Venom Fractions Research Articles

Assessing Anti-malaria Effect of Naja Naja Oxiana Snake Venom by Real-time Polymerase Chain Reaction Method

Objective: Malaria is one of the most important parasitic diseases and one of the important health issues especially in tropical and subtropical countries. The importance of this disease is due to its high prevalence and mortality, as well as drug resistance and side effects of common drugs used for its treatment. Snake venom is a complex mixture of active pharmaceutical ingredients. The present study aims to investigate the anti-Plasmodium falciparum activity of the purified fractions isolated from the venom of Iranian cobra snake (Naja Naja Oxiana) by real-time Polymerase Chain Reaction (PCR) method. The importance of this disease is due to its high prevalence, significant mortality, as well as drug resistance and, side effects of current drugs in treatment. Venom is a complex mixture of active pharmaceutical ingredients. The purpose of this study was to investigate the anti-Plasmodium falciparum activity of the purified fraction of Iranian cobra snake venom by Real -time PCR. Methods: After preparation and purification of lyophilized venom for determining the parasitic load, different fractions obtained from the venom of Naja Naja Oxiana put in different plates adjacent to Plasmodium falciparum (3D7) strain in the ring stage. The degree of parasitemia was determined by real-time PCR. Finally, the effective fraction with anti-malaria properties was identified. Results: The active fraction with a half maximal inhibitory concentration of 0.026 μg/mL was the most effective fraction on Plasmodium falciparum in vitro (P<0.001). Conclusion: The active fraction of Naja Naja Oxiana venom at the mentioned concentration has anti-malaria effect. This results can motivate the continuation of further research in this field.

Proteome Analysis of Toxic Fractions of Iranian Cobra (Naja naja Oxiana) Snake Venom Using Two-Dimensional Electrophoresis and Mass Spectrometry.

Snake venoms are mostly composed of various proteins and peptides with toxicity and pharmacological effects depending on their geographical sources. Naja naja oxiana is one of the most medically important venomous snakes in Iran and Central Asia. The bite of this type of snake can cause severe pain and swelling, as well as neurotoxicity. Without medical treatment, symptoms quickly worsen and death can occur soon. A detailed understanding of venom components can provide new insight into the production of antivenom against toxic agents instead of crude venom. Specific antibodies against toxic fractions are of utmost importance in neutralizing crude venom. Therefore, the proteome profile of these fractions of Naja naja oxidana venom was analyzed using fractionation by gel filtration, two-dimensional electrophoresis, mass spectrometry, and data mining. Base on the results, in total, 32 spots were detected and categorized into three protein families, namely three-finger toxin (3FTx), phospholipase, and Cysteine-rich secretory proteins (CRISP). These proteins consist of more than 70% crude venom all with a molecular weight below 25 kDa. The 3FTx as a highly diverse constituent in the venom of Naja species was in large quantity in this district. Short-chain neurotoxins, including short neurotoxin, cytotoxin, and muscarinic toxin-like protein, were in abundance, respectively. In conclusion, the recognition of toxic fractions of Naja naja oxiana in this region could be of great help in the production of an effective antivenom against similar compositions. It can also help the medical care department to find out the clinical sign of cobra venom. To the best of our knowledge, this was the first study to report the proteomic of toxic fractions of Naja naja oxiana in Iran.

Conus venom fractions inhibit the adhesion of Plasmodium falciparum erythrocyte membrane protein 1 domains to the host vascular receptors

