J Arthropod-Borne Dis, March 2019, 13(1): 104–115 O Ozkan and ME Alcigir: A Comparative … Original Article A Comparative Pathomorphological Findings Between Leiurus abdullahbayrami and Androctonus crassicauda (Scorpion: Buthidae) Envenomation in Rabbit Animal Model *Ozcan Ozkan1; Mehmet Eray Alcigir2 1Çankırı Karatekin University, Çankırı, Turkey 2Department of Pathology, Kirikkale University, Faculty of Veterinary Medicine, Kirikkale, Turkey (Received 15 May 2017; accepted 23 Dec 2018) Abstract Background: The aim of study was to compare macroscopical and histopathological findings between venoms be- longing to two scorpion species, Androdoctonus crassicauda, and the newly discovered Leirus abdullahbayrami. Methods: The animals used in this experimental study were fifteen New Zealand bred rabbits. Three groups were constituted as group I (L. abdullahbayrami group, n= 6), group II (A. crassicauda group, n= 6) and group III (control group, n= 3). The animals in the L. abdullahbayrami group and the A. crassicauda group were envenomed through an intravenous route. The rabbits were monitored for the first 24h following the envenomation. The animals dead within that time period were examined and all animals were sacrificed and standard necropsy process was performed at 24h. Results: The pathomorphological findings from group I were found to be more severe than those observed in group II. The venom from the newly identified L. abdullahbayrami has a greater effect than the venom from the A. crassicauda. Moreover, as this was a rabbit modeling study, the L. abdullahbayrami might pose the most serious health threat to infants in particular due to their smaller body weight. Conclusion: These findings will provide a better understanding of envenomation of human beings in terms of the possible consequences of scorpion toxication on the organs. Keywords: Leirus abdullahbayrami; Androctonus crassicauda; Venom; Pathomorphology; Rabbit Introduction In many parts of the world, millions of Africa, India, and the Middle Eastern countries people are stung by various scorpion species because those areas have scorpion species with yearly. These stings can result in death, par- the most potent venoms as they are classified as ticularly in children (1, 2). This is because of neurotoxic, hemotoxic, cardiotoxic, nephrotox- the serious health problems that result, such as ic, and myotoxic (3-5). cardiovascular, respiratory, and/or neurologic In Turkey, the most hazardous scorpions complications. Nowadays, despite advances in are the yellow scorpion, Leiurus abdul- medical science, scorpion envenomation cas- lahbayrami, which is endemic in Southeast- es still continue to be a current public health ern Anatolia, and a black scorpion, Androcto- problem all over the world, including in Tur- nus crassicauda, commonly found in South- key. Scorpions are considered to be life-threat- eastern Anatolia and in a part of Eastern Ana- ening venomous animals. Arguably, the med- tolia (Fig. 1) and neighbouring countries ical knowledge of the scorpion's species is most Iran, Iraq and Syria (2, 5-7). critical for the scorpion species that are typi- Furthermore, in these regions, the species cally found in Mexico, the middle and northern are responsible for the most deadly cases, par- regions of South America, North and South ticularly those involving children. According- 104 *Corresponding author: Dr Ozcan Ozkan, E-mail: [email protected] http://jad.tums.ac.ir Published Online: April 27, 2019 J Arthropod-Borne Dis, March 2019, 13(1): 104–115 O Ozkan and ME Alcigir: A Comparative … ly, these scorpion species are described as med- following a venom injection (20-22). In enven- ically significant to both the world and to Tur- omations from different scorpion species, the key (1, 3, 5, 7). lungs, heart, liver, intestines, and pancreas were In the toxicokinetic studies, radiolabeled documented as the most affected organs in ad- venom from the L. quinquestriatus and venom dition to the recently described effects on the from the A. crassicauda scorpion have reached brain following a Leiurus envenomation (23- the maximum value of total plasma radioac- 27). tivity in less than two minutes after a subcu- As a result of the effect of scorpion venom, taneous injection and begin distribution with- different clinical tables have reported varying in four to seven minutes after the injection with effects from localized signs to more serious au- a half-life of between 4–21h. At that time, the tonomic and neurologic findings as well as fatal degree of the clinical symptoms depends on cases due to multisystem organ failure. Ac- the amount of venom circulating in the body cording to medical records from Turkey, most (7-9). deaths were caused by the results of cardiopul- A low venom dose triggers an adrenergic monary complications, such as myocarditis and effect while high scorpion venom concentra- acute pulmonary edema following a scorpion’s tions result in venom cholinergic symptoms. sting (1, 2, 7). The venom results in an excessive acetylcho- In