Detection of scorpion venom by optical circular dichroism method.

Various efforts have been made to detect minimum amounts of any toxic materials in water or the neurotoxic effect of venom (Odontobuthus Doriae Scorpion) in the human's blood serum nerve by high-sensitivity, accurate, and low-cost sensors in order to enhance life style. Therefore, the present study was done to investigate reliability of two-dimensional plasmonic structure and circular dichroism (CD) in toxic samples in order to measure and determine venom concentrations and its neurotoxic effect on humans҆ blood serum Neurotransmitter analytes. Our results confirmed dependency of CD signal to neurotoxic effect of venom concentrations and good sensitivity of this sensor with the help of achiral plasmonic structure.

Novel Mutant Phospholipase D from Hemiscorpius lepturus Acts as A Highly Immunogen in BALB/c Mice Against the Lethality of Scorpion Venom.

Hemiscorpius lepturus (H. lepturus) which belongs to the Scorpionidae family, is the deadliest scorpion in Iran. It causes pathological manifestations like dermonecrosis, hemolysis, renal failure, necrotic ulcers, and in some cases, even death. The venom of this scorpion is well-known for its cytotoxic effects in comparison with the other venomous scorpions which show significant neurotoxic effects. Due to the painless nature of the sting of this scorpion, the clinical symptoms occur in victims 24 to 72 h post-sting. In our previous studies during the last decade, we demonstrated that the medical complications are attributable to the presence of phospholipase D (PLD) as a major toxin in the venom. With the purpose of designing and constructing a vaccine against H. lepturus for humans, animal model experiments were performed. To achieve this goal, non-toxic PLD was developed by mutation of two critical catalytic residues-His12 and His48-into alanines and the product was then denominated mut-rPLD1. The in-vivo tests showed that the mice immunized with interval doses of 10 µg of mut-rPLD1, were completely protected against 10× the LD100 of the venom. In conclusion, this mutant may be an effective vaccine candidate against scorpion envenomation by H. lepturus in future clinical studies.

Structural characterization of scorpion peptides and their bactericidal activity against clinical isolates of multidrug-resistant bacteria.

Scorpion venom peptides represent a novel source of antimicrobial peptides (AMPs) with broad-spectrum activity. In this study, we determined the minimum bactericidal concentration (MBC) of three scorpion AMPs, Uy234, Uy17, and Uy192, which are found in the venomous glands of the Urodacus yaschenkoi scorpion, against the clinical isolates of multidrug-resistant (MDR) bacteria. In addition, we tested the activity of a consensus AMP designed in our laboratory based on some previously reported IsCT-type (cytotoxic linear peptide) AMPs with the aim of obtaining higher antimicrobial activity. All peptides tested showed high antimicrobial activity against MDR clinical isolates, with the highest activity against ß-hemolytic Streptococcus strains. The hemolytic activity was determined against human red blood cells and was significantly lower than that of previously reported AMPs. The α-helical structure of the four AMPs was confirmed by circular dichroism (CD). These results suggest that the four peptides can be valuable tools for the design and development of AMPs for use in the inhibition of MDR pathogenic bacteria. A clear index of synergism and additivity was found for the combination of QnCs-BUAP + Uy234, which makes these peptides the most promising candidates against pathogenic bacteria.

Black Scorpion (Tityus obscurus) Fatalities in Guyana and a Literature Review.

BACKGROUND: More than 1500 scorpion species exist worldwide, with a few scorpion species potentially lethal to humans. About 1 million stings annually result in >3000 deaths, but the incidence and mortality vary greatly by species and location. Physicians working internationally must recognize that resulting toxidromes vary significantly by region. Over the past few decades, South America has reported relatively few deaths and low case mortality rates from envenomations. In Guyana, a small tropical country on its northeast coast, they have been extremely rare. A sudden fatal case cluster suggests an extension of the black scorpion's habitat, an increase in venom toxicity, or both. CASE REPORTS: During a 12-month period, Guyana experienced 3 deaths, including 1 adult, from black scorpion (Tityus obscurus) envenomation. The 30-year-old man and 2 young children experienced the same symptom complex, initially appearing well except for pain at the sting site. They soon developed persistent emesis and leukocytosis. All were flown from remote jungle areas to the only public tertiary care hospital where they received maximal available medical support. They gradually developed profound cardiopulmonary failure requiring ventilation and, eventually, dysrhythmias. None had hyperglycemia or pancreatitis, and they had no neurologic abnormalities until developing progressive obtundation immediately before intubation. WHY SHOULD AN EMERGENCY PHYSICIAN BE AWARE OF THIS?: Scorpion envenomation symptoms, outcomes, and treatment are geographically specific. Patients benefit when clinicians recognize the worldwide variations in grading systems and treatment options, which we discuss and compare to our patients.

