Sample records for agkistrodon piscivorus leucostoma

  1. Agkistrodon piscivorus conanti (Florida cottonmouth) Diet

    USGS Publications Warehouse

    Grajal-Puche, Alejandro; Josimovich, Jillian; Falk, Bryan; Reed, Robert


    Agkistrodon piscivorus is a generalist predator that feeds on a variety of prey, including snakes (Gloyd and Conant 1990. Snakes of the Agkistrodon Complex: A Monographic Review. Society for the Study of Amphibians and Reptiles, Oxford, Ohio. 614 pp.; Lillywhite et al. 2002. Herpetol. Rev. 33:259–260; Hill and Beaupre 2008. Copeia 2008:105–114). Cemophora coccinea (Scarletsnake) is not known as one of the 26 species of snakes consumed by A. piscivorus (Ernst and Ernst 2011. Venomous Reptiles of the United States, Canada, and Northern Mexico: Volume 1. Johns Hopkins University Press, Baltimore, Maryland. 193 pp.). On 16 June 2015, at 2210 h, we found a dead-on-road A. piscivorus (total length [TL] = 51.0 cm) in Everglades National Park on Main Park Road, 1.88 km S Pa-hay-okee, Miami-Dade Co., Florida, USA (25.414085°N, 80.78183146°W, WGS84; elev. 3 m). The snake had been killed by a vehicle and some internal organs were exposed. Visible stomach contents included a small (TL ca. 15 cm) C. coccinea. Photographic vouchers of the A. piscivorus (UF-Herpetology 177194) and C. coccinea (UF-Herpetology 177195) were deposited in the Division of Herpetology, Florida Museum of Natural History, University of Florida. Despite the fact that these species are sympatric over large areas of the southeastern United States, this is the first known documented predation of C. coccinea by A. piscivorus.

  2. Do warning displays predict striking behavior in a viperid snake, the cottonmouth (Agkistrodon piscivorus)?

    SciTech Connect

    Glaudas, X. and C.T. Winne


    Warning displays are defined as signals designed to intimidate predators or indicate a proclivity to fight. However, support for the idea that warning behaviors signal an intent to fight is largely based on anecdotes and isolated observations, and a complete understanding of antipredator behavior will only be achieved if specific hypotheses are experimentally tested. Herein, we tested in a North American viperid snake, the cottonmouth (Agkistrodon piscivorus), the hypothesis that warning displays serve as a reliable signal to potential predators that a snake will strike. The cottonmouth exhibits two stereotypical warning displays during predator confrontation, i.e., mouth gaping and tail vibrations, making it an ideal study organism to experimentally test the relationship between warning displays and defensive striking. To test this idea, we recorded the sequence of defensive behavior--gaping, tail vibrating, and striking--of cottonmouths towards a standardized predatory stimulus in the laboratory. As predicted, snakes that gaped during the trials were subsequently more likely to strike than snakes that did not. In contrast, striking behavior was independent of the occurrence of tail vibrations. Our results suggest that gaping behavior--but not tail-vibrating behavior--may provide an honest signal to would-be predators.

  3. Phylogenetic Analysis of Bacterial Communities in Different Regions of the Gastrointestinal Tract of Agkistrodon piscivorus, the Cottonmouth Snake

    PubMed Central

    Colston, Timothy J.; Noonan, Brice P.; Jackson, Colin R.


    Vertebrates are metagenomic organisms in that they are composed not only of their own genes but also those of their associated microbial cells. The majority of these associated microorganisms are found in the gastrointestinal tract (GIT) and presumably assist in processes such as energy and nutrient acquisition. Few studies have investigated the associated gut bacterial communities of non-mammalian vertebrates, and most rely on captive animals and/or fecal samples only. Here we investigate the gut bacterial community composition of a squamate reptile, the cottonmouth snake, Agkistrodon piscivorus through pyrosequencing of the bacterial 16S rRNA gene. We characterize the bacterial communities present in the small intestine, large intestine and cloaca. Many bacterial lineages present have been reported by other vertebrate gut community studies, but we also recovered unexpected bacteria that may be unique to squamate gut communities. Bacterial communities were not phylogenetically clustered according to GIT region, but there were statistically significant differences in community composition between regions. Additionally we demonstrate the utility of using cloacal swabs as a method for sampling snake gut bacterial communities. PMID:26039313

  4. Venomics of New World pit vipers: Genus-wide comparisons of venom proteomes across Agkistrodon

    PubMed Central

    Lomonte, Bruno; Tsai, Wan-Chih; Ureña-Diaz, Juan Manuel; Sanz, Libia; Mora-Obando, Diana; Sánchez, Elda E.; Fry, Bryan G.; Gutiérrez, José María; Gibbs, H. Lisle; Sovic, Michael G.; Calvete, Juan J.


    We report a genus-wide comparison of venom proteome variation across New World pit vipers in the genus Agkistrodon. Despite the wide variety of habitats occupied by this genus and that all its taxa feed on diverse species of vertebrates and invertebrate prey, the venom proteomes of copperheads, cottonmouths, and cantils are remarkably similar, both in the type and relative abundance of their different toxin families. The venoms from all the eleven species and subspecies sampled showed relatively similar proteolytic and PLA2 activities. In contrast, quantitative differences were observed in hemorrhagic and myotoxic activities in mice. The highest myotoxic activity was observed with the venoms of A. b. bilineatus, followed by A. p. piscivorus, whereas the venoms of A. c. contortrix and A. p. leucostoma induced the lowest myotoxic activity. The venoms of Agkistrodon bilineatus subspecies showed the highest hemorrhagic activity and A. c. contortrix the lowest. Compositional and toxicological analyses agree with clinical observations of envenomations by Agkistrodon in the USA and Central America. A comparative analysis of Agkistrodon shows that venom divergence tracks phylogeny of this genus to a greater extent than in Sistrurus rattlesnakes, suggesting that the distinct natural histories of Agkistrodon and Sistrurus clades may have played a key role in molding the patterns of evolution of their venom protein genes. Biological significance A deep understanding of the structural and functional profiles of venoms and of the principles governing the evolution of venomous systems is a goal of venomics. Isolated proteomics analyses have been conducted on venoms from many species of vipers and pit vipers. However, making sense of these large inventories of data requires the integration of this information across multiple species to identify evolutionary and ecological trends. Our genus-wide venomics study provides a comprehensive overview of the toxic arsenal across

  5. Management of Tissue Loss After Agkistrodon Snakebite: Appropriate Use of Crotalidae-Fab Antivenin.


    Larson, Kenneth W; Schaefer, Keith R; Austin, Cindy; Norton, Rhy; Finley, Phillip J


    Although initially created for the treatment of rattlesnake (genus: Crotalus) bites, Crotalidae-Fab antivenin is used to treat many different pit viper envenomations. However, the efficacy of Crotalidae-Fab in preventing tissue loss from copperhead (Agkistrodon contortrix) or cottonmouth (Agkistrodon piscivorus) snakebites remains unclear. Recent reports show that Agkistrodon-related bites rarely require treatment beyond simple observation and pain control. The purpose of this study was to examine the amount of tissue loss in patients who received Crotalidae-Fab compared with those who did not after an Agkistrodon bite. After institutional review board approval, a retrospective study was completed at a Level 1 trauma center. Between 2009 and 2013, a total of 57 snakebites were identified. Of the 57 bites, the snake species was documented in 36 cases including 31 copperheads, 1 cottonmouth, and 4 rattlesnakes. The other 21 bites were from unknown or nonvenomous species. Of the 32 Agkistrodon-related bites, 15 patients received Crotalidae-Fab (average of 3 vials administered) and 17 did not receive Crotalidae-Fab. None of the 32 patients, regardless of treatment option, had tissue loss or required surgical interventions. Only 1 patient received Crotalidae-Fab and debridement of a vesicle associated with the bite. No clinically significant differences were observed between the groups. These findings support previous literature that failed to show added benefit of Crotalidae-Fab treatment for Agkistrodon bites beyond patient comfort and pain control. Evaluation of current protocols for Agkistrodon envenomations is warranted. Snakebite wound education in trauma physicians and nurses may decrease unnecessary use of antivenom medication. PMID:27163225

  6. Envenomation by a juvenile Malayan pit viper (Agkistrodon rhodostoma).


    Vest, D K; Kardong, K V


    Following an accidental bite inflicted by a juvenile Malayan pit viper (Agkistrodon rhodostoma), the progress of envenomation was carefully monitored and subsequent laboratory work performed to determine the amount and quality of venom injected. Even a very small amount of venom from this species is capable of inducing noticeable local symptoms including edema, subcutaneous bleeding, and throbbing. Constitutional symptoms were present but minimal. The extent of inflammation present at any given time following envenomation was found to be a more accurate diagnostic signal than the speed with which it developed. This case is similar to that of bites by juveniles of other species of viperines and crotalines and may serve to aid in the diagnosis and treatment of bites by juvenile specimens of serpents in these families. PMID:7398218

  7. Viper metalloproteinase (Agkistrodon halys pallas) with antimicrobial activity against multi-drug resistant human pathogens.


    Samy, Ramar Perumal; Gopalakrishnakone, Ponnampalam; Chow, Vincent T K; Ho, Bow


    Metalloproteinases are abundant enzymes in crotalidae and viperidae snake venoms. Snake venom metalloproteinases (SVMPs) comprise a family of zinc-dependent enzymes, which display many different biological activities. A 23.1 kDa protein was isolated from Agkistrodon halys (pallas, Chinese viper) snake venom. The toxin is a single chain polypeptide with a molecular weight of 23146.61 and an N-terminal sequence (MIQVLLVTICLAVFPYQGSSIILES) relatively similar to that of other metalloprotein-like proteases isolated from the snake venoms of the Viperidae family. The antibacterial effect of Agkistrodon halys metalloproteinase (AHM) on Burkholderia pseudomallei (strains TES and KHW), Escherichia coli, Enterobacter aerogenes, Proteus vulgaris, Proteus mirabilis, Pseudomonas aeruginosa (Gram-negative bacteria) and Staphylococcus aureus (Gram-positive bacterium) was studied at a concentration 120 microM. Interestingly, we found that the metalloproteinase exhibited antibacterial properties and was more active against S. aureus, P. vulgaris, P. mirabilis and multi-drug resistant B. pseudomallei (strain KHW) bacteria. AHM variants with high bacteriostatic activity (MIC 1.875-60 microM) also tended to be less cytotoxic against U-937 human monocytic cells up to 1 mM concentrations. These results suggest that this metalloprotein exerts its antimicrobial effect by altering membrane packing and inhibiting mechanosensitive targets. PMID:18297685

  8. Studies on the coagulant enzyme from Agkistrodon rhodostoma venom. Isolation and some properties of the enzyme

    PubMed Central

    Hatton, M. W. C.


    1. Arvin, a commercial preparation of the coagulant activity from the venom of Agkistrodon rhodostoma, is shown to contain a non-coagulant caseinolytic fraction. 2. A method is described for the purification of the coagulant enzyme free from any detectable contaminating protein. 3. The coagulant enzyme is identified as a glycoprotein which probably consists of a single polypeptide chain containing approx. 29% by weight of carbohydrate. Amino acid and carbohydrate analyses are reported and the N- and C-terminal amino acid residues identified. 4. Electrophoresis on polyacrylamide gel reveals the polymorphic nature of the glycoprotein. Five forms of the enzyme are observed. 5. The coagulant action is correlated with an arginine esterase activity and kinetic properties are studied with both arginine and lysine esters as substrates. The inhibitory nature of guanidine and arginine toward the esterase activity is reported. PMID:4737326

  9. A catalog for transcripts in the venom gland of the Agkistrodon acutus: identification of the toxins potentially involved in coagulopathy.


