Serum Thyroid Hormone Values for African Spurred Tortoises (Centrochelys [Formerly Geochelone\ sulcata) Keri H. Franco1, DVM, Daniel J. Famini2, DVM, John P. Hoover1, DVM, DACVIM, DABVP, Mark E. Payton3, PhD

1. Center for Veterinary Health Sciences, Oklahoma State University, Stillwater, OK 74078, USA

2. Humane Society of Sonoma County, 5345 Highway 12 West, Santa Rosa, CA 95407, USA

3. Department of Statistics, College of Arts and Sciences, Oklahoma State University, Stillwater, OK 74078, USA Downloaded from http://meridian.allenpress.com/jhms/article-pdf/19/2/47/2209073/1529-9651_19_2_47.pdf by guest on 03 October 2021

Abstract: Hypothyroidism has been reported in Galapagos (Geochelone nigra [formerly G. elephantopus]) and Aldabra (G. gigantea) tortoises, and reference values for thyroxine (T4) are available for these two spe­ cies. Hypothyroidism also has been presumptively diagnosed in an African spurred tortoise (Centrochelys [formerly G.] sulcata) based on clinical disease and response to therapy, but no reference values were avail­ able to confirm the disease. Twelve clinically normal African spurred tortoises from a private collection were used to obtain blood for serum thyroid hormone analysis. The median values for the different thyroid hormones were as follows: T4, 4.0nmol/L (0.31 mcg/dl); free thyroxine (fT4), 4.0pmol/L (0.31 ng/dl); triiodothyronine (T3), 0.15 nmol/L (0.01 mcg/dl); and free triiodothyronine (fT3), 2.9 pmol/L (0.189 ng/dl). The descriptive measures for T4, fT4, T3, and fT3 from this study are proposed by us as initial thyroid hormone reference values for the African spurred tortoise. The serum T4 values of African spurred tortoises may be lower than serum T4 values reported for Galapagos and Aldabra tortoises.

Key W ords: Centrochelys, Geochelone, sulcata, thyroid, thyroxine (T4), triiodothyronine (T3).

Introduction digestive secretions, increased glucose metabolism, and increased rate of secretion of other hormones (Guyton, Hypothyroidism has been reported previously in the 1991). In tortoises and other reptiles, thyroid hormones may Galapagos tortoise (Geochelone nigra [formerly G. elephan­ be involved in regulation of ecdysis, growth, and develop­ topus]) and Aldabra tortoise (G. gigantea) (Frye and Dutra, ment, as well as in cycles of hibernation and breeding (Kohel 1974; Norton et al., 1989; DiGesualdo et al., 2004; et al., 2001). Although African spurred tortoises generally Donoghue, 2006), and is suspected to occur in African do not hibernate in captivity, it is unknown whether sea­ spurred tortoises (Centrochelys [formerly G.] sulcata) sonal changes play a role in their thyroid hormone levels. (Franco and Hoover, 2009). Although reference values for The objective of this study was to determine reference thyroid hormones are available for Galapagos and Aldabra values for serum total T4, total T3, fT4, and free triiodothy­ tortoises (Teare, 2002; DiGuesualdo et al., 2004), reference ronine (fT3) of captive African spurred tortoises. values have not been reported for African spurred tortoises, a commonly kept pet species. M ater ials a n d M e t h o d s In humans and other mammals, the most active thyroid hormone that binds to intracellular receptors is in the form Twelve (four male and eight female) privately owned and of triiodothyronine (T3), whereas most of the circulating maintained African spurred tortoises ranging in size from hormone is in the form of thyroxine (T4). Most circulating 1.9 to 28.6 kg (4.18-62.92 lbs) and estimated age from 3 to T4 and T3 are protein-bound, allowing for a slow release 17 yr were used for this study. Blood samples were obtained into the tissues (Guyton, 1991). In one study, desert tor­ in the spring and early summer by peripheral venipuncture toises (Gopherus agassizii) were found to lack a high-affinity of the jugular or subcarapacial vein. All tortoises were clini­ T4-binding protein, resulting in low total T4 levels and rela­ cally normal on physical examination. Sedation (medetomi- tively higher free thyroxine (fT4) levels compared with dine hydrochloride [Domitor, 1 mg/ml; Orion Corporation, mammals (Kohel et al., 2001). It has been suggested that T4 Espoo, Finland], 0.1 mg/kg intramuscularly (IM) and may be the more active hormone in reptiles (Kohel et al., ketamine hydrochloride [Ketaject, 100 mg/ml; Bioniche 2001). Teoranta, Inverin Co., Galway, Ireland], 10 mg/kg IM) was Thyroid hormones increase gene transcription in all cells required for some of the tortoises (all the males and four in the body, resulting in an increased basal metabolic rate, large females) to facilitate sample collection. Blood samples increased protein synthesis and catabolism, increased were placed in individually marked clot tubes (Vacutainer oxygen consumption, and increased mitochondrial activity. Serum Separator Tube; BD Biosciences, Franklin Lakes, These actions at the cellular level result in an increased New Jersey), labeled, and placed in a chilled cooler with cardiac output, increased blood pressure, increased frozen ice packs to maintain samples at approximately 4°C

