Proteomics, toxicity and antivenom neutralization of Sri Lankan and Indian Russell's viper (Daboia russelii) venoms

The western Russell's viper (Daboia russelii) is widely distributed in South Asia, and geographical venom variation is anticipated among distant populations. Antivenoms used for Russell's viper envenomation are, however, raised typically against snakes from Southern India. The present study investigated and compared the venom proteomes of D. russelii from Sri Lanka (DrSL) and India (DrI), the immunorecognition of Indian VINS Polyvalent Antivenom (VPAV) and its efficacy in neutralizing the venom toxicity. Methods: The venoms of DrSL and DrI were decomplexed with C18 high-performance liquid chromatography and SDS-polyacrylamide gel electrophoresis under reducing conditions. The proteins fractionated were identified through nano-ESI-liquid chromatography-tandem mass spectrometry (LCMS/MS). The immunological studies were conducted with enzyme-linked immunosorbent assay. The neutralization of the venom procoagulant effect was evaluated in citrated human plasma. The neutralization of the venom lethality was assessed in vivo in mice adopting the WHO protocol. Results: DrSL and DrI venom proteomes showed comparable major protein families, with phospholipases A2 (PLA2) being the most abundant (> 60% of total venom proteins) and diverse (six protein forms identified). Both venoms were highly procoagulant and lethal (intravenous median lethal dose in mice, LD50 = 0.24 and 0.32 µg/g, for DrSL and DrI, respectively), while lacking hemorrhagic and anticoagulant activities. VPAV was immunoreactive toward DrSL and DrI venoms, indicating conserved protein antigenicity in the venoms. The high molecular weight venom proteins were, however, more effectively immunorecognized than small ones. VPAV was able to neutralize the coagulopathic and lethal effects of the venoms moderately. Conclusion: Considering that a large amount of venom can be injected by Russell's viper during envenomation, the potency of antivenom can be further improved for optimal neutralization and effective treatment. Region-specific venoms and key toxins may be incorporated into the immunization procedure during antivenom production.(AU)

Proteomics, toxicity and antivenom neutralization of Sri Lankan and Indian Russell's viper (Daboia russelii) venoms

The western Russell's viper (Daboia russelii) is widely distributed in South Asia, and geographical venom variation is anticipated among distant populations. Antivenoms used for Russell's viper envenomation are, however, raised typically against snakes from Southern India. The present study investigated and compared the venom proteomes of D. russelii from Sri Lanka (DrSL) and India (DrI), the immunorecognition of Indian VINS Polyvalent Antivenom (VPAV) and its efficacy in neutralizing the venom toxicity. Methods: The venoms of DrSL and DrI were decomplexed with C18 high-performance liquid chromatography and SDS-polyacrylamide gel electrophoresis under reducing conditions. The proteins fractionated were identified through nano-ESI-liquid chromatography-tandem mass spectrometry (LCMS/MS). The immunological studies were conducted with enzyme-linked immunosorbent assay. The neutralization of the venom procoagulant effect was evaluated in citrated human plasma. The neutralization of the venom lethality was assessed in vivo in mice adopting the WHO protocol. Results: DrSL and DrI venom proteomes showed comparable major protein families, with phospholipases A2 (PLA2) being the most abundant (> 60% of total venom proteins) and diverse (six protein forms identified). Both venoms were highly procoagulant and lethal (intravenous median lethal dose in mice, LD50 = 0.24 and 0.32 µg/g, for DrSL and DrI, respectively), while lacking hemorrhagic and anticoagulant activities. VPAV was immunoreactive toward DrSL and DrI venoms, indicating conserved protein antigenicity in the venoms. The high molecular weight venom proteins were, however, more effectively immunorecognized than small ones. VPAV was able to neutralize the coagulopathic and lethal effects of the venoms moderately. Conclusion: Considering that a large amount of venom can be injected by Russell's viper during envenomation, the potency of antivenom can be further improved for optimal neutralization and effective treatment. Region-specific venoms and key toxins may be incorporated into the immunization procedure during antivenom production.(AU)

