A comparison of different methods to assess the hemorrhagic activity of Bothrops venoms.

The hemorrhagic activity of Bothrops (B.) alternatus, B. ammodytoides, B. jararaca, B. jararacussu, B. moojeni and B. neuwiedii venoms from specimens captured in Argentina was assayed after i.d. injection to mice. The hemorrhagic haloes produced by each venom had different color intensities, although no significant differences were observed by measurement of the average diameters or the weight of the excised hemorrhagic haloes. Conversely, important differences were found by measuring the amount of hemoglobin extracted from excised hemorrhagic haloes of similar size produced by different venoms. The relationship between the amount of hemoglobin extracted and the weight of the excised hemorrhagic haloes was linear, with a slope (hemoglobin released per gram of hemorrhagic halo) characteristic for each venom, and proportional to the potency. On this basis, the activity of B. alternatus, B. ammodytoides and B. jararaca is similar, about 1.5 times higher than that of B. jararacussu and B. moojeni venoms and threefold higher than that of B. neuwiedii venom. Thus, measurement of the of hemoglobin released provides additional information in comparative studies, and may be used to assess the antihemorrhagic potency of antivenoms.

Some toxic and enzymatic activities of Bothrops ammodytoides (yarará ñata) venom.

Bothrops ammodytoides, the smallest representative of this genus, is found only in Argentina. Venom was extracted from thirty adult specimens (35-70 cm in length, 90-300 g in weight) captured in the Province of Buenos Aires and kept in captivity. Venom yield was 3-30 mg. SDS-PAGE showed strong bands at 14.0; 23-25; 45; 54 and 63 kDa and weak bands at 17.0; 30.0; 40.0 and 85.0 kDa. Toxic activities were: LD50 (intravenous, mice) 0.5+/-0.2 microg/g; minimal procoagulant dose on human plasma (MPD-P) 35+/-2 mg/l; and minimal defibrinogenating dose (MDD, mice) 6-12 microg. Hemorrhagic and/or necrotic activities appear to play a major role in lethality; minimal hemorrhagic dose (MHD, mice) is 10+/-2 microg/g and minimal necrotizing dose (MND, mice) is 38+/-5 microg. The LD50, MPD-P and MND are among the lowest in venoms from Bothrops species found in Argentina. B. ammodytoides venom exhibited high proteolytic and phospholipase A2 activities. Most of the B. ammodytoides venom components cross-react with Bivalent Bothropic antivenom (Instituto Nacional de Producción de Biológicos ANLIS Dr. G. Malbrin, against B. alternatus and B. neuwiedii venoms). One ml of antivenom neutralizes 1.2 mg of B. ammodytoides venom.

[Cross neutralization of bothrops jararacussu venom by heterologous antivenoms]. / Neutralización cruzada de veneno de Bothrops jararacussu por sueros antiofidicos heterologos.

We have studied the immunochemical cross-reactivity and cross-neutralization of the lethal potency, hemorrhagic, necrotizing, procoagulant and (indirect) hemolytic activities of Bothrops jararacussu venom by the standard antivenoms produced in Argentina. These antivenoms are horse immunoglobulin F (ab')2 fragments from animals immunized with 1) Crotalus durissus terrificus venom (Monovalent Anticrotalic antivenom); 2) Bothrops alternatus and B. neuwiedii venoms (Bivalent Botropic antivenom); 3) B. alternatus, B. neuwiedii, B. jararaca and B. jararacussu venoms (Tetravalent Bothropic, or "Misiones" antivenom) and 4) B. alternatus, B. neuwiedii and C. d. terrificus venoms (Trivalent Botropic-Crotalic antivenom). In preincubation experiments, all the heterologous antivenoms neutralized the toxic and biological activities of B. jararacussu venom with a potency at least as high as the Tetravalent Botropic (i.e. the only homologous) antivenom, in which B. jararacussu venom was included as immunogen. These results suggest the possibility of using heterologous antibothropic antivenoms for the treatment of snake bites by B. jararacussu.

The immunochemical reactivity and neutralizing capacity of polyvalent Vipera (European) antivenom on enzymatic and toxic activities in the venoms of Crotalids from Argentina

The immunochemical reactivity and neutralizing capacity of polyvalent Vipera antivenom (Vipera ammodytes, Vipera aspis, Vipera berus, Vipera lebetina, and Vipera xanthina) were tested on the enzymatic and biological activities of Crotalus durissus terrificus and the following Bothrops venoms from Argentina (Bothrops alternatus, Bothrops ammodytoides, Bothrops neuwiedii, Bothrops jararaca, Bothrops jararacussu, and Bothrops moojeni). The Vipera antivenom reacted weakly when tested by double immunoprecipitation (DIP) and reacted with all the venoms when tested by ELISA. Several components in all the venoms studied were recognized in Western blots. Vipera antivenom deactivated to different degrees in vitro procoagulant, (indirect) hemolytic, and proteolytic activities in all the venoms studied. Preincubation of Bothrops alternatus venom with Vipera antivenom neutralized a lethal potency of 4.5 LD50 in mice with an ED50 of 1.25 ñ 0.25 µl per µg of venom, and with 1.0 µl/µg inhibited 54 per cent of the hemorragic activity and 48 per cent of necrotic activity. Vipera antivenom (2.0 µl per µg toxin) inhibited the phospholipase A2 activity of purified crotoxin and decreased its lethal potency by 60 per cent, while the neutralizing capacity on the lethal potency of crude Crotalus durissus terrificus venom was poor even at a level of 5.0 µl/µg of venom.

Neutralización cruzada de veneno de Bothrops jararacussu por sueros antiofidicos heterologos. / [Cross neutralization of bothrops jararacussu venom by heterologous antivenoms]

We have studied the immunochemical cross-reactivity and cross-neutralization of the lethal potency, hemorrhagic, necrotizing, procoagulant and (indirect) hemolytic activities of Bothrops jararacussu venom by the standard antivenoms produced in Argentina. These antivenoms are horse immunoglobulin F (ab)2 fragments from animals immunized with 1) Crotalus durissus terrificus venom (Monovalent Anticrotalic antivenom); 2) Bothrops alternatus and B. neuwiedii venoms (Bivalent Botropic antivenom); 3) B. alternatus, B. neuwiedii, B. jararaca and B. jararacussu venoms (Tetravalent Bothropic, or [quot ]Misiones[quot ] antivenom) and 4) B. alternatus, B. neuwiedii and C. d. terrificus venoms (Trivalent Botropic-Crotalic antivenom). In preincubation experiments, all the heterologous antivenoms neutralized the toxic and biological activities of B. jararacussu venom with a potency at least as high as the Tetravalent Botropic (i.e. the only homologous) antivenom, in which B. jararacussu venom was included as immunogen. These results suggest the possibility of using heterologous antibothropic antivenoms for the treatment of snake bites by B. jararacussu.

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

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