Bothrops envenomation in dogs: local and systemic manifestations

Background: Snakebite envenoming is a condition that affects humans and domestic animals worldwide. Identification of the snake species involved in the envenomation is infrequent. Bothrops envenomation presents typical clinicopathological features. This report describes epidemiological, clinical, and pathological data of 2 cases of Bothrops envenomation in dogs, including the first case of Bothrops moojeni snake striking a domestic animal in Brazil. Cases: Case 1. A dog was witnessed to have a Bothrops moojeni snakebite on a farm. In the first 24 h, acute lameness, pain, diffuse swelling, focal bleeding at the left forelimb, and increased whole-blood clotting time were observed in the envenomed dog. Polyvalent antivenom was administered in addition to fluid therapy, analgesics, corticosteroids, and antibiotics. On the 5th day, the animal presented spontaneous bleeding at the wound site, thrombocytopenia, and increased whole-blood clotting time. An additional dose of polyvalent antivenom was administered, and local treatment at the snakebite site was initiated. After 13 days, the dog showed no clinical or laboratory changes and recovered entirely. Case 2. A mongrel dog was taken for a necropsy to determine the cause of death. Grossly, major findings included swelling in the nasal plane that extended to the neck and dissecting hemorrhage in the subcutaneous tissue and adjacent musculature. Hemorrhages were observed in the heart, parietal pleura, left forelimb, lumbar region, and perirenal tissue. Marked necrosis and disruption of small blood vessels and lymphatics within the deep dermis and subcutaneous tissue were the main microscopic findings close to the snakebite site. Additionally, degeneration and necrosis of muscle fibers and dissecting hemorrhage were observed in the head and neck tissues surrounding the snakebite site. Kidneys showed marked interstitial hemorrhage and acute tubular nephrosis. Discussion: Bothrops envenoming is characterized by local (hemorrhage, dermonecrosis, and myonecrosis) and systemic (coagulative disorders, systemic hemorrhage, and acute kidney injury) changes due to the effect of the main venom components such as phospholipase A2 and metalloproteinases. These changes are hallmarks for the bothropic envenomation, supporting the diagnosis in cases 1 and 2. In case 1, the dog developed a Bothrops moojeni snakebite envenomation, but the immediate treatment with antivenom allowed a favorable outcome. In case 2, gross and microscopic findings supported the presumptive diagnosis of fatal bothropic envenomation. A marked local reaction such as swelling, pain, bleeding, bruising, and tissue necrosis was observed in case 1. In case 2, the most significant local changes were swelling and edema at the head and neck, hemorrhage in the subcutaneous tissue, and adjacent musculature. Systemic effects were observed clinically as spontaneous bleeding, thrombocytopenia, increased whole-blood clotting time (Case 1), systemic hemorrhages, and acute tubular nephrosis (Case 2). A proper treatment probably prevented the development of acute renal failure in Case 1. Herein, we show the first case of accidental snakebite envenomation by B. moojeni in a dog in Brazil. Information is scarce on the identification of venomous snake species striking domestic animals. Fast detection of well-determined clinical and pathological findings of Bothrops envenomation is essential for a correct diagnosis, therapeutics, and a good prognosis, even in cases with an unknown history.

Necrotic wound caused by jararaca (Bothrops jararaca) in a dog - hyperbaric oxygen therapy (HBTO)

Background: Snakebites are the main responsible for envenoming in dogs and the bothropic venom remains the mostcommon in Brazil, which can induce a necrotic skin wound. Hyperbaric oxygen therapy (HBOT) use 100% oxygen underhigh pressure and used to treat different wounds in human patients. To the authors’ knowledge, no reports regarding to usethe HBOT in skin wound caused by snakebite (Bothrops jararaca) are present in the literature. The present clinical caseaimed to describe the use of HBOT for the treatment of an extensive necrotic wound caused by jararaca snakebite in a dog.Case: A neutered 8-year-old mixed-breed dog, weighing 12 kg, was admitted with a 7-day history of extensive necroticwound was identified in the face and neck causing by a snakebite, and no sign of pain. The procedure of HBOT (singlesessions of 1.5 ATM, 45 min, repeated every 48 h, up to 12 sessions) was decided, and the complete blood cells, alanineaminotransferase, creatinine, creatine kinase, prothrombin time, activated partial thromboplastin time, wound clinicalevaluation were measured at the following time-points: 2nd, 5th, 10th, and 12th sessions. At the 5th session was identifiedleukopenia, neutropenia and lymphopenia. Wound re-epithelialization was initiated after the 5th session, and the completeepithelialization was identified at the 12th session of HBOT. During the HBOT no side effects were identified. Threemonths after the HBOT finished, the animal returned to the clinic and the clinical status evolved positively, and the woundwas completed healed.Discussion: This report described the treatment of an extensive necrotic skin wound caused by snakebite (Bothrops jararaca)in an 8-year-old, neutered, mixed-breed dog using the HBOT. The wound healing...

