Genome-wide identification, structural homology analysis, and evolutionary diversification of the phospholipase D gene family in the venom gland of three scorpion species.

BACKGROUND: Venom phospholipase D (PLDs), dermonecrotic toxins like, are the major molecules in the crude venom of scorpions, which are mainly responsible for lethality and dermonecrotic lesions during scorpion envenoming. The purpose of this study was fivefold: First, to identify transcripts coding for venom PLDs by transcriptomic analysis of the venom glands from Androctonus crassicauda, Hottentotta saulcyi, and Hemiscorpius lepturus; second, to classify them by sequence similarity to known PLDs and motif extraction method; third, to characterize scorpion PLDs; fourth to structural homology analysis with known dermonecrotic toxins; and fifth to investigate phylogenetic relationships of the PLD proteins. RESULTS: We found that the venom gland of scorpions encodes two PLD isoforms: PLD1 ScoTox-beta and PLD2 ScoTox-alpha I. Two highly conserved regions shared by all PLD1s beta are GAN and HPCDC (HX2PCDC), and the most important conserved regions shared by all PLD2s alpha are two copies of the HKDG (HxKx4Dx6G) motif. We found that PLD1 beta is a 31-43 kDa acidic protein containing signal sequences, and PLD2 alpha is a 128 kDa basic protein without known signal sequences. The gene structures of PLD1 beta and PLD2 alpha contain 6 and 21 exons, respectively. Significant structural homology and similarities were found between the modeled PLD1 ScoTox-beta and the crystal structure of dermonecrotic toxins from Loxosceles intermedia. CONCLUSIONS: This is the first report on identifying PLDs from A. crassicauda and H. saulcyi venom glands. Our work provides valuable insights into the diversity of scorpion PLD genes and could be helpful in future studies on recombinant antivenoms production.

Identification, classification, and characterization of alpha and beta subunits of LVP1 protein from the venom gland of four Iranian scorpion species.

Scorpion venoms contain bioactive peptides and proteins. Some, can be used for pharmaceutical purposes. So, identification of venom proteins matters because, in addition to determining the function of the toxins can also be an excellent guide to developing new drugs. Here, we got transcriptome of venom glands from four Iranian scorpion species, including Hemsicorpius lepturus, Mesobuthus eupeus, Andructunus crassicuada, and Hottentotta saulcyi using cDNA library synthesis and high-throughput transcriptomic analysis of the venom glands. In a comparative way, we identified the cDNA encoding isoforms of subunits (alpha and beta) of BotLVP1/BmLVP1-like protein in the venom gland of three species except for H. lepturus. Characterization and structure determination of the LVP1_like proteins combined with gene map analysis provided evidence of the existence of some isoforms of LVP1_like proteins, encoded by genes with two exons and one intron, which can be classified in CSαß superfamily in the venom gland of three Iranian scorpion species. According to the high similarity with BotLVP1 and BmLVP1, these proteins could also be potent to mediate cholesterol homeostasis. However, further research is needed to prove it, and this study just may lay the foundation lead to light up this way.

Identification, Characterization, and Modeling of a Bioinsecticide Protein Isolated from Scorpion Venom gland: A Three-Finger Protein.

Background: The majority of insecticides target sodium channels. The increasing emergence of resistance to the current insecticides has persuaded researchers to search for alternative compounds. Scorpion venom gland as a reservoir of peptides or proteins, which selectively target insect sodium channels. These proteins would be an appropriate source for finding new suitable anti-insect components. Methods: Transcriptome of venom gland of scorpion Mesobuthus eupeus was obtained by RNA extraction and complementary DNA library synthesis. The obtained transcriptome was blasted against protein databases to find insect toxins against sodium channel based on the statistically significant similarity in sequence. Physicochemical properties of the identified protein were calculated using bioinformatics software. The three-dimensional structure of this protein was determined using homology modeling, and the final structure was assessed by molecular dynamics simulation. Results: The sodium channel blocker found in the transcriptome of M. eupeus venom gland was submitted to the GenBank under the name of meuNa10, a stable hydrophilic protein consisting of 69 amino acids, with the molecular weight of 7721.77 g/mol and pI of 8.7. The tertiary structure of meuNa10 revealed a conserved LCN-type cysteine-stabilized alpha/beta domain stabilized by eight cysteine residues. The meuNa10 is a member of the 3FP superfamily consisting of three finger-like beta strands. Conclusion: This study identified meuNa10 as a small insect sodium channel-interacting protein with some physicochemical properties, including stability and water-solubility, which make it a good candidate for further in vivo and in vitro experiments in order to develop a new bioinsecticide.

