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PubMed2001

Fatal cerebral hemorrhage and acute renal failure after young Bothrops jararacussu snake bite.

An unusual case of a patient developing severe coagulopathy disorder and a clinical picture of cerebral hemorrhage and acute renal failure after young Bothrops jararacussu snake bite is reported. The mechanisms of snake venom-induced injury are discussed and similar cases in literature are revised and compared. The use of bothropic-Crotalus antivenom in severe B. jararacussu envenomation is discussed.

Acute Kidney Injury↗

PubMed1997

Snake bites by the jararacuçu (Bothrops jararacussu): clinicopathological studies of 29 proven cases in São Paulo State, Brazil.

The jararacuçu, one of the most dreaded snakes of Brazil, southern Bolivia, Paraguay and northeastern Argentina, is a heavily-built pit viper which may grow to a length of 2.2 m. Up to 1000 mg (dry weight) of highly-lethal venom may be milked from its venom glands on a single occasion. It has accounted for 0.8% to 10% of series of snake bites in São Paulo State, Brazil. We examined 29 cases of proven jararacuçu bites recruited over a 20-year period in two São Paulo hospitals. Severe signs of local and systemic envenoming, (local necrosis, shock, spontaneous systemic bleeding, renal failure) were seen only in patients bitten by snakes longer than 50 cm; bites by shorter specimens were more likely to cause incoagulable blood. Fourteen patients developed coagulopathy, six local necrosis (requiring amputation in one) and five local abscesses. Two became shocked and four developed renal failure. Three patients, aged 3, 11 and 65 years, died 18.75, 27.75 and 83 h after being bitten, with respiratory and circulatory failure despite large doses of specific antivenom and intensive-care-unit management. In two patients, autopsies revealed acute renal tubular necrosis, cerebral oedema, haemorrhagic rhabdomyolysis at the site of the bite and disseminated intravascular coagulation. In one survivor with chronic renal failure, renal biopsy showed bilateral cortical necrosis; the patient remains dependent on haemodialysis. Effects of polyspecific Bothrops antivenom were not impressive, and it has been suggested that anti-Bothrops and anti-Crotalus antivenoms should be given in combination.

Adult↗

PubMed1998

Crystallization and preliminary X-ray diffraction studies of piratoxin II, a phospholipase A2 isolated from the venom of Bothrops pirajai.

The phospholipases A2 (PLA2, E.C. 3.1.1.4, phosphatide sn2 acylhydrolases) are the major components of the venom of several snakes. They are responsible for several important pharmacological effects observed in ophidian incidents. PLA2 piratoxin II from Bothrops pirajai has been crystallized by the vapour-diffusion technique. X-ray diffraction data have been collected to 2.04 A resolution (90.2% complete, Rmerge = 0.070). The space group is P21 and the cell parameters are a = 46.19, b = 60.36, c = 58.74 A and beta = 96.05 degrees. The structure has been solved by molecular replacement using the crystallographic structure of PLA2 from Bothrops asper (PDB code 1CLP) as a search model.

Animals↗

PubMed2006

Preliminary X-ray crystallographic studies of BthTX-II, a myotoxic Asp49-phospholipase A2 with low catalytic activity from Bothrops jararacussu venom.

For the first time, a complete X-ray diffraction data set has been collected from a myotoxic Asp49-phospholipase A2 (Asp49-PLA2) with low catalytic activity (BthTX-II from Bothrops jararacussu venom) and a molecular-replacement solution has been obtained with a dimer in the asymmetric unit. The quaternary structure of BthTX-II resembles the myotoxin Asp49-PLA2 PrTX-III (piratoxin III from B. pirajai venom) and all non-catalytic and myotoxic dimeric Lys49-PLA2s. In contrast, the oligomeric structure of BthTX-II is different from the highly catalytic and non-myotoxic BthA-I (acidic PLA2 from B. jararacussu). Thus, comparison between these structures should add insight into the catalytic and myotoxic activities of bothropic PLA2s.

Animals↗

PubMed2004

Platelet dysfunction during Bothrops jararaca snake envenomation in rabbits.