Necrotic wound caused by jararaca (Bothrops jararaca) in a dog - hyperbaric oxygen therapy (HBTO)

Background: Snakebites are the main responsible for envenoming in dogs and the bothropic venom remains the mostcommon in Brazil, which can induce a necrotic skin wound. Hyperbaric oxygen therapy (HBOT) use 100% oxygen underhigh pressure and used to treat different wounds in human patients. To the authors knowledge, no reports regarding to usethe HBOT in skin wound caused by snakebite (Bothrops jararaca) are present in the literature. The present clinical caseaimed to describe the use of HBOT for the treatment of an extensive necrotic wound caused by jararaca snakebite in a dog.Case: A neutered 8-year-old mixed-breed dog, weighing 12 kg, was admitted with a 7-day history of extensive necroticwound was identified in the face and neck causing by a snakebite, and no sign of pain. The procedure of HBOT (singlesessions of 1.5 ATM, 45 min, repeated every 48 h, up to 12 sessions) was decided, and the complete blood cells, alanineaminotransferase, creatinine, creatine kinase, prothrombin time, activated partial thromboplastin time, wound clinicalevaluation were measured at the following time-points: 2nd, 5th, 10th, and 12th sessions. At the 5th session was identifiedleukopenia, neutropenia and lymphopenia. Wound re-epithelialization was initiated after the 5th session, and the completeepithelialization was identified at the 12th session of HBOT. During the HBOT no side effects were identified. Threemonths after the HBOT finished, the animal returned to the clinic and the clinical status evolved positively, and the woundwas completed healed.Discussion: This report described the treatment of an extensive necrotic skin wound caused by snakebite (Bothrops jararaca)in an 8-year-old, neutered, mixed-breed dog using the HBOT. The wound healing...(AU)

Venom composition of Trimeresurus albolabris, T. insularis, T. puniceus and T. purpureomaculatus from Indonesia

Background: Several studies have been published on the characterization of Trimeresurus venoms. However, there is still limited information concerning the venom composition of Trimeresurus species distributed throughout Indonesia, which contributes to significant snakebite envenomation cases. The present study describes a comparative on the composition of T. albolabris, T. insularis, T. puniceus, and T. purpureomaculatus venoms originated from Indonesia. Methods: Protein content in the venom of four Trimeresurus species was determined using Bradford assay, and the venom proteome was elucidated using one-dimension SDS PAGE nano-ESI- LCMS/MS shotgun proteomics. Results: The venom of T. albolabris contained the highest protein content of 11.1 mg/mL, followed by T. puniceus, T. insularis and T. purpureomaculatus venom with 10.7 mg/mL, 8.9 mg/mL and 5.54 mg/mL protein, respectively. In total, our venomic analysis identified 65 proteins belonging to 16 protein families in T. purpureomaculatus; 64 proteins belonging to 18 protein families in T. albolabris; 58 different proteins belonging to 14 protein families in T. puniceus; and 48 different proteins belonging to 14 protein familiesin T. insularis. Four major proteins identified in all venoms belonged to snake venom metalloproteinase, C-type lectin, snake venom serine protease, and phospholipase A2. There were 11 common proteins in all venoms, and T. puniceus venom has the highest number of unique proteins compared to the other three venoms. Cluster analysis of the proteins and venoms showed that T. puniceus venom has the most distinct venom composition. Conclusions: Overall, the results highlighted venom compositional variation of four Trimeresurus spp. from Indonesia. The venoms appear to be highly similar, comprising at least four protein families that correlate with venom's toxin properties and function. This study adds more information on venom variability among Trimeresurus species within the close geographic origin and may contribute to the development of optimum heterologous antivenom.(AU)

Guidelines for the diagnosis and differential diagnosis of snakebite envenoming of ruminants in Brazil / Diretrizes para o diagnóstico e diagnóstico diferencial de envenenamento por picadas de cobra em ruminantes no Brasil

In Brazil, snakebites are often cited as a cause of mortality in ruminants, but there are discrepancies in the literature regarding its actual prevalence, either by lack of diagnosis or by mistakes in the differential diagnosis. Among the factors that hinder the diagnosis are included the inconsistencies to distinguish between accidents caused by Bothrops and Crotalus, responsible for over 90% of the cases. For the diagnosis of accidents involving Lachesis muta, both the neurotropic and the proteolytic/hemolytic effects must be considered, similar to what is described in Crotalus scutulatus. This article describes the main clinical, pathological, and laboratory findings observed in envenoming by the aforementioned snakes and suggests procedures for establishing the diagnosis and differential diagnosis starting from a logical sequence, based on epidemiological evidence, clinical, laboratory, and pathological findings.

No Brasil, acidentes ofídicos são frequentemente citados como causa de mortalidade em ruminantes, mas existem discrepâncias em relação a sua atual prevalência, seja por falta de diagnóstico ou por erros no diagnóstico diferencial. Entre os fatores que dificultam o diagnóstico estão as inconsistências para distinguir entre os acidentes causados por Bothrops e Crotalus, responsáveis por mais de 90% dos casos. Para o diagnóstico de envenenamentos por Lachesis muta, devem ser considerados os efeitos neurotrópico e proteolítico/hemolíticos concomitantes, a exemplo do que ocorre com algumas cascavéis norte-americanas (Crotalus scutulatus, entre elas). Este artigo descreve os principais achados clinicopatológicos e laboratoriais observados em casos de envenenamento pelas serpentes citadas e sugere um roteiro simplificado para o estabelecimento do diagnóstico e diagnóstico diferencial, a partir de uma sequência lógica, baseada em evidências epidemiológicas e achados clínicos, laboratoriais e patológicos.

P-I metalloproteinases and L-amino acid oxidases from Bothrops species inhibit angiogenesis

