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Individual variability in Tityus serrulatus (Scorpiones, Buthidae) venom elicited by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Venom variability in specimens of Tityus serrulatus scorpion was assessed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOFMS) analyses. An expanded time lag venom extraction protocol was carried out using ten scorpions to study individual variations that might occur due to different rates in protein expression and/or processing. The first extraction of venom was made from the animals after 20 days of starvation, which allowed the venom gland to be filled up. The second extraction event was carried out 24 hours after the first one. The third was 8 days after the first extraction. By means of MALDI-TOF analyses, important variations were observed in venoms of a single specimen extracted at different times, especially in latter extraction events. These variations are most probably related to dynamics in cell gland production. Since T. serrulatus is a parthenogenetic species, sexual variations are naturally excluded and we did not expect intra-specific variations, which was confirmed. Knowledge of individual venom variability is extremely important to avoid misunderstandings in the use of venom proteomic analysis as a taxonomic tool.

Animals↗

Crystal structures of two alpha-like scorpion toxins: non-proline cis peptide bonds and implications for new binding site selectivity on the sodium channel.

The crystal structures of two group III alpha-like toxins from the scorpion Buthus martensii Karsch, BmK M1 and BmK M4, were determined at 1.7 A and 1.3 A resolution and refined to R factors of 0.169 and 0.166, respectively. The first high-resolution structures of the alpha-like scorpion toxin show some striking features compared with structures of the "classical" alpha-toxin. Firstly, a non-proline cis peptide bond between residues 9 and 10 unusually occurs in the five-member reverse turn 8-12. Secondly, the cis peptide 9-10 mediates the spatial relationship between the turn 8-12 and the C-terminal stretch 58-64 through a pair of main-chain hydrogen bonds between residues 10 and 64 to form a unique tertiary arrangement which features the special orientation of the terminal residues 62-64. Finally, in consequence of the peculiar orientation of the C-terminal residues, the functional groups of Arg58, which are crucial for the toxin-receptor interaction, are exposed and accessible in BmK M1 and M4 rather than buried as in the classical alpha-toxins. Sequence alignment and characteristics analysis suggested that the above structural features observed in BmK M1 and M4 occur in all group III alpha-like toxins. Recently, some group III alpha-like toxins were demonstrated to occupy a receptor site different from the classical alpha-toxin. Therefore, the distinct structural features of BmK M1 and M4 presented here may provide the structural basis for the newly recognized toxin-receptor binding site selectivity. Besides, the non-proline cis peptide bonds found in these two structures play a role in the formation of the structural characteristics and in keeping accurate positions of the functionally crucial residues. This manifested a way to achieve high levels of molecular specificity and atomic precision through the strained backbone geometry.

Amino Acid Sequence↗

Blocking of the squid axon K+ channel by noxiustoxin: a toxin from the venom of the scorpion Centruroides noxius.

We have studied the selective effects of noxiustoxin (NTX), a fraction of the venom of the scorpion Centruroides noxius, on the K currents of perfused squid giant axons using the voltage-clamp technique. At concentrations below 1.5 microM, NTX blocked K currents in a voltage-independent manner, with little effect on their turning-on and turning-off kinetics. Above 1.5 microM, the block by NTX became voltage-dependent and could be partially removed by repetitive pulsing and strong depolarizations. Long repolarizations and more negative holding potentials favoured the slow restoration of channel block. Reduction of K currents by internally perfusing the fibers with solutions of low K+ concentration (200 mM), affected very little the removal of NTX-block during repetitive pulsing, suggesting that block removal depended on membrane potential and not on outward movements of K+ ions through open channels. In high extracellular K+ (300 mM) the blocking action of NTX was reduced and the instantaneous I-V characteristics showed a marked outward rectification. At 20 microM NTX, inward tail currents measured on step repolarizations to -70 mV were fully blocked, suggesting a direct interaction of the toxin with the open channel. The effects of the total venom Centruroides noxius Hoffmann was also studied. External application of 0.25 mg/ml of the venom caused a marked reduction of both Na and K currents, an effect similar to that of other scorpion venoms.

Animals↗

Assessment of left ventricular function in severe scorpion envenomation: combined hemodynamic and echo-Doppler study.