Despite being well established that snake envenomation causes blood coagulation and fibrinolysis disturbances, scant information is available about blood platelet disorders. Herein we show that experimentally Bothrops jararaca-envenomed rabbits presented thrombocytopenia, hypofibrinogenemia, elevation of von Willebrand factor plasma levels, platelet hypoaggregation in platelet rich plasma and whole blood, normoaggregation in washed platelet suspensions, decreased platelet ATP secretion, normal plasma levels of platelet factor 4, and constant intraplatelet serotonin levels. Furthermore, by flow cytometric analyses, platelets displayed a significant decrease in the expression of a GPIIb-IIIa epitope recognized by P2 monoclonal antibody (p< 0.05) and an increased expression of a ligand-induced binding site (LIBS-1) of GPIIIa (p< 0.05), but total GPIIb-IIIa expression, evaluated with specific polyclonal antibodies, was normal. Fibrinogen and fibrin(ogen) degradation product (FfDP) expression on platelet surface showed no significant alteration. Nonetheless, significant elevations of platelet P-selectin were noticed on circulating platelets. The percentage of circulating reticulated platelets and the survival time of biotinylated platelets of envenomed rabbits were not statistically different from control animals. We suggest that thrombin engendered by procoagulating components of B. jararaca venom has an essential role in the pathogenesis of platelet and coagulation disorders in this experimental model. Increased expression of P-selectin in the experimental group demonstrates that platelets of envenomed rabbits are indeed activated in circulation, and that decreased fibrinogen or increased FfDP levels are not the primary cause of platelet dysfunction. These results imply the existence of an inhibitor in plasma that interferes with platelet aggregation in bothropic envenomation.

Animals↗

PubMed1997

Control of venom production and secretion by sympathetic outflow in the snake Bothrops jararaca.

Many studies have examined the morphological and biochemical changes in the secretory epithelium of snake venom glands after a bite or milking. However, the mechanisms of venom production and secretion are not yet well understood. The present study was undertaken to evaluate the role of the sympathetic nervous system in the control of venom production and secretion. Venom glands were obtained from Bothrops jararaca (Viperidae) snakes, either unmilked previously or milked 4, 7 or 15 days before they were killed. Levels of tyrosine-hydroxylase-like immunoreactivity were higher in venom glands collected 4 days after milking, coinciding with the maximal synthetic activity of the secretory cells. The only catecholamine detected by high-performance liquid chromatography was noradrenaline, indicating the presence of noradrenergic fibres in these glands. In reserpine-treated milked snakes, no venom could be collected, and electron microscopic analysis showed narrow rough endoplasmic reticulum cisternae, instead of wide cisternae, and less well-developed Golgi apparatus compared with milked untreated snakes, indicating impairment of protein synthesis and secretion. The administration of isoprenaline or phenylephrine (beta- and alpha-adrenoceptor agonists, respectively) to reserpine-treated milked snakes promoted the widening of the rough endoplasmic reticulum and restored venom production, but only phenylephrine restored the development of the Golgi apparatus and the formation of many secretory vesicles. These results provide the first evidence that the sympathetic nervous system plays an important role in venom production and secretion in the venom glands of Bothrops jararaca. Understanding the importance of noradrenergic stimulation in venom production may provide new insights for research into the treatment of snakebites.

Animals↗

PubMed2006

Cytotoxic action in myoblasts and myotubes (C2C12) and enzymatic characterization of a new phospholipase A2 isoform (Bj-V) from Bothrops jararacussu venom.

A new PLA2 Bj-V from Bothrops jararacussu (14039.49 Da determined by MALDI-TOF mass spectrometry) was isolated in only one chromatographic step by HPLC ion-exchange and its purity was confirmed by reverse phase. Amino acid analysis showed a high content of hydrophobic and basic amino acids as well as 14 half-cysteine residues. The N-terminal sequence (DLWQFGQMIL KETGKIPFPY YGAYGCYCGW GGRGGKPKDG TDRCCYVHD...) showed a high degree of homology with basic D49 PLA2 myotoxins from other Bothrops venoms. Bj V showed discrete sigmoidal enzymatic behavior, with maximal activity at pH 8.4 and 35-40 degrees C. Full PLA2 activity required Ca2+ (10 mM) and there was little catalytic activity in the presence of 1 mM Ca2+. The addition of Mn2+ or Mg2+ (10 mM) in the presence of low (1 mM) Ca2+ slightly increased the enzyme activity, whereas Zn2+ and Cu2+ (10 mM) diminished the activity. The substitution of Ca2+ for Mg2+ or Cu2+ also reduced the enzymatic activity. Bj V had PLA2 activity and produced cytotoxicity in murine C2C12 skeletal muscle myoblasts and myotubes. The isolation of these isoforms Bj-IV [1] and Bj-V (described herein) found in a fraction previously described as homogeneous shows us the importance of optimization in purification techniques in order to better understand their biological behavior.