Snake venoms are composed of pharmacologically active proteins that are evolutionarily diverse, stable and specific to targets. Hence, venoms have been explored as a source of bioactive molecules in treating numerous diseases. Recent evidences suggest that snake venom proteins may affect the formation of new blood vessels. Excessive angiogenesis has been implicated in several pathologies including tumours, diabetic retinopathy, arthritis, inter alia. In the present study, we have examined the effects of P-I metalloproteinases isolated from Bothrops moojeni (BmMP-1) and Bothrops atrox (BaMP-1) and L-amino acid oxidases (LAAO) isolated from B. moojeni (BmLAAO) and B. atrox (BaLAAO) on biochemical and functional aspects of angiogenesis. Methods: P-I metalloproteinases and LAAO were purified from venom by molecular size exclusion and ion-exchange chromatography and subsequently confirmed using mass spectrometry. The P-I metalloproteinases were characterized by azocaseinolytic, fibrinogenolytic and gelatinase activity and LAAO activity was assessed by enzyme activity on L-amino acids. Influence of these proteins on apoptosis and cell cycle in endothelial cells was analysed by flow cytometry. The angiogenic activity was determined by in vitro 3D spheroid assay, Matrigel tube forming assay, and in vivo agarose plug transformation in mice. Results: P-I metalloproteinases exhibited azocaseinolytic activity, cleaved α and partially β chain of fibrinogen, and displayed catalytic activity on gelatin. LAAO showed differential activity on L-amino acids. Flow cytometry analysis indicated that both P-I metalloproteinases and LAAO arrested the cells in G0/G1 phase and further induced both necrosis and apoptosis in endothelial cells. In vitro, P-I metalloproteinases and LAAO exhibited significant anti-angiogenic properties in 3D spheroid and Matrigel models by reducing sprout outgrowth and tube formation. Using agarose plug transplants in mice harbouring P-I metalloproteinases and LAAO we demonstrated a marked disruption of vasculature at the periphery. Conclusion: Our research suggests that P-I metalloproteinases and LAAO exhibit anti-angiogenic properties in vitro and in vivo.(AU)

P-I metalloproteinases and L-amino acid oxidases from Bothrops species inhibit angiogenesis

Snake venoms are composed of pharmacologically active proteins that are evolutionarily diverse, stable and specific to targets. Hence, venoms have been explored as a source of bioactive molecules in treating numerous diseases. Recent evidences suggest that snake venom proteins may affect the formation of new blood vessels. Excessive angiogenesis has been implicated in several pathologies including tumours, diabetic retinopathy, arthritis, inter alia. In the present study, we have examined the effects of P-I metalloproteinases isolated from Bothrops moojeni (BmMP-1) and Bothrops atrox (BaMP-1) and L-amino acid oxidases (LAAO) isolated from B. moojeni (BmLAAO) and B. atrox (BaLAAO) on biochemical and functional aspects of angiogenesis. Methods: P-I metalloproteinases and LAAO were purified from venom by molecular size exclusion and ion-exchange chromatography and subsequently confirmed using mass spectrometry. The P-I metalloproteinases were characterized by azocaseinolytic, fibrinogenolytic and gelatinase activity and LAAO activity was assessed by enzyme activity on L-amino acids. Influence of these proteins on apoptosis and cell cycle in endothelial cells was analysed by flow cytometry. The angiogenic activity was determined by in vitro 3D spheroid assay, Matrigel tube forming assay, and in vivo agarose plug transformation in mice. Results: P-I metalloproteinases exhibited azocaseinolytic activity, cleaved α and partially β chain of fibrinogen, and displayed catalytic activity on gelatin. LAAO showed differential activity on L-amino acids. Flow cytometry analysis indicated that both P-I metalloproteinases and LAAO arrested the cells in G0/G1 phase and further induced both necrosis and apoptosis in endothelial cells. In vitro, P-I metalloproteinases and LAAO exhibited significant anti-angiogenic properties in 3D spheroid and Matrigel models by reducing sprout outgrowth and tube formation. Using agarose plug transplants in mice harbouring P-I metalloproteinases and LAAO we demonstrated a marked disruption of vasculature at the periphery. Conclusion: Our research suggests that P-I metalloproteinases and LAAO exhibit anti-angiogenic properties in vitro and in vivo.(AU)

Design, synthesis, and evaluation of Bothrops venom serine protease peptidic inhibitors

In Central and South America, snakebite envenomation is mainly caused by Bothrops spp. snakes, whose venoms feature significant biochemical richness, including serine proteases. The available bothropic antivenoms are efficient in avoiding fatalities, but do not completely neutralize venom serine proteases, which are co-responsible for some disorders observed during envenomation. Methods: In order to search for tools to improve the antivenom's, 6-mer peptides were designed based on a specific substrate for Bothrops jararaca venom serine proteases, and then synthesized, with the intention to selectively inhibit these enzymes. Results: Using batroxobin as a snake venom serine protease model, two structurally similar inhibitor peptides were identified. When tested on B. jararaca venom, one of the new inhibitors displayed a good potential to inhibit the activity of the venom serine proteases. These inhibitors do not affect human serine proteases as human factor Xa and thrombin, due to their selectivity. Conclusion: Our study identified two small peptides able to inhibit bothropic serine proteases, but not human ones, can be used as tools to enhance knowledge of the venom composition and function. Moreover, one promising peptide (pepC) was identified that can be explored in the search for improving Bothrops spp. envenomation treatment.(AU)

Design, synthesis, and evaluation of Bothrops venom serine protease peptidic inhibitors

In Central and South America, snakebite envenomation is mainly caused by Bothrops spp. snakes, whose venoms feature significant biochemical richness, including serine proteases. The available bothropic antivenoms are efficient in avoiding fatalities, but do not completely neutralize venom serine proteases, which are co-responsible for some disorders observed during envenomation. Methods: In order to search for tools to improve the antivenom's, 6-mer peptides were designed based on a specific substrate for Bothrops jararaca venom serine proteases, and then synthesized, with the intention to selectively inhibit these enzymes. Results: Using batroxobin as a snake venom serine protease model, two structurally similar inhibitor peptides were identified. When tested on B. jararaca venom, one of the new inhibitors displayed a good potential to inhibit the activity of the venom serine proteases. These inhibitors do not affect human serine proteases as human factor Xa and thrombin, due to their selectivity. Conclusion: Our study identified two small peptides able to inhibit bothropic serine proteases, but not human ones, can be used as tools to enhance knowledge of the venom composition and function. Moreover, one promising peptide (pepC) was identified that can be explored in the search for improving Bothrops spp. envenomation treatment.(AU)