Using high-throughput BioPlex assays, we determined that six fractions from the venom of Conus nux inhibit the adhesion of various recombinant PfEMP-1 protein domains (PF08_0106 CIDR1α3.1, PF11_0521 DBL2β3, and PFL0030c DBL3X and DBL5e) to their corresponding receptors (CD36, ICAM-1, and CSA, respectively). The protein domain-receptor interactions permit P. falciparum-infected erythrocytes (IE) to evade elimination in the spleen by adhering to the microvasculature in various organs including the placenta. The sequences for the main components of the fractions, determined by tandem mass spectrometry, yielded four T-superfamily conotoxins, one (CC-Loop-CC) with I-IV, II-III connectivity and three (CC-Loop-CXaaC) with a I-III, II-IV connectivity. The 3D structure for one of the latter, NuxVA = GCCPAPLTCHCVIY, revealed a novel scaffold defined by double turns forming a hairpin-like structure stabilized by the two disulfide bonds. Two other main fraction components were a miniM conotoxin, and a O2-superfamily conotoxin with cysteine framework VI/VII. This study is the first one of its kind suggesting the use of conotoxins for developing pharmacological tools for anti-adhesion adjunct therapy against malaria. Similarly, mitigation of emerging diseases like AIDS and COVID-19, can also benefit from conotoxins as inhibitors of protein-protein interactions as treatment. Biological significanceAmong the 850+ species of cone snail species there are hundreds of thousands of diverse venom exopeptides that have been selected throughout several million years of evolution to capture prey and deter predators. They do so by targeting several surface proteins present in target excitable cells. This immense biomolecular library of conopeptides can be explored for potential use as therapeutic leads against persistent and emerging diseases affecting non-excitable systems.We aim to expand the pharmacological reach of conotoxins/conopeptides by revealing their in vitro capacity to disrupt protein-protein and protein-polysaccharide interactions that directly contribute to pathology of Plasmodium falciparum malaria. This is significant for severe forms of malaria, which might be deadly even after treated with current parasite-killing drugs because of persistent cytoadhesion of P. falciparum infected erythrocytes even when parasites within red blood cells are dead. Anti-adhesion adjunct drugs would de-sequester or prevent additional sequestration of infected erythrocytes and may significantly improve survival of malaria patients. These results provide a lead for further investigations into conotoxins and other venom peptides as potential candidates for anti-adhesion or blockade-therapies.This study is the first of its kind and it suggests that conotoxins can be developed as pharmacological tools for anti-adhesion adjunct therapy against malaria. Similarly, mitigation of emerging diseases like AIDS and COVID-19, can also benefit from conotoxins as potential inhibitors of protein-protein interactions as treatment.

The Effect of Various Fractions of Hemiscorpius lepturus Scorpion (Scorpionida: Hemiscorpiidae) Venom on Hemostatic System in Peripheral Blood of Rats in Comparison to Whole Venom

Background: Hemiscorpius lepturus is the most critical scorpion in Khuzestan, Iran, responsible for the majority of deaths caused by scorpion stings. Objectives: This prospective randomized experimental study was performed from June to August 2019 to investigate the effects of various venom fractions on the hemostatic system. Methods: Lyophilized venom of H. lepturus was separated into six fractions by Sephadex G-50 gel chromatography. A total of 96 Albino male rats were treated in eight equal groups; Control: 0.5 ml normal saline (IP); Whole venom group: H. lepturus Lyophilized venom, one mg/kg, IP; F1-F6 groups: Isolated venom fraction 1-6, 0.12 mg/kg, 0.43 mg/kg, 0.08 mg/kg, 0.18 mg/kg, 0.06 mg/kg and 0.13 mg/kg, IP, respectively. Blood sampling was performed at 1, 3, and 24 hours after injection. Results: Plasma fibrinogen was significantly elevated in the whole venom group, while it was significantly decreased in F3, F5, and F6 groups compared to the control group. Prothrombin time (PT) and activated partial thromboplastin time (APTT) assessment revealed that there was a significant prolongation of both tests in whole venom and fraction 5 and 6 receiving groups while fraction 2 only caused a significant PT extension, compared to the control group. Conversely, fraction 3 and 4 resulted in PT and APTT in comparison to the control group. The concentration of FDP and D-dimer was significantly increased in whole venom and F2 group compared to the control group at all sampling times. Conclusions: It seems that H. lepturus venom has both procoagulant and anticoagulant properties which are distinct characteristics of it. Further purification and sequencing of the amino acids in the fraction of peptides can lead to a more precise identification of the mechanisms of venom-induced coagulopathies.