Proteomics and antivenom immunoprofiling of Russell's viper (Daboia siamensis) venoms from Thailand and Indonesia

The Eastern Russell's viper, Daboia siamensis, is a WHO Category 1 medically important venomous snake. It has a wide but disjunct distribution in Southeast Asia. The specific antivenom, D. siamensis Monovalent Antivenom (DsMAV-Thailand) is produced in Thailand but not available in Indonesia, where a heterologous trivalent antivenom, Serum Anti Bisa Ular (SABU), is used instead. This study aimed to investigate the geographical venom variation of D. siamensis from Thailand (Ds-Thailand) and Indonesia (Ds-Indonesia), and the immunorecognition of the venom proteins by antivenoms. Methods: The venom proteins were decomplexed with reverse-phase high-performance liquid chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis, followed by in-solution tryptic digestion, nano-liquid chromatography-tandem mass spectrometry and protein identification. The efficacies of DsMAV-Thailand and SABU in binding the various venom fractions were assessed using an enzyme-linked immunosorbent assay optimized for immunorecognition profiling. Results: The two most abundant protein families in Ds-Thailand venom are phospholipase A2 (PLA2) and Kunitz-type serine protease inhibitor (KSPI). Those abundant in Ds-Indonesia venom are PLA2 and serine protease. KSPI and vascular endothelial growth factor were detected in Ds-Thailand venom, whereas L-amino acid oxidase and disintegrin were present in Ds-Indonesia venom. Common proteins shared between the two included snaclecs, serine proteases, metalloproteinases, phosphodiesterases, 5'nucleotidases and nerve growth factors at varying abundances. DsMAV-Thailand exhibited strong immunorecognition of the major protein fractions in both venoms, but low immunoreactivity toward the low molecular weight proteins e.g. KSPI and disintegrins. On the other hand, SABU was virtually ineffective in binding all fractionated venom proteins. Conclusion: D. siamensis venoms from Thailand and Indonesia varied geographically in the protein subtypes and abundances. The venoms, nevertheless, shared conserved antigenicity that allowed effective immunorecognition by DsMAV-Thailand but not by SABU, consistent with the neutralization efficacy of the antivenoms. A specific, appropriate antivenom is needed in Indonesia to treat Russell's viper envenomation.(AU)

Systemic vascular leakage induced in mice by Russell's viper venom from Pakistan.

Envenomings by some populations of the Russell's viper (Daboia russelii) are characterized by a systemic capillary leak syndrome (CLS) which causes hemoconcentration, and is associated with the severity of envenoming. We adapted a model of CLS in mice by assessing hemoconcentration. The venom of D. russelii from Pakistan, but not that of another viperid, Bothrops asper, induced hemoconcentration and an increment in vascular permeability, being devoid of hemorrhagic activity at the doses tested. These findings reveal a dichotomous pattern of vasculotoxicity in viperid snake venoms. This difference might depend on variations in venom composition, especially regarding metalloproteinases (SVMPs), which are low in Pakistani D. russelii and high in B. asper. Inhibition of SVMPs and phospholipases A2 in D. russelii venom did not abrogate hemoconcentration. An hemoconcentration-inducing fraction was obtained by chromatography, which contains vascular endothelial growth factor (VEGF), a known potent inducer of increment in vascular permeability. Exudates collected from tissue injected with venom also induced hemoconcentration, and the effect was inhibited by antivenom. However, the amount of venom in exudate required to induce the effect is low, as compared with venom dissolved in saline solution, hence suggesting that endogenous proteins present in the exudate, probably inflammatory mediators, potentiate the effect.

