Journal of Zoo and Wildlife Medicine 38(2): 245–251, 2007 Copyright 2007 by American Association of Zoo Veterinarians HEMOLYMPH BIOCHEMISTRY REFERENCE RANGES FOR WILD- CAUGHT GOLIATH BIRDEATER SPIDERS (THERAPHOSA BLONDI) AND CHILEAN ROSE SPIDERS (GRAMMOSTOLA ROSEA) Trevor T. Zachariah, D.V.M., Mark A. Mitchell, D.V.M., Ph.D., Clare M. Guichard, and Rimme S. Singh, B.A. Abstract: Theraphosid spiders have become increasingly popular for private and public uses in the United States. However, little is known about their physiology from a medical standpoint. This study represents the first attempt to establish reference hemolymph values for two common species of theraphosids, the goliath birdeater spider (Theraphosa blondi) and the Chilean rose spider (Grammostola rosea). Eleven T. blondi and twelve G. rosea, all wild-caught subadults, were obtained after importation and hemolymph was collected for biochemical analysis. After 8 wk of captivity, hemolymph was again collected from the spiders and analyzed. The biochemical analytes measured in the study included aspartate transferase, creatine kinase, glucose, total protein, albumin, uric acid, blood urea nitrogen, phosphorous, calcium, potassium, and sodium. The osmolality of the hemolymph was estimated for each spider using two different formulae. There were significant differences in body weight, sodium, potassium, and osmolality between the sampling times for both species. There were also significant differences in creatine kinase, calcium, total protein, and blood urea nitrogen between sampling periods for T. blondi. The results of this study suggest that serial hemolymph samples may be used to assess the hydration status of theraphosid spiders. In addition, the differences in hemolymph analytes between spiders suggest that there may be differences between species that should be addressed in future studies. Key words: Biochemistry, Grammostola rosea, hemolymph, spider, Theraphosidae, Theraphosa blondi. INTRODUCTION been made for spiders of any taxonomic group.1,2,5,7,12,13 These previous attempts focused on Invertebrates comprise the majority of the analyzing different organic and inorganic hemo- world’s fauna, and are popular as research, display, lymph constituents using relatively slow, technical- and pet animals. In the pet trade, spiders represent ly involved procedures with equipment that would a significant portion of the invertebrate specimens not be found in the average veterinary clinic or available for sale, and most of these are from the zoological hospital. The primary objective of this family Theraphosidae. It has been estimated that research was to establish reference ranges for the more than 100,000 theraphosid spiders are import- biochemical components of goliath birdeater spider ed into the United States each year, and approxi- (Theraphosa blondi) and Chilean rose spider mately the same number are bred in captivity (Grammostola rosea) hemolymph using a standard (Breene, pers. comm.). Currently, there are 22 spe- analytical machine that could be commonly found cies of theraphosid spiders listed with the Conven- in general veterinary practice. The secondary ob- tion on International Trade in Endangered Species jective of this study was to compare paired hemo- 15 of Wild Fauna and Flora (CITES) Appendix II. lymph samples from spiders post-import and after The listing of these invertebrates under CITES pro- 8 wk of captivity to assess the effect of captivity tection suggests that there are natural and/or un- on biochemical parameters. natural influences that can affect the long-term sur- The specific biological hypotheses tested in this vival of these spiders. To protect the future of these study were to determine if significant differences theraphosids, research to increase our understand- existed in body weight, sodium (Na), and total pro- ing of their physiologic functions is required. tein (TP) between the paired hemolymph samples Reference values for blood biochemical and gas and if there were differences in the biochemical an- constituents have long been established for domes- alytes between male and female T. blondi. tic species, but only a few limited attempts have MATERIALS AND METHODS From the Department of Veterinary Clinical Sciences, Theraphosa blondi School of Veterinary Medicine, Louisiana State Univer- sity, Skip Bertman Drive, Baton Rouge, Louisiana 70803, Eleven (six male and five female) wild-caught, USA. Correspondence should be directed to Dr. Zachariah subadult T. blondi were obtained from an inverte- ([email protected]). brate importer in Florida (LASCO, Naples, Florida 245 246 JOURNAL OF ZOO AND WILDLIFE MEDICINE 34119, USA). The spiders were housed individually Formula 1.17 (NA ϩ K) ϫ 2 ϩ BUN ϩ GLU in rectangular, 5.7-L plastic