human health concern as encounters with these snakes increase.

Acetylcholinesterase↗

The detection of hemorrhagic proteins in snake venoms using monoclonal antibodies against Virginia opossum (Didelphis virginiana) serum.

Most snakes and a few warm-blooded animals have a resistance to snake venoms because of naturally occurring antihemorrhagins found in their sera. The antihemorrhagins in serum of Virginia opossum (Didelphis virginiana) neutralize hemorrhagic activity by binding to hemorrhagins in snake venoms. The binding characteristic of antihemorrhagins in D. virginiana serum was used to develop a five-step western blot. The detection of hemorrhagic proteins were measured indirectly with antihemorrhagins in Virginia opossum serum and with DV-2LD#2, a monoclonal antibody specific for Virginia opossum antihemorrhagins. Snake venoms were separated by native-PAGE, transferred to a Millipore Immobilon-P membrane and then incubated with crude Virginia opossum serum. The hemorrhagins in snake venom bind to antihemorrhagins in Virginia opossum serum which react with DV-2LD#2 a monoclonal antibody that is specific for Virginia opossum antihemorrhagins. DV-2LD#2 monoclonal antibody inhibits antihemorrhagic activity in Virginia opossum serum when mixed in equal amounts. The inhibition of antihemorrhagins by DV-2LD#2 monoclonal antibody suggests specificity. DV-2LD#2 monoclonal antibody does not recognize antihemorrhagins in gray woodrat (Neotoma micropus) serum. The five-step western blot reveals two well-defined bands which represent hemorrhagins found in Western diamondback rattlesnake (Crotalus atrox) venom. Venoms from 15 different snake species were examined to determine the usefulness of the five-step western blot. Other hemorrhagic venoms (Great Basin rattlesnake (C. viridis lutosus), Prairie rattlesnake (C. viridis viridis), Tancitaran dusky rattlesnake (C. pusillus), Northern Mojave rattlesnake (C. scutulatus scutulatus type B) and Northern Pacific rattlesnake (C. v. oreganus)) had one single band in the five-step western blot. DV-2LD#2 did not bind to the non-hemorrhagic venoms and reacted with 50% of the hemorrhagic venoms used in this study. The monoclonal antibody, CAH, reacted with all the hemorrhagic venoms except for the venom of the King cobra (Ophiophagus hannah) and did not react with the non-hemorrhagic venoms. The hemorrhagic binding site of CAH monoclonal antibody and the antihemorrhagin in Virginia opossum are different binding sites. The five-step western blot will be a very useful assay for determining hemorrhagic activity without using live animals.

Animals↗

A study on the venom yield of venomous snake species from Argentina.

A study on the venom yield of snakes from Argentina over a three year period was carried out on adult specimens of Bothrops alternatus (n = 74); Bothrops neuwiedii (n = 127); Bothrops ammodytoides (n = 30); Bothrops moojeni (n = 14); Bothrops jararaca (n = 14); B. jararacussu (n = 6); Crotalus durissus terrificus (n = 120) and Micrurus spp. (n = 6) as well as with 12 specimens of newborn C. d. terrificus kept in captivity. While for each species there was a positive correlation between venom yield and number of snakes milked, the correlation with the snake's body weights after individual milkings was even better, suggesting that the size of the snakes is more important in determining the venom yield than the number of snakes milked or the specimen's sex. Individual milkings indicated that, in addition to the snake size, when the amount of venom is normalized per 100 g body weight there is a species specific difference in venom yield. It follows the order B. jararacussu > B. moojeni approximately = B. jararaca approximately = B. alternatus > B. neuwiedii> Micrurus spp approximately = B. ammodytoides> C. d. terrificus. Although the venom yield per 100 g body weight of newborn C. d. terrificus specimens is 2-fold higher than that of adults, no correlation was observed between venom yield and body weight.

Age Factors↗

Bites by foreign venomous snakes in Britain.