Volume 19, No. 2,2009 Journal of Herpetological Medicine and Surgery 47 Table 1. The median, 10th and 90th percentiles, and minimum these values may be much higher than for African spurred (Min) and maximum (max) values for serum thyroxine (T4), tortoises (median, 2.9 pmol/L) for all ages and both sexes free thyroxine (fT4), triiodothyronine (T3), and free triiodothy­ combined. The fact that the International Species Informa­ ronine (fT3) from African spurred tortoises. tion System value reports fT3 in nmol/L rather than the more standard picomoles per liter suggests either that Gala­ Sample size Median 10-90% Min-max values pagos tortoises have a much larger amount of circulating fT3 than African spurred tortoises or that the units reported T4 (nmol/L) 12 4.0 2.0-9.0 1.0-9.0 in that study, based on only two individuals, may be incor­ rect. Statistical comparisons were not made for the possible fT4 (pmol/L) 11 4.0 3.0-6.0 3.0-12.0 differences in serum thyroid hormones values between the T3 (nmol/L) 12 0.15 0.0-0.8 0.0-1.0 three tortoise species because of the small sample sizes used in the three studies. The small sample sizes for Galapagos fT3 (pmol/L) 11 2.9 1.5-4.8 1.2-5.5 and Aldabra tortoises, as well as the limited number of samples in the study of this report, may actually magnify the apparent differences in the respective thyroid hormone

(40°F) for transportation back to the clinic (<3 h). Clotted values. Additional samples will be necessary before mean­ Downloaded from http://meridian.allenpress.com/jhms/article-pdf/19/2/47/2209073/1529-9651_19_2_47.pdf by guest on 03 October 2021 blood samples were centrifuged at 3,200 rpm for 15min, ingful statistical comparisons can be made between these and the sera were pipetted into individually marked tubes three species of large land tortoises. It also remains to be for each tortoise. Samples were frozen and maintained at - determined whether there are differences in serum hormone 18°C (0°F) until shipped on wet ice to the laboratory for values between sexes, age groups, or sampling seasons in the hormone analyses (Diagnostic Center for Population and serum thyroid hormones of African spurred tortoises. Anim al Health, Endocrine Diagnostic Section, Michigan Although the actual incidence of hypothyroidism in large State University, Lansing, Michigan). All analyses were land tortoises is unknown, there are published anecdotal done using radioimmunoassay (RIA). Commercial RIAs reports of hypothyroidism in Geochelone species (Frye and (DiaSorin Inc., Stillwater, Minnesota) were used to measure Dutra, 1974; Guyton, 1991; DiGesualdo etal., 2004). A case total T4, fT4, and fT3 assays. Total T3 was measured using of presumed hypothyroidism in an African spurred tortoise an in-house RIA at Michigan State University. One tortoise with myxedema, and what seemed to be a low serum T4 blood sample was of insufficient quantity to run a complete value, responded clinically to oral levothyroxine therapy hormone panel and provided data for total T4 and T3 (Franco and Hoover, 2009). At the time of the case report, only. a reference range for thyroid hormones of African spurred Due to the small sample size, all data (both sexes and all tortoises was not available for comparison, and treatment sizes) were combined. Study data were analyzed with PC was initiated based on reported T4 values of Galapagos tor­ SAS version 9.1 (SAS Institute, Cary, North Carolina) using toises (Norton, 1989). This study seems to provide the initial the PROC UNIVARIATE procedure. The data were found data necessary to establish reference ranges for thyroid to not have a normal distribution, so results are reported as hormones in the African spurred tortoise. The clinical sig­ median, 10th and 90th percentiles, and minimum-maximum nificance of and reason for the apparently much lower serum values. fT3 in the African spurred tortoises than in Galapagos tortoises is not known. Additional studies using larger R e su l t s sample sizes and perhaps animals from different climates and seasons are warranted to substantiate the T4, fT4, T3 Table 1 presents the median, 10th and 90th percentiles, and and fT3 initial reference values for African spurred tortoises minimum-maximum values for total T4 (« = 12), total T3 reported here (Table 1), and to determine if any differences (« = 12), fT4 (n = 11), and fT3 («= 11) from African spurred in serum thyroid hormones exist for tortoises based on tortoises of all ages (sizes) and both sexes combined. different age, sex, or environmental/season variables. We propose that the median, 10th and 90th percentiles, D is c u s s io n and minimum-maximum values for T4, fT4, T3, and fT3 obtained from the tortoises in this study be used as baseline The tortoises of this study had median total serum T4 values thyroid hormone references ranges for African spurred (n = 12; median = 4.0 nmol/L) that seemed to be lower than tortoises. Serum T4 values for African spurred tortoises the serum T4 values reported for Galapagos (n = 5; mean do seem to be lower than those reported for Galapagos or + SD: 10.39 +18.72 nmol/L) and Aldabra tortoises (n = 5; Aldabra tortoises. mean + SD: 9.0 ±3.0 nmol/L) for both sexes and all ages combined; however, it is important to note that the distribu­ Acknowledgments: This project was funded through the tions of the data were not stated in the references (Teare, Department of Veterinary Clinical Sciences, Center for 2002; DiGesualdo et al., 2004). The large standard devia­ Veterinary Health Science, Oklahoma State University, tion noted in the Galapagos tortoise reference suggests that the data did not follow a Gaussian distribution and that the Stillwater, Oklahoma 74078, USA. We acknowledge Susan mean is skewed to the right; in that case, the median would Beyerlein at Diagnostic Center for Population and Animal serve as the better estimate of central tendency and is likely Health, Endocrine Diagnostic .Section, Michigan State closer in value to the median of the African spurred tor­ University, PO Box 30076, Lansing, Michigan 48909-7576, toises. Other than T4 values, only fT3 have been reported USA, for performing the hormone analyses, and Al Wolf of for Galapagos tortoises (« = 2; mean + SD: 393 + 75 nmol/L Sonoma County Reptile Rescue for the contribution of the for both sexes and all ages combined) (Teare, 2002), and tortoise blood samples used in this study.