Hemostatic evaluation of rabbits envenomed with Bothrops alternatus treated with anti-bothropic serum, desmopressin and tranexamin acid / Avaliação hemostática de coelhos envenenados com Bothrops alternatus e tratados com soro antibotrópico, desmopressina e ácido tranexâmico

In Brazil, snakes from the Bothrops genus are responsible for thousands of accidents, and their venoms are mainly composed of proteolytic enzymes. Although the antibothropic serum produced by the Brazilian Institutes is remarkably efficient, more studies are necessary, especially in veterinary medicine. The venom contain enzymes and non-enzymatic proteins that interfere with hemostasis leading to hemorrhage or even thrombosis. Possible treatment associations with known bothropic antivenom were the reason for the development of the present study. The aim of this study was to evaluate hemostasis alterations caused by Bothrops alternatus venom in rabbits followed by treatments with anti-bothropic serum, tranexamic acid and desmopressin. Twenty New Zealand rabbits were distributed into five groups (n=4) that were experimentally envenomed with 150mcg/kg of B. alternatus venom via intramuscular injection and treated as follow: Group 1 (G1) was the positive control and received venom and PBS/BSA; Group 2 (G2) was treated with tranexamic acid; Group 3 (G3) with desmopressin; Group 4 (G4) with tranexamic acid and anti-bothropic serum; and Group 5 (G5) with anti-bothropic serum and desmopressin. Blood samples were collected before venom administration, and one, four, eight and 12 hours after, for Partial activated partial thromboplastin time, Prothrombin Time, Thrombin Time and fibrinogen evaluation. Thrombin generation (TG) test was carried out with a pool of samples from final times (8 and 12h). At the end of 12h, all animals were euthanized and necropsy was conducted. Samples from muscle tissue, heart, lungs and kidney were analyzed. Classic coagulation tests showed no significant differences amongst groups and times. However, TG indicated that the venom causes a hypocoagulability state, which was not reversed by proposed treatments. Histology showed muscle inflammation, hemorrhage and necrosis, as well as hemorrhage in other tissues with no differences amongst groups. B. alternatus envenomation causes hypocoagulability detected by TG assay, but not through classical coagulation tests. The use of tranexamic acid and desmopressin for hemostasis stabilization after inoculation of the venom did not show advantage in coagulation restoration.(AU)

No Brasil, as serpentes do gênero Bothrops são responsáveis por milhares de acidentes, e seus venenos são compostos principalmente de enzimas proteolíticas. Embora o soro antiofídico produzido pelos institutos brasileiros seja notavelmente eficiente, mais estudos são necessários, especialmente na medicina veterinária. O veneno contem enzimas e proteínas não-enzimáticas que interferem com a hemostasia levando a hemorragias ou trombose. A associação de outros tratamentos ao soro antibotrópico foi a razão para o desenvolvimento do presente estudo. O objetivo deste estudo foi avaliar as alterações da hemostasia causadas pelo veneno de Bothrops alternatus em coelhos, após tratamento com soro antibotrópico, ácido tranexâmico e desmopressina. Vinte coelhos da Nova Zelândia foram distribuídos em cinco grupos (n = 4) que foram submetidos a experimentos com 150mcg/kg de veneno de B. alternatus por injeção intramuscular. O Grupo 1 (G1) foi o controle positivo e recebeu veneno e PBS / BSA, enquanto o Grupo 2 (G2) foi tratado com ácido tranexâmico, o Grupo 3 (G3) com desmopressina, o Grupo 4 (G4) com ácido tranexâmico e soro antibotrópico, e o Grupo 5 (G5) com soro antibotrópico e desmopressina. As amostras de sangue foram coletadas antes da administração do veneno, e uma, quatro, oito e 12 horas após os tratamentos para realização de tempo de tromboplastina parcial ativada parcial (TTPa), tempo de protrombina (TP), tempo de trombina (TT) e mensuração de fibrinogênio. Para o ensaio de geração de trombina (TG) foi realizado com um pool de amostras nos tempos finais (8 e 12h). Ao final das 12h, todos os animais foram sacrificados e a necropsia foi realizada. Amostras de tecido muscular, coração, pulmões e rins foram analisadas. Os testes TTPa, TP, TT e fibrinogênio não mostraram diferenças significativas entre os grupos e os tempos. No entanto, o TG indicou que o veneno causa um estado de hipocoagulabilidade, que não foi revertido pelos tratamentos propostos. Na histologia, foram observadas inflamação muscular, hemorragia e necrose, além de hemorragia em outros tecidos, sem diferenças entre os grupos. O envenenamento por B. alternatus causa hipocoagulabilidade detectada mais precocemente pelo teste de geração de trombina. O uso de ácido tranexâmico e desmopressina para estabilização da hemostasia após a inoculação do veneno não mostrou vantagem na restauração da coagulação.(AU)

Hemostatic evaluation of rabbits envenomed with Bothrops alternatus treated with anti-bothropic serum, desmopressin and tranexamin acid / Avaliação hemostática de coelhos envenenados com Bothrops alternatus e tratados com soro antibotrópico, desmopressina e ácido tranexâmico