    Qinghua, Liu; Xiaowei, Zhang; Wei, Yin; Chenji, Li; Yijun, Huang; Pengxin, Qiu; Xingwen, Su; Songnian, Hu; Guangmei, Yan


    Agkistrodon acutus is a special agkistrodon halys, only distributed in Southern China, with a few exceptions in Vietnam. It is a cherished element used in traditional Chinese medicine. In order to produce a global panorama of gene expression in the Agkistrodon acutus venom gland, a non-normalized cDNA library was constructed, and 8696 high quality 5' end expressed sequenced tags (ESTs) were sequenced and analyzed. The initial sequences were assembled into 2855 clusters. Of these clusters, only 45.60% clusters matched known sequence and 54.40% had no match to any known sequence in GenBank. Except for putative cellular proteins (1184 clusters), the remaining 118 clusters (40.16% of all ESTs) corresponded to sequences associated with diverse toxin function. According to expression abundance, the major toxin components were metalloproteinases (32.08%) and C-type lectin (5.22%), and other components including bradykinin-potentiating peptide (0.90%), serine proteases (0.51%), nucleotidase and nuclease (0.41%), phospholipase A2 (0.30%), disintegrin (0.05%), cytokine-like molecules (0.06%), and other proteins (0.63%). The majority of these components are thought to be responsible for coagulopathy after A. acutus bites. We have therefore generated a comprehensive catalog of the A. acutus venom gland described so far. Gene expression from the very specialized secretory tissue, especially for those involved in coagulopathy, can be surveyed and provide important information in finding novel toxins. PMID:16438937

  10. Development of Snake Fungal Disease after Experimental Challenge with Ophidiomyces ophiodiicola in Cottonmouths (Agkistrodon piscivorous)

    PubMed Central

    Allender, Matthew C.; Baker, Sarah; Wylie, Daniel; Loper, Daniel; Dreslik, Michael J.; Phillips, Christopher A.; Maddox, Carol; Driskell, Elizabeth A.


    Snake fungal disease (SFD) is a clinical syndrome associated with dermatitis, myositis, osteomyelitis, and pneumonia in several species of free-ranging snakes in the US. The causative agent has been suggested as Ophidiomyces ophiodiicola, but other agents may contribute to the syndrome and the pathogenesis is not understood. To understand the role of O. ophiodiicola in SFD, a cottonmouth snake model of SFD was designed. Five cottonmouths (Agkistrodon piscivorous) were experimentally challenged by nasolabial pit inoculation with a pure culture of O. ophiodiicola. Development of skin lesions or facial swelling at the site of inoculation was observed in all snakes. Twice weekly swabs of the inoculation site revealed variable presence of O. ophiodiicola DNA by qPCR in all five inoculated snakes for 3 to 58 days post-inoculation; nasolabial flushes were not a useful sampling method for detection. Inoculated snakes had a 40% mortality rate. All inoculated snakes had microscopic lesions unilaterally on the side of the swabbed nasolabial pit, including erosions to ulcerations and heterophilic dermatitis. All signs were consistent with SFD; however, the severity of lesions varied in individual snakes, and fungal hyphae were only observed in 3 of 5 inoculated snakes. These three snakes correlated with post-mortem tissue qPCR evidence of O. ophiodiicola. The findings of this study conclude that O. ophiodiicola inoculation in a cottonmouth snake model leads to disease similar to SFD, although lesion severity and the fungal load are quite variable within the model. Future studies may utilize this model to further understand the pathogenesis of this disease and develop management strategies that mitigate disease effects, but investigation of other models with less variability may be warranted. PMID:26469977

  11. A new Agkistrodon halys venom-purified protein C activator prevents myocardial fibrosis in diabetic rats

    PubMed Central

    Li, Shu; Hong, Yun; Jin, Xin; Zhang, Genbao; Hu, Zaichang; Nie, Liuwang


    Aim To assess the effects of protein C activator (PCA) from Agkistrodon halys snake venom on cardiac fibrosis in streptozotocin (STZ) induced diabetic rat model, and investigate the mechanisms of its action. Methods PCA was identified by one-dimensional reversed phase liquid chromatography – mass spectrometry/mass spectrometry. Male Sprague-Dawley rats (120-140 g) were randomly assigned to negative control (NC) and diabetic group. Diabetes was induced by STZ in high-fat diet fed rats. Diabetic group was subdivided into three groups: diabetic group (DM), diabetic group treated with PCA (0.5, 2, and 8 mg/kg), and diabetic group treated with metformin (5 mg/kg, positive control). NC and DM groups received the same volume of distilled water. Left ventricular mass index (LVWI) and collagen volume fraction were measured by hematoxylin and eosin and Masson staining. Transforming growth factor beta-1 (TGF-β1) and interleukin 1 beta (IL-1β) levels were determined by enzyme-linked immunosorbent assay. Results The diabetic rat model was successfully established by STZ induction and high-fat diet. Glucose level, LVWI, TGF-β1 and IL-1β level, and collagen volume fraction were significantly reduced in diabetic rats treated by PCA in a dose-dependent manner (P < 0.050), especially in the high dose (8 mg/kg) group (P < 0.010), compared to diabetes group. The high dose PCA had the same effect as metformin positive control in reducing the level of fasting blood glucose. PCA decreased the expression of MMP-2 and reduced that of TIMP-2. Conclusion Our results indicate that PCA has anti-fibrotic effects and that it may be used to treat myocardial fibrosis. PMID:26526881

  12. A novel plasminogen activator from Agkistrodon blomhoffii Ussurensis venom (ABUSV-PA): Purification and characterization

    SciTech Connect

    Liu Shuqing; Sun Mingzhong . E-mail:; Greenaway, Frederick T.


    A plasminogen activator with arginine ester hydrolysis activity (ABUSV-PA) has been identified and purified to homogeneity from Chinese Agkistrodon blomhoffii Ussurensis snake venom. ABUSV-PA, a monomeric protein with molecular mass of 27815.2 Da, was purified 180-fold with 0.02% recovery for protein and 3.6% recovery for esterase activity. ABUSV-PA reacts optimally with its substrate N {sub {alpha}}-tosyl-L-arginine-methyl ester (TAME) at {approx}pH 7.5 and at 51 {sup o}C. Measurement from inductively coupled plasma-atomic emission spectroscopy (ICP-AES) reveals that ABUSV-PA is a Zn{sup 2+}-containing protein with a stoichiometry of 1:1 [Zn{sup 2+}]:[ABUSV-PA]. Analyses of esterase hydrolysis and UV absorption and CD spectra indicate that Zn{sup 2+} plays an important role in maintaining the structural integrity rather than the esterase activity of ABUSV-PA. Divalent metal ions, including Ca{sup 2+}, Mg{sup 2+}, Cu{sup 2+}, Ni{sup 2+}, Mn{sup 2+}, and Co{sup 2+}, increase the TAME hydrolysis activity of ABUSV-PA. A red-shift of the emission wavelengths of the synchronous fluorescence of ABUSV-PA, compared to those of free Tyr and Trp, indicates a conformation where the Tyr and Trp residues are in exposed hydrophilic environments. The presence of zinc increases the hydrophobicity of the conformational environments surrounding the Trp residues of ABUSV-PA and affects the secondary structure of ABUSV-PA, as proved by UV absorption and CD spectroscopy.

  13. [Fibrinogen/fibrin-specific enzymes from copperhead (Agkistrodon halys halys) and cobra (Naja oxiana eichwald) snake venoms].


    Yunusova, E S; Sadykov, E S; Sultanalieva, N M; Shkinev, A V


    Ability of fractions of cobra's (Naja oxiana Eichwald) and copperhead snake's (Agkistrodon halys halys) venoms to hydrolyze fibrinogen/fibrin was studied. In cobra's snake a component with molecular mass of nearly 60 kDa was found to hydrolyze a-chain of fibrinogen but failed to hydrolyze casein/azocasein and fibrin. A fibrinogen-specific metalloproteinase, the enzyme was inhibited by EDTA. Cobra's venom reduced the mass of donor's fresh blood clots. The copperhead snake's venom and the fractions obtained by gel-filtration (HW-50) and ion exchange chromatography (DEAE-650) were found to hydrolyze casein/azocasein, a- and b-chains of fibrinogen/fibrin and donor's blood clots. The results from the study of the venom and proteolytically active fractions are the evidence for a thrombolytic potential in a copperhead snake's venom. PMID:27420616

  14. Effect of Iron and Carbon Monoxide on Fibrinogenase-like Degradation of Plasmatic Coagulation by Venoms of Six Agkistrodon Species.


    Nielsen, Vance G; Redford, Daniel T; Boyle, Patrick K


    Annually, thousands suffer poisonous snakebite, often from defibrinogenating species. It has been demonstrated that iron and carbon monoxide change the ultrastructure of plasma thrombi and improve coagulation kinetics. Thus, this investigation sought to determine whether pre-treatment of plasma with iron and carbon monoxide could attenuate venom-mediated catalysis of fibrinogen obtained from Agkistrodon species with fibrinogenase activity. Human plasma was pre-treated with ferric chloride (0-10 μM) and carbon monoxide-releasing molecule-2 (CORM-2, 0-100 μM) prior to exposure to 0.5-11 μg/ml of six different Agkistrodon species' venom. The amount of venom used for experimentation needed to decrease coagulation function of one or more kinetic parameters by at least 50% of normal values for (e.g. half the normal speed of clot formation). Coagulation kinetics were determined with thrombelastography. All six snake venoms degraded plasmatic coagulation kinetics to a significant extent, especially prolonging the onset to clot formation and diminishing the speed of clot growth. Pre-treatment of plasma with iron and carbon monoxide attenuated these venom-mediated coagulation kinetic changes in a species-specific manner, with some venom effects markedly abrogated while others were only mildly decreased. Further in vitro investigation of other pit viper venoms that possess fibrinogenolytic activity is indicated to identify species amenable to or resistant to iron and carbon monoxide-mediated attenuation of venom-mediated catalysis of fibrinogen. Lastly, future pre-clinical investigation with animal models (e.g. rabbit ear-bleed model) is planned to determine whether iron and carbon monoxide can be used therapeutically after envenomation. PMID:26467642

  15. Purification, characterization and bactericidal activities of basic phospholipase A2 from the venom of Agkistrodon halys (Chinese pallas).