OBJECTIVE: To assess left ventricular function in patients presenting with pulmonary edema following scorpion envenomation. DESIGN: Cohort study. SETTING: Medical intensive care unit of a teaching hospital. PATIENTS: Nine consecutive adult patients stung by Androctonus australis and presenting with pulmonary edema entered the study. Fourteen normal volunteers comprised the control group. INTERVENTIONS: Upon admission, all patients had right heart catheterization and, within the first 8 h, a Doppler echocardiographic study. Results of Doppler echocardiographic studies were compared to those of controls. MEASUREMENTS AND RESULTS: Usual hemodynamic information (heart and vascular pressures, derived data and tissue oxygenation parameters), left ventricular dimensions and indicators of systolic function, and Doppler-derived parameters of left ventricular filling and diastolic function were obtained upon admission. Serial echocardiographic measurements were repeated daily until full clinical recovery (eight patients) or death (one patient). All patients had a hemodynamic profile of acute congestive heart failure (mean PAOP = 24 +/- 2 mmHg; mean SVI = 22 +/- 7 ml/m2; mean CI = 2.5 +/- 0.5 l/min/m2). However, SVR were not increased (mean = 22 +/- 3 U/m2). Left ventricle was hypokinetic in all patients with transient mitral regurgitation present in five patients. Left ventricular systolic function was markedly depressed (FS = 12 +/- 6%; EF = 26 +/- 12%). An associated diastolic dysfunction is suggested by Doppler records of mitral inflow. Left ventricular systolic function evolved toward normalization within 6 +/- 2 days preceded by full clinical recovery. CONCLUSIONS: These data suggest that pulmonary edema in scorpion envenomation is of hemodynamic origin and is related to a severe and prominent impairment of left ventricular systolic function.

Adolescent↗

Toxin III of the scorpion Androctonus australis Hector: proton nuclear magnetic resonance assignments and secondary structure.

1H NMR has been applied to a 3.5 mM, pH 5.4, solution of toxin III (64 amino acids) from venom of the scorpion Androctonus australis Hector. The resonance assignment strategy began by applying a generalized main-chain directed method for rapid identification and resonance assignments of secondary structures. The remaining resonances were assigned by the sequential method. Major structural features include a helix of 2 1/2 turns (residues 20-28) which is linked by two disulfide bridges to the central strand of a triple-stranded anti-parallel beta-sheet. Turns were identified at residues 15-17, 47-49 and also at residues 51-53. Numerous NOEs have been observed between hydrophobic residues which suggest the presence of a hydrophobic core; these include Leu37, Leu23, Val47, Tyr14, Trp45 and Tyr5. The Trp45 and Tyr5 rings lie orthogonal to one another. No crystal structure has been solved for this AaH III toxin. Comparisons are made with other members of the scorpion toxin family.

Amino Acid Sequence↗

Outcome of scorpion sting envenomation after a protocol guided therapy.

OBJECTIVE: Scorpion sting (SS) envenomation is a life threatening emergency in children, though not so severe in adults. Attempt to develop protocol using prazosin and dobutamine and few other drugs to treat SS. METHODS: Children aged 0-13 years with a history of scorpion sting were studied. Clinical features, complications, drug therapy and outcome of the cases for the period 1992-97(N = 186) was collected by the authors and also from the medical records department (RETROSPECTIVE GROUP). Cases treated during 1997-2000 (N = 198) as per the protocol were recorded as PROSPECTIVE GROUP. All the cases were observed for at least for 24 hours. Cases coming within 4 hours of a sting were given a dose of Prazosin (30 mic.gm/Kg/dose) and were observed. Those who came after 4 hours & were asymptomatic received only symptomatic treatment. Cases with signs of envenomation received Prazosin every 6 hourly till recovery. Cases having acute pulmonary edema (APE) were treated with dobutamine and sodium nitroprusside drip. Complicated cases were monitored in PICU as per the protocol. RESULT: Complications associated with excessive parasympathetic and sympathetic stimulation were observed. Myocarditis was observed due to the toxin and excessive catecholamine, which complicated in left ventricular failure (LVF) and APE. Nearly half of the children with acute myocarditis developed APE. Death was mainly due to myocarditis and APE, with or without encephalopathy. Mortality was high in children who received steroid and antihistaminics outside and who came late (> 4 hours). CONCLUSION: Complication rate remained almost same in both the groups. There was a significant reduction in overall mortality (P = < 0.0155) and in deaths associated with APE (P = < 0.0001) after the protocol guided therapy. There was also a reduction in mortality in encephalopathy group though not statistically significant. This treatment protocol and aggressive management of APE reduced the mortality due to SS significantly.