Amino Acid Sequence↗

PubMed1999

Histologic and functional renal alterations caused by Bothrops moojeni snake venom in rats.

Acute renal failure (ARF) is the main cause of death following snake bites by Bothrops species. In this study, we investigated the morphologic and functional renal disturbances caused by Bothrops moojeni venom in rats. Renal function was assessed based on creatinine and lithium clearances and on histologic examination of renal tissue 5 hr after the intravenous administration of 0.2 mg of venom/kg and 5 hr, 16 hr, and 48 hr after 0.4 mg of venom/ kg. A venom dose of 0.4 mg/kg produced renal tubule disturbances, including acute impairment of proximal and post-proximal tubule sodium handling associated with acute tubule necrosis. The glomerular filtration rate (GFR) decreased significantly and was accompanied by severe morphologic disturbances in the renal glomeruli. These functional and morphologic findings were observed in the absence of any change in mean arterial blood pressure. The decrease in GFR was not related to the presence of fibrin deposits in the glomerular capillary loops. These results suggest an early nephrotoxic action of B. moojeni venom involving significant morphologic and functional changes similar to those observed in snakebite-induced ARF in humans.

Acute Kidney Injury↗

PubMed2002

Bothrops moojeni snake venom-induced renal glomeruli changes in rat.

The venom of Bothrops moojeni has potent proteolytic and phospholipase A2 activities. In previous work, we showed that intravenous injection of this venom in rats decreased creatinine clearance and caused tubular dysfunction and histopathological changes with no alterations in blood pressure. The current study used scanning and transmission electron microscopy to assess the ultrastructural changes caused by B. moojeni venom (0.4 mg/kg i.v.) in rat renal glomeruli and correlated these alterations with the severity of proteinuria 5 hours, 16 hours, and 48 hours after venom injection. The changes included mesangiolysis, glomerular microaneurysms, and glomerular basement membrane (GBM) abnormalities. In addition, there was a reduction in the number and width of podocyte pedicels, which caused a reduction in the number of filtration slits. Electron-dense amorphous material, which may be proteinaceous in origin, was found in the pedicels. The severity of the ultrastructural abnormalities correlated with the level of proteinuria. These morphophysiological changes were attributed to biochemical and physiological disturbances in the components of the GBM and mesangial matrix as well as in cytoskeleton-associated proteins of podocytic processes, and could account for the breakdown of optimal glomerular filtration barrier functioning. These results, together with the absence of appreciable glomerular fibrin deposits, support the hypothesis of a direct activity of B. moojeni venom on rat kidneys. Proteolytic activity of the venom on renal glomeruli could then contribute to the onset of acute renal failure, and would explain the clinical manifestations of renal injury after bites by this and other Bothrops species.

Animals↗

PubMed1999

[Hemorrhagic, edema-forming, proteolytic and myonecrotic activities of viper venoms of Bothrops alternatus (Vibora de la Cruz)].

A study of hemorrhagic, edema-forming, proteolytic and myonecrotic activities induced by newborn Bothrops alternatus snake venoms from Argentina was performed. The analyzed venom had these activities with markedly differences when compared with the venom of adult specimens. Hemorrhagic activity was four times higher, while edema forming activity was ten times or more. But proteolytic activity was similar in both groups. Hystological analysis showed intense hemorrhage and muscular fiber myolisis after 60 minutes of venom injection. Myonecrosis and inflammatory exudate were higher in the following hours. Hemorrhage decreased after 24 hours and muscular fiber regeneration started after the first week with granulation tissue formation. On the fourth week regenerating muscular fiber areas and granulation tissue were still observed. As a conclusion, we can assume that the injury induced in the snake accident by newborn Bothrops alternatus snakes would be more intense than those induced by the adult ones.

Animals↗

PubMedDate not supplied

ELISA assays for the detection of Bothrops lanceolatus venom in envenomed patient plasmas.

