In vivo and in vitro effects of scorpion venoms in Turkey: a mini-review

Scorpion stings constitute a common type of accident in Turkey due to the geographic location, climate and socioeconomic structure of this country. Moreover, envenomation cases are considered a public health problem throughout Turkey. Based on data in the literature, the main clinical features of scorpion envenomations are defined as systemic manifestations including sweating, hypertension and vomiting, and among prominent causes of death are cardiorespiratory manifestations, toxic myocarditis and pulmonary edema. The most important health-threatening scorpions found in Turkey are: Androctonus crassicauda, Leiurus quinquestriatus, Mesobuthus gibbosus and Mesobuthus eupeus, all of which belong to the Buthidae family. This study indicates that there is no data related to venom toxicity and in vivo effects of other scorpions found in Turkey – such as Mesobuthus caucasicus, Mesobuthus nigrocinctus and Hottentotta saulcyi – and that further research should be performed to determine their health effects.


INTRODUCTION
Of an estimated 1,500 distinct scorpion species found worldwide, only 30 types of scorpion venoms have been characterized.Although about 100,000 different polypeptides constituting the venoms of all these species have been estimated, only 0.02% have been defined (1).Scorpion venom contains simple proteins with short neurotoxin and low-molecular-weight polypeptides; it also presents lethal and paralytic effects.The scorpion toxins that affect sodium channels (NaCh) gating in excitable cells are divided into α and β classes.Alpha-toxins have been found in scorpions throughout the world, whereas anti-mammalian β-toxins have been assigned to New World scorpions and anti-insect selective β-toxins (depressant and excitatory) have been described only in the Old World (1)(2)(3)(4).
Scorpions can cause serious health problems by stinging humans; most of the time they use their venom to protect themselves.Several studies on scorpion sting cases have emphasized various clinical pictures, ranging from local to serious autonomic and central nervous system symptoms, including death due to cardiac and respiratory failure, especially in children (5).However, scorpions do not harbor any agent capable of causing disease.
In spite of the limitations on epidemiological and clinical data, in this paper we inform about both in vitro and in vivo effects of scorpionism in Turkey, based on previously conducted studies.

Androctonus crassicauda
The species A. crassicauda from Turkey has been studied for toxin determination, and five toxins were described by Calıskan et al. (15), two of which (Acra 1, 6496.8 Da and Acra 2, 7848.6 Da) were lethal to mice (15).Furthermore, Ozkan et al. (16) employed high-performance liquid chromatography (HPLC) to obtain the Adiguzel S. In vivo and in vitro effects of scorpion venoms in Turkey: a mini-review.J Venom Anim Toxins incl Trop Dis.2010;16(2):201 chromatographic profile of A. crassicauda venom and about 25 compounds were separated.Forty-two major components in the venom were within 437 to 44.737 Da according to mass spectrometry (LC-ESI-HRMS) of [MH] + (16).

Mesobuthus gibbosus
Ucar et al. (17) analyzed a peptide from Mesobuthus gibbosus venom and described two of its five fractions and their lethality on mice and Musca domestica larvae by using in vivo toxicity tests (fractions 4 and 5).