storage containers with ϭ milliosmoles/L lids. A 50:50 mixture of potting soil and vermicu- lite was used for the substrate. The spiders were Formula 2.3 1.86 ϫ (NA ϩ K) ϩ (GLU/18) provided a 12-hr light cycle; half of a plastic flower ϩ (BUN/2.8) ϩ 9 pot served as a hiding space. The temperature and humidity in the enclosures were maintained at ap- ϭ milliosmoles/L Њ Њ proximately 23.9 C (75 F) and 80%, respectively. The spiders were held under the conditions de- Each spider was placed into a square, 3-L plastic scribed earlier for an additional 8 wk. The spiders storage container that was modified into a gas an- had constant access to chlorinated tap water and esthetic chamber. The container was modified by were fed five adult crickets weekly. The crickets drilling a hole in one side and inserting an endotra- had access to a high-Ca cricket food and water cheal tube adapter. Each spider was anesthetized source (High Calcium Cricket Diet and Cricket with 5% isoflurane10 (Isoflo, Abbott Laboratories, Quencher Calcium, Fluker’s Farms, Port Allen, North Chicago, Illinois 60064, USA) at a flow rate Louisiana 70767, USA) until being offered to the of 1 L/min oxygen. Once the spider had lost its spiders. A second hemolymph sample was collect- ability to right itself, it was removed from the an- ed 8 wk after the initial sample. Sample collection, esthetic chamber and weighed. A 26-ga, ¾-inch handling, and analysis were repeated as described, needle fastened to a 3-ml syringe was used to col- with the exception that immediately after sample lect an intracardiac hemolymph sample.16 A volume collection, 0.15 ml of the hemolymph sample was of 0.5 ml was collected from each individual by analyzed using a CG8ϩ blood gas cartridge with inserting the needle at approximately a 45Њ angle the i-STAT analyzer (Abbott Laboratories, North through the exoskeleton at the midpoint of the dor- Chicago, Illinois 60064, USA). Results were col- sal midline of the opisthosoma. This method was lected for the following additional hemolymph val- used to facilitate the rapid collection of a relatively ues: total carbon dioxide, ionized Ca, pH, partial pressure of carbon dioxide, partial pressure of ox- large amount of hemolymph. Hemolymphstasis was ygen, bicarbonate, and oxygen saturation. accomplished by applying a small amount of Nex- aband glue (Veterinary Products Laboratories, Grammostola rosea Phoenix, Arizona 85067, USA) to the collection site. The spiders were recovered in 100% oxygen, Twelve wild-caught G. rosea were obtained from and recovery from anesthesia was uneventful in all a vendor at a reptile show in Mandeville, Louisiana. spiders. The spiders used for this study were all subadults Each sample was placed in a lithium heparin Mi- of unknown gender. The spiders were housed in the crotainer tube (Becton Dickinson, Franklin Lakes, same type of containers described for T. blondi. Bed-A-Beast ground coconut hull was used as the New Jersey 07417, USA) and centrifuged for 10 substrate (Pet-Tech Products LLC, Van Nuys, Cal- min at 1,411 g. After centrifugation, the supernatant ifornia 91406, USA). The temperature and humid- was removed and analyzed using an avian/reptile ity in the enclosures were maintained at approxi- rotor for the VetScan Analyzer (Abaxis Inc., Union mately 23.9ЊC (75ЊF) and 70%, respectively. The City, California 94587, USA). Results were col- spiders were provided a 12-hr light cycle; half of a lected for the following biochemical analytes: as- plastic flower pot was provided as a hiding space. partate transferase (AST), creatine kinase (CK), The spiders were anesthetized using the same glucose (GLU), TP, albumin, uric acid (UA), blood technique described for T. blondi. Hemolymph urea nitrogen (BUN), phosphorous, calcium (Ca), samples were collected using a 30-U insulin syringe potassium (K), and Na. All samples were analyzed fitted with a 30-ga needle. A total of 0.15 ml he- within 30 min of sample collection. Na levels for molymph was collected from each individual. Be- the VetScan Analyzer have a maximum measurable cause of the spiders’ small size, a bolus of 0.5 ml limit of 180 mmol/L. Because Na levels for these 0.9% saline (Hospira Inc., Lake Forest, Illinois spiders were often greater than that level, an aliquot 60045, USA) was administered into the opisthoso- of each sample was tested using the Olympus ma after the hemolymph was collected. Recovery AU600 Clinical Chemistry Analyzer (Olympus from anesthesia was uneventful for all spiders. America Inc., Melville, New York 11747, USA). After the initial hemolymph collection, the spi- An estimated osmolality was calculated for each ders were provided constant access to water and spider using the following formulae: were fed three adult crickets weekly for 8 wk. The