A double antibody sandwich enzyme linked immunosorbant assay (ELISA) was carried out to detect Bothrops Ianceolatus venom in plasma from envenomed patients at various time intervals (0, 6, 12, 18 and 24 hrs). The test could detect Bothrops lanceolatus levels up to 12 ng/mL of envenomed patient plasmas. Elaboration of an easy, fast and species-diagnostic based on this ELISA technique useful to physicians is discussed.

Animals↗

PubMed2004

[Renal injury in rats poisoned by venom of Bothrops neuwiedii diporus from Argentina].

Acute renal failure is one of the systemic complications that can be found in bothropic accidents. In this study the effects on male Wistar rats induced by the venom of Bothrops neuwiedii diporus were evaluated. The histopathology revealed acute tubular necrosis, lesions firstly were observed 3 hours post inoculation of 700 microg of venom. Cortical kidney congestion and granulohialin degeneration of tubular epithelial cells were observed, these lesions achieved a maximum at 24 hours after inoculation. Tubular epithelial hidropic degeneration and dilatation of tubular lumen with hyalin casts were present inclusive up to 4 weeks after inoculation. Biochemical parameter values associated with kidney renal failure were increased 6 hours after venom inoculation (urea: 1.10+/-0.22 g/dl; creatinine: 19.60+/-1.51 mg/dl), but at the end of the first week they decreased till normal values. The urinary density was lower than normal value: 1.005+/-0.001 (p<0.001) and at the end of the first month they oscillated between 1.005 and 1.060 (p<0.001). Renal injury induced by B. neuwiedii diporus could be better appreciated by histopathology than by the routine laboratory assays.

Animals↗

PubMed1994

Clinical and epidemiological aspects of the 'urutu' lance-headed viper (Bothrops alternatus) bite in a Brazilian hospital.

Data on 32 cases of Bothrops alternatus (the 'urutu' lance-headed viper) bite admitted to a Brazilian teaching hospital from 1985 to 1992 were retrospectively surveyed. The majority of the bites occurred within the household area, i.e. in the house or its environs (56.3%), in the 15-49 years age group (74.2%), and in the lower limbs (84.3%). The male to female ratio was 1 to 1. 59.4% of the patients were given the antivenom within two hours of the bite. Bites in the household area were commonest in old ladies, whereas all the cases associated with leisure activities, and the majority of the bites associated with labour in the field occurred in men. All patients had local pain and oedema; prolonged clothing time was present in 96.9% of the cases, haemorrhage in 40.6%, blisters in 21.9% and necrosis in 9.4%. A tourniquet was used by 43.8% of the patients. A median of 4 ampoules of antivenom were administered. There was no death in this series. There was no significant trend between increasing age and necrosis, nor between time interval between bite and antivenom administration and occurrence of blisters or necrosis. Presence of blister(s) was associated with necrosis (p = 0.007), but use of a tourniquet, altered clotting time or presence of haemorrhage were not. These findings, compared with those of case series of bites of other species of Bothrops, and contrary to the popular belief, indicate that B. alternatus bite is not always severe, and has even a lower rate of complications.(ABSTRACT TRUNCATED AT 250 WORDS)

Adolescent↗

PubMed1995

Hematological changes induced by Bothrops jararaca venom in dogs.

The mechanism of consumption coagulopathy observed in cases of human envenomation by Bothrops jararaca is well established. However, this mechanism may vary according to the animal species studied. In order to study both the clinical and laboratory aspects of bothropic envenomation in dogs, a sublethal defibrinating dose of venom (100 micrograms/kg) was given intravenously. A coagulopathy similar to that observed in humans - including fibrinogen depletion, consumption of factors II, X, V and antithrombin III, and moderate thrombocytopenia - was observed. The presence of circulating activated platelets was also noted. Neutrophilic leukocytosis, lymphopenia, and monocytosis occurred at different times. Erythrocytic values remained normal in dogs treated with B. jararaca venom compared with those treated with saline alone. The erythrocyte sedimentation rate fell rapidly after venom administration and this fall was correlated logarithmically with fibrinogen concentration. Since the effects of envenomation in dogs is similar to that in humans, it was concluded that the dog can be used as a good animal model for studying human venom-induced coagulopathy.

Analysis of Variance↗

PubMed1995

Determination of the neutralizing potency of horse antivenom against bothropic and crotalic venoms by indirect enzyme immunoassay.