In vivo and in vitro effects
The same species present a range of LD 50 values in the lethality test of venoms.
Signs and symptoms depend on various factors including genus, species, age, weight, feeding state and structure of the scorpion, as well as the amount of venom injected, the number of stings, the stung site, the patient's susceptibility, the time spent between the sting and the first medical aid, and the climate of the region (18,19).
Scorpion toxins can change neuronal action potentials and cause the release of neurotransmitters from cholinergic and adrenergic neurons (4,(19)(20)(21)(22)(23)(24).Neurotoxins also disrupt synaptic vesicles, leading to the cessation of acetylcholine (ACh) release and to the blockage of neuromuscular transmission (18,23,25).In vertebrates, ACh is the major transmitter at neuromuscular junctions, autonomic ganglia, parasympathetic effector junctions, sympathetic effector junctions, and at many sites in the central nervous system.It is released by motor nerve cells and once attached to its receptors on muscle fibers, stimulates the fibers to contract.The glands that receive impulses from the parasympathetic part of the autonomic nervous system are stimulated in the same way (18,21).Scorpion venom stimulates neuronal sodium channels, resulting thus in autonomic storm.Stimulation of both branches of the autonomic system leads to sweating, salivation, vomiting, fasciculation, hypotension, hypertension, bradycardia or tachycardia, ventricular premature contraction, cool extremities, pulmonary edema, shock, and priapism in men.The majority of envenomation symptoms are due to a massive release of catecholamines, which play an important role in the pathogenesis of scorpion sting; they are released from the adrenal glands and postganglionic nerve Adiguzel S. In vivo and in vitro effects of scorpion venoms in Turkey: a mini-review.J Venom Anim Toxins incl Trop Dis.2010;16(2):202 endings and act either on β-adrenergic receptors, increasing the peripheral resistance, or on α-adrenergic receptors, increasing the cardiac contractility or causing renin release from the kidneys.Pulmonary edema, as a result of scorpion envenomation, is due to myocardial dysfunction.Bradykinin-induced secretory pulmonary edema is secondary and results of kallikrein stimulation due to the tissue damage caused by anoxia and accumulation of oxygen free radicals when cardiogenic manifestations are not managed early enough with prazosin, which enhances insulin secretion by blocking α-receptors on β cells in the pancreas.
Rats experimentally envenomed with A. crassicauda venom developed acute renal failure, liver dysfunction and cell destruction.Androctonus crassicauda venom showed its toxicity by altering the electrolyte balance, especially sodium and chlorine ion levels in the serum, rather than by inhibiting AchE activity (28).
Sahin et al. (29) emphasized that the levels of aspartate aminotransferase, gammaglutamyl transpeptidase, malondialdehyde, reduced glutathione and blood urea nitrogen notably increased in rats after experimental envenomation with M. eupeus venom in a province of eastern Turkey.
Mice experimentally envenomed with M. gibbosus venom manifested tremor, hypersalivation, mouth and nose bleeding, paralysis and death (30).Ucar and Tas (31) indicated that M. gibbosus scorpion venom contains peptides with specific cholinesterase inhibitory activities, which may be responsible for some of the neurotoxic effects of the venom on animals and humans.However, those authors also demonstrated that M. gibbosus venom has a peptide showing specific MAO-A inhibitory activity, which may be responsible for the anxiogenic effects of the scorpion venoms (31).

Epidemiological and Clinical Data on Scorpionism in Turkey
There are only a few epidemiological and clinical records of case reports involving Mesobuthus eupeus and Androctonus crassicauda, as well as scorpionism in children in the western and southeastern parts of Turkey (5,(32)(33)(34)(35)(36)(37).
Adiguzel S. In vivo and in vitro effects of scorpion venoms in Turkey: a mini-review.J Venom Anim Toxins incl Trop Dis.2010;16(2):203

Epidemiological aspect
In Turkey, epidemiological studies have shown that most scorpion stings affect females.The highest incidence of scorpion stings was recorded for the warmest months of the year since scorpions become more active in the warm season.
Scorpion sting was mostly seen in extremities, especially upper limbs.Such a high ratio of scorpion sting in extremities is due to the country's socioeconomic structure, which depends on agriculture in rural areas, where people wear sandals or walk barefoot (especially children), put on shoes without pre-shaking them, hand search for scorpions in their homes, lift up stones in a non-controlled manner and move their hands during sleep or rest to get away of scorpions.Stings in the head, neck and other body sites mostly occur at sleep because bed mattresses are not checked and sleeping clothes are put on without checking (5,32,33,35,37).