In Brazil, snakes from the Bothrops genus are responsible for thousands of accidents, and their venoms are mainly composed of proteolytic enzymes. Although the antibothropic serum produced by the Brazilian Institutes is remarkably efficient, more studies are necessary, especially in veterinary medicine. The venom contain enzymes and non-enzymatic proteins that interfere with hemostasis leading to hemorrhage or even thrombosis. Possible treatment associations with known bothropic antivenom were the reason for the development of the present study. The aim of this study was to evaluate hemostasis alterations caused by Bothrops alternatus venom in rabbits followed by treatments with anti-bothropic serum, tranexamic acid and desmopressin. Twenty New Zealand rabbits were distributed into five groups (n=4) that were experimentally envenomed with 150mcg/kg of B. alternatus venom via intramuscular injection and treated as follow: Group 1 (G1) was the positive control and received venom and PBS/BSA; Group 2 (G2) was treated with tranexamic acid; Group 3 (G3) with desmopressin; Group 4 (G4) with tranexamic acid and anti-bothropic serum; and Group 5 (G5) with anti-bothropic serum and desmopressin. Blood samples were collected before venom administration, and one, four, eight and 12 hours after, for Partial activated partial thromboplastin time, Prothrombin Time, Thrombin Time and fibrinogen evaluation. Thrombin generation (TG) test was carried out with a pool of samples from final times (8 and 12h). At the end of 12h, all animals were euthanized and necropsy was conducted. Samples from muscle tissue, heart, lungs and kidney were analyzed. Classic coagulation tests showed no significant differences amongst groups and times. However, TG indicated that the venom causes a hypocoagulability state, which was not reversed by proposed treatments. Histology showed muscle inflammation, hemorrhage and necrosis, as well as hemorrhage in other tissues with no differences amongst groups. B. alternatus envenomation causes hypocoagulability detected by TG assay, but not through classical coagulation tests. The use of tranexamic acid and desmopressin for hemostasis stabilization after inoculation of the venom did not show advantage in coagulation restoration.(AU)

No Brasil, as serpentes do gênero Bothrops são responsáveis por milhares de acidentes, e seus venenos são compostos principalmente de enzimas proteolíticas. Embora o soro antiofídico produzido pelos institutos brasileiros seja notavelmente eficiente, mais estudos são necessários, especialmente na medicina veterinária. O veneno contem enzimas e proteínas não-enzimáticas que interferem com a hemostasia levando a hemorragias ou trombose. A associação de outros tratamentos ao soro antibotrópico foi a razão para o desenvolvimento do presente estudo. O objetivo deste estudo foi avaliar as alterações da hemostasia causadas pelo veneno de Bothrops alternatus em coelhos, após tratamento com soro antibotrópico, ácido tranexâmico e desmopressina. Vinte coelhos da Nova Zelândia foram distribuídos em cinco grupos (n = 4) que foram submetidos a experimentos com 150mcg/kg de veneno de B. alternatus por injeção intramuscular. O Grupo 1 (G1) foi o controle positivo e recebeu veneno e PBS / BSA, enquanto o Grupo 2 (G2) foi tratado com ácido tranexâmico, o Grupo 3 (G3) com desmopressina, o Grupo 4 (G4) com ácido tranexâmico e soro antibotrópico, e o Grupo 5 (G5) com soro antibotrópico e desmopressina. As amostras de sangue foram coletadas antes da administração do veneno, e uma, quatro, oito e 12 horas após os tratamentos para realização de tempo de tromboplastina parcial ativada parcial (TTPa), tempo de protrombina (TP), tempo de trombina (TT) e mensuração de fibrinogênio. Para o ensaio de geração de trombina (TG) foi realizado com um pool de amostras nos tempos finais (8 e 12h). Ao final das 12h, todos os animais foram sacrificados e a necropsia foi realizada. Amostras de tecido muscular, coração, pulmões e rins foram analisadas. Os testes TTPa, TP, TT e fibrinogênio não mostraram diferenças significativas entre os grupos e os tempos. No entanto, o TG indicou que o veneno causa um estado de hipocoagulabilidade, que não foi revertido pelos tratamentos propostos. Na histologia, foram observadas inflamação muscular, hemorragia e necrose, além de hemorragia em outros tecidos, sem diferenças entre os grupos. O envenenamento por B. alternatus causa hipocoagulabilidade detectada mais precocemente pelo teste de geração de trombina. O uso de ácido tranexâmico e desmopressina para estabilização da hemostasia após a inoculação do veneno não mostrou vantagem na restauração da coagulação.(AU)

Seasonal variations of male reproductive parameters of Tomodon dorsatus from Southeastern Brazil / Variações sazonais dos parâmetros reprodutivos do macho de Tomodon dorsatus do sudeste brasileiro

The morphology of the male reproductive tract of Tomodon dorsatus was described in the austral seasons of the year considering macroscopic and microscopic variables. For this purpose, 56 specimens from the herpetological collection of the "Instituto Butantan" were used. Fragments of the testes, kidneys and ductus deferens were collected and submitted to histological routine. The peak of the testicular volume was observed in the summer and the epithelium of the seminiferous tubules had higher height in the summer (p=0.001). The testes were active throughout the year, however, the spermiogenesis peaked in the summer. There were spermatozoa in the lumen of the ductus deferens in all seasons of the year. Renal length was higher in autumn (p=0.027), and renal width did not show a significant increase (p=0.237). The diameter and epithelial height of the sexual segment of the kidney (SSK) showed hypertrophy in winter and spring, coinciding with the mating period. Based on findings of this study, we can suggest that, at the population level, the reproductive cycle of T. dorsatus can be considered seasonal semi-synchronous, due to the peak of spermiogenic activity in the hot season, and discontinuous at the individual level.(AU)

A morfologia do trato reprodutivo do macho de Tomodon dorsatus foi descrita nas estações climáticas do ano com base em variáveis macroscópicas e microscópicas. Para isto, foram usados 56 espécimes oriundos da coleção herpetológica do Instituto Butantan. Fragmentos dos testículos, rins e ductos deferentes foram coletados e submetidos à rotina histológica. O volume testicular foi maior no verão e o epitélio dos túbulos seminíferos mostrou uma maior altura no verão (p=0.001). Os testículos estavam ativos durante todo o ano, contudo, a espermiogênese foi maior no verão. Espermatozoides foram encontrados no lúmen do ducto deferente em todas as estações do ano. O comprimento renal foi maior no outono (p=0.027), e a largura renal não mostrou um aumento significativo (p=0.237). O diâmetro e a altura epitelial do segmento sexual do rim (SSR) mostrou hipertrofia nas estações inverno e primavera, coincidindo com o período reprodutivo. Com base nestes resultados, pode-se sugerir que, em nível populacional, o ciclo reprodutivo de T. dorsatus possa ser considerado semi-sincrônico sazonal, devido à atividade espermiogênica na estação quente, e descontínuo em nível individual.(AU)