Adolescent↗

Indian red scorpion envenoming.

The clinical course and treatment outcome of scorpion envenoming in 293 children was studied in a hospital at Mahad in Raigad district of Maharastra. 111 (38%) children who reported 1-10 hours (mean 3.5 hours) after sting had hypertension, 87 (29.6%) with tachycardia reported within 1-24 hours (mean 6.7 hours) of being envenomed and 72 (24.5%) children developed acute pulmonary edema after 6-24 hours (mean 8 hours) of sting. Six victims were brought dead, while 17 (6%) died later owing to multiorgan failure with loss of consciousness and convulsions (who reported after 24 hours of sting). Early administration of prazosin (125-250 ug orally) improved the clinical symptoms. Morbidity and mortality due to scorpion envenoming depends upon time lapse between sting and administration of post synaptic alpha-1 blocker, prazosin hydrochloride.

Adolescent↗

Design and evaluation of a diabody to improve protection against a potent scorpion neurotoxin.

Diabodies are recombinant, dimeric, antibody-based molecules composed of two non-covalently associated single-chain antibody fragments that bind to an antigen in a divalent manner. In an attempt to develop more effective therapeutic molecules against scorpion venoms, we designed a diabody derived from monoclonal antibody 9C2, which neutralizes the toxicity of scorpion neurotoxin AahI in mammals. The recombinant diabody produced in the periplasm of Escherichia coli was purified to homogeneity in a single step by protein L-agarose affinity chromatography. It was functional, and possessed a high binding affinity to AahI (8 x 10(-11) M). The bivalence of the diabody was confirmed by size-exclusion chromatography, isoelectrofocussing and electron microscopic observations. Finally, the diabody showed high thermal stability in serum and demonstrated protective activity when injected intraperitoneally in mice experimentally envenomed with toxin AahI. In conclusion, the diabody format gives the 9C2 molecule advantageous properties that are particularly important for potential clinical applications in the treatment of envenomations.

Animals↗

Functional analysis of the alpha-neurotoxin, BmalphaTX14, derived from the Chinese scorpion, Buthus martensii Karsch.

The gene encoding the BmalphaTX14 (alpha-neurotoxin TX14) protein, derived from the cDNA library of the Chinese scorpion Buthus martensii Karsch, was expressed in Pichia pastoris. The recombinant protein was purified by metal chelate affinity chromatography and gel filtration chromatography. Using patch-clamp technique, electrophysiological activity of rBmalphaTX14 was identified. In the neurons isolated from mice trigeminal root ganglion, the Na+ current amplitude was reduced by 80% under whole cell patch-clamp recording. There were no apparent modifications to the gating mechanism in the presence of rBmalphaTX14. Although BmalphaTX14 shared a high amino acid sequence similarity with other typical alpha-toxins, it has different effects on neurons. Further electrophysiological analysis suggested that rBmalphaTX14 selectively blocked Na+ channels and is a member of a new group of scorpion toxins.

Animals↗

Amino acid sequence of toxin VII, a beta-toxin from the venom of the scorpion Tityus serrulatus.

The sequence of the 61 amino acids of toxin VII, a beta-toxin from the venom of the South American scorpion Tityus serrulatus, has been determined by automatic sequencing of the reduced and S-[14C] carboxymethylated protein and of tryptic peptides obtained before or after citraconylation of this protein. This toxin, the most active beta-toxin from this venom, is the first Tityus toxin to be fully sequenced. The results clearly show that toxin VII belongs to the structural group of scorpion toxins originating from Central and North America.

Amino Acid Sequence↗

Photoaffinity labeling of scorpion toxin receptors associated with insect synaptosomal Na+ channels.

Photoreactive and radioiodinated derivatives of several scorpion toxins acting on insect Na+ channels were prepared without loss of their pharmacological activities. Photoaffinity experiments were carried out on a synaptosomal fraction from the nerve cord of the cockroach Periplaneta americana: with all toxin derivatives, a single specifically labeled band was obtained with a molecular weight of 188,000 +/- 12,000 (n = 17). These results indicate for the first time the molecular weight of the scorpion toxin receptor from the insect nervous system which is probably associated with voltage sensitive Na+ channels. One of these toxins, toxin VII from Tityus serrulatus venom, has been previously shown to be active both in mammals and in insects, in rat brain synaptosomes this toxin labeled a Mr = 31,000 +/- 4,000 band in contrast, to observations in the insect preparation.