Antibacterial potential of a basic phospholipase A (VRV-PL-VIIIa) from Daboia russelii pulchella (Russell's viper) venom

Background: Microbial/bacterial resistance against antibiotics poses a serious threat to public health. Furthermore, the side effects of these antibiotics have stimulated tremendous interest in developing new molecules from diverse organisms as therapeutic agents. This study evaluates the antibacterial potential of a basic protein, Vipera russellii venom phospholipase A2 fraction VIIIa (VRV-PL-VIIIa), from Daboia russelii pulchella venom against gram-positive and gram-negative bacteria. Methods: The antibacterial potential of VRV-PL-VIIIa in the presence and absence of an inhibitor (p-bromophenacyl bromide) was tested against gram-positive and gram-negative bacteria and the minimum inhibitory concentration was determined by microdilution tests. Results: VRV-PL-VIIIa demonstrated potent antibacterial activities against all the human pathogenic strains tested. It more effectively inhibited such gram-positive bacteria as Staphylococcus aureus and Bacillus subtilis, when compared to the gram-negative bacteria Escherichia coli, Vibrio cholerae, Klebsiella pneumoniae and Salmonella paratyphi. It inhibited bacterial growth at minimum inhibitory concentration values ranging from 11.1 to 19.2 μg/mL. The anti-bacterial potential of VRV-PL-VIIIa was comparable to the standards gentamycin, chlorophenicol and streptomycin. The PLA2's hemolytic and antibacterial activities were strongly correlated. Furthermore, even in the presence of p-bromophenacyl bromide, intense antibacterial activity was observed, suggesting a dissociation or partial overlapping of the bactericidal/antimicrobial domains. Conclusion: VRV-PL-VIIIa demonstrated potent antibacterial activities against all the human pathogenic strains tested. The study shows that despite a strong correlation between enzymatic and antimicrobial activities of VRV-PL-VIIIa, it may possess additional properties that mimic the bactericidal/membrane permeability-increasing protein. This study encourages further in-depth studies on the molecular mechanisms of antibacterial properties of VRV-PL-VIIIa, which would thereby facilitate development of this protein into a possible therapeutic lead molecule for treating bacterial infections.(AU)

Antibacterial potential of a basic phospholipase A (VRV-PL-VIIIa) from Daboia russelii pulchella (Russell's viper) venom

Background: Microbial/bacterial resistance against antibiotics poses a serious threat to public health. Furthermore, the side effects of these antibiotics have stimulated tremendous interest in developing new molecules from diverse organisms as therapeutic agents. This study evaluates the antibacterial potential of a basic protein, Vipera russellii venom phospholipase A2 fraction VIIIa (VRV-PL-VIIIa), from Daboia russelii pulchella venom against gram-positive and gram-negative bacteria. Methods: The antibacterial potential of VRV-PL-VIIIa in the presence and absence of an inhibitor (p-bromophenacyl bromide) was tested against gram-positive and gram-negative bacteria and the minimum inhibitory concentration was determined by microdilution tests. Results: VRV-PL-VIIIa demonstrated potent antibacterial activities against all the human pathogenic strains tested. It more effectively inhibited such gram-positive bacteria as Staphylococcus aureus and Bacillus subtilis, when compared to the gram-negative bacteria Escherichia coli, Vibrio cholerae, Klebsiella pneumoniae and Salmonella paratyphi. It inhibited bacterial growth at minimum inhibitory concentration values ranging from 11.1 to 19.2 μg/mL. The anti-bacterial potential of VRV-PL-VIIIa was comparable to the standards gentamycin, chlorophenicol and streptomycin. The PLA2's hemolytic and antibacterial activities were strongly correlated. Furthermore, even in the presence of p-bromophenacyl bromide, intense antibacterial activity was observed, suggesting a dissociation or partial overlapping of the bactericidal/antimicrobial domains. Conclusion: VRV-PL-VIIIa demonstrated potent antibacterial activities against all the human pathogenic strains tested. The study shows that despite a strong correlation between enzymatic and antimicrobial activities of VRV-PL-VIIIa, it may possess additional properties that mimic the bactericidal/membrane permeability-increasing protein. This study encourages further in-depth studies on the molecular mechanisms of antibacterial properties of VRV-PL-VIIIa, which would thereby facilitate development of this protein into a possible therapeutic lead molecule for treating bacterial infections.