Bradykinin-Potentiating Activity of a Gamma-Irradiated Bioactive Fraction Isolated from Scorpion (Leiurus quinquestriatus) Venom in Rats with Doxorubicin-Induced Acute Cardiotoxicity: Favorable Modulation of Oxidative Stress and Inflammatory, Fibrogenic and Apoptotic Pathways.

Although doxorubicin (Dox) is a backbone of chemotherapy, the search for an effective and safe therapy to revoke Dox-induced acute cardiotoxicity remains a critical matter in cardiology and oncology. The current study was the first to explore the probable protective effects of native and gamma-irradiated fractions with bradykinin-potentiating activity (BPA) isolated from scorpion (Leiurus quinquestriatus) venom against Dox-induced acute cardiotoxicity in rats. Native or irradiated fractions (1μg/g) were administered intraperitoneally (i.p.) twice per week for 3 weeks, and Dox (15mg/kg, i.p.) was administered on day 21 at 1h after the last native or irradiated fraction treatment. Electrocardiographic (ECG) aberrations were ameliorated in the Dox-treated rats pretreated with the native fraction, and the irradiated fraction provided greater amelioration of ECG changes than that of the native fraction. The group pretreated with native protein with BPA also exhibited significant improvements in the levels of oxidative stress-related, inflammatory, angiogenic, fibrogenic, and apoptotic markers compared with those of the Dox group. Notably, the irradiated fraction restored these biomarkers to their normal levels. Additionally, the irradiated fraction ameliorated Dox-induced histological changes and alleviated the severity of cardiac injury to a greater extent than that of the native fraction. In conclusion, the gamma-irradiated detoxified fraction of scorpion venom elicited a better cardioprotective effect than that of the native fraction against Dox-induced acute cardiotoxicity in rats.

Characterization of a cell death‐inducing endonuclease‐like venom protein from the parasitoid wasp Pteromalus puparum (Hymenoptera: Pteromalidae)

Parasitoid wasps are valuable natural enemies for controlling pests. To ensure successful parasitism, these wasps inject venoms along with their eggs that are deposited either into or on their hosts. Parasitoid venoms regulate host behaviors, development, metabolism and immune responses. Pteromalus puparum is a pupal endoparasitoid that parasitizes a number of butterflies, including the worldwide pest cabbage butterfly, Pieris rapae. Venom from P. puparum has a variety of effects on host hemocytes, including alteration of absolute and relative hemocyte counts, and inhibition of hemocyte spreading and encapsulation. In particular, P. puparum venom causes hemocyte cell death in vivo and in vitro. Using assay-guided chromatography, a cell death-inducing venom fraction was identified and defined as P. puparum endonuclease-like venom protein (PpENVP). It belongs to the DNA/RNA nonspecific endonuclease family, which contains two conserved endonuclease activation sites. We analyzed its expression profiles and demonstrated that PpENVP inhibits gene expression in transfected cells relying on two activation sites. However, RNA interference of PpENVP did not significantly reduce P. puparum venom cytotoxicity, suggesting that PpENVP may not be the sole cytotoxic factor present. Our results provide novel insight into the function of the P. puparum venom cocktail and identify a promising insecticide candidate endonuclease that targets insect hemocytes.

The Venom of Philippine Tarantula (Theraphosidae) Contains Peptides with Pro-Oxidative and Nitrosative-Dependent Cytotoxic Activities against Breast Cancer Cells (MCF-7) In Vitro.