Affinity Labels↗

Characterization of a new leiurotoxin I-like scorpion toxin. PO5 from Androctonus mauretanicus mauretanicus.

Three novel peptide inhibitors of the SKCa channels were purified to homogeneity from the venom of the scorpion Androctonus mauretanicus mauretanicus using one step of RP-HPLC and competition assays with [125I]apamin to rat brain synaptosomes. PO1, PO2 and PO5 have K0.5 of 100, 100 and 0.02 nM, respectively, for the apamin binding site. The sequence of PO5 was established and compared to that of other scorpion toxins active on K+ channels: it contains 31 residues and has a free carboxyl end. it shares sequence similarity with apamin and leiurotoxin I.

Amino Acid Sequence↗

Stings by red scorpions (Buthotus tamulus) in Maharashtra State, India: a clinical study.

Of 33 cases of scorpion sting admitted to hospital in Mahad, Maharashtra State, India, 10 had a mean blood pressure between 100 and 137 mm/Hg and 11 had a heart rate between 130 and 215 per min. Seven patients developed acute pulmonary oedema; there were 2 deaths. Three patients had local pain at the site of sting. The role of vasodilators such as prazosin hydrochloride, sodium nitroprusside and nifedipine were investigated in addition to digoxin, diuretics and aminophylline to alleviate refractory myocardial failure due to scorpion sting.

Adolescent↗

Epidemiological and clinical characteristics of the scorpion envenomation in Tunisia.

This work deals with a clinical and epidemiological study of the scorpion envenomation in Tunisia, from 717 scorpion stings recorded during a year at the Sfax Hospital with a mortality rate of 0.35%, all the patients died being less 15 years old. The serotherapy with a specific antivenin made in the Institut Pasteur of Tunis is the main element of the therapeutics.

Adolescent↗

Introduction of the scorpion Centruroides exilicauda into California and its public health significance.

The bark scorpion, Centruroides exilicauda, formerly C. sculpturatus, is a native of Arizona and adjacent parts of New Mexico in the United States, and Baja California and Sonora in Mexico. A few specimens have been captured on the California side of the Colorado River. The general biology of this scorpion is discussed and the introduction of the arthropod into three California counties, probably in trailer paraphernalia and dune buggies, is described. Four probable stings by C. exilicauda in California and their public health significance are presented.

Adult↗

Antigenic cross-reactivity among the venoms from several species of Brazilian scorpions.

The venoms of seven species of scorpions living in different regions of Brazil were analysed with regard to their lethality, antigenic cross-reactivity and ability to induce antibody production. In mice, the tested scorpion venoms can be grouped as: (a) highly toxic: Tityus stigmurus Thorell (LD50 = 0.773 mg/kg), Tityus bahiensis (Perty) (LD50 = 1.062 mg/kg), Tityus serrulatus Lutz and Mello (LD50 = 1.160 mg/kg), and Tityus costatus (Karsch) (LD50 = 1.590 mg/kg); (b) moderately toxic: Tityus cambridgei Pocock (LD50 = 12.136 mg/kg); and (c) practically nontoxic: Rhopalurus agamemnon (Koch) (LD50 = 36.363 mg/kg), and Brotheas amazonicus Lourenço (LD50 = 90.909 mg/kg). On electrophoresis the venoms showed many protein bands displayed along the chromatogram, most of them cross-reacting in immunoelectrophoresis and immunoblotting using horse anti-T. serrulatus, anti-T. bahiensis or anti-T. serrulatus+T. bahiensis sera as probes. The antibodies present in these antivenoms combine with venom components as measured in vitro by the ELISA assay, and neutralize their lethal effects in vivo. These results indicate that horse anti-venoms against a mixture of T. serrulatus and T. bahiensis venoms or only against T. serrulatus venom yield an antibody population able to neutralize the toxic effects found in all venoms studied.

Animals↗

Cloning and characterization of the genomic region encoding toxin IV-5 from the scorpion Tityus serrulatus Lutz and Mello.

By means of PCR and using synthetic oligonucleotides designed from the reported cDNA, we amplified the gene that codes for toxin IV-5 from the Brazilian scorpion Tityus serrulatus. The analysis of the nucleotide sequence shows that the amplified genomic region is composed of 659 base pairs (bp) comprising two exons (28 and 284 bp) and an intron of 347 bp interrupting the region that encodes the signal peptide of the precursor toxin. Based on these findings a model for the structural organization of scorpion toxin genes is proposed.

Amino Acid Sequence↗