Volume 19, No. 2,2009 48 Journal of Herpetological Medicine and Surgery L itera tur e C ited Frye FL, Dutra FR. 1974. Hypothyroidism in turtles and tortoises. Vet Med Small Animal Clin, 69(8):990-993. DiGesualdo CL, West G, Brown TR, Hoover JP. 2004. Gauvin J. 1993. Drug therapy in reptiles. Sem Avian Exotic Pet Determining normal thyroid hormone status in Galapagos Med, 2(1):48—59. tortoises, then comparing normal levels to thyroid levels Guyton A. 1991. The thyroid metabolic hormones. In Wonsie- of Galapagos tortoises (Geochelone elephantopus) suspected wicz MJ (ed.): Textbook of Medical Physiology, 8th ed. WB of hypothyroidism. Proc AAZV, AAWV, WDA Joint Saunders Co., Philadelphia, PA:831-839. Conference:550-551. Kohel KA, MacKenzie DS, Rostal DC, Grumbles JS, Lance Donoghue S. 2006. Nutrition. In Mader DR (ed): Reptile VA. 2001. Seasonality in plasma thyroxine in the desert Medicine and Surgery. 2nd ed. Saunders Elsevier, St. Louis, tortoise, Gopherus agassizii. Gen Comp Endocrinol, 121: MO:251-298. 214-222. Fleming GJ, Heard DJ, Uhl EW, Johnson CM. 2004. Thymic Norton TM, Jacobson ER, Caligiuri R, Kollias GV. 1989. hyperplasia in subadult Galapagos tortoises, Geochelone Medical management of a Galapagos tortoise (Geochelone nigra. J Herp Med Surg, 14(l):24-27. elephantopus) with hypothyroidism. J Zoo Wildl Med, Franco KH, Hoover JP. 2009. Levothyroxine as a treatment 20(2):212—216. for presumed hypothyroidism in an adult male African Teare JA. 2002. Reference ranges for physiological values of Downloaded from http://meridian.allenpress.com/jhms/article-pdf/19/2/47/2209073/1529-9651_19_2_47.pdf by guest on 03 October 2021 spurred tortoise (Centrochelys [formerly Geochelone] captive wildlife. International Species Inventory System, sulcata). J Herp Med Surg, 19(2):42-44. Apple Valley, MN.

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