Male of Centruroides sculpturatus. Photo: G. Giribet.

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1 Prashant P. Sharma Scorpion maintenance and dissection protocol Maintenance of scorpions for developmental work: General care and dissection of gravid females INTRODUCTION The Arizona bark scorpion (Centruroides sculpturatus) is a highly venomous North American buthid scorpion. Gravid females are aggressive and will attempt to sting repeatedly when offended. Working with this species requires serious precautions and complete concentration at all times. Male of Centruroides sculpturatus. Photo: G. Giribet. Page 1 of 5

2 General basic handling and maintenance 1. NEVER insert bare hands into scorpion chamber 2. NEVER directly handle a scorpion 3. ALWAYS use 12 forceps 4. ALWAYS wear work gloves 5. ALWAYS wear closed- toed shoes 6. ALWAYS keep tetanus shots and boosters current 7. ALWAYS keep scorpion chamber locked unless it is current use 8. ALWAYS keep warning signs prominently displayed in and around scorpion chamber In the event of escapees: 1. Isolate with adhesive tape 2. Immobilize with acetone 3. Destroy escapees 4. If an animal reaches an inaccessible hiding place, inform senior lab personnel Symptoms of scorpion stings: Mild signs and symptoms include: 1. Pain, which can be intense 2. Numbness and tingling in the area around the sting 3. Slight swelling in the area around the sting More severe signs and symptoms include: 1. Muscle twitching or thrashing 2. Unusual head, neck and eye movements 3. Drooling 4. Sweating 5. Vomiting 6. High blood pressure (hypertension) or low blood pressure (hypotension) 7. Accelerated heart rate (tachycardia) or irregular heart beat (arrhythmia) 8. Restlessness or excitability or inconsolable crying (in children) In the event of a scorpion sting: 1. INFORM other lab personnel IMMEDIATELY. 2. WASH area with soap and water 3. APPLY a cool compress on sting area 4. ELEVATE stung area to heart level 5. CALL Poison Control and Prevention Services for MA and RI: FOLLOW assessment and directions of Poison Control Page 2 of 5

3 Sexing Centruroides sculpturatus Males of C. sculpturatus have narrower bodies and longer, thinner tail segments. Females of C. sculpturatus have wider bodies and thicker, shorter tail segments. Other sexual dimorphisms in this species include the pectines on the underside of the body and the shape of the sternum. Male of C. sculpturatus. Photo: bugguide.net. Female of C. sculpturatus. Photo: bugguide.net. Page 3 of 5

4 General care of Centruroides sculpturatus 1. This species is communal. Keep up to 50 adults in a single 15- gallon container, lined with coconut fiber and wood chips. Add egg carton cardboard and/or pieces of bark as resting places. 2. Keep on a 16 h : 8 h day/night cycle or use natural light cycle. Scorpions are nocturnal and very light sensitive. All lights must be extinguished during night cycle. 3. Add large, high- walled container of water to one corner of the container and keep aperture of container small in order to maintain humidity, especially for molting animals. Sprinkle container with distilled water periodically. 4. Keep smaller subadults and independent scorplings (early instars) in separate containers to avoid cannibalism. 5. Feed scorpions once per week by adding live crickets (1.2 x the number of scorpions in the container) just before black out. Crickets should be approximately the size of the scorpion body, not including the tail. Adult Acheta domesticus or penultimate molts of Gryllus bimaculatus will suffice for adult C. sculpturatus. 6. Remove partially eaten carcasses of crickets (if any) hours after feeding. 7. If promenade à deux (mating dance) is observed, avoid disturbing the pair. Page 4 of 5

5 Dissection of gravid scorpions 1. Transfer one scorpion to Ptep cylinder with 12 forceps. Cover with Ptep cylinder foam. 2. Anesthetize: flush cylinder with CO2 for 15 s; wait 5 min. 3. With 6 forceps and scissors, FIRST pick up with the tail and separate last five segments from body. QUICKLY separate head from body next. 4. Pin to Silgard plate. Flush with 1x PBS. 5. Remove prosomal appendages. 6. With scalpel or fine scissors, slice pleural membranes on both sides of body. Remove tergites. 7. Separate caeca from ovariuterus. 8. Working carefully, remove embryos from ovariuterus. 9. Working carefully, remove external membranes from embryos. 10. Make longitudinal cut along dorsal midline. Open and separate as much yolk as possible from embryo. 11. Flush embryo with fresh PBS and proceed to fixation. Page 5 of 5