    Perumal Samy, R; Gopalakrishnakone, P; Ho, Bow; Chow, Vincent T K


    Agkistrodon snake venoms contain a variety of phospholipases (PLA2), some of which are myotoxic. In this study, we used reverse-phase HPLC to purify PLA2 from the venom of Agkistrodon halys. The enzyme named as AgkTx-II, a basic Asp49 PLA2, has a molecular masses of 13,869.05. The amino acid sequence and molecular mass of AgkTx-II was identical to those of an Asp49 basic myotoxic PLA2 previously isolated from this venom. Antibacterial activities were tested by susceptibility and broth-dilution assays. AgkTx-II exerted a potent antibacterial activity against Staphylococcus aureus, Proteus vulgaris, Proteus mirabilis, and Burkholderia pseudomallei. The MIC values of AgkTx-II ranged between 85 and 2.76microM and was most effective against S. aureus, P. vulgaris, P. mirabilis (MIC of 21.25microM) and B. pseudomallei (MIC of 10.25microM). This AgkTx-II rapidly killed S. aureus, P. vulgaris and B. pseudomallei in a dose-dependent manner. The effect of the AgkTx-II on bacterial membranes was evaluated by scanning and transmission electron microscopy. AgkTx-II caused morphological alterations apparent on their cellular surfaces, suggesting a killing mechanism based on membrane permeabilization and damage. Cytotoxicity was measured by XTT tetrazolium (2,3-bis[2-methoxy-4-nitro-5-sulfophenyl]-2H-tetrazolium-5-carboxanilide) and lactate dehydrogenase (LDH) assays using U-937 cells (monocytes). The AgkTx-II did not affect cell viability up to 500microM concentrations but cell death was evident at 1000microM concentration after 24 and 48h. Furthermore, the repeated exposure of AgkTx-II (2-14microM) treated mice showed different tissue alterations, mainly at the brain and kidney; the toxicological potential of AgkTx-II remains to be elucidated. The AgkTx-II exhibits no hemolytic action even at high doses (10-100microM) in human erythrocytes. However, the AgkTx-II is believed to exert its bactericidal effect by permeabilizing the bacterial membrane by forming pores. In addition

  16. The characterization of trans-pecos copperhead (Agkistrodon contortrix pictigaster) venom and isolation of two new dimeric disintegrins.


    Lucena, Sara; Rodríguez-Acosta, Alexis; Grilli, Elyse; Alfonso, Andrea; Goins, Amber; Ogbata, Ifunanya; Walls, Robert; Suntravat, Montamas; Uzcátegui, Nestor L; Guerrero, Belsy; Sánchez, Elda E


    The vast amounts of toxins within the venom of snakes, while known to cause medical emergencies, display various biological functions. Trans-pecos copperhead (Agkistrodon contortrix pictigaster) crude venom separated by cation-exchange chromatography showed several fractions with fibrinolytic, hemorrhagic, gelatinase and platelet activities. Venom fractions 1, 2, 4, 5, and 12-17 contained fibrinolytic activity. Venom fractions 1, 2, 5 and 12-14 had hemorrhagic activity. Fractions 1, 2, 12, 13 and 17 contained gelatinase activity. Reverse-Phase C18 High Performance Liquid Chromatography was also used to purify and isolated disintegrins from this venom. Anti-platelet aggregation activity of the C18 fractions collected and performed on whole human blood showed that they inhibited platelet aggregation in presence of several agonists. Results from both SDS-PAGE and N-terminal sequencing determined that pictistatin 1 obtained from the Trans-Pecos copperhead venom was a dimeric disintegrin, and pictistatin 2 was a heterodimeric disintegrin. The molecules with anti-platelet activity could be considered in the development of more effective drugs, for numerous blood-related diseases such as stroke, heart attacks, thrombosis, and other medical conditions. In this study, we are presenting the first report of the purification, isolation, and partial characterization of two new dimeric disintegrins isolated from the venom of trans-pecos copperhead. PMID:27068364

  17. Rapid identification of thousands of copperhead snake (Agkistrodon contortrix) microsatellite loci from modest amounts of 454 shotgun genome sequence.


    Castoe, Todd A; Poole, Alexander W; Gu, Wanjun; Jason de Koning, A P; Daza, Juan M; Smith, Eric N; Pollock, David D


    Optimal integration of next-generation sequencing into mainstream research requires re-evaluation of how problems can be reasonably overcome and what questions can be asked. One potential application is the rapid acquisition of genomic information to identify microsatellite loci for evolutionary, population genetic and chromosome linkage mapping research on non-model and not previously sequenced organisms. Here, we report on results using high-throughput sequencing to obtain a large number of microsatellite loci from the venomous snake Agkistrodon contortrix, the copperhead. We used the 454 Genome Sequencer FLX next-generation sequencing platform to sample randomly ∼27 Mbp (128 773 reads) of the copperhead genome, thus sampling about 2% of the genome of this species. We identified microsatellite loci in 11.3% of all reads obtained, with 14 612 microsatellite loci identified in total, 4564 of which had flanking sequences suitable for polymerase chain reaction primer design. The random sequencing-based approach to identify microsatellites was rapid, cost-effective and identified thousands of useful microsatellite loci in a previously unstudied species. PMID:21565030

  18. Recombinant expression and affinity purification of snake venom gland parvalbumin in Escherichia coli.


    Jia, Ying; Pérez, John C


    Parvalbumins (PV) are small, acidic, water soluble and calcium-binding proteins generally present in muscular and nervous tissues. In the present study, we identified and characterized a cDNA clone encoding PV, named AplPV, from a snake (Agkistrodon piscivorus leucostoma) venom gland cDNA library. AplPV belongs to EF-hand proteins with six alpha-helices constituting three EF-hand domains. The deduced amino acid sequence of AplPV is 91% and 68% identical to the previously characterized PVs of Boa constrictor and Cyprinus carpio, respectively. The full-length cDNA was subcloned into the expression vector pGEX and transformed into Escherichia coli (E.coli) to produce recombinant protein. The bacterially expressed GST-AplPV fusion protein was highly expressed, and effectively purified by Glutathione-Sepharose affinity chromatography. A high concentration of thrombin protease specifically cleaved and removed the GST tag from fusion protein, and further purified by Benzamidine column for removal of thrombin protease. As a result, the 12 kDa AplPV recombinant protein alone was purified. To investigate the tissue-specific biological occurrence of AplPV, a polyclonal antibody (anti-AplPV-antibody) was raised against GST-AplPV fusion protein in rabbit. Western blot analysis revealed that immunoreactive bands were exhibited in both recombinant protein and samples of venom glands, but not in any crude venom. This specific occurrence indicates a specialized function of AplPV in snake venom glands. PMID:19275943



    Clancy, Meredith M; Newton, Alisa L; Sykes, John M


    An adult female Taylor's cantil (Agkistrodon bilineatus taylori) presented with marked spinal and mandibular osteomyelitis that cultured positive for Salmonella enterica subsp. houtenae, serovar IV 43:z4,z32:-. Progression of osteomyelitis was arrested by treatment using amikacin (0.026 mg/kg per hour) delivered via subcutaneous osmotic pump for 10 mo, replacing the pump every 4 wk. No adverse effects on renal function were appreciated throughout the course of therapy. Amikacin therapy was discontinued after improvement with treatment, but 5 mo later, bony lesions worsened, and an additional abscess formed at the previous pump site. The animal's condition declined and euthanasia was elected. Postmortem examination confirmed marked osteomyelitis with Salmonella infection of same serovar as the initial biopsy. This report highlights the pathogenicity of the S. enterica subsp. houtenae serovar and the ability to deliver effective amikacin dosage via osmotic pump to arrest osteomyelitis due to salmonellosis in a venomous snake. PMID:27468053

  20. [Clinical study of 48 cases of viper (Agkistrodon halys blomhoffii "Mamu shi") bite--Does the swelling at the first medical examination reflect the seriousness of the envenomation?].


    Naito, Hiromichi; Matsumoto, Kazuyuki; Nagae, Masaharu; Kasai, Shinya; Shiraishi, Kensuke; Morimoto, Naoki; Hagioka, Shingo; Sugiyama, Masatoshi


    We have evaluated 48 cases of viper (Agkistrodon halys blomhoffii "Mamushi") bite treated in our hospital from 2001 to 2005. Of all the patients, 21 were bitten in rice field/farm, 10 were in path, 9 were in yard and 8 were in other places. Most cases occurred between July and September. The most frequently bitten regions were fingers and toes. Because swelling reached its maximum at 1.1+/-0.5 day, it was difficult to judge the severity of Mamushi bite by the degree of swelling at the first examination. There was no significant relation between the grade classification of swelling and CPK values. Value of CPK became its peek at 2.0+/-1.1 days after bites. Some reports recommend giving antivenin judging from the grade classification. However we could not estimate the severity of Mamushi bite from the degree of swelling and CPK values at the first examination. The degree of swelling and CPK values at the first examination will not be an index to determine the choice of the treatment including use of the antivenin. PMID:17319499

  1. Wide distribution of cysteine-rich secretory proteins in snake venoms: isolation and cloning of novel snake venom cysteine-rich secretory proteins.


    Yamazaki, Yasuo; Hyodo, Fumiko; Morita, Takashi


    Cysteine-rich secretory proteins (CRISPs) are found in epididymis and granules of mammals, and they are thought to function in sperm maturation and in the immune system. Recently, we isolated and obtained clones for novel snake venom proteins that are classified as CRISP family proteins. To elucidate the distribution of snake venom CRISP family proteins, we evaluated a wide range of venoms for immuno-cross-reactivity. Then we isolated, characterized, and cloned genes for three novel CRISP family proteins (piscivorin, ophanin, and catrin) from the venom of eastern cottonmouth (Agkistrodon piscivorus piscivorus), king cobra (Ophiophagus hannah), and western diamondback rattlesnake (Crotalus atrox). Our results show the wide distribution of snake venom CRISP family proteins among Viperidae and Elapidae from different continents, indicating that CRISP family proteins compose a new group of snake venom proteins. PMID:12646276

  2. Apparent fatal snakebite in three hawks.


    Heckel, J O; Sisson, D C; Quist, C F


    Based on histories and gross and histologic findings, snakebite was diagnosed in three dead raptors submitted for post-mortem examination from northern Florida and southern Georgia (USA). Two immature red-tailed hawks (Buteo jamaicensis) were found next to dead and partially eaten venomous snakes. An adult Cooper's hawk (Accipiter cooperii) was found adjacent to a den containing a cotton-mouth (Agkistrodon piscivorus) and an eastern diamondback rattlesnake (Crotalus adamanteus). Gross findings in all birds consisted of hemorrhage and gangrenous necrosis of one limb. Severe muscular degeneration was seen histologically. PMID:7760504

  3. Facultative parthenogenesis in a critically endangered wild vertebrate.


    Fields, Andrew T; Feldheim, Kevin A; Poulakis, Gregg R; Chapman, Demian D


    Facultative parthenogenesis - the ability of sexually reproducing species to sometimes produce offspring asexually - is known from a wide range of ordinarily sexually reproducing vertebrates in captivity, including some birds, reptiles and sharks [1-3]. Despite this, free-living parthenogens have never been observed in any of these taxa in the wild, although two free-living snakes were recently discovered each gestating a single parthenogen - one copperhead (Agkistrodon contortrix) and one cottonmouth (Agkistrodon piscivorus) [1]. Vertebrate parthenogens are characterized as being of the homogametic sex (e.g., females in sharks, males in birds) and by having elevated homozygosity compared to their mother [1-3], which may reduce their viability [4]. Although it is unknown if either of the parthenogenetic snakes would have been carried to term or survived in the wild, facultative parthenogenesis might have adaptive significance [1]. If this is true, it is reasonable to hypothesize that parthenogenesis would be found most often at low population density, when females risk reproductive failure because finding mates is difficult [5]. Here, we document the first examples of viable parthenogens living in a normally sexually reproducing wild vertebrate, the smalltooth sawfish (Pristis pectinata). We also provide a simple approach to screen any microsatellite DNA database for parthenogens, which will enable hypothesis-driven research on the significance of vertebrate parthenogenesis in the wild. PMID:26035783

  4. Facultative parthenogenesis discovered in wild vertebrates

    PubMed Central

    Booth, Warren; Smith, Charles F.; Eskridge, Pamela H.; Hoss, Shannon K.; Mendelson, Joseph R.; Schuett, Gordon W.