Peptides with Diverse Functions from Scorpion Venom: A Great Opportunity for the Treatment of a Wide Variety of Diseases.

The venom glands are a rich source of biologically important peptides with a wide range of pharmaceutical properties. Scorpion venoms have been identified as a reservoir for the components which might be considered as great candidates for drug development. These components are usually used by a scorpion to capture prey and defense; however, pharmacological properties of the venom compounds have been confirmed in the treatment of different disorders, including cardiac diseases, autoimmune diseases, infections, and varied cancer types. Ion channel blockers, antimicrobial peptides and proteins, are the main groups of scorpion venom components. Despite several studies exist on the subject of scorpion peptides, there are still valuable components to be discovered. Additionally, owing to the improvement of proteomics and transcriptomics, the number of peptide drugs is steadily increasing, which reflects the importance of it. This review evaluates the available literature about some important scorpion venom peptides with pharmaceutical activities. Given that the last three years have been dominated by the COVID-19 from the medical/pharmaceutical perspective, scorpion compounds with potential against the coronavirus 2 (SARS-CoV-2) are also discussed in this review.

Intraspecific differences in Androctunus crassicauda venom and envenomation symptoms.

Envenomation by Androctunus crassicauda is very frequent in Iran, especially in the south-west. This scorpion is one of the six scorpions whose venom is used to prepare anti-venom. Using HPLC, we discovered venom components of A. crassicauda varies from one specimen to another depending on geographical location, and this result is confirmed by those first found in various symptoms of A. crassicauda sting in envenomed persons from two separate geographical places (north and south of Khuzestan province). There was a significant relationship between symptoms and location of envenomation by A. crassicauda. Muscle spasm was more dominant in envenomed people from Northern cities, and venom chromatogram analysis showed the presence of at least six main sharp peaks in Northern A. crassicauda rather than Southern A. crassicauda. It shows intraspecific differences in venom of A. crassicauda that must be considered in treatment of stung people from different geographical locations as well as in the preparation of anti-venom. See also Figure 1(Fig. 1).

Effects of climate variables on the incidence of scorpion stings in Iran for five years.

BACKGROUND: Although scorpionism is recorded worldwide, some regions such as Iran present a higher incidence. Due to the great prevalence of scorpion stings in Khuzestan province, southwestern Iran, the present study examined the relationship between different climate parameters and the scorpion sting rate in this area from April 2010 to March 2015. METHODS: In this cross-sectional descriptive-analytical study, we considered all scorpion sting cases recorded in the Department of Infectious Diseases, Ahvaz Jundishapur University of Medical Sciences. Data were analyzed using statistics, frequency distribution and Pearson's correlation coefficient. RESULTS: A total of 104,197 cases of scorpion stings was recorded from 2010 to 2015. The cumulative incidence of scorpion sting was 2.23%. The spatial distribution of scorpion stings showed that most cases occurred in the Dehdez district (4,504 scorpion stings/100,000 inhabitants) and the Masjed Soleyman county (4,069 scorpion stings/100,000 inhabitants). A significant association was found between climate factors (temperature, evaporation rate, sunshine duration, humidity, and precipitation) and the scorpion sting rate. An increase in rainfall and humidity coincided with a reduction in scorpion stings whereas an increase in temperature, evaporation, and sunshine duration was accompanied by a growth of scorpion stings. No significant correlation was found between wind velocity/direction and the incidence rate of stings. Moreover, the seasonal peak incidence of scorpion stings was recorded in summer (an average of 8,838 cases) and the lowest incidence was recorded during winter (an average of 1,286 cases). The annual trend of scorpion sting cases decreased during the period from 2010 to 2015. CONCLUSION: Climate variables can be a good index for predicting the incidence of scorpion stings in endemic regions. Since they occur mostly in the hot season, designing preventive measures in the counties and districts with a high incidence of scorpion stings such as Dehdez and Masjed Soleyman can minimize mortality and other burdens.