Clinical aspect
Based on recent data in the literature on scorpionism and its clinical aspects in Turkey, envenomation has been characterized by local and systemic symptoms.
Such effects could be related to the scorpion species and its feeding state; the venom structure and amount; the number of stings; sensitivity, age and weight of the patient; the time of admission to the hospital; and the climate of the region (20,22,38).

Envenomation by Androctonus crassicauda
Venoms from species of the Androctonus genus are highly toxic (10).Androctonus crassicauda venom has a 0.32 mg/kg LD 50 (dose killing 50% of animals) when intravenously injected into mice, which makes this scorpion species one of the most toxic in the world (21,43).In addition, Androctonus crassicauda venom can specifically stimulate acetylcholine receptors throughout the body; therefore, it can be considered neurotoxic.Similarly to the venoms of other scorpion species from different parts of the world, A. crassicauda venom can directly increase ACh concentration (18,20,44,45).Radmanesh (20) noticed adrenergic signs, pain and no clinical symptoms using low concentrations of this venom, which cannot reach a threshold to stimulate ACh receptors.On the other hand, cholinergic signs only occur when high venom concentrations are used.Most symptoms are caused either by the release of catecholamines from adrenal glands (sympathetic nerves) or by the release of acetylcholine from postganglionic parasympathetic neurons (18,20,44,45).Ozkan et al. (35) noticed that, in A. crassicauda toxicity cases, the patients had parasympathetic and local failures, which are characterized by pain, hyperemia and edema.Bosnak et al. (37) stated that pulmonary edema developed in 9.6% and dyspnea in 23.0% of the victims; the predominant signs of cardiovascular system compromising were tachycardia (36.5%), dyspnea (23.0%), paleness (15.3%), hypertension (7.6%) and hypotension (3.8%), and manifestations of cholinergic stimulation, including excessive sweating (32.6%) and vomiting (3.8%), were seen after scorpion sting in children in southeast Turkey.In addition, Al et al. (39) reported 120 cases of scorpion envenomation in Batman province.
Moreover, parasympathetic effects were more common than sympathetic effects and no deaths were reported.This species plays a major role in 50.8% of scorpionism cases in Sanliurfa province in Turkey (35).On the other hand, Adiguzel et al. (5) stated that child patients presented signs of local and autonomic nervous system failures, but no lethality.

Envenomation by Mesobuthus sp.
In Turkey, stings by Mesobuthus species are more frequent than those by other scorpion species since one of their most common habitats is from the west to the east of Anatolia (46,42).Mesobuthus gibbosus is generally seen from the Aegean region of Central Anatolia to the middle parts of East Anatolia (6,8,9).

Figure 2 .
Figure 2. Overall annual incidence is about 36 scorpion stings per 100,000 inhabitants and the estimated annual mortality is 0.01 death per 100,000 inhabitants (42).X: Location of previously designed studies or case reports.
Soker and Haspolat (32) reported 64 cases of scorpion sting in children admitted to the Pediatric Emergency Department in Dicle University Hospital at Diyarbakir from the provinces of Mardin, Sirnak, Batman and Siirt in the southeastern part of Turkey from 1995 to 1999.Those authors also recorded eight lethal cases (12.5%) which Adiguzel S. In vivo and in vitro effects of scorpion venoms in Turkey: a mini-review.J Venom Anim Toxins incl Trop Dis.2010;16(2):205 were due to cardiac and respiratory failure during the first 24 hours of hospitalization.

Suzek
et al. (47) stated that 262 patients stung by scorpions (M.gibbosus) required the Emergency Service of the State Hospital in Mugla province between 1999 and 2001.Those authors also reported that two cases (1.17%) resulted in death in 1999.Akcay et al. (48) investigated envenomation cases in Denizli including the previous five years and reported that 1.5% of these cases were scorpionisms.Altinkaynak et al. (33) investigated 24 children stung by scorpions in Marmaris, district of Mugla province, and stated that the patients had clinical symptoms such as abdominal pain,