- Email: [email protected]
Study on biological properties of Bothrops erythromelas venom
9th World Congress
83
Culture and toxicity of dinoflagellates from ciguatera endemic regions of the world. DONALD R. TINDALL,I DONALD M. M m L ~ 2 and JEFFREY W. BOMBERl (1Department of Botany and 2Department of Physiology, Southern Illinois University, Carbondale, Illinois 62901, U.S.A.). THE SOUTm~N Illinois University Culture Collection of Dinoflagellates houses 163 strains or clones representing 23 species from major centers of ciguatera in the Atlantic and Pacific Oceans and the Caribbean Sea. All stock cultures are maintained in a natural seawater medium supplemented with ES or K enrichments under a 16 : 8 light : dark cycle (3200 lux, cool-white fluorescent) at 27°C. We are systematically rearing clones of each species in large-scale cultures and screening them for toxicity using 20 g mice and guinea pig ileum preparations. One or more crude toxins have been extracted from each of 10 separate species representing ? genera: Amphidinium, Coolia, Gambierdiscus, Gonyaulax, Gymnodinium, Ostreopsis and Prorocentrum. However, we have concentrated our efforts on those species which dominate natural habitats which have a long history of producing ciguatoxic fish, namely Gain. toxicus, O. lenticularis, P. concavum, P. lima, P. mexicanum and P. cassubieum. Several clones of each species have been subjected to physiological examination and toxin extraction, purification, and bioassay. Crude toxins and their respective potencies (LDso, mg/kg mouse) which have been identified are as follows: Gam. toxieus--maitotoxin (1.1 - 2.75) and ciguatoxin (1.5 - 4.95); O. lenticularis--2 unnamed toxins (0.34 - 0.55 and 0.28 - 0.56); P. concavum--3 unnamed toxins (23.6, 8.3 and 4.0) and a fast-acting toxin (FAT) (1.5 - 7.18); P. lima--3 unnamed toxins (6.45, 7.25 and 10.95) and a FAT (2.5); P. mexicanum--1 FAT (not determined) and P. eassubicum--2 unnamed toxins (0.38 and 0.55). Progress has been made toward the purification of maitotoxin and FAT from P. concavum. The number and variety of toxins found in these species suggest that an explanation of the ciguatera syndrome is going to be far more complex than previously believed. Sponsored by the Sea Grant College Program, the U.S. Army Medical Research Institute of Infectious Diseases, and Mr Myron Hokin.
Functional importance of cysteine and histidine residues in the alpha toxin (phospholipase C) of Clostridium perfringens. RICHARD W. TITBALL and TIM RUmIX~E (Chemical Defence Establishment, Porton Down, Salisbury, Wiltshire SP4 0JQ, U.K.). THE ALPHAtoxin (phospholipase C) of C. perfringens is thought to play a key role in the pathogenesis of several diseases, including gas gangrene in man. Prior immunisation with alpha toxoid has been reported to induce some protection against disease, however, such studies have been hampered by the impurity of toxin and toxoid preparations. In order to study prophylactic strategies, we have initially investigated the mode of action of the alpha toxin at a molecular level. Chemical modification of histadine residues with diethylpyrocarbonate completely inactivated the toxin whilst modification of the single cysteine residue with p-hydroxy mercuribenzoate did not affect phospholipase C activity. These results are discussed in relation to the amino acid sequence of the alpha toxin and its mode of action.
Study on biologicalproperties ofBothrops erythromelas venom. SANDRAC. TOMY,1 MASUGI MARUYAMA,1 LUIZ ROBERTO C. GON~ALVES,2 AURA S. KAMIGUTI1 and MAmO MARIANO2 (tLaboratory of Hematology and 2Laboratory of Experimental Physiopathology, Instituto Butantan, Silo Paulo, Brazil). THE SPECIESB. erythromelas is commonly found in north-eastern Brazil and is assumed to be a common cause of snake envenomation in that area. In the present study, several biological activities of this venom were estimated in comparison to those of B. jararaca, showing that its minimum coagulant dose (MCD) was extremely higher. The main coagulant activity was suggested to be due to a procoagulant property, either by activating Factor X or prothrombin. The minimum defibrinogenating dose (MDD) estimated in mouse was 175/~g/kg for B. erythromelas venom and 80 #g/kg for B. jararaca venom. However, MDD of B. erythromelas venom using rats was 75/~g/kg which demonstrates the low sensitivity of mice. One hour after administration of one MDD of B. erythromelas venom in rats, the fibrinogen levels were 37.6+21.5 mg/dl, then following the time course the fibrinogen levels were within normal range by 24 hr after. We may conclude that the B. erythromelas snakes could severely affect blood coagulation system of snake bitten patients.