The use of ELISA to determine antisnake venom potency of horse immune sera should provide benefits of costs and reproducibility compared to in vivo assays. In the present investigation we evaluated the correlation between ELISA antibody levels and in vivo neutralization assays. For the indirect ELISA method, 0.016 micrograms/well of Bothrops jararaca or Crotalus durissus terrificus venom were used to coat the plates and 100 microliters/well of each sample of antibothropic or anticrotalic venom sera were used at 1:10,000 dilution. Sheep anti-horse IgG conjugated to peroxidase was added and the substrate H2O2/o-phenylenediamine produced the color that was read at 492 nm. A correlation coefficient of r = 0.97 was found for anticrotalic venom antibodies and no significant correlation was observed for antibothropic venom sera using 16 serum samples from immunized horses. However, when three antibothropic venom sera showing high in vivo neutralization potency and low absorbance in ELISA or high absorbance values and low in vivo protection were not included in the correlation analysis the coefficient value was r = 0.88. The correlation coefficient did not improve for all 16 antibothropic sera when a partially purified Bothrops jararaca venom fraction was used to coat the ELISA plates. The results indicate that ELISA could be used to determine the neutralizing potency of anticrotalic venom sera. For the antibothropic venom sera further studies are needed.

Animals↗

PubMed1998

Blister formation and skin damage induced by BaP1, a haemorrhagic metalloproteinase from the venom of the snake Bothrops asper.

Blister formation and skin damage can be induced by BaP1, a haemorrhagic metalloproteinase from the venom of the snake Bothrops asper. Pathological changes in the skin were investigated after intramuscular injections of Bothrops asper haemorrhagic metalloproteinase BaP1. Blisters developed within the first hour, with separation of epidermis from the dermal-epidermal junction, whereas acantholysis of epithelial cells was not observed. After the third hour there was ulceration with formation of a proteinaceous scab and inflammatory infiltrate. By 7 to 14 days there was evidence of a regenerative process in dermis and epidermis. Haemorrhage occurred in both dermis and hypodermis as a consequence of BaP1 injection, together with damage of sebaceous glands and an inflammatory reaction in which enlarged macrophages were the predominant cell type. Zymography assays showed the presence of several endogenous metalloproteinases in the exudate, skin homogenates and plasma. In addition, BaP1 was detected in exudates and plasma by immunoblotting. This technique also demonstrated the presence of components immunologically related to laminin and collagen type IV in exudates. It is suggested that BaP1, and probably endogenous matrix metalloproteinases, degrade some protein components at the dermal-epidermal junction, inducing the formation of blisters.

Animals↗

PubMed1995

Purification and characterization of myotoxin IV, a phospholipase A2 variant, from Bothrops asper snake venom.

A basic myotoxic protein was purified from Bothrops asper venom. Like other basic Bothrops myotoxins, myotoxin IV induces acute muscle damage after intramuscular injection in mice and disrupts negatively charged liposomes but not positively charged ones. Furthermore, this protein exerts a weak anticoagulant effect only at concentrations of 40 micrograms/ml or higher, and is devoid of phospholipase A2 activity. Rabbit polyclonal antibodies to B. asper myotoxin II, a related lysine-49 isoform, cross-react strongly with myotoxin IV by enzyme immunoassay, indicating a high degree of antigenic similarity between these two proteins. The fact that both toxins have similar amino acid compositions and amino-terminal sequences, including leucine-5 and glutamine-11, 2 amino acid residues conserved in all lysine-49 phospholipase A2 variants purified, strongly suggests that B. asper myotoxin IV is a lysine-49 phospholipase A2.

Amino Acid Sequence↗

PubMed1978

Components of the renin-angiotensin system in the plasma of Bothrops jararaca.

By means of high voltage electrophoresis experiments it could be demonstrated that the dipeptide hydrolase present in the plasma of Bothrops jararaca is similar to the angiotensin I converting enzyme of human plasma. Therefore, angiotensin I can be considered as a probable natural substrate for this potent snake peptidase in contrast to bradykinin, which is excluded in that case, since this snake plasma was previously found to be deficient in intrinsic kinin releasing system. On the other hand, the presence of angiotensinase activity in this snake plasma could also be demonstrated. Through the pharmacological comparison of angiotensin II with the pressor peptide released from the Bothrops jararaca plasma by chymotrypsin, an indirect indication of the presence of angiotensinogen in the plasma of this reptile was obtained.

Amino Acids↗