Synergistic effect of serine protease inhibitors and a bothropic antivenom in reducing local hemorrhage and coagulopathy caused by Bothrops jararaca venom.

Snakebite accidents are a public health problem that affects the whole world, causing thousands of deaths and amputations each year. In Brazil, snakebite envenomations are caused mostly by snakes from the Bothrops genus. The local symptoms are characterized by pain, swelling, ecchymosis, and hemorrhages. Systemic disturbances can lead to necrosis and amputations. The present treatment consists of intravenous administration of bothropic antivenom, which is capable of reversing most of the systemic symptoms, while presenting limitations to treat the local effects, such as hemorrhage and to neutralize the snake venom serine protease (SVSP). In this context, we aimed to evaluate the activity of selective serine protease inhibitors (pepC and pepB) in combination with the bothropic antivenom in vivo. Further, we assessed their possible synergistic effect in the treatment of coagulopathy and hemorrhage induced by Bothrops jararaca venom. For this, we evaluated the in vivo activity in mouse models of local hemorrhage and a series of in vitro hemostasis assays. Our results showed that pepC and pepB, when combinated with the antivenom, increase its protective activity in vivo and decrease the hemostatic disturbances in vitro with high selectivity, possibly by inhibiting botropic proteases. These data suggest that the addition of serine protease inhibitor to the antivenom can improve its overall potential.

Maintaining rear-fanged snakes for venom production: an evaluation of mortality and survival rates for Philodryas olfersii and P. patagoniensis in captivity

This study evaluates the mortality and average survival rates of captive female Philodryas olfersii and Philodryas patagoniensis snakes maintained for venom production. Also, two factors likely to reduce captive survival were studied - body condition at admission and seasonality. Mortality peaks occurred during the second month in captivity. More than half the individuals were dead at the end of the third month. This suggests that the first three months in captivity are the most critical in terms of survival and adaptation. Females collected and admitted during spring and summer lived less time than those collected in autumn and winter. As gravidity and egg-laying occur during spring and summer, we suggest that the lower survival rates in these seasons may be due to high costs and stress involved in these reproductive events. Unexpectedly, body mass and body condition were poor predictors of survival in captivity. Our results have important implications in maintaining snakes for venom production. We propose some prophylactic measures to minimize the deleterious impacts of captivity during the adaptation period.(AU)

Maintaining rear-fanged snakes for venom production: an evaluation of mortality and survival rates for Philodryas olfersii and P. patagoniensis in captivity

This study evaluates the mortality and average survival rates of captive female Philodryas olfersii and Philodryas patagoniensis snakes maintained for venom production. Also, two factors likely to reduce captive survival were studied - body condition at admission and seasonality. Mortality peaks occurred during the second month in captivity. More than half the individuals were dead at the end of the third month. This suggests that the first three months in captivity are the most critical in terms of survival and adaptation. Females collected and admitted during spring and summer lived less time than those collected in autumn and winter. As gravidity and egg-laying occur during spring and summer, we suggest that the lower survival rates in these seasons may be due to high costs and stress involved in these reproductive events. Unexpectedly, body mass and body condition were poor predictors of survival in captivity. Our results have important implications in maintaining snakes for venom production. We propose some prophylactic measures to minimize the deleterious impacts of captivity during the adaptation period.(AU)

Analysis of the ontogenetic variation in the venom proteome/peptidome of bothrops jararaca reveals different strategies to deal with prey

Previous studies have demonstrated that the pharmacological activities displayed by Bothrops jararaca venom undergo a significant ontogenetic shift. Variation in the venom proteome is a well-documented phenomenon; however, variation in the venom peptidome is poorly understood. We report a comparative proteomic and peptidomic analysis of venoms from newborn and adult specimens of B. jararaca and correlate it with the evaluation of important venom features. We demonstrate that newborn and adult venoms have similar hemorrhagic activities, while the adult venom has a slightly higher lethal activity in mice; however, the newborn venom is extremely more potent to kill chicks. The coagulant activity of newborn venom upon human plasma is 10 times higher than that of adult venom. These differences were clearly reflected in their different profiles of SDS-PAGE, gelatin zimography, immunostaining using specific antibodies, glycosylation pattern, and concanavalin A-binding proteins. Furthermore, we report for the first time the analysis of the peptide fraction of newborn and adult venoms by MALDI-TOF mass spectrometry and LC-MS/MS, which revealed different contents of peptides, while the bradykinin potentiating peptides (BPPs) showed rather similar profiles and were detected in the venoms showing their canonical sequences and also novel sequences corresponding to BPPs processed from their precursor protein at sites so far not described. As a result of these studies, we demonstrated that the ontogenetic shift in diet, from ectothermic prey in early life to endothermic prey in adulthood, and in animal size are associated with changes in the venom proteome in B. jararaca species.

Sexual dimorphism in venom of Bothrops jararaca(Serpentes: Viperidae).

Bothrops jararaca is an abundant snake in Brazil, and its venom has been studied exhaustively. The species exhibits adult size dimorphism in which female are larger. We registered the growth in Snout-Vent Length and weight of one litter (with 11 females and 12 males). We compared growth curves and venom profile between male and female of B. jararaca in order to establish the relationship of those characters and sex. Their venoms were analyzed when they were 36 months old, concerning SDS PAGE, protein content, proteolytic, hyaluronidasic, phospholipasic, blood-clotting, edematogenic, hemorrhagic, myotoxic activities, and lethality. Differences in the growth curves of the females and the males were significantly different after the 12th month of age, with the females growing faster. Females produced five times more venom than males. The electrophoretic patterns were variable: the venom from males had more protein bands than females. Venom composition varied significantly between males and females. Venom from females is more potent for hyaluronidasic, hemorrhagic, and lethality activities, whereas venom from males is more potent for coagulant, phospholipasic, and myotoxic activities. The variability of proteolytic and edematogenic activities were not significant. The important sexual dimorphism in body size and mass, amount of venom produced, and venom composition in B. jararaca may reflect a divergence in niche partitioning.

Some aspects of the venom proteome of the Colubridae snake Philodryas olfersii revealed from a Duvernoy's (venom) gland transcriptome

We investigated the putative toxins of Philodryas olfersii (Colubridae), a representative of a family of snakes neglected in venom studies despite their growing medical importance. Transcriptomic data of the venom gland complemented by proteomic analysis of the gland secretion revealed the presence of major toxin classes from the Viperidae family, including serine proteases, metalloproteases, C-type lectins, Crisps, and a C-type natriuretic peptide (CNP). Interestingly, the phylogenetic analysis of the CNP precursor showed it as a linker between two related precursors found in Viperidae and Elapidae snakes. We suggest that these precursors constitute a monophyletic group derived from the vertebrate CNPs.