Seasonal variations of male reproductive parameters of Tomodon dorsatus from Southeastern Brazil / Variações sazonais dos parâmetros reprodutivos do macho de Tomodon dorsatus do sudeste brasileiro

The morphology of the male reproductive tract of Tomodon dorsatus was described in the austral seasons of the year considering macroscopic and microscopic variables. For this purpose, 56 specimens from the herpetological collection of the "Instituto Butantan" were used. Fragments of the testes, kidneys and ductus deferens were collected and submitted to histological routine. The peak of the testicular volume was observed in the summer and the epithelium of the seminiferous tubules had higher height in the summer (p=0.001). The testes were active throughout the year, however, the spermiogenesis peaked in the summer. There were spermatozoa in the lumen of the ductus deferens in all seasons of the year. Renal length was higher in autumn (p=0.027), and renal width did not show a significant increase (p=0.237). The diameter and epithelial height of the sexual segment of the kidney (SSK) showed hypertrophy in winter and spring, coinciding with the mating period. Based on findings of this study, we can suggest that, at the population level, the reproductive cycle of T. dorsatus can be considered seasonal semi-synchronous, due to the peak of spermiogenic activity in the hot season, and discontinuous at the individual level.(AU)

A morfologia do trato reprodutivo do macho de Tomodon dorsatus foi descrita nas estações climáticas do ano com base em variáveis macroscópicas e microscópicas. Para isto, foram usados 56 espécimes oriundos da coleção herpetológica do Instituto Butantan. Fragmentos dos testículos, rins e ductos deferentes foram coletados e submetidos à rotina histológica. O volume testicular foi maior no verão e o epitélio dos túbulos seminíferos mostrou uma maior altura no verão (p=0.001). Os testículos estavam ativos durante todo o ano, contudo, a espermiogênese foi maior no verão. Espermatozoides foram encontrados no lúmen do ducto deferente em todas as estações do ano. O comprimento renal foi maior no outono (p=0.027), e a largura renal não mostrou um aumento significativo (p=0.237). O diâmetro e a altura epitelial do segmento sexual do rim (SSR) mostrou hipertrofia nas estações inverno e primavera, coincidindo com o período reprodutivo. Com base nestes resultados, pode-se sugerir que, em nível populacional, o ciclo reprodutivo de T. dorsatus possa ser considerado semi-sincrônico sazonal, devido à atividade espermiogênica na estação quente, e descontínuo em nível individual.(AU)

In vivo and in vitro prospection of the anti-ophidic properties exercised by the extracts of Jacaranda decurrens L

The research and development of alternative treatments for snakebites (e.g., medicinal plants) is necessary due to the high costs of the existing ones. The effects of the aqueous extracts from Jacaranda decurrens leaves, roots, and xylopodium were analyzed upon the venom-induced (Bothrops spp. and Crotalus spp.) systemic and local toxicity. The extracts were able to partially inhibit the phospholipase activity of the venoms from Bothrops jararacussu and Crotalus durissus terrificus. The myotoxic, edema-inducing, coagulant, and hemorrhagic activities were also inhibited. The SDS-PAGE showed that the venom proteins were intact after their incubation with the extracts. This suggests that the possible mechanism of inhibition is not related to the degradation of the protein but rather to their binding to specific sites of the enzymes. The extracts significantly prolonged the survival time of animals in the lethality assay performed with Crotalus durissus terrificus venom and its toxin (crotoxin). The anti-ophidic activity of medicinal plants may aid in the management of snakebites in distant locations by reducing the victim’s local effects and time to heal.

In vivo and in vitro prospection of the anti-ophidic properties exercised by the extracts of Jacaranda decurrens L

The research and development of alternative treatments for snakebites (e.g., medicinal plants) is necessary due to the high costs of the existing ones. The effects of the aqueous extracts from Jacaranda decurrens leaves, roots, and xylopodium were analyzed upon the venom-induced (Bothrops spp. and Crotalus spp.) systemic and local toxicity. The extracts were able to partially inhibit the phospholipase activity of the venoms from Bothrops jararacussu and Crotalus durissus terrificus. The myotoxic, edema-inducing, coagulant, and hemorrhagic activities were also inhibited. The SDS-PAGE showed that the venom proteins were intact after their incubation with the extracts. This suggests that the possible mechanism of inhibition is not related to the degradation of the protein but rather to their binding to specific sites of the enzymes. The extracts significantly prolonged the survival time of animals in the lethality assay performed with Crotalus durissus terrificus venom and its toxin (crotoxin). The anti-ophidic activity of medicinal plants may aid in the management of snakebites in distant locations by reducing the victims local effects and time to heal.(AU)

Antimicrobial effect of bothropstoxin-i in broilers / Atividade antimicrobiana da bothropstoxin-i em frangos de corte