In 1970-7 17 people in Britain were the victims of 32 bites by foreign venomous snakes. Crotalus atrox caused eight of these bites, Bitis arietans five, and the remaining 19 bites were caused by 12 different species. All the victims were bitten while handling the snake, and 24 bites were incurred by private individuals in their own homes. Poisoning was negligible in 17 of the 32 bites but life-threatening in at least two cases. Thus in the early stages snake bite may be unpredictable as a clinical problem. All victims of snake bite should be observed for at least 12 hours to assess the severity of poisoning and to ensure rational treatment. Local necrosis developed in six cases and resulted in prolonged illness in five of these cases; local incision was carried out and many have been a casual factor. Comprehensive stocks of antivenoms for treating bites by foreign venomous snakes are held by the National Health Services in Liverpool and London. Antivenom is indicated (a) for potentially serious systemic poisoning, as evidenced by hypotension, electrocardiographic changes, neurtrophilia, and acidosis (after viper or elapid bites), abnormal bleeding or non-clotting blood after viper bites; and ptosis or glossopharyngeal palsy after elapid bites; and (b) for bites from snakes whose venom causes local necrosis, to prevent or minimise this unpleasant complication. For effective antivenom treatment intravenous infusion is mandatory.

Adolescent↗

[Snakes from the urban area of Cuiabá, Mato Grosso: ecological aspects and associated snakebites].

This study presents data on snakes recorded in the urban area of Cuiabá, Mato Grosso, Brazil. Sources of information included specimens captured by local residents (1986-1993) and turned over to the Mato Grosso Regional Ophiological Center (Normat), and data from the Anti-Venom Information Center (Ciave), regarding urban snake bites (1988-1993). Thirty-seven species of snakes from 25 genera and three families were recorded. Diurnal and terrestrial habits predominated, as well as a diet based on amphibians and/or lizards. From a total of 307 snake bites recorded, some 56% were of no clinical importance, caused by non-venomous snakes, whereas 44% were clinically relevant. Approximately 99% of the latter were attributed to vipers of the genus Bothrops, and especially the Bothrops moojeni and Bothrops neuwiedi species The colubrids Philodryas olfersii and Waglerophis merremii were probably responsible for most of the non-venomous snake bites.

Animals↗

Impact of snake bites and determinants of fatal outcomes in southeastern Nepal.

Current available data on snake bites in Nepal are based solely on hospital statistics. This community-based study aimed at evaluating the impact of snake bites and determining the risk factors associated with a fatal outcome in southeastern Nepal. A total of 1,817 households, selected by a random proportionate sampling method, were visited by trained field workers in five villages. Extensive data from snake bite victims during the 14 previous months were recorded and analyzed. One hundred forty-three snake bites including 75 bites with signs of envenoming were reported (annual incidence = 1,162/100,000 and 604/100,000, respectively), resulting in 20 deaths (annual mortality rate = 162/100,000). Characteristics of krait bites such as bites occurring inside the house, while resting, and between midnight and 6:00 am were all factors associated with an increased risk of death, as were an initial consultation with a traditional healer, a long delay before transport, and a lack of available transport. An initial transfer to a specialized treatment center and transport by motorcycle were strong protective factors. Among the 123 survivors, wounds required dressing and surgery in 30 (24%) and 10 (8%) victims, respectively, the mean working incapacity period was 15 days, and the mean out-of-pocket expense was 69 U.S. dollars. Snake bite is a major but neglected public health problem in southeastern Nepal. Public health interventions should focus on improving victims' rapid access to anti-snake venom serum by promoting immediate and fast transport to adequate treatment centers, particularly for bites occurring at night.

Adult↗

The cochlear nuclei of snakes.