Background:Breast cancer is a multifactorial disease that affects women worldwide. Its progression is likely to be executed by oxidative stress wherein elevated levels of reactive oxygen and nitrogen species drive several breast cancer pathologies. Spider venom contains various pharmacological peptides which exhibit selective activity to abnormal expression of ion channels on cancer cell surface which can confer potent anti-cancer activities against this disease. Methods:Venom was extracted from a Philippine tarantula by electrostimulation and fractionated by reverse phase-high performance liquid chromatography (RP-HPLC). Venom fractions were collected and used for in vitro analyses such as cellular toxicity, morphological assessment, and oxidative stress levels. Results:The fractionation of crude spider venom generated several peaks which were predominantly detected spectrophotometrically and colorimetrically as peptides. Treatment of MCF-7 cell line of selected spider venom peptides induced production of several endogenous radicals such as hydroxyl radicals (•OH), nitric oxide radicals (•NO), superoxide anion radicals (•O2−) and lipid peroxides via malondialdehyde (MDA) reaction, which is comparable with the scavenging effects afforded by 400 µg/mL vitamin E and L-cysteine (p<0.05). Concomitantly, the free radicals produced decrease the mitochondrial membrane potential and metabolic activity as detected by rhodamine 123 and tetrazolium dye respectively (p>0.05). This is manifested by cytotoxicity in MCF-7 cells as seen by increase in membrane blebbing, cellular detachment, caspase activity and nuclear fragmentation. Conclusion:These data suggest that the Philippine tarantula venom contains peptide constituents exhibiting pro-oxidative and nitrosative-dependent cytotoxic activities against MCF-7 cells and can indicate mechanistic insights to further explore its potential application as prooxidants in cancer therapy.

Development of high-throughput screening assays for profiling snake venom phospholipase A2 activity after chromatographic fractionation

Many organisms, ranging from plants to mammals, contain phospholipase A2 enzymes (PLA2s), which catalyze the production of lysophospholipids and fatty acid proinflammatory mediators. PLA2s are also common constituents of animal venoms, including bees, scorpions and snakes, and they cause a wide variety of toxic effects including neuro-, myo-, cyto-, and cardio-toxicity, anticoagulation and edema. The aim of this study was to develop a generic method for profiling enzymatically active PLA2s in snake venoms after chromatographic separation. For this, low-volume high-throughput assays for assessment of enzymatic PLA2 activity were evaluated and optimized. Subsequently, the assays were incorporated into a nanofractionation platform that combines high-resolution fractionation of crude venoms by liquid chromatography (LC) with bioassaying in 384-well plate format, and parallel mass spectrometric (MS) detection for toxin identification. The miniaturized assays developed are based on absorbance or fluorescence detection (respectively, using cresol red or fluorescein as pH indicators) to monitor the pH drop associated with free fatty acid formation by enzymatically active PLA2s. The methodology was demonstrated for assessment of PLA2 activity profiles of venoms from the snake species Bothrops asper, Echis carinatus, Echis coloratus, Echis ocellatus, Oxyuranus scutellatus and Daboia russelii russelii.

Venom components of the scorpion Centruroides limpidus modulate cytokine expression by T helper lymphocytes: Identification of ion channel-related toxins by mass spectrometry

The study of the effector mechanisms of T helper cells has revealed different phenotypic characteristics that can be manipulated for designing new therapeutic schemes in different pathological scenarios. Ion channels are significant targets in T lymphocyte modulation since they are closely related to their effector activity. Remarkably, some toxins produced by scorpions specifically affect the function of these membrane proteins. For that reason, these toxins are important candidates in the search for new immunomodulators. Here, the effect of two venom fractions of the scorpion Centruroides limpidus was assessed on T lymphocyte proliferation and cytokine production. The venom fractions ClF8 and ClF9 were separated by reversed-phase high-performance liquid chromatography (RP-HPLC) and cultured at 25 and 35 µg/ml with murine T lymphocytes. The results indicate that the fraction ClF8 increased both production and secretion levels of IFN-γ, IL-4, IL-17A and IL-10 by CD4+ T cells at 24 h. In contrast, fraction ClF9 only promoted the secretion of IL-17A and IL-10 at its highest concentration (35 µg/ml). Both fractions did not show any effect on T cell proliferation. Subsequent analyses by liquid chromatography-tandem mass spectrometry (LC-MS/MS) revealed seventeen toxins in the fraction ClF8 and five toxins in the fraction ClF9, most of them with voltage-gated sodium (NaScTx) and potassium (KScTx) channels as molecular targets. These toxins might probably interact with ion channels involved in T lymphocyte activity. Our findings suggest that the difference in composition between the two fractions could be related to the observed effects, and the components identified could be isolated to search for possible immunomodulatory molecules.