Bacterial resistance is a sanitary issue explained by indiscriminate use of nonprescription drugs, and antimicrobial use in food production for growth promotion. Bothropstoxin-I (BthTx-I) is a phospholipase A2 (PLA2) from Bothrops jararacussu venom, which has a known antimicrobial effect. The goal of this study was the unprecedented evaluation of in vivo antimicrobial activity of BthTx-I in broilers. Microbiological, biochemical, and histological parameters were determined using 84 21-day old broilers that were kept in cages with four birds each at a density of 625 cm2/broiler. The experiment was randomized by three treatments with seven repetitions of four broilers each that lasted seven days. The treatments were: 1) bacitracin zinc diet; 2) PLA2-BthTx-I; 3) without additives. The data obtained from the studied variables was subjected to analysis of variance and an F-test at the 5% significance level. Averages of each variable in each treatment were compared by Tukey’s test. Broiler bacterial cloacal counts showed that BthTx-I decreased the microbial population without reducing body weight, intestinal morphology, or liver or kidney histopathological damage. The toxin showed in vivo activity, being an alternative for better performance in the production of broiler chickens, because it acted by decreasing the microbial load of potentially pathogenic bacteria in the intestinal(AU)

A resistência bacteriana é uma questão sanitária, explicada pelo uso indiscriminado de medicamentos sem receita médica e pelo uso de antimicrobianos na produção de alimentos para promover o crescimento. Bothropstoxin-I (BthTx-I) é uma fosfolipase A2 (PLA2) obtida do veneno da Bothrops jararacussu. A PLA2 do veneno de cobra tem efeito antimicrobiano conhecido. Objetivou-se com este estudo avaliar sem precedentes a atividade antimicrobiana in vivo de BthTx-I em frangos de corte. Os parâmetros microbiológicos, bioquímicos e histológicos foram realizados em 84 frangos de corte com 21 dias de idade mantidos em gaiolas com quatro animais cada e densidade de 625 cm2/frango. O experimento foi dividido em três tratamentos com sete repetições de quatro frangos cada um, com duração de sete dias. Os tratamentos foram: 1) dieta com bacitracina de zinco; 2) PLA2-BthTx-I; 3) sem aditivos. Os dados obtidos das variáveis estudadas foram submetidos à análise de variância e teste F ao nível de significância de 5%. As médias dos tratamentos em cada variável foram comparadas pelo teste de Tukey. A contagem cloacal bacteriana de frangos de corte mostrou que o BthTx-I diminuiu a população microbiana sem comprometer o peso corporal, a morfologia intestinal ou causar danos histopatológico no fígado e rins. Concluiu-se que a toxina apresentou atividade in vivo, sendo uma alternativa para um melhor desempenho na produção de frangos de corte, pois agiu diminuindo a carga microbiana de bactérias potencialmente patogênicas na microbiota intestinal das aves e não causou danos musculares, hepáticos ou renais na dosagem avaliada.(AU)

Antimicrobial effect of bothropstoxin-i in broilers / Atividade antimicrobiana da bothropstoxin-i em frangos de corte

Bacterial resistance is a sanitary issue explained by indiscriminate use of nonprescription drugs, and antimicrobial use in food production for growth promotion. Bothropstoxin-I (BthTx-I) is a phospholipase A2 (PLA2) from Bothrops jararacussu venom, which has a known antimicrobial effect. The goal of this study was the unprecedented evaluation of in vivo antimicrobial activity of BthTx-I in broilers. Microbiological, biochemical, and histological parameters were determined using 84 21-day old broilers that were kept in cages with four birds each at a density of 625 cm2/broiler. The experiment was randomized by three treatments with seven repetitions of four broilers each that lasted seven days. The treatments were: 1) bacitracin zinc diet; 2) PLA2-BthTx-I; 3) without additives. The data obtained from the studied variables was subjected to analysis of variance and an F-test at the 5% significance level. Averages of each variable in each treatment were compared by Tukey’s test. Broiler bacterial cloacal counts showed that BthTx-I decreased the microbial population without reducing body weight, intestinal morphology, or liver or kidney histopathological damage. The toxin showed in vivo activity, being an alternative for better performance in the production of broiler chickens, because it acted by decreasing the microbial load of potentially pathogenic bacteria in the intestinal

A resistência bacteriana é uma questão sanitária, explicada pelo uso indiscriminado de medicamentos sem receita médica e pelo uso de antimicrobianos na produção de alimentos para promover o crescimento. Bothropstoxin-I (BthTx-I) é uma fosfolipase A2 (PLA2) obtida do veneno da Bothrops jararacussu. A PLA2 do veneno de cobra tem efeito antimicrobiano conhecido. Objetivou-se com este estudo avaliar sem precedentes a atividade antimicrobiana in vivo de BthTx-I em frangos de corte. Os parâmetros microbiológicos, bioquímicos e histológicos foram realizados em 84 frangos de corte com 21 dias de idade mantidos em gaiolas com quatro animais cada e densidade de 625 cm2/frango. O experimento foi dividido em três tratamentos com sete repetições de quatro frangos cada um, com duração de sete dias. Os tratamentos foram: 1) dieta com bacitracina de zinco; 2) PLA2-BthTx-I; 3) sem aditivos. Os dados obtidos das variáveis estudadas foram submetidos à análise de variância e teste F ao nível de significância de 5%. As médias dos tratamentos em cada variável foram comparadas pelo teste de Tukey. A contagem cloacal bacteriana de frangos de corte mostrou que o BthTx-I diminuiu a população microbiana sem comprometer o peso corporal, a morfologia intestinal ou causar danos histopatológico no fígado e rins. Concluiu-se que a toxina apresentou atividade in vivo, sendo uma alternativa para um melhor desempenho na produção de frangos de corte, pois agiu diminuindo a carga microbiana de bactérias potencialmente patogênicas na microbiota intestinal das aves e não causou danos musculares, hepáticos ou renais na dosagem avaliada.