    Facultative parthenogenesis (FP)—asexual reproduction by bisexual species—has been documented in a variety of multi-cellular organisms but only recently in snakes, varanid lizards, birds and sharks. Unlike the approximately 80 taxa of unisexual reptiles, amphibians and fishes that exist in nature, FP has yet to be documented in the wild. Based on captive documentation, it appears that FP is widespread in squamate reptiles (snakes, lizards and amphisbaenians), and its occurrence in nature seems inevitable, yet the task of detecting FP in wild individuals has been deemed formidable. Here we show, using microsatellite DNA genotyping and litter characteristics, the first cases of FP in wild-collected pregnant females and their offspring of two closely related species of North American pitviper snakes—the copperhead (Agkistrodon contortrix) and cottonmouth (Agkistrodon piscivorus). Our findings support the view that non-hybrid origins of parthenogenesis, such as FP, are more common in squamates than previously thought. With this confirmation, FP can no longer be viewed as a rare curiosity outside the mainstream of vertebrate evolution. Future research on FP in squamate reptiles related to proximate control of induction, reproductive competence of parthenogens and population genetics modelling is warranted. PMID:22977071

  5. Neurotropic effects of venoms and other factors that promote prey acquisition.


    Gennaro, Joseph Francis; Hall, Hollis Pace; Casey, Ernest Raymond; Hayes, William Kent


    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. PMID:17620305

  6. Environmental contaminants in Texas, USA, wetland reptiles: Evaluation using blood samples

    USGS Publications Warehouse

    Clark, D.R., Jr.; Bickham, J.W.; Baker, D.L.; Cowman, D.F.


    Four species of reptiles (diamondback water snake [Nerodia rhombifer], blotched water snake [N. erythrogaster], cottonmouth [Agkistrodon piscivorus], and red-eared slider [Trachemys scripta]) were collected at two contaminated and three reference sites in Texas, USA. Old River Slough has received intensive applications of agricultural chemicals since the 1950s. Municipal Lake received industrial arsenic wastes continuously from 1940 to 1993. Blood samples were analyzed for organochlorines, potentially toxic elements, genetic damage, and plasma cholinesterase (ChE). Dichlorodiphenyldichloroethylene (DDE) concentrations reached as high as 3.0 ppm (wet weight) in whole blood of a diamondback water snake at Old River Slough, a level probably roughly equivalent to the maximum concentration found in plasma of peregrine falcons (Falco peregrinus) in 1978 to 1979 when DDE peaked in this sensitive species. Possible impacts on diamondback water snakes are unknown, but at least one diamondback water snake was gravid when captured, indicating active reproduction. Arsenic was not found in red-eared sliders (only species sampled) from Municipal Lake. Red-eared sliders of both sexes at Old River Slough showed declining levels of ChE with increasing mass, suggesting a life-long decrease of ChE levels. Possible negative population consequences are unknown, but no evidence was found in body condition (mass relative to carapace length) that red-eared sliders at either contaminated site were harmed.


    PubMed Central



    Uranotaenia sapphirina, Culex erraticus, and Cx. peccator were collected in an enzootic eastern equine encephalomyelitis (EEE) virus focus in central Alabama (Tuskegee National Forest) from 2001 to 2003 and analyzed for virus as well as host selection. EEE virus was detected in each species every year except 2003, when pools of Cx. peccator were negative. Most (97%) of the 130 Cx. peccator blood meals identified were from ectothermic hosts; 3% were from birds. Among blood meals from reptiles (approximately 75% of the total), 81% were from Agkistrodon piscivorus (cottonmouth); all amphibian blood meals (approximately 25%) were from Rana spp. with > 50% taken from the bullfrog R. catesbeiana. Host identifications were made from 131 of 197 Cx. erraticus, but only 3 (2%) were derived from ectothermic species. Identification of Ur. sapphirina blood meals proved difficult and only 2 of 35 hosts were determined. Both were from R. catesbeiana. Ectothermic species are possible EEE virus reservoirs in the southeastern United States where species such as Cx. peccator and Ur. sapphirina occur with large, diverse reptilian, amphibian, and avian populations such as those at the Tuskegee site. PMID:15381805

  8. Comparative mitochondrial genomics of snakes: extraordinary substitution rate dynamics and functionality of the duplicate control region

    PubMed Central

    Jiang, Zhi J; Castoe, Todd A; Austin, Christopher C; Burbrink, Frank T; Herron, Matthew D; McGuire, Jimmy A; Parkinson, Christopher L; Pollock, David D


    Background The mitochondrial genomes of snakes are characterized by an overall evolutionary rate that appears to be one of the most accelerated among vertebrates. They also possess other unusual features, including short tRNAs and other genes, and a duplicated control region that has been stably maintained since it originated more than 70 million years ago. Here, we provide a detailed analysis of evolutionary dynamics in snake mitochondrial genomes to better understand the basis of these extreme characteristics, and to explore the relationship between mitochondrial genome molecular evolution, genome architecture, and molecular function. We sequenced complete mitochondrial genomes from Slowinski's corn snake (Pantherophis slowinskii) and two cottonmouths (Agkistrodon piscivorus) to complement previously existing mitochondrial genomes, and to provide an improved comparative view of how genome architecture affects molecular evolution at contrasting levels of divergence. Results We present a Bayesian genetic approach that suggests that the duplicated control region can function as an additional origin of heavy strand replication. The two control regions also appear to have different intra-specific versus inter-specific evolutionary dynamics that may be associated with complex modes of concerted evolution. We find that different genomic regions have experienced substantial accelerated evolution along early branches in snakes, with different genes having experienced dramatic accelerations along specific branches. Some of these accelerations appear to coincide with, or subsequent to, the shortening of various mitochondrial genes and the duplication of the control region and flanking tRNAs. Conclusion Fluctuations in the strength and pattern of selection during snake evolution have had widely varying gene-specific effects on substitution rates, and these rate accelerations may have been functionally related to unusual changes in genomic architecture. The among-lineage and

  9. A Novel Direct Factor Xa Inhibitory Peptide with Anti-Platelet Aggregation Activity from Agkistrodon acutus Venom Hydrolysates

    PubMed Central

    Chen, Meimei; Ye, Xiaohui; Ming, Xin; Chen, Yahui; Wang, Ying; Su, Xingli; Su, Wen; Kong, Yi


    Snake venom is a natural substance that contains numerous bioactive proteins and peptides, nearly all of which have been identified over the last several decades. In this study, we subjected snake venom to enzymatic hydrolysis to identify previously unreported bioactive peptides. The novel peptide ACH-11 with the sequence LTFPRIVFVLG was identified with both FXa inhibition and anti-platelet aggregation activities. ACH-11 inhibited the catalytic function of FXa towards its substrate S-2222 via a mixed model with a Ki value of 9.02 μM and inhibited platelet aggregation induced by ADP and U46619 in a dose-dependent manner. Furthermore, ACH-11 exhibited potent antithrombotic activity in vivo. It reduced paralysis and death in an acute pulmonary thrombosis model by 90% and attenuated thrombosis weight in an arterio-venous shunt thrombosis model by 57.91%, both at a dose of 3 mg/kg. Additionally, a tail cutting bleeding time assay revealed that ACH-11 did not prolong bleeding time in mice at a dose of 3 mg/kg. Together, our results reveal that ACH-11 is a novel antithrombotic peptide exhibiting both FXa inhibition and anti-platelet aggregation activities, with a low bleeding risk. We believe that it could be a candidate or lead compound for new antithrombotic drug development. PMID:26035670

  10. Valsalnicola D. Walker & Rossman, gen. nov.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Valsonectria is based on a species that was described in the genus Valsa. Although it resembles Valsa in having allantoid ascospores, the ascospores of Valsalnicola are one-septate while the majority of species of Valsa and the closely related Leucostoma and Valsella have non-septate ascospores. How...

  11. Unusual snail species involved in the transmission of Fasciola hepatica in watercress beds in central France.


    Dreyfuss, G; Vignoles, P; Abrous, M; Rondelaud, D


    Four freshwater pulmonate species (Lymnaea ovata, L. stagnalis, Physa acuta, Planorbis leucostoma) were living in several watercress beds known for their relationships with human cases of fasciolosis, whereas L. truncatula was never found. The aims of these studies were to determine the prevalence of natural infections with Fasciola hepatica in snails and to verify if these species might ensure the full larval development of this trematode (with cercarial shedding) when they were experimentally subjected to F. hepatica only, or to co-infections with an other trematode species. Investigations were so carried out in six snail populations living in watercress beds (including three for P. acuta) and in four others originating from three brooks or a pond (as controls). Snails naturally infected with F. hepatica were found in two watercress beds inhabited by L. ovata (prevalence of infection: 1.4%) and P. leucostoma (0.1%), respectively. The L. ovata from the watercress bed could be infected at a higher size than those from the control population and the prevalence of this infection was greater in the bed population. Similar findings were noted for L. stagnalis. Despite single or dual infections, the results obtained with the four populations of P. acuta were unsuccessful. In contrast, the co-infections of young P. leucostoma with Paramphistomum daubneyi and F. hepatica resulted in the shedding of some F. hepatica cercariae. According to the authors, the occurrence of fasciolosis in these watercress beds would be the consequence of frequent natural encounters between parasite and snails (L. ovata, L. stagnalis), or of co-infections with P. daubneyi and F. hepatica (P. leucostoma). In watercress beds only colonized by P. acuta, a lymnaeid species would have ensured the larval development of F. hepatica but it would have been eliminated by P. acuta, as this last species was known to be invasive and could colonize open drainage ditches on siliceous soil. PMID:12116856

  12. First record of the genus Acropimpla Townes, 1960 (Hymenoptera: Ichneumonidae: Pimplinae) from Thailand, with descriptions of three new species.


    Varga, Oleksandr; Reshchikov, Alexey


    Nine species of Acropimpla Townes, 1960 are recorded from Thailand with three species described as new: A. leoni sp.n., A. natae sp.n., A. patellalutea sp.n. Six species, A. aspera Gupta & Tikar, 1976, A. hapaliae (Rao, 1953), A. leucostoma (Cameron, 1907), A. nakula Gupta & Tikar, 1976, A. taishunensis Liu, He & Chen, 2010, and A. uchidai (Cushman, 1933) are recorded from Thailand for the first time. A key to the females of Acropimpla species in South East Asia is also provided. PMID:26623916

  13. [Expression of snake venom thrombin-like enzyme calobin in Pichia pastoris].


    Yuan, Shengling; Wang, Peng; Tao, Haoxia; Zhan, Dewen; Wang, Yanchun; Wang, Lingchun; Liu, Chunjie; Zhang, Zhaoshan


    Thrombin-like enzymes (TLEs) are studied widely because of their therapeutic potential in myocardial infarction and thrombotic diseases. We synthesized the DNA fragment encoding thrombin-like enzyme calobin from Agkistrodon caliginosus (Korean Viper) venom by fusion PCR and expressed it in Pichia pastoris. After induction by 0.5% methanol for 48 h, the expression level of recombinant calobin reached 3.5 g/L in medium. The recombinant calobin was purified by Q-Sepharose Fast Flow ion-exchange chromatography and Sephacryl-S-100 gel filtration chromatography. Purified sample had a molecular weight of 32 kD shown in SDS-PAGE. It hydrolyzed fibrinogen and formed a light white hydrolysis circle in fibrinogen plate. SDS-PAGE analysis showed that recombinant calobin cleaved Aalpha-chain of fibrinogen specifically, and produced an appropriately 40 kD new band. However, we failed to find its fibrin-clot formation activity. PMID:19637626

  14. The proteolytic action of Arvin on human fibrinogen

    PubMed Central

    Ewart, M. R.; Hatton, M. W. C.; Basford, J. M.; Dodgson, K. S.