In Vitro Studies for the Evaluation of Insecticidal Potential of the Venom of Endoparasitic Wasp Aenasius arizonensis (Girault) (Hymenoptera, Encyrtidae)

Aenasius arizonensis (Hymenoptera: Encyrtidae) a nymphal, solitary endoparasitoid is a potent biocontrol agent of cotton mealybug, Phenacoccus solenopsis Tinsley (Hemiptera: Pseudococcidae). Female wasp injects a blend of secretions in its host during parasitization which alters the host physiology. Insecticidal potential of crude venom of A. arizonensis was evaluated along with its chromatographic fractions by microinjections bioassays in its natural host insect P. solenopsis at 3rd nymphal instars and adult. Host insects were treated with two different volumes of crude venom (0.3 and 0.5 µL) and RP-HPLC fractions of venom, while host insects with saline injections were used as control. Biological activity of the crude venom was also evaluated after heat and protease treatment. Maximum mortality (84.0 ± 2.91) of the host insects was observed at highest dose rate (0.5 µL) of the crude venom; while lowest mortality (71.0 ± 2.31) was recorded at lowest dose (0.3 µL) of venom applied on 3rd instar life stage. On the other hand, significant difference in mortality of four peaks of RP-HPLC venom fractions were recorded against 3rd instar larvae and adult when microinjections containing crude venom and fractionated venom. Our data confirms that crude venom of the wasp induces significant mortality against its host P. solenopsis. Whereas, no significant difference in mortality was observed for heat and protease treated venom, suggesting that bioactive component of the venom secretions of the wasp are proteins in nature, which lost their biological activity upon heat and protease treatments.

Australian Scorpion Hormurus waigiensis Venom Fractions Show Broad Bioactivity Through Modulation of Bio-Impedance and Cytosolic Calcium.

Scorpion venoms are a rich source of bioactive molecules, but characterisation of toxin peptides affecting cytosolic Ca2+, central to cell signalling and cell death, is limited. We undertook a functional screening of the venom of the Australian scorpion Hormurus waigiensis to determine the breadth of Ca2+ mobilisation. A human embryonic kidney (HEK293) cell line stably expressing the genetically encoded Ca2+ reporter GCaMP5G and the rabbit type 1 ryanodine receptor (RyR1) was developed as a biosensor. Size-exclusion Fast Protein Liquid Chromatography separated the venom into 53 fractions, constituting 12 chromatographic peaks. Liquid chromatography mass spectroscopy identified 182 distinct molecules with 3 to 63 components per peak. The molecular weights varied from 258 Da—13.6 kDa, with 53% under 1 kDa. The majority of the venom chromatographic peaks (tested as six venom pools) were found to reversibly modulate cell monolayer bioimpedance, detected using the xCELLigence platform (ACEA Biosciences). Confocal Ca2+ imaging showed 9/14 peak samples, with molecules spanning the molecular size range, increased cytosolic Ca2+ mobilization. H. waigiensis venom Ca2+ activity was correlated with changes in bio-impedance, reflecting multi-modal toxin actions on cell physiology across the venom proteome.

Analysis of the active fraction of Iranian Naja naja oxiana snake venom on the metabolite profiles of the malaria parasite by 1HNMR in vitro.