Cerebrospinal inflammatory response following scorpion envenomation: role of histamine H1 and H3 receptors.

BACKGROUND: The mechanism of the inflammatory process induced by scorpion venom in the cerebrospinal tissues has not yet been completely elucidated. Therefore, we aimed to investigate the role of histamine through its H1 and H3 receptors in this process. METHODS: Histamine H1 and H3 receptor antagonists, Hydroxyzine (10 mg/kg) and Betaserc (20 mg/kg), respectively, were administered by intraperitoneal route to mice 1 h before subcutaneous envenomation with a subletal dose (0.5 mg/kg) of Androctonus australis hector venom. Cerebrospinal inflammation response was assessed 24 h after envenomation by evaluating the vascular permeability changes, inflammatory cell infiltration, oxidative/nitrosative stress marker levels (hydrogen peroxide, nitric oxide, malondialdehyde, glutathione and catalase) and by histological examination of cerebrospinal tissue. RESULTS: Envenomed mice displayed an installation of an inflammatory response marked by increased vascular permeability (76% and 68% in brain and spinal cord, respectively, in comparison to controls), inflammatory cell infiltration, increased pro-oxidant levels and decreased anti-oxidant markers (p  < 0.05 to p  < 0.001). Scorpion venom also induced structural changes in brain and spinal cord tissues. Hydroxyzine seemed to be more efficient than Betaserc in the prevention of the induced cerebrospinal inflammation response, as evidenced by the decreased vascular permeability, inflammatory cell infiltration, pro-oxidant levels, increased anti-oxidant defense (p  < 0.05 to p  < 0.001) and a reduction of the anatomo-pathological alterations. CONCLUSION: The results showed that the histamine H1 receptor is more involved in the induced central nervous system inflammatory response during scorpion envenomation.

Effects of Medium Molecular Weight Heparinyl Arginine on Scorpion Venom-induced Pulmonary Edema in Rats.

BACKGROUND/AIM: The aim of this study was to examine the inhibitory action of medium molecular weight heparinyl amino acid derivatives (MHADs) on scorpion venom (SV)-induced acute pulmonary edema. MATERIALS AND METHODS: SV was injected into the tail vein of rats after MHAD pre-treatment. An hour later, lungs were removed from each experimental animal, followed by measurement of the lung/body index (LBI) and Na+/K+ ratio of the pulmonary tissue as indices for acute pulmonary edema. RESULTS: Medium molecular weight heparinyl arginine (MHR)-administered rats exhibited significantly lower LBI and Na+/K+ ratios compared to control rats. Although the mechanism of inhibitory action of MHR on pulmonary edema is unclear, MHR inhibited the vascular permeability increase by SV because both LBI and Na+/K+ ratio of the pulmonary tissue remained at almost normal values. CONCLUSION: MHR may prevent scorpion venom-induced acute pulmonary edema and thus makes a good candidate for clinical use.

Inverted Takotsubo syndrome in Androctonus australis scorpion envenomation.

CONTEXT: The nature of scorpion-related cardiomyopathy is still a matter of debate where specific toxin-induced cardiomyopathy, ischemic, or catecholaminergic cardiomyopathy is advocated as well. We report two cases of Takotsubo syndrome following envenomation by Androctonus australis, bringing new evidence for the fundamental role of catecholamines in the pathogenesis of this cardiomyopathy. Case 1: A woman aged 36 presented with pulmonary edema and shock following scorpion envenomation. Echocardiography-Doppler showed a LVEF at 30%. Cardiac magnetic resonance (CMR) imaging showed a basal ballooning of the left and right ventricles suggestive of an inverted biventricular Takotsubo syndrome. A second CMR performed after recovery was normal. Case 2: A woman aged 45 was admitted for pulmonary edema and shock consecutive to scorpion envenomation. Echocardiography showed a LVEF at 35%. CMR showed a basal ballooning. The patient was discharged four days following admission with a normal LV function on repeat echocardiography examination. CONCLUSIONS: Cardiomyopathy in these cases, following scorpion envenomation by Androctonus australis, fulfills the criteria of Takotsubo syndrome. These observations contribute to our understanding of the mechanism, prognosis, and treatment of scorpion-related cardiomyopathy.

Centruroides sculpturatus envenomation in three adult patients requiring treatment with antivenom.