Antibacterial potential of a basic phospholipase A2(VRV-PL-VIIIa) from Daboia russelii pulchella (Russell's viper) venom

Background:Microbial/bacterial resistance against antibiotics poses a serious threat to public health. Furthermore, the side effects of these antibiotics have stimulated tremendous interest in developing new molecules from diverse organisms as therapeutic agents. This study evaluates the antibacterial potential of a basic protein, Vipera russellii venom phospholipase A2 fraction VIIIa (VRV-PL-VIIIa), from Daboia russelii pulchella venom against gram-positive and gram-negative bacteria.Methods:The antibacterial potential of VRV-PL-VIIIa in the presence and absence of an inhibitor (p-bromophenacyl bromide) was tested against gram-positive and gram-negative bacteria and the minimum inhibitory concentration was determined by microdilution tests.Results:VRV-PL-VIIIa demonstrated potent antibacterial activities against all the human pathogenic strains tested. It more effectively inhibited such gram-positive bacteria as Staphylococcus aureus and Bacillus subtilis, when compared to the gram-negative bacteria Escherichia coli, Vibrio cholerae, Klebsiella pneumoniae and Salmonella paratyphi. It inhibited bacterial growth at minimum inhibitory concentration values ranging from 11.1 to 19.2 μg/mL. The anti-bacterial potential of VRV-PL-VIIIa was comparable to the standards gentamycin, chlorophenicol and streptomycin. The PLA2's hemolytic and antibacterial activities were strongly correlated. Furthermore, even in the presence of p-bromophenacyl bromide, intense antibacterial activity was observed, suggesting a dissociation or partial overlapping of the bactericidal/antimicrobial domains.Conclusion:VRV-PL-VIIIa demonstrated potent antibacterial activities against all the human pathogenic strains tested. The study shows that despite a strong correlation between enzymatic and antimicrobial activities of VRV-PL-VIIIa, it may possess additional properties that mimic the bactericidal/membrane permeability-increasing protein. This study encourages further in-depth studies on the molecular mechanisms of antibacterial properties of VRV-PL-VIIIa, which would thereby facilitate development of this protein into a possible therapeutic lead molecule for treating bacterial infections.(AU)

Shape Variation and Allometry in the Precloacal Vertebral Series of the Snake Daboia russelli (Viperidae) / Variación Morfológica y Alometría de las Vértebras Precloacales en el Ofidio Daboia russelli (Viperidae)

Understanding the variation of the ophidian vertebral morphology is an essential tool in snake paleobiology, but so far this field remains hardly investigated. A major problematic is the still scarce knowledge about the basis of homogeneity of intracolumnar shape variation along the vertebrae of the precloacal region in these animals. For instance, this variation can be overwhelmingly low in cases such as in vipers, for which it seems almost impossible to describe a concrete regionalization of the precloacal region without ambiguity. This study has applied geometric morphometrics to analyze if the shape variation of the vertebrae of the precloacal vertebrae of an adult specimen of Daboia russelli allows differentiating any sort of parcellation within the column of this organism. We have also explored if size is associated with the organization of vertebral shape along the axial skeleton. The multivariate analyses showed that the main pattern of vertebral shape variation in D. russelli concerns the neural spine and the hypapophysis, whereas the shape of the vertebral centrum appears to be nearly invariant along the series. Our analysis also showed that the precloacal region can be sudivided into two portions that merge in a transitional boundary of largest vertebrae in the middle of the column. From this middle region towards the distal ends of the column vertebrae become smaller changing their shapes in two antithetical ways.

Entender la variación en la morfología vertebral de los ofidios es crucial para la paleobiología del grupo pero, hasta ahora, este campo está poco investigado. Uno de los principales problemas es el escaso conocimiento sobre las bases de la homogeneidad en la variación de la forma a lo largo de la región precloacal en estos animales. Por ejemplo, en el caso de las víboras, dicha variación puede ser muy pequeña lo cual hace casi imposible la descripción de una regionalización precisa sin ambigüedad. En este estudio se ha aplicado morfometría geométrica para analizar si la variación de la forma vertebral de un individuo adulto de la especie Daboia russelli permite subdividir la región precloacal. Además, hemos explorado si el tamaño está asociado con la organización vertebral a lo largo del esqueleto axial. Los análisis multivariantes han demostrado que el patrón principal de la variación de la forma vertebral está determinado por la espina neural y la hipapofisis, mientras que el centro vertebral varía poco a lo largo de la serie. Nuestro análisis ha mostrado que la región precloacal puede ser dividida en dos series cuya separación está marcada por las vértebras más grandes, posicionadas aproximadamente en la mitad de la columna. Tomando como referencia la mitad de la columna, hacia los extremos distales, las vértebras tienden a ser más pequeñas cambiando su forma de modo antitético.