Objective(s):Malaria is an important parasitic disease with high morbidity and mortality in tropical areas. Resistance to most antimalarial drugs has encouraged the development of new drugs including natural products. Venom is a complex mixture of active pharmaceutical ingredients. The purpose of this study was to investigate the antimalarial activity of purified fractions of Naja naja oxiana. Materials and Methods:Lyophilized venom was purified with a Sephacryl S-200 HR column and the fractions lyophilized and inhibitory concentration 50% against Plasmodium falciparum 3D7 in vitro obtained. The 4th fraction was run on a Mono Q column, and activity against P. falciparum was detected by lactate dehydrogenase assay and purity by SDS PAGE. Large scale culture of the parasite was carried out with and without the active fraction on the ring stage for 48 hr. The parasites were collected and lyophilized and analyzed by 1HNMR. Chemometrics studies were performed using MATLAB, differentiating metabolites were identified by Human Metabolic Database, and metabolic pathways by the Metaboanalyst online package. Results:The active fraction from the ion exchange column had a 50% inhibitory concentration of 0.026 µg/ml on P. falciparum in vitro (P<0.001) with molecular weight of 63 kDa by SDS-PAGE and no hemolytic activity. Metabolomics studies on the two groups with and without the fraction identified 5 differentiating metabolites and a number of related pathways.Conclusion:The metabolites were succinic acid, l-glutamic acid, pyruvic acid, cholesterol, and NAD. The changes in the Krebs cycle and metabolism pathways of nicotinamide and pyruvate were noticeable.

High throughput screening and identification of coagulopathic snake venom proteins and peptides using nanofractionation and proteomics approaches.

Snakebite is a neglected tropical disease that results in a variety of systemic and local pathologies in envenomed victims and is responsible for around 138,000 deaths every year. Many snake venoms cause severe coagulopathy that makes victims vulnerable to suffering life-threating haemorrhage. The mechanisms of action of coagulopathic snake venom toxins are diverse and can result in both anticoagulant and procoagulant effects. However, because snake venoms consist of a mixture of numerous protein and peptide components, high throughput characterizations of specific target bioactives is challenging. In this study, we applied a combination of analytical and pharmacological methods to identify snake venom toxins from a wide diversity of snake species that perturb coagulation. To do so, we used a high-throughput screening approach consisting of a miniaturised plasma coagulation assay in combination with a venom nanofractionation approach. Twenty snake venoms were first separated using reversed-phase liquid chromatography, and a post-column split allowed a small fraction to be analyzed with mass spectrometry, while the larger fraction was collected and dispensed onto 384-well plates. After fraction collection, any solvent present in the wells was removed by means of freeze-drying, after which it was possible to perform a plasma coagulation assay in order to detect coagulopathic activity. Our results demonstrate that many snake venoms simultaneously contain both procoagulant and anticoagulant bioactives that contribute to coagulopathy. In-depth identification analysis from seven medically-important venoms, via mass spectrometry and nanoLC-MS/MS, revealed that phospholipase A2 toxins are frequently identified in anticoagulant venom fractions, while serine protease and metalloproteinase toxins are often associated with procoagulant bioactivities. The nanofractionation and proteomics approach applied herein seems likely to be a valuable tool for the rational development of next-generation snakebite treatments by facilitating the rapid identification and fractionation of coagulopathic toxins, thereby enabling specific targeting of these toxins by new therapeutics such as monoclonal antibodies and small molecule inhibitors.

Venom Peptide Repertoire of the European Myrmicine Ant Manica rubida: Identification of Insecticidal Toxins.

Using an integrated transcriptomic and proteomic approach, we characterized the venom peptidome of the European red ant, Manica rubida. We identified 13 "myrmicitoxins" that share sequence similarities with previously identified ant venom peptides, one of them being identified as an EGF-like toxin likely resulting from a threonine residue modified by O-fucosylation. Furthermore, we conducted insecticidal assays of reversed-phase HPLC venom fractions on the blowfly Lucilia caesar, permitting us to identify six myrmicitoxins (i.e., U3-, U10-, U13-, U20-MYRTX-Mri1a, U10-MYRTX-Mri1b, and U10-MYRTX-Mri1c) with an insecticidal activity. Chemically synthesized U10-MYRTX-Mri1a, -Mri1b, -Mri1c, and U20-MYRTX-Mri1a irreversibly paralyzed blowflies at the highest doses tested (30-125 nmol·g-1). U13-MYRTX-Mri1a, the most potent neurotoxic peptide at 1 h, had reversible effects after 24 h (150 nmol·g-1). Finally, U3-MYRTX-Mri1a has no insecticidal activity, even at up to 55 nmol·g-1. Thus, M. rubida employs a paralytic venom rich in linear insecticidal peptides, which likely act by disrupting cell membranes.