CONTEXT: Envenomation by Centruroides sculpturatus can manifest with cranial nerve dysfunction and neuromuscular hyperactivity. While these symptoms are most commonly seen in young children, they may also be seen in adults. CASE DETAILS: Three cases of adult patients are presented with grades III & IV scorpion envenomation. They reported symptoms including disconjugate, roving eye movements, and motor involvement. Also reported were hyposmia, difficulty with fine motor movements, and dysgeusia. All were first treated with benzodiazepines with little to no effect. They then received a three vial antivenom bolus with resolution of severe symptoms within 30-60 min. DISCUSSION: Severe Centruroides envenomation can occur in adults as well as children. These three cases demonstrate the usefulness, safety, and effectiveness of antivenom therapy to quickly relieve symptoms in adult patients with grades III & IV envenomations.

Reactogenicity and safety assessment of an attenuated nanovaccine against scorpion envenomation: Preclinical study.

An attenuated nanovaccine (Nps - V∗) has been developed to protect humans from fatal scorpion envenomation in at-risk regions. This study was conducted to evaluate the toxicity and the local reactogenicity of the Nps - V∗ nanovaccine developed against Androctonus australis hector (Aah) venom. Assessment of the systemic inflammatory response and serum cytokine levels were evaluated in vaccinated mice with 100µg of irradiated Aah venom (V∗) encapsulated or not into polymeric calcium-alginate nanoparticles (Nps) and injected by subcutaneous (s.c) route. The local reactogenicity was evaluated by dermal Draize observations and skin tissue analysis at the injection site of vaccinated rabbits with 250 or 500µg of V∗-loaded into Nps. All animals gained weight and had normal food consumption during the study. Additionally, results showed that the nanoformulation Nps - V∗ did not cause clinical evidence of systemic toxicity in mice or rabbits, a transient edema/erythema at the injection site was only recorded as treatment-related reactogenicity. These results indicated a favorable safety profile for Nps - V∗ and supported its use in superior animal tests, then in a Phase 1 clinical trial.

A mutant of the Buthus martensii Karsch antitumor-analgesic peptide exhibits reduced inhibition to hNav1.4 and hNav1.5 channels while retaining analgesic activity.

Scorpion toxins can kill other animals by inducing paralysis and arrhythmia, which limits the potential applications of these agents in the clinical management of diseases. Antitumor-analgesic peptide (AGAP), purified from Buthus martensii Karsch, has been proved to possess analgesic and antitumor activities. Trp38, a conserved aromatic residue of AGAP, might play an important role in mediating AGAP activities according to the sequence and homology-modeling analyses. Therefore, an AGAP mutant, W38G, was generated, and effects of both AGAP and the mutant W38G were examined by whole-cell patch clamp techniques on the sodium channels hNav1.4 and hNav1.5, which were closely associated with the biotoxicity of skeletal and cardiac muscles, respectively. The data showed that both W38G and AGAP inhibited the peak currents of hNav1.4 and hNav1.5; however, W38G induced a much weaker inhibition of both channels than AGAP. Accordingly, W38G exhibited much less toxic effect on both skeletal and cardiac muscles than AGAP in vivo The analgesic activity of W38G and AGAP were verified in vivo as well, and W38G retained analgesic activity similar to AGAP. Inhibition to both Nav1.7 and Nav1.8 was involved in the analgesic mechanism of AGAP and W38G. These findings indicated that Trp38 was a key amino acid involved in the biotoxicity of AGAP, and the AGAP mutant W38G might be a safer alternative for clinical application because it retains the analgesic efficacy with less toxicity to skeletal and cardiac muscles.

A bimodal activation mechanism underlies scorpion toxin-induced pain.

Venomous animals use peptide toxins for hunting and self-defense. To achieve these goals, toxins need to bind to their targets with high affinity due to the small amount that a single bite or sting can deliver. The scorpion toxin BmP01 is linked to sting-induced excruciating pain; however, the reported minimum concentrations for activating TRPV1 channel or inhibiting voltage-gated potassium (Kv) channels (both in the micromolar range) appear too high to be biologically relevant. We show that the effective concentration of BmP01 is highly pH-dependent-it increases by about 10-fold in inhibiting Kv channels upon a 1-U drop in pH but decreases more than 100-fold in activating TRPV1. Mechanistic investigation revealed that BmP01 binds to one of the two proton-binding sites on TRPV1 and, together with a proton, uses a one-two punch approach to strongly activate the nociceptive channel. Because most animal venoms are acidic, proton-facilitated synergistic action may represent a general strategy for maximizing toxin potency.