Effects of climate variables on the incidence of scorpion stings in Iran for five years

Background: Although scorpionism is recorded worldwide, some regions such as Iran present a higher incidence. Due to the great prevalence of scorpion stings in Khuzestan province, southwestern Iran, the present study examined the relationship between different climate parameters and the scorpion sting rate in this area from April 2010 to March 2015. Methods: In this cross-sectional descriptive-analytical study, we considered all scorpion sting cases recorded in the Department of Infectious Diseases, Ahvaz Jundishapur University of Medical Sciences. Data were analyzed using statistics, frequency distribution and Pearson's correlation coefficient. Results: A total of 104,197 cases of scorpion stings was recorded from 2010 to 2015. The cumulative incidence of scorpion sting was 2.23%. The spatial distribution of scorpion stings showed that most cases occurred in the Dehdez district (4,504 scorpion stings/100,000 inhabitants) and the Masjed Soleyman county (4,069 scorpion stings/100,000 inhabitants). A significant association was found between climate factors (temperature, evaporation rate, sunshine duration, humidity, and precipitation) and the scorpion sting rate. An increase in rainfall and humidity coincided with a reduction in scorpion stings whereas an increase in temperature, evaporation, and sunshine duration was accompanied by a growth of scorpion stings. No significant correlation was found between wind velocity/direction and the incidence rate of stings. Moreover, the seasonal peak incidence of scorpion stings was recorded in summer (an average of 8,838 cases) and the lowest incidence was recorded during winter (an average of 1,286 cases). The annual trend of scorpion sting cases decreased during the period from 2010 to 2015. Conclusion: Climate variables can be a good index for predicting the incidence of scorpion stings in endemic regions. Since they occur mostly in the hot season, designing preventive measures in the counties and districts with a high incidence of scorpion stings such as Dehdez and Masjed Soleyman can minimize mortality and other burdens.(AU)

Scorpion stings in pregnancy: an analysis of outcomes in 66 envenomed pregnant patients in Iran.

BACKGROUND: Scorpionism is one of the most important health problems in tropical regions, which unfortunately results in thousands of deaths annually. Pregnant women are potential victims in areas with high scorpion-sting prevalence. Limited medical data are available on the effects of scorpion envenomation in pregnant women. This study aimed to examine the effect of scorpion envenomation on pregnancy outcomes in 66 cases. METHODS: The present descriptive/analytical cross-sectional study was performed on 66 scorpion-envenomed pregnant women referred to the clinical toxicology unit of Ahvaz Razi Hospital in Iran during 2015-2017. The variables assessed in all cases, via questionnaire and hospital medical records, were: age, patient residency, gestational week, status of the fetus, laboratory anomalies, clinical severity of envenomation, sting site and scorpion species. Pregnancy outcome (miscarriage, stillbirth, preterm birth, normal delivery) and status of the newborns were also evaluated. Data were analyzed using SPSS ® software (version 24.0). RESULTS: The following pregnancy outcomes were recorded from envenomed pregnant women: miscarriage = 1.5% (n = 1), stillbirth = 4.5% (n = 3), preterm birth = 10.6% (n = 7), normal birth = 83% (n = 55). Among participants whose pregnancy led to birth, 11(17.7%) cases had prenatal-neonatal complications. Neonatal complications, including Apgar score less than 8 points at 5 min, were found in 7 (11.3%) preterm birth cases and in 4 (6.4%) normal birth cases, along with birth weight below 2500 g in normal births. A significant relationship was found between adverse pregnancy outcomes and bite location, as well as scorpion species, but no relationship was found with other variables. CONCLUSION: Envenomation significantly contributes to preterm birth. Moreover, the location of bites and the type of scorpion species have a decisive role in the pregnancy outcome of scorpion-envenomed pregnant women.

Scorpion stings in pregnancy: an analysis of outcomes in 66 envenomed pregnant patients in Iran

Scorpionism is one of the most important health problems in tropical regions, which unfortunately results in thousands of deaths annually. Pregnant women are potential victims in areas with high scorpion-sting prevalence. Limited medical data are available on the effects of scorpion envenomation in pregnant women. This study aimed to examine the effect of scorpion envenomation on pregnancy outcomes in 66 cases. Methods: The present descriptive/analytical cross-sectional study was performed on 66 scorpion-envenomed pregnant women referred to the clinical toxicology unit of Ahvaz Razi Hospital in Iran during 2015-2017. The variables assessed in all cases, via questionnaire and hospital medical records, were: age, patient residency, gestational week, status of the fetus, laboratory anomalies, clinical severity of envenomation, sting site and scorpion species. Pregnancy outcome (miscarriage, stillbirth, preterm birth, normal delivery) and status of the newborns were also evaluated. Data were analyzed using SPSS ® software (version 24.0). Results: The following pregnancy outcomes were recorded from envenomed pregnant women: miscarriage = 1.5% (n = 1), stillbirth = 4.5% (n = 3), preterm birth = 10.6% (n = 7), normal birth = 83% (n = 55). Among participants whose pregnancy led to birth, 11(17.7%) cases had prenatal-neonatal complications. Neonatal complications, including Apgar score less than 8 points at 5 min, were found in 7 (11.3%) preterm birth cases and in 4 (6.4%) normal birth cases, along with birth weight below 2500 g in normal births. A significant relationship was found between adverse pregnancy outcomes and bite location, as well as scorpion species, but no relationship was found with other variables. Conclusion: Envenomation significantly contributes to preterm birth. Moreover, the location of bites and the type of scorpion species have a decisive role in the pregnancy outcome of scorpion-envenomed pregnant women.(AU)

Three New Scorpion Chloride Channel Toxins as Potential Anti-Cancer Drugs: Computational Prediction of The Interactions With Hmmp-2 by Docking and Steered Molecular Dynamics Simulations.