    1. Human fibrinogen was subjected to proteolysis by enzyme preparations (clinical Arvin and IRC-50 Arvin) from the venom of Agkistrodon rhodostoma. 2. IRC-50 Arvin releases three peptides from fibrinogen, and these were identified as fibrinopeptides AP, AY and A. 3. The less purified `clinical' Arvin releases, in addition to fibrinopeptides AP, AY and A, small amounts of two heptapeptides derived from fibrinopeptides AP and A, probably because it contains another enzyme as well as Arvin. 4. No fibrinopeptide B is released by either Arvin preparation. 5. Thus, although Arvin is known to differ from `reptilase' from Bothrops jararaca in that it does not activate the enzyme that cross-links fibrin (fibrin-stabilizing factor), it is identical with reptilase with respect to the peptides that it liberates from fibrinogen. PMID:5529716

  15. Identification and functional analysis of a novel bradykinin inhibitory peptide in the venoms of New World Crotalinae pit vipers

    SciTech Connect

    James Graham, Robert Leslie . E-mail:; Graham, Ciaren; McClean, Stephen; Chen, Tianbao; O'Rourke, Martin; Hirst, David; Theakston, David; Shaw, Chris


    A novel undecapeptide has been isolated and structurally characterized from the venoms of three species of New World pit vipers from the subfamily, Crotalinae. These include the Mexican moccasin (Agkistrodon bilineatus), the prairie rattlesnake (Crotalus viridis viridis), and the South American bushmaster (Lachesis muta). The peptide was purified from all three venoms using a combination of gel permeation chromatography and reverse-phase HPLC. Automated Edman degradation sequencing and MALDI-TOF mass spectrometry established its peptide primary structure as: Thr-Pro-Pro-Ala-Gly-Pro-Asp-Val-Gly-Pro-Arg-OH, with a non-protonated molecular mass of 1063.18 Da. A synthetic replicate of the peptide was found to be an antagonist of bradykinin action at the rat vascular B2 receptor. This is the first bradykinin inhibitory peptide isolated from snake venom. Database searching revealed the peptide to be highly structurally related (10/11 residues) with a domain residing between the bradykinin-potentiating peptide and C-type natriuretic peptide domains of a recently cloned precursor from tropical rattlesnake (Crotalus durissus terrificus) venom gland. BIP thus represents a novel biological entity from snake venom.

  16. Interaction of toxic venoms with the complement system

    PubMed Central

    Birdsey, Vanessa; Lindorfer, Jean; Gewurz, H.


    Thirty-nine venoms from various vertebrate and invertebrate species were tested for their ability to consume haemolytic complement (C) activity upon incubation in fresh guinea-pig serum. Nineteen had `anti-complementary' activity, and these were provisionally sorted into the following groups: Pattern I—exemplified by the Naja haje (Egyptian cobra) and six other Elapidae species (all cobras), which induced selective consumption of C3—C9, and led to formation of a stable C3—C9-consuming intermediate; Pattern II—exemplified by the Agkistrodon rhodostoma (Malayan pit viper), Bitis arietans (puff adder), Bothrops jararaca (South American pit viper), Bothrops atrox (Fer de Lance) and three other species, which induced marked consumption of C4 and C2, as well as C3—C9, but did not form a stable C3—C9-consuming intermediate; and individual animals, e.g. the Lachesis muta (bushmaster), which induced other patterns (III—VI) of complement component consumption. Active fractions of representative venoms were partially purified by ion exchange and gel filtration chromatography and their interactions with the complement system characterized further. It is anticipated that these enzymes, with a capacity to activate the complement system in unique ways, will prove to be of further experimental usefulness. PMID:4398349

  17. Balancing the Expression and Production of a Heterodimeric Protein: Recombinant Agkisacutacin as a Novel Antithrombotic Drug Candidate.


    Guo, Yugang; Wu, Jing; Jia, Hao; Chen, Wei; Shao, Changsheng; Zhao, Lei; Ma, Jiajia; Li, Rui; Zhong, Yongjun; Fang, Fang; Wang, Dong; Sun, Jie; Qian, Fang; Dai, Xiangrong; Zhang, Guohui; Tian, Zhigang; Xiaoyi Li, Benjamin; Xiao, Weihua


    Agkisacucetin extracted from the venom of Agkistrodon acutus has been demonstrated to be a promising antithrombotic drug candidate in clinical studies due to its function as a novel platelet membrane glycoprotein (GP) Ib inhibitor. Agkisacucetin is a heterodimeric protein composed of α- and β-subunits with seven disulphide bonds. Both subunits form inactive homodimeric products, which cause difficulties for recombinant production. In this study, Agkisacucetin α- and β-subunits were inserted sequentially into the chromosome of Pichia pastoris at the mutant histidinol dehydrogenase gene and ribosomal DNA repeat sites, respectively. By optimizing the gene copies and productivity of each subunit by drug screening, we successfully obtained a recombinant strain with balanced expression of the two subunits. Using this strain, a yield greater than 100 mg/L recombinant Agkisacucetin in fed-batch fermentation was reached. The recombinant Agkisacucetin possessed extremely similar binding affinity to recombinant GPIb and human platelets in in vitro assays, and its ristocetin-induced platelet aggregation activity ex vivo was identical to that of the extracted native Agkisacucetin, demonstrating that the yeast-derived Agkisacucetin could be an effective alternative to native Agkisacucetin. Moreover, this study provides an effective strategy for balancing the expression and production of heterodimeric proteins in P. pastoris. PMID:26144864

  18. Co-overexpression of PpPDI enhances secretion of ancrod in Pichia pastoris.


    Zhang, Shou-Tao; Fang, Hui-Min; Zhao, Li; Tian, Qing-Nan; Qin, Yun-Fei; Lu, Ping; Chen, San-Jun; Bao, Zhen-Xia; Liang, Feng


    Ancrod, a serine protease purified from the venom of Agkistrodon rhodostoma, is highly specific for fibrinogen. It causes anticoagulation by defibrinogenation and has been used as a therapeutic anticoagulant for the treatment of moderate to severe forms of peripheral arterial circulatory disorders in a variety of countries. The DNA of ancrod was amplified by recursive PCR with a yeast bias codon and cloned into the pGEM-T Easy vector. In order to achieve a high level secretion and a full activity expression of ancrod in Pichia pastoris (P. pastoris), the P. pastoris protein disulfide bond isomerase (PpPDI) was co-overexpressed in the strain. The secretion characteristics of ancrod with and without PpPDI were examined. With co-overexpression of PpPDI, the production of recombinant ancrod (rAncrod) was increased to 315 mg/L in the culture medium, which is twofold higher than the control strain carrying only the ancrod gene. Through purified by Ni²⁺ affinity chromatography and phenyl Sepharose column, the purity of rAncrod was found to be as high as 95.2%. The fibrinogenolytic and zymographic activities of the rAncrod were determined and found to be similar to that of the native protein. This improved expression system can facilitate further studies and the industrial production of ancrod. PMID:21340538

  19. Discovery of Larval Gnathostoma nipponicum in Frogs and Snakes from Jeju-do (Province), Republic of Korea

    PubMed Central

    Woo, Ho-Choon; Oh, Hong-Shik; Cho, Shin-Hyeong; Na, Byoung-Kuk


    A survey was performed to find out the intermediate hosts of Gnathostoma nipponicum in Jeju-do (Province), the Republic of Korea. In August 2009 and 2010, a total of 82 tadpoles, 23 black-spotted pond frogs (Rana nigromaculata), 7 tiger keelback snakes (Rhabdophis tigrinus tigrinus), 6 red-tongue viper snakes (Agkistrodon ussuriensis), and 2 cat snakes (Elaphe dione) were collected in Jeju-do and examined by the pepsin-HCl digestion method. Total 5 gnathostome larvae were detected in 3 (50%) of 6 A. ussuriensis, 70 larvae in 3 of 7 (42.9%) R. tigrinus tigrinus, and 2 larvae in 2 of 82 (8.7%) frogs. No gnathostome larvae were detected in tadpoles and cat snakes. The larvae detected were a single species, and 2.17×0.22 mm in average size. They had characteristic head bulbs, muscular esophagus, and 4 cervical sacs. Three rows of hooklets were arranged in the head bulbs, and the number of hooklets in each row was 29, 33, and 36 posteriorly. All these characters were consistent with the advanced third-stage larvae of G. nipponicum. It has been first confirmed in Jeju-do that R. nigromaculata, A. ussuriensis, and R. tigrinus tigrinus play a role for intermediate and/or paratenic hosts for G. nipponicum. PMID:22355217

  20. Purification and cloning of cysteine-rich proteins from Trimeresurus jerdonii and Naja atra venoms.


    Jin, Yang; Lu, Qiumin; Zhou, Xingding; Zhu, Shaowen; Li, Rui; Wang, Wanyu; Xiong, Yuliang


    Three 26 kDa proteins, named as TJ-CRVP, NA-CRVP1 and NA-CRVP2, were isolated from the venoms of Trimeresurus jerdonii and Naja atra, respectively. The N-terminal sequences of TJ-CRVP and NA-CRVPs were determined. These components were devoid of the enzymatic activities tested, such as phospholipase A(2), arginine esterase, proteolysis, L-amino acid oxidase, 5'nucleotidase, acetylcholinesterase. Furthermore, these three components did not have the following biological activities: coagulant and anticoagulant activities, lethal activity, myotoxicity, hemorrhagic activity, platelet aggregation and platelet aggregation-inhibiting activities. These proteins are named as cysteine-rich venom protein (CRVP) because their sequences showed high level of similarity with mammalian cysteine-rich secretory protein (CRISP) family. Recently, some CRISP-like proteins were also isolated from several different snake venoms, including Agkistrodon blomhoffi, Trimeresurus flavoviridis, Lanticauda semifascita and king cobra. We presumed that CRVP might be a common component in snake venoms. Of particular interest, phylogenetic analysis and sequence alignment showed that NA-CRVP1 and ophanin, both from elapid snakes, share higher similarity with CRVPs from Viperidae snakes. PMID:14529736

  1. Quantitative investigation of two metallohydrolases by X-ray absorption spectroscopy near-edge spectroscopy

    NASA Astrophysics Data System (ADS)

    Zhao, W.; Chu, W. S.; Yang, F. F.; Yu, M. J.; Chen, D. L.; Guo, X. Y.; Zhou, D. W.; Shi, N.; Marcelli, A.; Niu, L. W.; Teng, M. K.; Gong, W. M.; Benfatto, M.; Wu, Z. Y.