Cardiorespiratory alterations in rodents experimentally envenomed with Hadruroides lunatus scorpion venom

Abstract Background Hadruroides lunatus is the most abundant scorpion species in the Peruvian central coast, where most of the accidents involving humans are registered. In spite of its prevalence, there are only very few studies on H. lunatus envenomation. The aim of the present study was to analyze the cardiorespiratory alterations caused by H. lunatus envenomation in rodents. Methods Wistar rats injected with H. lunatus scorpion venom were submitted to electrocardiography. After euthanasia, rat lungs were collected and histopathologically analyzed. Mouse cardiomyocytes were used to perform immunofluorescence and calcium transient assays. Data were analyzed by ANOVA or Student’s t-test. The significance level was set at p< 0.05. Results It was observed that H. lunatus venom increased heart rate and caused arrhythmia, thereby impairing the heart functioning. Lungs of envenomed animals showed significant alterations, such as diffuse hemorrhage. In addition, immunofluorescence showed that H. lunatus venom was capable of binding to cardiomyocytes. Furthermore, mouse ventricular cardiomyocytes incubated with H. lunatus venom showed a significant decrease in calcium transient, confirming that H. lunatus venom exerts a toxic effect on heart. Conclusion Our results showed that H. lunatus venom is capable of inducing cardiorespiratory alterations, a typical systemic effect of scorpionism, stressing the importance of medical monitoring in envenomation cases.

Spinal 5-HT3AR contributes to BmK I-induced inflammatory pain in rats.

Subcutaneous injection of BmK I could be adopted to well establish a novel pain model. Moreover, 5-hydroxytryptamine (serotonin, 5-HT) receptor is involved in regulating animal pain-related behaviors. However, the underlying mechanism of 5-HT3R on BmK I-induced pain remains unclear. Animal behavioral testing, RT-PCR and Western blotting were used to yield the following results: first, intraplantar (i.pl.) injection of BmK I (10 µg) induced elevated mRNA and protein levels of 5-HT3AR in bilateral L4-L5 spinal cord; Second, intrathecal (i.t.) injection of ondansetron (a specific antagonist of 5-HT3AR) reduced spontaneous pain responses, attenuated unilateral thermal and bilateral mechanical hypersensitivity elicited by BmK I; Microglia could be activated by BmK I (i.pl.) in both sides of L4-L5 spinal cord, and this effect was reversed by intrathecal pre-treatment with 5-HT3AR antagonist. Meanwhile, the 5-HT3AR in L4-L5 spinal cord was almost co-localized with NeuN (a marker of nerve cell), but not co-expressed with Iba-1 (a marker of microglia). Finally, the expression level of CX3CL1 and CX3CR1 was reduced by intrathecal pre-treatment with ondansetron. Our results indicate that both 5-HT3AR signaling pathway and microglia are activated in the process of induction and maintenance of BmK I-induced pain nociception. Meanwhile, our results suggest that the neuronal 5-HT3AR may communicate with microglia indirectly via CX3CL1 which is involved in regulating the BmK I-induced hyperalgesia and sensitization.

Complement system and immunological mediators: Their involvements in the induced inflammatory process by Androctonus australis hector venom and its toxic components.

Androctonus australis hector scorpion venom is well known by its high toxicity, it induces massive release of neurotransmitters that lead to pathophysiological disorders in cardiovascular, neuro-hormonal and immune systems. Previous studies have shown the relationship between the severity of scorpion envenoming and immune system activation. This study was assessed to investigate the involvement of complement system and inflammatory mediators after sublethal injection of Aah venom, its toxic fraction (FtoxG50) and its main toxins (AahI and AahII) into NMRI mice. The Activation complement system by the venom is also compared to that induced of lipopolysaccharides (LPS). Obtained results showed that seric complement system (CS) is activated by the venom and by its toxic components; this activation is more pronounced into liver tissue when toxic components (FtoxG50, AahI or AahII) are used. Increase of cytokine levels (IL1ß, TNFα and ICAM) into hepatic tissue induced by AahI or AahII neurotoxins is correlated with tissue alterations. Aprotinin, a non specific inhibitor of complement system seems to be able to reduce CS consumption and to restore partially the induced tissue damage by venom. The mechanisms by which toxic fraction or LPS induced the activation of complement system seem to be different. Sensitivity of hepatic tissue is more pronounced after FtoxG50 injection; however lung tissue is more sensible to LPS than FoxG50.

Androctonus australis hector venom contributes to the interaction between neuropeptides and mast cells in pulmonary hyperresponsiveness.