Effect of configuration on lethality and in vivo effects on the neuromuscular junction of anatoxin-a. WILLIAM M. VALENTINE,I JEROMEC. BEHRENS,2 RICHARD A. AMOS,~ PETER H. DuQUETTE,2 ANDREW M. DAHLEM1 and VAL R. BEASLEY1 (*Department of Veterinary Biosciences, College of Veterinary Medicine, University of Illinois, Urbana, IL 61801; and 2Bio Metrics Systems Inc., 9932 West 74th St., Eden Prairie, MN 55344, U.S.A.). ANATOXlN-a hydrochloride has been determined to be stereospecific in regard to lethality following intraperitoneal injection in male Balb/C mice. The LDs0and standard error for the isomers are: (+)anatoxin-a
83
Culture and toxicity of dinoflagellates from ciguatera endemic regions of the world. DONALD R. TINDALL,I DONALD M. M m L ~ 2 and JEFFREY W. BOMBERl (1Department of Botany and 2Department of Physiology, Southern Illinois University, Carbondale, Illinois 62901, U.S.A.). THE SOUTm~N Illinois University Culture Collection of Dinoflagellates houses 163 strains or clones representing 23 species from major centers of ciguatera in the Atlantic and Pacific Oceans and the Caribbean Sea. All stock cultures are maintained in a natural seawater medium supplemented with ES or K enrichments under a 16 : 8 light : dark cycle (3200 lux, cool-white fluorescent) at 27°C. We are systematically rearing clones of each species in large-scale cultures and screening them for toxicity using 20 g mice and guinea pig ileum preparations. One or more crude toxins have been extracted from each of 10 separate species representing ? genera: Amphidinium, Coolia, Gambierdiscus, Gonyaulax, Gymnodinium, Ostreopsis and Prorocentrum. However, we have concentrated our efforts on those species which dominate natural habitats which have a long history of producing ciguatoxic fish, namely Gain. toxicus, O. lenticularis, P. concavum, P. lima, P. mexicanum and P. cassubieum. Several clones of each species have been subjected to physiological examination and toxin extraction, purification, and bioassay. Crude toxins and their respective potencies (LDso, mg/kg mouse) which have been identified are as follows: Gam. toxieus--maitotoxin (1.1 - 2.75) and ciguatoxin (1.5 - 4.95); O. lenticularis--2 unnamed toxins (0.34 - 0.55 and 0.28 - 0.56); P. concavum--3 unnamed toxins (23.6, 8.3 and 4.0) and a fast-acting toxin (FAT) (1.5 - 7.18); P. lima--3 unnamed toxins (6.45, 7.25 and 10.95) and a FAT (2.5); P. mexicanum--1 FAT (not determined) and P. eassubicum--2 unnamed toxins (0.38 and 0.55). Progress has been made toward the purification of maitotoxin and FAT from P. concavum. The number and variety of toxins found in these species suggest that an explanation of the ciguatera syndrome is going to be far more complex than previously believed. Sponsored by the Sea Grant College Program, the U.S. Army Medical Research Institute of Infectious Diseases, and Mr Myron Hokin.
Functional importance of cysteine and histidine residues in the alpha toxin (phospholipase C) of Clostridium perfringens. RICHARD W. TITBALL and TIM RUmIX~E (Chemical Defence Establishment, Porton Down, Salisbury, Wiltshire SP4 0JQ, U.K.). THE ALPHAtoxin (phospholipase C) of C. perfringens is thought to play a key role in the pathogenesis of several diseases, including gas gangrene in man. Prior immunisation with alpha toxoid has been reported to induce some protection against disease, however, such studies have been hampered by the impurity of toxin and toxoid preparations. In order to study prophylactic strategies, we have initially investigated the mode of action of the alpha toxin at a molecular level. Chemical modification of histadine residues with diethylpyrocarbonate completely inactivated the toxin whilst modification of the single cysteine residue with p-hydroxy mercuribenzoate did not affect phospholipase C activity. These results are discussed in relation to the amino acid sequence of the alpha toxin and its mode of action.
Study on biologicalproperties ofBothrops erythromelas venom. SANDRAC. TOMY,1 MASUGI MARUYAMA,1 LUIZ ROBERTO C. GON~ALVES,2 AURA S. KAMIGUTI1 and MAmO MARIANO2 (tLaboratory of Hematology and 2Laboratory of Experimental Physiopathology, Instituto Butantan, Silo Paulo, Brazil). THE SPECIESB. erythromelas is commonly found in north-eastern Brazil and is assumed to be a common cause of snake envenomation in that area. In the present study, several biological activities of this venom were estimated in comparison to those of B. jararaca, showing that its minimum coagulant dose (MCD) was extremely higher. The main coagulant activity was suggested to be due to a procoagulant property, either by activating Factor X or prothrombin. The minimum defibrinogenating dose (MDD) estimated in mouse was 175/~g/kg for B. erythromelas venom and 80 #g/kg for B. jararaca venom. However, MDD of B. erythromelas venom using rats was 75/~g/kg which demonstrates the low sensitivity of mice. One hour after administration of one MDD of B. erythromelas venom in rats, the fibrinogen levels were 37.6+21.5 mg/dl, then following the time course the fibrinogen levels were within normal range by 24 hr after. We may conclude that the B. erythromelas snakes could severely affect blood coagulation system of snake bitten patients.
Effect of configuration on lethality and in vivo effects on the neuromuscular junction of anatoxin-a. WILLIAM M. VALENTINE,I JEROMEC. BEHRENS,2 RICHARD A. AMOS,~ PETER H. DuQUETTE,2 ANDREW M. DAHLEM1 and VAL R. BEASLEY1 (*Department of Veterinary Biosciences, College of Veterinary Medicine, University of Illinois, Urbana, IL 61801; and 2Bio Metrics Systems Inc., 9932 West 74th St., Eden Prairie, MN 55344, U.S.A.). ANATOXlN-a hydrochloride has been determined to be stereospecific in regard to lethality following intraperitoneal injection in male Balb/C mice. The LDs0and standard error for the isomers are: (+)anatoxin-a