    The last several years have witnessed a tremendous increase in biological applications using X-ray absorption spectroscopy (BioXAS), thanks to continuous advancements in synchrotron radiation (SR) sources and detector technology. However, XAS applications in many biological systems have been limited by the intrinsic limitations of the Extended X-ray Absorption Fine Structure (EXAFS) technique e.g., the lack of sensitivity to bond angles. As a consequence, the application of the X-ray absorption near-edge structure (XANES) spectroscopy changed this scenario that is now continuously changing with the introduction of the first quantitative XANES packages such as Minut XANES (MXAN). Here we present and discuss the XANES code MXAN, a novel XANES-fitting package that allows a quantitative analysis of experimental data applied to Zn K-edge spectra of two metalloproteins: Leptospira interrogans Peptide deformylase ( LiPDF) and acutolysin-C, a representative of snake venom metalloproteinases (SVMPs) from Agkistrodon acutus venom. The analysis on these two metallohydrolases reveals that proteolytic activities are correlated to subtle conformation changes around the zinc ion. In particular, this quantitative study clarifies the occurrence of the LiPDF catalytic mechanism via a two-water-molecules model, whereas in the acutolysin-C we have observed a different proteolytic activity correlated to structural changes around the zinc ion induced by pH variations.

  2. Thousands of microsatellite loci from the venomous coralsnake (Micrurus fulvius) and variability of select loci across populations and related species

    PubMed Central

    Castoe, Todd A.; Streicher, Jeffrey W.; Meik, Jesse M.; Ingrasci, Matthew J.; Poole, Alexander W.; de Koning, A.P. Jason; Campbell, Jonathan A.; Parkinson, Christopher L.; Smith, Eric N.; Pollock, David D.


    Studies of population genetics increasingly use next-generation DNA sequencing to identify microsatellite loci in non-model organisms. There are, however, relatively few studies that validate the feasibility of transitioning from marker development to experimental application across populations and species. North American coralsnakes of the Micrurus fulvius species complex occur in the United States and Mexico, and little is known about their population structure and phylogenetic relationships. This absence of information and population genetics markers is particularly concerning because they are highly venomous and have important implications on human health. To alleviate this problem in coralsnakes, we investigated the feasibility of using 454 shotgun sequences for microsatellite marker development. First, a genomic shotgun library from a single individual was sequenced (~7.74 megabases; 26,831 reads) to identify potentially amplifiable microsatellite loci (PALs). We then hierarchically sampled 76 individuals from throughout the geographic distribution of the species complex and examined whether PALs were amplifiable and polymorphic. Approximately half of the loci tested were readily amplifiable from all individuals, and 80% of the loci tested for variation were variable and thus informative as population genetic markers. To evaluate the repetitive landscape characteristics across multiple snakes, we also compared microsatellite content between the coralsnake and two other previously sampled snakes, the venomous copperhead (Agkistrodon contortrix) and Burmese python (Python molurus). PMID:22938699

  3. Bioenergetic components of reproductive effort in viviparous snakes: costs of vitellogenesis exceed costs of pregnancy.


    Van Dyke, James U; Beaupre, Steven J


    Reproductive effort has been defined as the proportion of an organism's energy budget that is allocated to reproduction over a biologically meaningful time period. Historically, studies of reproductive bioenergetics considered energy content of gametes, but not costs of gamete production. Although metabolic costs of vitellogenesis (MCV) fundamentally reflect the primary bioenergetic cost of reproductive allocation in female reptiles, the few investigations that have considered costs of reproductive allocation have focused on metabolic costs of pregnancy (MCP) in viviparous species. We define MCP as energetic costs incurred by pregnant females, including all costs of maintaining gestation conditions necessary for embryogenesis. MCP by our definition do not include fetal costs of embryogenesis. We measured metabolic rates in five species of viviparous snakes (Agkistrodon contortrix, Boa constrictor, Eryx colubrinus, Nerodia sipedon, and Thamnophis sirtalis) during vitellogenesis and pregnancy in order to estimate MCV and MCP. Across all species, MCV were responsible for 30% increases in maternal metabolism. Phylogenetically-independent contrasts showed that MCV were significantly greater in B. constrictor than in other species, likely because B. constrictor yolk energy content was greater than that of other species. Estimates of MCP were not significantly different from zero in any species. In viviparous snakes, MCV appear to represent significant bioenergetic expenditures, while MCP do not. We suggest that MCV, together with yolk energy content, represent the most significant component of reptilian reproductive effort, and therefore deserve greater attention than MCP in studies of reptilian reproductive bioenergetics. PMID:21884815

  4. Agglucetin, a tetrameric C-type lectin-like venom protein, regulates endothelial cell survival and promotes angiogenesis by activating integrin {alpha}v{beta}3 signaling

    SciTech Connect

    Wang, W.-J.


    Agglucetin, a platelet glycoprotein (GP)Ib binding protein from Formosan Agkistrodon acutus (A. acutus) venom, could sustain human umbilical vein endothelial cell (HUVEC) proliferation and HUVEC adhering to immobilized agglucetin showed extensive spreading, which was strongly abrogated by integrin antagonists 7E3 and triflavin. Flow cytometric analyses confirmed the expression of GPIb complex on HUVEC is absent and fluorescein isothiocyanate (FITC)-agglucetin binds to HUVEC in a dose-dependent and saturable manner. Furthermore, native agglucetin specifically and dose-dependently inhibited the binding of FITC-23C6, an anti-{alpha}v{beta}3 monoclonal antibody (mAb), but not antibodies against {alpha}2 and {alpha}5, toward HUVEC and purified {alpha}v{beta}3 also bound to immobilized agglucetin-{beta} in a dose-dependent manner. Moreover, agglucetin exhibited a pro-angiogenic effect in vitro, as well as the focal adhesion kinase (FAK)-associated signaling molecules responsible for HUVEC activation were initiated by agglucetin. In conclusion, agglucetin, acting as a survival factor, promotes endothelial adhesion and angiogenesis by triggering {alpha}v{beta}3 signaling through FAK/phosphatidylinositol 3-kinase (PI3K)/Akt pathway.

  5. Stable isotope tracer reveals that viviparous snakes transport amino acids to offspring during gestation.


    Van Dyke, James U; Beaupre, Steven J


    Viviparity and placentation have evolved from oviparity over 100 times in squamate reptiles (lizards and snakes). The independent origins of placentation have resulted in a variety of placental morphologies in different taxa, ranging from simple apposition of fetal and maternal tissues to endotheliochorial implantation that is homoplasious with mammalian placentation. Because the eggs of oviparous squamates transport gases and water from the environment and calcium from the eggshell, the placentae of viviparous squamates are thought to have initially evolved to accomplish these functions from within the maternal oviduct. Species with complex placentae have also been shown to rely substantially, or even primarily, on placental transport of organic nutrients for embryonic nutrition. However, it is unclear whether species with only simple placentae are also capable of transporting organic nutrients to offspring. Among viviparous squamates, all of the snakes that have been studied thus far have been shown to have simple placentae. However, most studies of snake placentation are limited to a single lineage, the North American Natricinae. We tested the abilities of four species of viviparous snakes - Agkistrodon contortrix (Viperidae), Boa constrictor (Boidae), Nerodia sipedon (Colubridae: Natricinae) and Thamnophis sirtalis (Colubridae: Natricinae) - to transport diet-derived amino acids to offspring during gestation. We fed [(15)N]leucine to pregnant snakes, and compared offspring (15)N content with that of unlabeled controls. Labeled females allocated significantly more (15)N to offspring than did controls, but (15)N allocation did not differ among species. Our results indicate that viviparous snakes are capable of transporting diet-derived amino acids to their offspring during gestation, possibly via placentation. PMID:22323198

  6. Landscape-level influences of terrestrial snake occupancy within the southeastern United States.


    Steen, David A; McClure, Christopher J W; Brock, Jean C; Rudolph, D Craig; Pierce, Josh B; Lee, James R; Humphries, W Jeffrey; Gregory, Beau B; Sutton, William B; Smith, Lora L; Baxley, Danna L; Stevenson, Dirk J; Guyer, Craig


    Habitat loss and degradation are thought to be the primary drivers of species extirpations, but for many species we have little information regarding specific habitats that influence occupancy. Snakes are of conservation concern throughout North America, but effective management and conservation are hindered by a lack of basic natural history information and the small number of large-scale studies designed to assess general population trends. To address this information gap, we compiled detection/nondetection data for 13 large terrestrial species from 449 traps located across the southeastern United States, and we characterized the land cover surrounding each trap at multiple spatial scales (250-, 500-, and 1000-m buffers). We used occupancy modeling, while accounting for heterogeneity in detection probability, to identify habitat variables that were influential in determining the presence of a particular species. We evaluated 12 competing models for each species, representing various hypotheses pertaining to important habitat features for terrestrial snakes. Overall, considerable interspecific variation existed in important habitat variables and relevant spatial scales. For example, kingsnakes (Lampropeltis getula) were negatively associated with evergreen forests, whereas Louisiana pinesnake (Pituophis ruthveni) occupancy increased with increasing coverage of this forest type. Some species were positively associated with grassland and scrub/shrub (e.g., Slowinski's cornsnake, Elaphe slowinskii) whereas others, (e.g., copperhead, Agkistrodon contortrix, and eastern diamond-backed rattlesnake, Crotalus adamanteus) were positively associated with forested habitats. Although the species that we studied may persist in varied landscapes other than those we identified as important, our data were collected in relatively undeveloped areas. Thus, our findings may be relevant when generating conservation plans or restoration goals. Maintaining or restoring landscapes that

  7. Southern copperhead venom enhances tissue-type plasminogen activator induced fibrinolysis but does not directly lyse human plasma thrombi.


    Nielsen, Vance G


    In addition to degrading fibrinogen as a source of consumptive coagulopathy, purified fractions of southern copperhead (Agkistrodon contortrix contortrix; A. c. contortrix) venom has been demonstrated to enhance fibrinolysis. The goal of this investigation was to characterize the kinetic fibrinolytic profile of A. c. contortrix venom in the absence and presence of tissue-type plasminogen activator (tPA) to determine if intact venom had tPA independent fibrinolytic properties. Utilizing thrombelastographic methods, the coagulation and fibrinolytic kinetic profiles of human plasma exposed to A. c. contortrix venom (0-6 μg/ml) were determined in the absence or presence of tPA (0-100 IU/ml). Then, plasma was exposed to 0-6 μg/ml of venom without tPA added and coagulation observed for 3 h. Venom significantly prolonged the onset of coagulation, decreased the velocity of thrombus growth but did not significantly decrease clot strength. In the presence of tPA, venom significantly decreased clot strength, shortened the time of onset of fibrinolysis, decreased clot lysis time but did not significantly affect the maximum rate of lysis. Lastly, while venom exposure in the absence of tPA significantly prolonged the onset of coagulation and decreased the velocity of clot growth, venom exposure did not result in detectable fibrinolysis over the 3 h observation period. A. c. contortrix venom enhances tPA mediated fibrinolysis by degrading plasma coagulation kinetics. Intact A. c. contortrix venom does not possess sufficient fibrinolytic activity to cause fibrinolysis in human plasma at the concentration tested. PMID:26407681

  8. Multi-host Model-Based Identification of Armillifer agkistrodontis (Pentastomida), a New Zoonotic Parasite from China

    PubMed Central

    Chen, Shao-Hong; Liu, Qin; Zhang, Yong-Nian; Chen, Jia-Xu; Li, Hao; Chen, Ying; Steinmann, Peter; Zhou, Xiao-Nong


    Background Pentastomiasis is a rare parasitic infection of humans. Pentastomids are dioecious obligate parasites requiring multiple hosts to complete their lifecycle. Despite their worm-like appearance, they are commonly placed into a separate sub-class of the subphylum Crustacea, phylum Arthropoda. However, their systematic position is not uncontested and historically, they have been considered as a separate phylum. Methodology/Principal Findings An appraisal of Armillifer agkistrodontis was performed in terms of morphology and genetic identification after its lifecycle had been established in a multi-host model, i.e., mice and rats as intermediate hosts, and snakes (Agkistrodon acutus and Python molurus) as definitive hosts. Different stages of the parasite, including eggs, larvae and adults, were isolated and examined morphologically using light and electron microscopes. Phylogenetic and cluster analysis were also undertaken, focusing on the 18S rRNA and the Cox1 gene. The time for lifecycle completion was about 14 months, including 4 months for the development of eggs to infectious larvae in the intermediate host and 10 months for infectious larvae to mature in the final host. The main morphological difference between A. armillatus and Linguatula serrata is the number of abdominal annuli. Based on the 18S rRNA sequence, the shortest hereditary distance was found between A. agkistrodontis and Raillietiella spp. The highest degree of homology in the Cox 1 nucleic acid sequences and predicted amino acid sequences was found between A. agkistrodontis and A. armillatus. Conclusion This is the first time that a multi-host model of the entire lifecycle of A. agkistrodontis has been established. Morphologic and genetic analyses supported the notion that pentastomids should be placed into the phylum Arthropoda. PMID:20386597

  9. Genetic Identification of Spirometra decipiens Plerocercoids in Terrestrial Snakes from Korea and China

    PubMed Central

    Jeon, Hyeong-Kyu; Park, Hansol; Lee, Dongmin; Choe, Seongjun; Kim, Kyu-Heon; Sohn, Woon-Mok; Eom, Keeseon S.