animal models, scorpion envenomation line (Ach) release or a decrease in the destruc- symptoms were similar to those described by tion of Ach, which is acting on the postgan- the clinical findings in humans. Therefore, to glionic nerve endings. Thus, cholinergic activ- mimic human victims, especially children, rab- ity begins. On the other side, catecholamine is bits were chosen as the animal model for ex- released into the peripheral sympathetic nerve perimental scorpion envenomation to mimic endings from the adrenal medulla in response a natural sting accident. Thus, in this study, to the venom. Then, an autonomic storm begins different pathomorphological findings devel- because of the renin secretion by the alpha ad- oped by two scorpion venoms were revealed renergic receptor. The receptor stimulation out L. abdullahbayrami and A. crassicauda, plays a significant role because it increases which species were most encountered in south- blood pressure and ultimately results in the east region of Turkey and also were showed pathogenesis of pulmonary edema. Addition- L. abdullahbayrami having unusual harmful ally, it causes an inflammatory reaction in the effects onto several organs during long term. vital organs (8, 10-13). Scorpion envenomation has been reported to mainly culminate in a syndrome of fuel-en- Materials and Methods ergy deficiency. This situation develops as a deficiency during the usage of existing meta- Scorpions Origin bolic substrates. As an aftermath, a failure oc- Scorpions were collected using an ultra- curs within vital organs including a multi-or- violet lamp at night in the Sanliurfa and Ga- gan system deficiency that can lead to death ziantep Provinces, southeastern part of Tur- (14, 15). The main reason for this failure is the key. The animals were kept in plastic boxes triggering of an inflammatory response cascade at the Department of Entomology, Faculty of that results in a release of several mediators, Veterinary Medicine, Ankara. The scorpions such as prostaglandins, cytokines, and nitric ox- were received fresh water daily and fed crick- ide, and an infiltration of inflammatory cells ets or cockroaches weekly. (16-19). The highest venom concentrations can be found in the kidneys, liver, heart, and lungs 105 http://jad.tums.ac.ir Published Online: April 27, 2019 J Arthropod-Borne Dis, March 2019, 13(1): 104–115 O Ozkan and ME Alcigir: A Comparative … Scorpion venom and the median lethal dium Pentobarbital injection (100mg/kg Intra- dose (LD50) venous route) were done and then cervical dis- The venom of scorpions was milked by location were performed for each one. Necrop- electrical stimulation of 24 Volt. The venoms sy process was done systematically and all or- were added sterile double-distilled water, dis- gans and tissues were examined routinely. Le- solved and centrifuged at 14000rpm for 15 sions were photographed. For histopathologi- min at 4 °C. Supernatant was dissolved in PSS. cal examination, tissue samples were taken sys- The median lethal doses (LD50) of the scor- tematically from each organ in 10% buffered pions were determined as in previous studies formalin. After fixation for 48h, tissue samples (28, 29). were processed routinely through alcohol and xylol series and mounted in paraffin. Processed Animals tissues were embedded in paraffin wax. Five- This study was conducted on 15 of healthy micron thickness-sections were cut from par- New Zealand rabbits 12 months old of both affin blocks and Haematoxylin-Eosin (H and sexes and between 2.4±0.1kg body weight for E) staining method. this experiment. The rabbits were fed with a special rabbit pellet diet ad libitum until en- Results venomation. The animals were kept in room temperature set to 19±1 °C and with 12h light/ Macroscopical Findings 12h dark schedule. The experimental protocol In all groups, cadavers were well-fed, and was approved by the local Laboratory Animal rigor mortis did not happen. However, the Ethics Committee. After 24h all animals were blood in group I was lately clotted in contrast subjected to euthanasia for macroscopical and to the other groups. Conjunctiva was hyper- histopathological findings. emic in two of the animals from group I. The gut walls were thickened and filled with a yel- Experimental envenomation of the animals lowish content in all of the members of group The animals were envenomed by the ven- I and II. The liver, kidneys, and lungs were con- om of L. abdullahbayrami in group I (GI, n: gested in all members of both groups I and II. 6), and the venom of A. crassicauda in group However, there was haemorrhagia that hap- II (GII, n: 6). Finally, three rabbits were se- pened in the kidneys in the members of group lected as a control group and will be referred II, and also atrophia and capsular contraction to as GIII. The venoms (2xLD50 for both) were in the spleen was noticed in all animals within injected into the marginal ear vein by intrave- group II. In group I, a meningeal vessel was nous (IV) route after dissolving the venoms congested in one animal.