The cochlear nuclei of three burrowing snakes (Xenopeltis unicolor, Cylindrophis rufus, and Eryx johni) and three non-burrowing snakes (Epicrates cenchris, Natrix sipedon, and Pituophis catenifer) were studied. The posterior branch of the statoacoustic nerve and its posterior ganglion were destroyed and the degenerated nerve fibers and terminals traced to primary cochlear nuclei in 13 specimens of Pituophis catenifer. All these snake species possess three primary and one secondary cochlear nuclei. The primary cochlear nuclei consist of a small nucleus angularis located at the cerebello-medullary junction and a fairly large nucleus magnocellularis forming a dorsal cap over the cephalic end of the alar eminence. Nucleus magnocellularis may be subdivided into a medially placed group of rounder cells, nucleus magnocellularis medialis, and a laterally placed group of more ovate and paler-staining cells, nucleus magnocellularis lateralis. A small but well-defined secondary nucleus which showed no degenerated nerve terminals after nerve root section, nucleus laminaris, underlies the cephalic part of both nucleus magnocellularis medialis and nucleus magnocellularis lateralis. Larger and better-developed cochlear nuclei were found in burrowing species than in non-burrowing species of snakes. Of the three burrowing species studied, Xenopeltis showed the greatest development of cochlear nuclei; Eryx cochlear nuclei were not quite as large but were better differentiated than in Xenopeltis; and Cylindrophis cochlear nuclei were fairly large but not as well developed nor as well differentiated as in either Xenopeltis or Eryx. The cochlear nuclei of the three non-burrowing snakes, Epicrates, Natrix, and Pituophis, were not as large nor as well developed as those of the burrowing snakes. There is some, but not complete, correlation between cochlear development and papilla basilaris length and number of hair cells. Thus, Xenopeltis and Eryx, with well-developed cochlear nuclei, have relatively long papillae basilares; but the boid, Epicrates, with less well-developed cochlear nuclei, has a fairly well-developed papilla basilaris. Cylindrophis, a burrowing species, shows only a moderate degree of cochear nuclei and papilla basilaris development. The non-burrowers, Natrix and Pituophis, have both small cochlear nuclei and relatively short papillae basilares.

Animals↗

Functional markers of the major histocompatibility gene complex of snakes.

In optimal seasonal conditions, outbred adult snakes Psammophis sibilans displayed the major immunological functions related, in mammals, to the presence of the major histocompatibility gene complex (MHC). Thus, out of 30 snake random pairs that exchanged skin transplants 72.9% rejected their allograft in an acute or subacute manner. Strongly significant proliferative response was recorded in 67.3% of 168 separate one-way mixed leukocyte reaction (MLR) cultures. Lymphocytes from 6/11 snakes immunized by skin allografting displayed, after secondary stimulation in vitro, cell-mediated lympholysis (CML) in vitro of 51Cr-labeled lymphoblasts derived from the donor snake. Finally, cytotoxic alloantibodies were readily generated after snake priming with skin allograft and blood cells. Snakes did not only exhibit the major cell- and humoral-mediated immune functions, but these functions appeared to be linked with the degree of MLR disparity. Thus, animals with different MLR rejected skin allografts acutely and produced cytotoxic effector cells. In contrast, MLR-identical animals rejected the skin allograft of their partner chronically and failed to produce killer cells in CML. This significant positive correlation between MLR disparity, graft rejection and CML suggests that the responsible antigens are encoded, as in other vertebrates, by the same genetic system, the MHC.

Animals↗

Ontogenetic change in molecular and functional properties of blood of garter snakes, Thamnophis sirtalis.

At birth the blood of garter snakes (Thamnophis sirtalis) has high oxygen affinity (P50 at 25 degrees C and pH 7.2 = 21 mm Hg), low hematocrit (14%), and a high proportion of inactive hemoglobin (44%). Growth of the snakes is accompanied by a steady decrease in oxygen affinity (P50 = 44 mm Hg in adults), increased hematocrit (to 44%), and decreased inactive hemoglobin (to 3%). These adult values are reached in the second or third year of a snake's life. There is a continuous change in the electrophoretically separable hemoglobin fractions during this period, but there is no ontogenetic change in the oxygen affinity of phosphate-stripped hemoglobin in solution. Organic triphosphates do not fully explain the ontogenetic change in blood oxygen affinity. Similar ontogenetic changes in blood oxygen affinity were found in two other species of snakes. The unusual morphology of snake lungs and consequent changes in pulmonary pO2 and pCO2 during ontogeny may be the basis for the unusual relationship between body size and blood oxygen affinity in snakes.