Mycoplasma spp. in captive snakes (Boa constrictor and Bothrops atrox) from Brazil / Mycoplasma spp. em cobras de cativeiro (Boa constrictor e Bothrops atrox) do Brasil

Although rare, mycoplasmas are included among the causes of respiratory diseases in reptiles and, in the order Squamata, three reports of these microorganisms causing diseases in pythons have already been reported. This study aimed to evaluate the occurrence of Mycoplasma species in captive snakes. A total of 26 snakes of the families Pythonidae (13), Boidae (7), Viperidae (5) and Colubridae (1) from RioZoo, Brazil, were evaluated. Animals were examined to determine clinical signs consistent with any infectious disease. Tracheal swab samples from snakes were collected in Frey medium and analyzed for the presence of Mycoplasma spp.by isolation and a genus-specific PCR. DNA sequencing analyses of six positive samples by PCR were carried out to identify the species. Using isolation 19.23% (5/26) was positive, while 65.38% (17/26) of the animals were positive by PCR. Based on the analyses of the six sequences obtained, there was similarity with a Mycoplasma spp. previously described in a phyton and, M. agassizii and M. testudineum reported in chelonians. This is the first report of Mycoplasma spp. in animals of the families Boidae and Viperidae. Mycoplasma spp. were detected in snakes with and without clinical signs. The mycoplasmas reported resented identity (range, 95% to 100%) to others already described in reptiles. There was no relationship between the presence of Mycoplasma spp. and clinical signs.(AU)

Embora raros, os micoplasmas estão incluídos entre as causas de doenças respiratórias em répteis e, na ordem Squamata, já foram realizados três relatos destes microrganismos causando doença em pítons. Este estudo teve como objetivo avaliar a ocorrência de espécies de Mycoplasma em serpentes em cativeiro. Foram avaliadas 26 serpentes das famílias Pythonidae (13), Boidae (7), Viperidae (5) e Colubridae (1) do RioZoo, Brasil. Os animais foram examinados para determinar sinais clínicos consistentes com qualquer doença infecciosa. Amostras de swab traqueal de cobras foram coletadas em meio Frey e analisadas por isolamento microbiológico e pela técnica da PCR para identificar Mycoplasma spp. As amostras positivas para o gênero Mycoplasma spp. foram submetidas ao sequenciamento genético para identificação das espécies. No isolamento, 19,23% (5/26) foram positivos, enquanto 65,38% (17/26) dos animais foram positivos por PCR. Com base nas análises das seis sequências obtidas, houve similaridade com o Mycoplasma spp. descrito anteriormente em um píton e M. agassizii e M. testudineum encontrados em quelônios. Este é o primeiro relato de Mycoplasma spp. em animais das famílias Boidae e Viperidae. Mycoplasma spp. foi detectado em serpentes com e sem sinais clínicos. Os micoplasmas encontrados apresentaram semelhança genética com outros já descritos em répteis. Não houve relação entre a presença de Mycoplasma spp. e sinais clínicos.(AU)

Maintenance of venomous snakes in captivity for venom production at Butantan Institute from 1908 to the present: a scoping history

Maintenance of snakes at Butantan Institute started in the last century, intending to produce a different antivenom serum to reduce death caused by snakebites. Through a successful campaign coordinated by Vital Brazil, farmers sent venomous snakes to Butantan Institute by the railway lines with no cost. From 1908 to 1962, the snakes were kept in an outdoor serpentarium, where venom extraction was performed every 15 days. During this period, the snake average survival was 15 days. In 1963, the snakes were transferred to an adapted building, currently called Laboratory of Herpetology (LH), to be maintained in an intensive system. Although the periodicity of venom extraction remained the same, animal average survival increased to two months. With the severe serum crisis in 1983, the Ministry of Health financed remodeling for the three public antivenom producers, and with this support, the LH could be improved. Air conditioning and exhausting systems were installed in the rooms, besides the settlement of critical hygienic-sanitary managements to increase the welfare of snakes. In the early 1990s, snake survival was ten months. Over the years to the present day, several improvements have been made in the intensive serpentarium, as the establishment of two quarantines, feeding with thawed rodents, an interval of two months between venom extraction routines, and monitoring of snake health through laboratory tests. With these new protocols, average snake survival increased significantly, being eight years for the genus Bothrops, ten years for genus Crotalus and Lachesis, and four years for the genus Micrurus. Aiming the production of venoms of good quality, respect for good management practices is essential for the maintenance of snakes in captivity. New techniques and efficient management must always be sought to improve animal welfare, the quality of the venom produced, and the safety of those working directly with the venomous snakes.(AU)

Maintenance of venomous snakes in captivity for venom production at Butantan Institute from 1908 to the present: a scoping history

Maintenance of snakes at Butantan Institute started in the last century, intending to produce a different antivenom serum to reduce death caused by snakebites. Through a successful campaign coordinated by Vital Brazil, farmers sent venomous snakes to Butantan Institute by the railway lines with no cost. From 1908 to 1962, the snakes were kept in an outdoor serpentarium, where venom extraction was performed every 15 days. During this period, the snake average survival was 15 days. In 1963, the snakes were transferred to an adapted building, currently called Laboratory of Herpetology (LH), to be maintained in an intensive system. Although the periodicity of venom extraction remained the same, animal average survival increased to two months. With the severe serum crisis in 1983, the Ministry of Health financed remodeling for the three public antivenom producers, and with this support, the LH could be improved. Air conditioning and exhausting systems were installed in the rooms, besides the settlement of critical hygienic-sanitary managements to increase the welfare of snakes. In the early 1990s, snake survival was ten months. Over the years to the present day, several improvements have been made in the intensive serpentarium, as the establishment of two quarantines, feeding with thawed rodents, an interval of two months between venom extraction routines, and monitoring of snake health through laboratory tests. With these new protocols, average snake survival increased significantly, being eight years for the genus Bothrops, ten years for genus Crotalus and Lachesis, and four years for the genus Micrurus. Aiming the production of venoms of good quality, respect for good management practices is essential for the maintenance of snakes in captivity. New techniques and efficient management must always be sought to improve animal welfare, the quality of the venom produced, and the safety of those working directly with the venomous snakes.(AU)