The Influence of the Different Disposition Characteristics of Snake Toxins on the Pharmacokinetics of Snake Venom.

Snake venom is comprised of a combination of different proteins and peptides with a wide range of molecular weights and different disposition processes inherent to each compound. This causes venom to have a complex exposure profile. Our study investigates 1) how each molecular weight fraction (toxin) of venom contributes to the overall time course of the snake venom, and 2) the ability to determine toxin profiles based on the profile of the overall venom only. We undertook an in silico simulation and modelling study. Sixteen variations of venom, comprising of two to nine toxins with different molecular weights were investigated. The pharmacokinetic parameters (i.e., clearance, , and volume of distribution, ) of each toxin were generated based on a log-linear relationship with molecular weight. The concentration–time data of each toxin were simulated for 100 virtual patients using MATLAB and the total concentration–time data of each toxin were modelled using NONMEM. We found that the data of sixteen mixtures were best described by either two- or three-compartment models, despite the venom being made up of more than three different toxins. This suggests that it is generally not possible to determine individual toxin profiles based on measurements of total venom concentrations only.

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

BackgroundThe 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.

Mitochondrial activity disruption and local muscle damage induced in mice by Scolopendra polymorpha venom.

Background: Scolopendra polymorpha (S. polymorpha) is a predatory centipede whose venom contains a multiplicity of biochemical effectors that can cause muscle damage and cumulative cell destruction in its prey. Despite previous investigations of S. polymorpha and other centipede venoms, there is a lack of information on the morphological and biochemical patterns elicited by their myotoxic effects. To elucidate these processes, this paper presents evidence of skeletal muscle damage, and alterations in key biochemical mediators that appear only after exposure to centipede venom.Methods:Venom was collected and fractionated using RP-HPLC; mouse extensor digitorum longus (EDL) muscle was exposed to whole venom and venom fractions to evaluate myotoxicity by means of creatine kinase (CK) - a muscle damage marker - activity measurements and histochemical analysis.Results:CK activity was higher in EDL muscle exposed to venom than in unexposed muscle. This increase was observed after 15 min of venom incubation, and remained stable up to 45 min. Venom-exposed EDL muscle showed signs of muscle damage including necrosis, loss of fascicular structure as well as mitochondrial accumulations and ragged red fibers (RRF), suggesting an impairment in the normal mitochondrial arrangement. Nicotinamide adenine dinucleotide (NADH) and cytochrome oxidase (COX) tests also indicate that respiratory complexes might be affected.Conclusion:Our results suggest a different biochemical composition of S. polymorpha venom, based on the different effects of four venom fractions on the cells tested, according to statistical evidence. Fractions F6 and F7 caused the most important alterations.

Phlogiellus bundokalbo spider venom: cytotoxic fractions against human lung adenocarcinoma (A549) cells.

Background:Spider venom is a potential source of pharmacologically important compounds. Previous studies on spider venoms reported the presence of bioactive molecules that possess cell-modulating activities. Despite these claims, sparse scientific evidence is available on the cytotoxic mechanisms in relation to the components of the spider venom. In this study, we aimed to determine the cytotoxic fractions of the spider venom extracted from Phlogiellus bundokalbo and to ascertain the possible mechanism of toxicity towards human lung adenocarcinoma (A549) cells.Methods:Spider venom was extracted by electrostimulation. Components of the extracted venom were separated by reversed-phase high performance liquid chromatography (RP-HPLC) using a linear gradient of 0.1% trifluoroacetic acid (TFA) in water and 0.1% TFA in 95% acetonitrile (ACN). Cytotoxic activity was evaluated by the MTT assay. Apoptotic or necrotic cell death was assessed by microscopic evaluation in the presence of Hoechst 33342 and Annexin V, Alexa FluorTM 488 conjugate fluorescent stains, and caspase activation assay. Phospholipase A2 (PLA2) activity of the cytotoxic fractions were also measured.Results:We observed and isolated six fractions from the venom of P. bundokalbo collected from Aurora, Zamboanga del Sur. Four of these fractions displayed cytotoxic activities. Fractions AT5-1, AT5-3, and AT5-4 were found to be apoptotic while AT5-6, the least polar among the cytotoxic components, was observed to induce necrosis. PLA2 activity also showed cytotoxicity in all fractions but presented no relationship between specific activity of PLA2 and cytotoxicity.Conclusion:The venom of P. bundokalbo spider, an endemic tarantula species in the Philippines, contains components that were able to induce either apoptosis or necrosis in A549 cells.