    Human sparganosis is a zoonotic disease caused by infection with larval forms (procercoid/plerocercoid) of Spirometra spp. The purpose of this study was to identify Spirometra spp. of infected snakes using a multiplex PCR assay and phylogenetic analysis of mitochondrial DNA sequence data from the spargana of terrestrial snakes obtained from Korea and China. A total of 283 snakes were obtained that included 4 species of Colubridae comprising Rhabdophis tigrinus tigrinus (n=150), Dinodon rufozonatum rufozonatum (n=64), Elaphe davidi (n=2), and Elaphe schrenkii (n=7), and 1 species of Viperidae, Agkistrodon saxatilis (n=60). The snakes were collected from the provinces of Chungbuk, Chungnam, and Gyeongbuk in Korea (n=161), and from China (n=122). The overall infection rate with spargana was 83% (235/283). The highest was recorded for D. rufozonatum rufozonatum (100%), followed by A. saxatilis (85%) and R. tigrinus tigrinus (80%), with a negative result for E. davidi (0%) and E. schrenkii (0%). The sequence identities between the spargana from snakes (n=50) and Spirometra erinaceieuropaei (KJ599680) or S. decipiens (KJ599679) control specimens were 90.8% and 99.2%, respectively. Pairwise genetic distances between spargana (n=50) and S. decipiens ranged from 0.0080 to 0.0107, while those between spargana and S. erinaceieuropaei ranged from 0.1070 to 0.1096. In this study, all of the 904 spargana analyzed were identified as S. decipiens either by a multiplex PCR assay (n=854) or mitochondrial cox1 sequence analysis (n=50). PMID:27180576

  10. Increase of the cytotoxic effect of Bothrops jararacussu venom on mouse extensor digitorum longus and soleus by potassium channel blockers and by Na(+)/K(+)-ATPase inhibition.


    Tomaz, Marcelo A; Fernandes, Fabrício F A; El-Kik, Camila Z; Moraes, Raphael A M; Calil-Elias, Sabrina; Saturnino-Oliveira, Jeison; Martinez, Ana Maria B; Ownby, Charlotte L; Melo, Paulo A


    We investigated the myotoxicity of Bothrops jararacussu crude venom and other cytolytic agents on mouse isolated extensor digitorum longus (EDL) and soleus (SOL) muscles, which present distinct properties: EDL is a fast-twitch, white muscle with predominantly glycolytic fibers, while SOL is slow-twitch, red muscle with predominantly oxidative fibers. Muscles were exposed to B. jararacussu crude venom (25 microg/ml) and other crotaline venoms (Agkistrodon contortrix laticinctus; Crotalus viridis viridis; Crotalus durissus terrificus) at the same concentration. Basal creatine kinase (CK) release to bathing solution was 0.43+/-0.06 for EDL and 0.29+/-0.06 for SOL (U g(-)(1) h(-)(1), n=36 for each muscle). Sixty minutes after exposure to B. jararacussu venom, EDL presented higher increase in the rate of CK release than SOL, respectively, 13.2+/-1.5 and 2.9+/-0.7 U g(-)(1)h(-)(1), n=10-12. Muscle denervation, despite decreasing CK content, did not affect sensitivities to B. jararacussu venom. Ouabain and potassium channel blockers (TEA; clotrimazole; glibenclamide) increased the rate of CK release by B. jararacussu in EDL and SOL muscles, decreasing and almost abolishing the different sensitivity. When we exposed EDL or SOL muscles to Naja naja, Apis mellifera venoms (25 microg/ml), or Triton X-100 (0.01%), they showed similar rate of CK release. Our present data suggest that a mechanism involving intracellular calcium regulation or potassium channels may participate in the different sensitivity of EDL and SOL to B. jararacussu venom. PMID:18675839

  11. Akbu-LAAO exhibits potent anti-tumor activity to HepG2 cells partially through produced H2O2 via TGF-β signal pathway

    PubMed Central

    Guo, Chunmei; Liu, Shuqing; Dong, Panpan; Zhao, Dongting; Wang, Chengyi; Tao, Zhiwei; Sun, Ming-Zhong


    Previously, we characterized the biological properties of Akbu-LAAO, a novel L-amino acid oxidase from Agkistrodon blomhoffii ussurensis snake venom (SV). Current work investigated its in vitro anti-tumor activity and underlying mechanism on HepG2 cells. Akbu-LAAO inhibited HepG2 growth time and dose-dependently with an IC50 of ~38.82 μg/mL. It could induce the apoptosis of HepG2 cells. Akbu-LAAO exhibited cytotoxicity by inhibiting growth and inducing apoptosis of HepG2 as it showed no effect on its cell cycle. The inhibition of Akbu-LAAO to HepG2 growth partially relied on enzymatic-released H2O2 as catalase only partially antagonized this effect. cDNA microarray results indicated TGF-β signaling pathway was linked to the cytotoxicity of Akbu-LAAO on HepG2. TGF-β pathway related molecules CYR61, p53, GDF15, TOB1, BTG2, BMP2, BMP6, SMAD9, JUN, JUNB, LOX, CCND1, CDK6, GADD45A, CDKN1A were deregulated in HepG2 following Akbu-LAAO stimulation. The presence of catalase only slightly restored the mRNA changes induced by Akbu-LAAO for differentially expressed genes. Meanwhile, LDN-193189, a TGF-β pathway inhibitor reduced Akbu-LAAO cytotoxicity on HepG2. Collectively, we reported, for the first time, SV-LAAO showed anti-tumor cell activity via TGF-β pathway. It provides new insight of SV-LAAO exhibiting anti-tumor effect via a novel signaling pathway. PMID:26655928

  12. A new modified animal model of myosin-induced experimental autoimmune myositis enhanced by defibrase

    PubMed Central

    Wen-Jing, Luo; Hong-Hua, Li; Xiang-Hui, Lu; Jie-Xiao, Liu


    Introduction We investigated the effect of defibrase (a proteolytic enzyme extraction of Agkistrodon halys venom) on experimental autoimmune myositis (EAM) in guinea pigs and explored the option of using a modified pig model of EAM to enhance the study of this disease. Material and methods Guinea pigs were divided into 3 groups: group A (control group) was immunized with complete Freund adjuvant (CFA), then received 6 injections of saline weekly; group B (EAM group) was immunized with partially purified rabbit myosin emulsified with CFA, then received an injection of saline; group C (EAM + defibrase group) was immunized with purified rabbit myosin emulsified with CFA, then received an injection of defibrase. The animals were observed for their general health condition and the body weight was measured daily. Plasma levels of fibrinogen and creatine kinase (CK) were determined. Muscle tissues were examined histologically. Results After immunizations for 6 weeks, incidence of EAM in groups A, B and C was 0 (0/7), 83.3% (10/12) and 100% (15/15), respectively. Guinea pigs with EAM presented angeitis symptoms of muscle weakness. Histological analysis revealed a significant difference. Muscles with EAM had scattered or diffuse inflammatory manifestations, which are also common pathological features of human idiopathic polymyositis (IPM). Defibrase-treated animals displayed extensive inflammation and fiber necrosis compared with the EAM group (histological score: 2.80 ±1.15 vs. 1.88 ±1.32, p < 0.05). Severity of inflammation of group B was mainly mild to moderate; 16.7% (2/12) of animals developed severe inflammation. Incidence of severe inflammation with a score up to 4 in group C was 40% (6/15). Conclusions Defibrase can exacerbate myosin-induced EAM; thus a new modified model was generated. PMID:26788090

  13. Traditional Chinese Medicine Induced Liver Injury.


    Teschke, Rolf


    Traditional Chinese Medicine (TCM) is popular around the world and encompasses many different practices with particular emphasis on herbal TCM. Using the PubMed database, a literature search was undertaken to assess the extent herbal TCM products exert rare hepatotoxicity. Analysis of reported cases revealed numerous specified herbal TCM products with potential hepatotoxicity. Among these were An Shu Ling, Bai Fang, Bai Xian Pi, Ban Tu Wan, Bo He, Bo Ye Qing Niu Dan, Bofu Tsu Sho San, Boh Gol Zhee, Cang Er Zi, Chai Hu, Chaso, Chi R Yun, Chuan Lian Zi, Ci Wu Jia, Da Chai Hu Tang, Da Huang, Du Huo, Gan Cao, Ge Gen, Ho Shou Wu, Hu Bohe You, Hu Zhang, Huang Qin, Huang Yao Zi, Hwang Geun Cho, Ji Gu Cao, Ji Ji, Ji Xue Cao, Jiguja, Jin Bu Huan, Jue Ming Zi, Kamishoyosan, Kudzu, Lei Gong Teng, Long Dan Xie Gan Tang, Lu Cha, Ma Huang, Mao Guo Tian Jie Cai, Onshido, Polygonum multiflorum, Qian Li Guang, Ren Shen, Sairei To, Shan Chi, Shen Min, Shi Can, Shi Liu Pi, Shou Wu Pian, Tian Hua Fen, White flood, Wu Bei Zi, Xi Shu, Xiao Chai Hu Tang, Yin Chen Hao, Zexie, Zhen Chu Cao, and various unclassified Chinese herbal mixtures. Causality was firmly established for a number of herbal TCM products by a positive reexposure test result, the liver specific scale of CIOMS (Council for International Organizations of Medical Sciences), or both. Otherwise, the quality of case data was mixed, especially regarding analysis of the herb ingredients because of adulteration with synthetic drugs, contamination with heavy metals, and misidentification. In addition, non-herbal TCM elements derived from Agaricus blazei, Agkistrodon, Antelope, Bombyx, Carp, Fish gallbladder, Phellinus, Scolopendra, Scorpio, and Zaocys are also known or potential hepatotoxins. For some patients, the clinical course was severe, with risks for acute liver failure, liver transplantation requirement, and lethality. In conclusion, the use of few herbal TCM products may rarely be associated with hepatotoxicity in some

  14. Serine proteinase inhibition by the active site titrant N alpha-(N, N-dimethylcarbamoyl)-alpha-azaornithine p-nitrophenyl ester. A comparative study.