Lung injury and respiratory distress syndrome are frequent symptoms observed in the most severe cases of scorpion envenomation. The uncontrolled transmigration of leukocyte cells into the lung interstitium and alveolar space and pulmonary edema may be the cause of death. Mast cells can release various inflammatory mediators known to be involved in the development of lung edema following scorpion venom injection. The present study was designed to determine the evidence of neurokinin 1 (NK1) receptor and the involvement of mast cell activation to induce pulmonary edema and to increase vascular permeability after Androctonus australis hector (Aah) venom administration. To this end, mast cells were depleted using compound 48/80 (C48/80). Furthermore, the involvement of tachykinin NK1 receptors expressed on mast cell membranes was elucidated by their blocking with an antagonist. On the other hand, the ability of Aah venom to increase vascular permeability and to induce edema was also assessed by measuring the amount of Evans blue dye (EBD) extravasation in bronchoalveolar lavage (BAL) fluid and in the lungs of mice. Pulmonary edema, as assessed by the levels of EBD extravasation, was completely inhibited in compound 48/80-treated animals. Depletion by stimuli non-immunological C48/80 component markedly reduced induced inflammatory response following the venom administration. The mast cells seem to play an important role in the development of lung injury and the increase of vascular permeability in mice following the subcutaneous administration of Aah scorpion venom through the NK1 receptor.

The importance of early bedside echocardiography in children with scorpion envenomation.

Scorpion sting may cause myocardial injury and heart failure (HF). Clinical signs of failure may develop several hours or even days after the sting, while electrocardiography (ECG) and blood examination soon after the sting may be normal. We sought to examine whether normal echocardiographic (echo) examination performed shortly after hospital arrival would exclude subsequent HF. We also sought to check if blood troponin and natriuretic peptide values measured shortly after arrival may predict or exclude subsequent HF. Natriuretic peptide activities have not been measured in scorpion sting victims. We also wanted to check if HF occurs in envenomated young infants. In a 3-year prospective study we looked at the demographic, clinical, laboratory, ECG, and echo data of all patients with general envenomation who arrived at the emergency department (ED) after scorpion sting. Clinical, laboratory, ECG, and echo results on arrival and 24 h after arrival were checked and compared between groups of patients with normal and abnormal echo on arrival. We then looked for differences in clinical course, therapy, and outcome between groups. The study included 98 children aged 80 days to 19 years (median 53.1 months), 25 were below the age of 2 years. Envenomation by the "yellow scorpion"Leiurus quinquestriatus was suspected in 74 cases. Median time between sting and ED arrival was 80 min. Echo was performed on arrival in 93 of the 98 patients, (in 5 occasions it was not performed or not recorded) 74 were normal and 19 were abnormal. Abnormal echo included hypokinesia and low fractional shortening and ejection fraction of the left ventricle. Clinical signs, abnormal ECG, and laboratory results were not discriminative between groups on arrival. Mean troponin T was higher in patients with abnormal echo, but within normal range in 13 of the 19 patients with abnormal echo and above normal in 2 of the 74 patients with normal echo-missing sensitivity and specificity. Mean N-terminal pro B-type natriuretic peptide was above normal in both groups but within normal range in 5 patients with abnormal echo and above normal range in 24 patients with normal echo-missing sensitivity and specificity. None of the patients with normal echo had subsequent HF and none of the children younger than 2 years of age had HF. All patients survived the intoxication and were discharged home without sequel. We conclude that early echo examination is an important procedure. In our study, normal examination excluded subsequent HF. Abnormal examination accelerated cardiac therapy which might have contributed to our favorable outcome. HF did not occur in infants younger than two years of age.

Pulmonary edema following scorpion envenomation: mechanisms, clinical manifestations, diagnosis and treatment.

Scorpion envenomation is common in tropical and subtropical regions. Cardio-respiratory manifestations, mainly cardiogenic shock and pulmonary edema, are the leading causes of death after scorpion envenomation. The mechanism of pulmonary edema remains unclear and contradictory conclusions were published. However, most publications confirm that pulmonary edema has been attributed to acute left ventricular failure. Cardiac failure can result from massive release of catecholamines, myocardial damage induced by the venom or myocardial ischemia. Factors usually associated with the diagnosis of pulmonary edema were young age, tachypnea, agitation, sweating, or the presence of high plasma protein concentrations. Treatment of scorpion envenomation has two components: antivenom administration and supportive care. The latter mainly targets hemodynamic impairment and cardiogenic pulmonary edema. In Latin America, and India, the use of Prazosin is recommended for treatment of pulmonary edema because pulmonary edema is associated with arterial hypertension. However, in North Africa, scorpion leads to cardiac failure with systolic dysfunction with normal vascular resistance and dobutamine was recommended. Dobutamine infusion should be used as soon as we have enough evidence suggesting the presence of pulmonary edema, since it has been demonstrated that scorpion envenomation can result in pulmonary edema secondary to acute left ventricular failure. In severe cases, mechanical ventilation can be required.