Scorpion venom is a rich source of toxins which have great potential to develop new therapeutic agents. Scorpion chloride channel toxins (ClTxs), such as Chlorotoxin selectively inhibit human Matrix Methaloproteinase-2 (hMMP-2). The inhibitors of hMMP-2 have potential use in cancer therapy. Three new ClTxs, meuCl14, meuCl15 and meuCl16, derived from the venom transcriptome of Iranian scorpion, M. eupeus (Buthidea family), show high sequence identity (71.4%) with Chlorotoxin. Here, 3-D homology model of new ClTxs were constructed. The models were optimized by Molecular Dynamics simulation based on MDFF (molecular dynamics flexible fitting) method. New ClTxs indicate the presence of CSαß folding of other scorpion toxins. A docking followed by steered molecular dynamics (SMD) simulations to investigate the interactions of meuCl14, meuCl15, and meuCl16 with hMMP-2 was applied. The current study creates a correlation between the unbinding force and the inhibition activities of meuCl14, meuCl15 and meuCl16 to shed some insights as to which toxin may be used as a drug deliverer. To this aim, SMD simulations using Constant Force Pulling method were carried out. The SMD provided useful details related to the changes of electrostatic, van de Waals (vdW), and hydrogen-bonding (H-bonding) interactions between ligands and receptor during the pathway of unbinding. According to SMD results, the interaction of hMMP-2 with meuCl14 is more stable. In addition, Arginine residue was found to contribute significantly in interaction of ClTxs with hMMP-2. All in all, the present study is a dynamical approach whose results are capable of being implemented in structure-based drug design.

First Transcriptome Analysis of Iranian Scorpion, Mesobuthus Eupeus Venom Gland.

Scorpions are generally an important source of bioactive components, including toxins and other small peptides as attractive molecules for new drug development. Mesobuthus eupeus, from medically important and widely distributed Buthidae family, is the most abundant species in Iran. Researchers are interesting on the gland of this scorpion due to the complexity of its venom. Here, we have analyzed the transcriptome based on expressed sequence tag (EST) database from the venom tissue of Iranian M. eupeus by constructing a cDNA library and subsequent Sanger sequencing of obtained inserts. Sixty-three unique transcripts were identified, which were grouped in different categories, including Toxins (44 transcripts), Cell Proteins (9 transcripts), Antimicrobial Peptides (4 transcripts) and Unknown Peptides (3 transcripts). The analysis of the ESTs revealed several new components categorized among various toxin families with effect on ion channels. Sequence analysis of a new precursor provides evidence to validate the first CaTxs from M. eupeus. The results are exploration of the diversity of precursors expressed of Iranian M. eupeus venom gland. We further described comparative analysis of venom components of Iranian M. eupeus with other sibling species.

A Novel Defensin-Like Peptide Associated with Two Other New Cationic Antimicrobial Peptides in Transcriptome of the Iranian Scorpion Venom.

INTRODUCTION: Scorpion venom is a source of bioactive peptides, and some antimicrobial peptides (AMPs) have been found in the venom gland of scorpions. Therefore, the discovery of new anti-infective agents is an essential need to overcome the problem of antibiotic resistance of clinical isolates. Here, we describe three new cationic AMPs, including meuVAP-6, meuAP-18-1, and meuPep34 from the venom gland of the Iranian scorpion, Mesobuthus eupeus. METHODS: The cDNA sequences encoding all the three peptides were obtained from the cDNA library of scorpion venom gland and were deposited in the GenBank database. RESULTS: MeuVAP-6 and meuAP-18-1 are non-disulphide-bridged antimicrobial peptides, while meuPep34 is a cysteine-rich defensin-like peptide. DISCUSSION: All three identified AMPs are rich in arginine and tryptophan. The overall results from the length, net charge, and hydrophobicity index suggested that meuPep34 could be the most active AMPs with the potential ability of biofilm inhibition. The data from molecular characterization of identified AMPs can provide a platform for further investigations in the drug design.