Significant Anticancer Activity of a Venom Fraction Derived from the Persian Gulf Sea Anemone, Stichodactyla haddoni.

Chemotherapy is still one of the main therapeutic regimens in cancer patients but its toxicity is a hard challenge for every patient yet. One of the available solutions is tracing for non-toxic anticancer agents from natural resources. Numerous proteins and peptides in the venom of sea anemones are potentially useful agents with pharmacological properties. Concerning to significance of this issue, the current study was aimed to finding a non-toxic anticancer fraction from the venom of the Persian Gulf sea anemone, Stichodactyla haddoni. Anticancer and hemolytic activity of crude venom was evaluated and followed by fractionation using RP-HPLC. Breast, Brain, and Colon cancer cell lines were selected to assessment of anticancer activity and toxicity. IC50 of crude venom on the abovementioned cancer cell lines was as 4.13, 6.58, and 31.54 µg, respectively. According to the results obtained by paired sample t-test and comparison of toxicity of the fractions in normal cell line, F10, designated as hadonin, was determined as the candidate anti-cancer fraction. The non-toxic dose of F10 was 20 ng in which showed respectively 66, 29, and 7 anticancer activities on breast, brain, and colon cancer cell lines. According to results, anticancer activity of hadonin is of high pharmaceutical value to follow its therapeutic potency in animal model. In conclusion, the venom of the Persian Gulf sea anemone contains a potential anticancer agent with reasonable activity at nanogram level against three kinds of cancer cells with no toxicity on normal cells.

Bradykinin-Potentiating Factors of Venom from Iranian Medically Important Scorpions.

The venom of animals, including snakes, scorpions, and spiders is a complex combination of proteins, peptides, and other biomolecules as well as some minerals. Among the biomolecules of some animal&amp;rsquo;s venom, small peptides that lack disulfide bands known as Non-Disulfide Bridge Peptides (NDBPs) potentiate the bradykinin by preventing the conversion of angiotensin 1 to angiotensin 2 using the mechanism of inhibiting the Angiotensin-Converting Enzyme activity and finally reducing the blood pressure in the victims. This feature of the NDBPs of animal&amp;rsquo;s venom is suggested as the potential of biological drugs. This study aimed to isolate venom components of three species of Iranian medically important scorpions and study the bradykinin potentiating effect of them. The scorpion specimens were collected from the venomous animals and antivenom production department of Razi Vaccine and Serum Research Institute, Karaj, Iran. Moreover, venom extraction was performed by electrical shock (5 volts). The obtained liquid venom of three species specimens was frozen and lyophilized immediately and then preserved in a cool and dried place. The isolation of the venom components for each scorpion was carried out using high-performance liquid chromatography. The obtained ranges of venom fractions (zones) were tested on isolated tissues of guinea-pig ileum and rat uterus using organ bath instrumentation in several replicates. The bioassays resulted in the peptides, including Z1 and Z2 regions in the venom fractionsof the Hottentotta saulcyi, Z2 in Odontobuthus doriae, as well as Z2 and Z3 in Mesobuthus eupeus demonstrated bradykinin potentiating effect. It is concluded that Bradykinin Potentiating Factors were traceable in the venom of all three scorpion species. Therefore, these venoms have the therapeutic potential to exploit biological-based drugs.