Adenosine Triphosphate↗

Anatomical and gravitational influences on cardiac displacement in snakes (Lepidosauria, Serpentes).

Radiographs of live, unanesthetized snakes were used to document the position of the heart in the body cavity during horizontal, head-up, and head-down postures. The extent of cardiac displacement observed during these postural changes differed substantially among the snakes examined, ranging from virtually none in a thin-bodied arboreal snake to as much as three vertebral lengths (= half the length of the heart) in a heavy-bodied terrestrial Crotalus. The basis of this differential cardiac displacement is attributed to the anatomical "packaging" of the pericardial sac. In some snakes the pericardial sac is loosely suspended in the body cavity by the great vessels and connective tissue sheets. In contrast, in other snakes the pericardical sac is buttressed against the body wall, the lung, or the liver. We hypothesize that cardiac displacement during postural change may alter the pattern of blood flow in the aortae of snakes.

Animals↗

Sea snake (Microcephalophis gracilis) hemoglobin: primary structure and relationships to other forms.

The hemoglobin of the sea snake Microcephalophis gracilis was purified and the primary structure of the alpha and beta chains determined. This is the first sea snake hemoglobin structure characterized, and apparently also the first complete structure of any snake hemoglobin (an alpha chain of a viper was known), allowing judgments of reptilian variants. Variations between the sea snake form and other reptilian forms are large (52-65 differences for the alpha chains), of similar order as those between the sea snake and avian (56-65 differences) or human (58 differences) forms. Functionally, 19 residues at alpha/beta contact areas and 7 at heme contacts are exchanged in relation to the human alpha and beta chains. Four positions of the sea snake hemoglobin contain residues thus far unique to this form. However, all replacements appear compatible with conserved overall functional properties.

Amino Acid Sequence↗

Identification of a locality in snake venom alpha-neurotoxins with a significant compositional similarity to marine snail alpha-conotoxins: implications for evolution and structure/activity.

alpha-neurotoxins from elapid snake venoms and alpha-conotoxins from marine snails bind specifically and with high affinity to nicotinic cholinoceptors. Although both types of toxin are polypeptides, there is more than a fourfold difference in size between the two and no clear sequence homology is evident. A systematic computer search of the three-dimensional structure of erabutoxin b (an alpha-neurotoxin from the false sea snake Laticauda semifasciata) was performed to identify the locality that most closely matched the amino acid compositions of the smaller alpha-conotoxins (from the marine snails Conus magus and Conus geographus). The area of greatest similarity centered on residue position 25 of erabutoxin b, a locale that is conserved throughout the snake alpha-neurotoxins and their homologues. Six proteins unrelated to erabutoxin b were compared to the alpha-conotoxins to show that the extent of the erabutoxin b/alpha-conotoxin match was too high to be coincidental. Homologues of erabutoxin b, namely alpha-cobratoxin from Naja naja siamensis and cytotoxin VII4 from Naja mossambica mossambica, were also analyzed. The extent of the matching with the alpha-conotoxins decreased in the series erabutoxin b greater than alpha-cobratoxin greater than cytotoxin VII4, and this also relates the order of similarity to the pharmacological properties of the alpha-conotoxins. The alpha-conotoxin-like area of the snake alpha-neurotoxins is peripheral to the site previously considered important for binding to the cholinoceptor, even though it seems to represent the focus of evolutionary convergence between the two types of neurotoxin. The area of resemblance does, however, have strong associations with the conformational behavior of the snake toxins. Hence, the outcome of this study has important consequences for the current ideas on snake alpha-neurotoxin structure/activity relationships and the evolutionary origins of neurotoxicity.

Amino Acid Sequence↗