Posterior fossa infarct following Viper bite: a paradox

Cerebral infarction after a viper bite is relatively uncommon. A combination of factors has been implicated in the pathophysiology of infarct following snakebite. In this case report, the clinical outcome after a posterior circulation infarct and various possibilities that could lead to such a catastrophic event are discussed. The present study stresses the need to keep hydration, blood pressure and central venous pressure optimal in all snakebite patients. Cerebral infarction should be considered a differential diagnosis, in any patient with neurological deterioration following snakebite. Prognosis of such patients with posterior circulation stroke remains poor and decompressive craniectomy has not been found to be helpful.(AU)

Unusual visual loss after snakebite

Snakebites are endemic in some parts of Thailand, being associated with several complications. Ocular disturbances are uncommon, except in cases of corneal or conjunctival injury, when the eye is directly exposed to the snake venom. The present study presents a case of combined ophthalmic artery occlusion and transient central retinal artery occlusion with macular ischemia after a Russells viper bite.(AU)

Unusual visual loss after snakebite

Snakebites are endemic in some parts of Thailand, being associated with several complications. Ocular disturbances are uncommon, except in cases of corneal or conjunctival injury, when the eye is directly exposed to the snake venom. The present study presents a case of combined ophthalmic artery occlusion and transient central retinal artery occlusion with macular ischemia after a Russell's viper bite.(AU)

Novel role of antiplatelet agents (aspirin plus clopidogrel) in an incoagulable blood of a victim of russell's viper snakebite

Snake antivenom is a specific antidote to the venom action, neutralizing the circulating venom. However, it fails to neutralize the venom fixed to target organs such as platelets, renal tubules, etc. Russell's viper venom initiates rapid coagulation in a victim by activating blood platelets, factors V, X, and anticoagulant cofactors. Activation of thrombin, resulting in formation of micro-thrombi, fibrinolysis, and a vicious cascade, sets in. Inhibition of activated platelets by aspirin (cyclooxygenase inhibitor) and clopidogrel (ADP receptor inhibitor) helps to break this vicious circle induced by Russell's venom and may initiate the natural physiological clotting mechanism. They can be utilized as an adjuvant treatment.(AU)

Anticoagulante lúpico en preeclampsia e hipertensión gestacional no proteinúria en primigestas / Lupus anticoagulant in preeclampsia and non-proteinuric gestational hypertensión in primigravids

El propósito de este estudio fue determinar la relación entre el Anticoagulante Lúpico y la Hipertensión Gestacional Proteinúria y no Proteinúria en primigestas sin patología subyacente. Se estudiaron 65 pacientes primigestas, con embarazo de 20 o más semanas de duración. Treinta y cuatro pacientes (Grupo A) fueron embarazadas normotensas y 31 (Grupo B) presentaron Enfermedad Hipertensiva desarrollada en el embarazo según la clasificación de Davey y MacGillivray. A todas se les practicó una prueba para determinar la presencia del Anticoagulante Lúpico según el método de veneno de víbora de Russell diluido. Ninguna de las pacientes presentó otra patología subyacente sintomática. El 55 por ciento de las pacientes del Grupo B desarrolló Hipertensión Gestacional Proteinúrica (Preeclampsia) y el 45 por ciento Hipertensión Gestacional no Proteinúria. Las pruebas para la determinación del Anticoagulante Lúpico resultaron negativas en todas las pacientes de ambos grupos. Los resultados sugieren que en la Hipertensión Gestacional Proteinúrica y no Proteinúrica en primigestas sin patología subyacente, es poco probable la presencia de Anticoagulante Lúpico, por lo que persiste como un área de estudio controversial