    Ascenzi, P; Balliano, G; Gallina, C; Polticelli, F; Bolognesi, M


    Kinetics for the hydrolysis of the chromogenic active-site titrant N alpha-(N,N-dimethylcarbamoyl)-alpha-azaornithine p-nitrophenyl ester (Dmc-azaOrn-ONp) catalysed by bovine beta-trypsin, bovine alpha-thrombin, bovine Factor Xa, human alpha-thrombin, human Factor Xa, human Lys77-plasmin, human urinary kallikrein, Mr 33 000 and Mr 54 000 species of human urokinase, porcine pancreatic beta-kallikrein-A and -B and Ancrod (the coagulating serine proteinase from the Malayan pit viper Agkistrodon rhodostoma venom) have been obtained between pH 6.0 and 8.0, at 21.0 degrees C, and analysed in parallel with those for the enzymatic cleavage of N alpha-(N,N-dimethylcarbamoyl)-alpha-azalysine p-nitrophenyl ester (Dmc-azaLys-ONp). The enzyme kinetics are consistent with the minimum three-step catalytic mechanism of serine proteinases, the rate-limiting step being represented by the deacylation process. Bovine beta-trypsin kinetics are modulated by the acid-base equilibrium of the His57 catalytic residue (pKa approximately 6.9). Dmc-azaOrn-ONp and Dmc-azaLys-ONp bind stoichiometrically to the serine proteinase active site, and allow the reliable determination of the active enzyme concentration between 1.0 x 10-6 M and 3.0 x 10-4 M. The affinity and the reactivity for Dmc-azaOrn-ONp (expressed by Ks and k+2/Ks, respectively) of the serine proteinases considered are much lower than those for Dmc-azaLys-ONp. The very different affinity and reactivity properties for Dmc-azaOrn-ONp and Dmc-azaLys-ONp have been related to the different size of the ornithine/lysine side chains, and to the ensuing different positioning of the active-site titrants upon binding to the enzyme catalytic centre (i.e. to P1-S1 recognition). These data represent the first detailed comparative investigation on the catalytic properties of serine proteinases towards an ornithine derivative (i. e. Dmc-azaOrn-ONp). PMID:10672036

  15. Traditional Chinese Medicine Induced Liver Injury

    PubMed Central


    Traditional Chinese Medicine (TCM) is popular around the world and encompasses many different practices with particular emphasis on herbal TCM. Using the PubMed database, a literature search was undertaken to assess the extent herbal TCM products exert rare hepatotoxicity. Analysis of reported cases revealed numerous specified herbal TCM products with potential hepatotoxicity. Among these were An Shu Ling, Bai Fang, Bai Xian Pi, Ban Tu Wan, Bo He, Bo Ye Qing Niu Dan, Bofu Tsu Sho San, Boh Gol Zhee, Cang Er Zi, Chai Hu, Chaso, Chi R Yun, Chuan Lian Zi, Ci Wu Jia, Da Chai Hu Tang, Da Huang, Du Huo, Gan Cao, Ge Gen, Ho Shou Wu, Hu Bohe You, Hu Zhang, Huang Qin, Huang Yao Zi, Hwang Geun Cho, Ji Gu Cao, Ji Ji, Ji Xue Cao, Jiguja, Jin Bu Huan, Jue Ming Zi, Kamishoyosan, Kudzu, Lei Gong Teng, Long Dan Xie Gan Tang, Lu Cha, Ma Huang, Mao Guo Tian Jie Cai, Onshido, Polygonum multiflorum, Qian Li Guang, Ren Shen, Sairei To, Shan Chi, Shen Min, Shi Can, Shi Liu Pi, Shou Wu Pian, Tian Hua Fen, White flood, Wu Bei Zi, Xi Shu, Xiao Chai Hu Tang, Yin Chen Hao, Zexie, Zhen Chu Cao, and various unclassified Chinese herbal mixtures. Causality was firmly established for a number of herbal TCM products by a positive reexposure test result, the liver specific scale of CIOMS (Council for International Organizations of Medical Sciences), or both. Otherwise, the quality of case data was mixed, especially regarding analysis of the herb ingredients because of adulteration with synthetic drugs, contamination with heavy metals, and misidentification. In addition, non-herbal TCM elements derived from Agaricus blazei, Agkistrodon, Antelope, Bombyx, Carp, Fish gallbladder, Phellinus, Scolopendra, Scorpio, and Zaocys are also known or potential hepatotoxins. For some patients, the clinical course was severe, with risks for acute liver failure, liver transplantation requirement, and lethality. In conclusion, the use of few herbal TCM products may rarely be associated with hepatotoxicity in some

  16. Annotated catalogue of the Tachinidae (Insecta, Diptera) of the Afrotropical Region, with the description of seven new genera

    PubMed Central

    O’Hara, James E.; Cerretti, Pierfilippo


    regalis Cerretti & O’Hara, sp. n. (South Africa), and Myxophryxe satanas Cerretti & O’Hara, sp. n. (South Africa) (Exoristinae, Goniini); and Stiremania Cerretti & O’Hara with type species Stiremania karoo Cerretti & O’Hara, gen. n. and sp. n. (South Africa), and Stiremania robusta Cerretti & O’Hara, sp. n. (South Africa) (Exoristinae, Goniini). Paraclara Bezzi, 1908 is transferred from the Cylindromyiini to the Hermyini, comb. n. Sarrorhina Villeneuve, 1936 is transferred from the Minthoini to the Graphogastrini, comb. n. Three genera are newly recorded from the Afrotropical Region: Madremyia Townsend, 1916 (Eryciini); Paratrixa Brauer & Bergenstamm, 1891 (Blondeliini); and Simoma Aldrich, 1926 (Goniini). Three genera previously recorded from the Afrotropical Region are no longer recognized from the region: Calozenillia Townsend, 1927 (Palaearctic, Oriental and Australasian regions); Eurysthaea Robineau-Desvoidy, 1863 (Palaearctic, Oriental and Australasian regions); and Trixa Meigen, 1824 (Palaearctic and Oriental regions). Two species are newly recorded from the Afrotropical Region: Amnonia carmelitana Kugler, 1971 (Ethiopia, Kenya); and Simoma grahami Aldrich, 1926 (Namibia). Three species previously recorded from the Afrotropical Region are no longer recognized from the region: Euthera peringueyi Bezzi, 1925 (Oriental Region); Hamaxia incongrua Walker, 1860 (Palaearctic, Oriental and Australasian regions); Leucostoma tetraptera (Meigen, 1824) (Palaearctic Region). New replacement names are proposed for five preoccupied names of Afrotropical species: Billaea rubida O’Hara & Cerretti for Phorostoma rutilans Villeneuve, 1916, preoccupied in the genus Billaea Robineau-Desvoidy, 1830 by Musca rutilans Fabricius, 1781, nom. n.; Cylindromyia braueri O’Hara & Cerretti for Ocyptera nigra Villeneuve, 1918, preoccupied in the genus Cylindromyia Meigen, 1803 by Glossidionophora nigra Bigot, 1885, nom. n.; Cylindromyia rufohumera O’Hara & Cerretti for Ocyptera

  17. Annotated catalogue of the Tachinidae (Insecta, Diptera) of the Afrotropical Region, with the description of seven new genera.


    O'Hara, James E; Cerretti, Pierfilippo


    . n. (South Africa), and Myxophryxe satanas Cerretti & O'Hara, sp. n. (South Africa) (Exoristinae, Goniini); and Stiremania Cerretti & O'Hara with type species Stiremania karoo Cerretti & O'Hara, gen. n. and sp. n. (South Africa), and Stiremania robusta Cerretti & O'Hara, sp. n. (South Africa) (Exoristinae, Goniini). Paraclara Bezzi, 1908 is transferred from the Cylindromyiini to the Hermyini, comb. n. Sarrorhina Villeneuve, 1936 is transferred from the Minthoini to the Graphogastrini, comb. n. Three genera are newly recorded from the Afrotropical Region: Madremyia Townsend, 1916 (Eryciini); Paratrixa Brauer & Bergenstamm, 1891 (Blondeliini); and Simoma Aldrich, 1926 (Goniini). Three genera previously recorded from the Afrotropical Region are no longer recognized from the region: Calozenillia Townsend, 1927 (Palaearctic, Oriental and Australasian regions); Eurysthaea Robineau-Desvoidy, 1863 (Palaearctic, Oriental and Australasian regions); and Trixa Meigen, 1824 (Palaearctic and Oriental regions). Two species are newly recorded from the Afrotropical Region: Amnonia carmelitana Kugler, 1971 (Ethiopia, Kenya); and Simoma grahami Aldrich, 1926 (Namibia). Three species previously recorded from the Afrotropical Region are no longer recognized from the region: Euthera peringueyi Bezzi, 1925 (Oriental Region); Hamaxia incongrua Walker, 1860 (Palaearctic, Oriental and Australasian regions); Leucostoma tetraptera (Meigen, 1824) (Palaearctic Region). New replacement names are proposed for five preoccupied names of Afrotropical species: Billaea rubida O'Hara & Cerretti for Phorostoma rutilans Villeneuve, 1916, preoccupied in the genus Billaea Robineau-Desvoidy, 1830 by Musca rutilans Fabricius, 1781, nom. n.; Cylindromyia braueri O'Hara & Cerretti for Ocyptera nigra Villeneuve, 1918, preoccupied in the genus Cylindromyia Meigen, 1803 by Glossidionophora nigra Bigot, 1885, nom. n.; Cylindromyia rufohumera O'Hara & Cerretti for Ocyptera scapularis Villeneuve, 1944, preoccupied in

  18. Alfimeprase.



    Alfimeprase, a fibrolase derivative with thrombolytic activity produced by recombinant DNA technology, was discovered by Amgen and was in development with Nuvelo for the treatment of stroke and catheter occlusion. However, development has been discontinued. Fibrolase is a zinc-containing metalloendopeptidase that was first isolated from the venom of the Southern copperhead snake, Agkistrodon contortrix contortrix. Alfimeprase directly degrades fibrin to break down clots. Alfimeprase is infused directly into the thrombus (side-hole catheter pushed through the entire clot) via multiple manual pulsed infusions. Alfimeprase degrades fibrin directly and entrapped blood cells are freed. Excess alfimeprase is rapidly inactivated by alpha-2 macroglobulin through an irreversible, covalent interaction. Phase III development of alfimeprase for peripheral arterial occlusion was discontinued based on poor results from the NAPA-2 and SONOMA-2 trials; however, Nuvelo resumed development of alfimeprase in 2007 for stroke and catheter occlusion.Alfimeprase's thrombolytic activity appears to be localized to the site of delivery because it is rapidly inactivated by alpha-2 macroglobulin, a naturally occurring protein in the blood, as it moves away from the site of delivery and into general blood circulation. In January 2002, Amgen and Hyseq Pharmaceuticals (now Nuvelo) entered into a collaboration to develop and commercialize alfimeprase. Nuvelo is to develop the product through clinical trials and Amgen was to be responsible for its manufacture. Both companies were to participate in commercial activities; Amgen was to have the option to lead these. Full financial terms were not disclosed. Further to this agreement, in November 2004 Amgen granted Nuvelo worldwide rights to develop and commercialize alfimeprase in exchange for milestone and royalty payments. In August 2007, Nuvelo decided to focus on core development programmes that it believed would produce the nearest-term proof