Z. Jagdwiss. 47 (2001), 241–252 Submitted: 25 June 2001 © 2001, Blackwell Wissenschafts-Verlag, Berlin Accepted: 24 August 2001 ISSN 0044-2887 I. ABHANDLUNGEN University of Guelph Cryptorchism, hypogonadism and antler malformations in black-tailed deer (Odocoileus hemionus sitkensis) of Kodiak island1 By G. A. BUBENIK, Guelph, J. P. JACOBSON, Kodiak, D. SCHAMS, Freising, and L. BARTOSˇ, Prague 1 Introduction Since the early nineteen nineties, an ever-increasing number of antler abnormalities were observed among males of Sitka black-tailed deer (Odocoileus hemionus sitkensis) of Kodiak Island, (Alaska, USA). These antler malformations included incomplete growth of points, serious distortion of antler form (asymmetric antlers), incomplete polishing of velvet, and extremely sharp antler points. Furthermore, some incompletely mineralized antlers were cast above the coronet exhibiting conspicuously concave seals (Fig. 2). Such casting and other malformations such as diagonal sequestration of antlers or „cauliflower-like“ antlers (Fig. 3), was associated with an excessive growth of the antler base which partly resembles the „helmet-perukes”. These wig-like structures are formed in castrates or hypogonadic deer by the proliferation of antler bone and velvet tissues without the regular mineraliza- Fig. 1. Left side of a Sitka black-tailed deer mandible showing severe periodontal disease as indicated by deep periodontal pockets (arrow). Photos 1–9: J. JACOBSON 1 Eingesetzt wurde ein Druckkostenzuschuss des Hessischen Ministeriums für Umwelt, Landwirt- schaft und Forsten, für dessen Gewährung verbindlich gedankt wird. – Die Schriftleitung U.S. Copyright Clearance Center Code Statement: 0044–2887/2001/4704–0241 $15.00/0 242 G. A. Bubenik, J. P. Jacobson, D. Schams, L. Bartoˇs Fig. 2. Head and antlers (which dropped off when the deer was shot) of a bilateral cryptorchid. Note the severely concave seal (arrow) and remnants of old antlers. The pre- sence of coronets and the remaining antler tissue can result in the for- mation of “helmet-peruke” – like antlers grown in the next antler cycle (see Fig. 4) Fig. 3. Skull of the bilateral cryp- torchid deer with the “cauliflower” antlers Cryptorchism, hypogonadism and antler malformations in black-tailed deer of Kodiak island 243 tion and casting (BUBENIK, A., 1966; HARTWIG, 1981). In addition, supernumerary pedicles with incomplete antlers and a severe periodontal disease were observed. Many of these and other physiological parameters (such as the lack of swollen neck during the rut and the female type of fat reserves) were indicative of a moderate or severe hypogonadism. An ade- quate seasonal concentration of male sex hormones (androgens) is essential for the normal development of deer antlers (BUBENIK, G., 1990). The lack of sex hormones could have also contributed to the fight-induced separation of frontal bones, a feature observed in several bucks. The suspicion of hypogonadism was strengthened when the culled bucks with mal- formed antlers were more closely examined. Many of these animals were lacking one or both testes in the scrotum and therefore a unilateral or bilateral cryptorchism was suspected. In order to elucidate the causes of the suspected hypogonadism, samples of blood and testes were taken from affected and control bucks. Serum concentrations of Luteinizing Hormone (LH) and testosterone (T) were determined by radioimmunoassay and the sper- matogenic activity was investigated in histological section of the gonads. An extensive pho- tographic documentation of antler and skull malformations was prepared and catalogued. We tested the hypothesis that serum concentrations of testosterone will be lower and LH concentrations higher in bilateral cryptorchids as compared to intact deer. 2 Materials and Methods Blood and testes samples: Five to ten ml of blood was taken from the abdominal cavity or from the heart of bucks shot during the rut by a rifle. The samples were kept in a cool place until the serum separated from the coagulum. Then the serum samples were frozen at – 20 °C. Between October 23 and November 11 (the peak period of seasonal T concentra- tions in blood), blood samples were taken from nine bilateral cryptorchid bucks, two uni- lateral cryptorchid and nine control bucks. Testicular samples were taken from two intact and one unilaterally cryptorchid bucks. Hormone assays: Concentrations of LH were determined by a homologous bovine radio- immunoassay (RIA) with no cross reactivity to other pituitary hormones (SCHAMS et al., 1980). The reference preparation was bovine pituitary preparation (LH-DSA; biological activity 1.0 I.U. of NIH-bLH-S1). The sensitivity was 0.2 ng/ml; intra-assay coefficient of variation (CV) averaged 7.4%, and inter-assay CV was 9.2–13.8%. The assay was validat- ed for white-tailed deer (Odocoileus virginianus) by recovery studies. Testosterone was measured by a highly sensitive enzyme immunoassay after extraction with tertiary butylmethylether/petrolether 30/70 (V/V) (KARG et al., 1976). The sensitiv- ity was 50 pg/ml. The inter-assay CV was 7.8–13.8%. Statistic: Since our data did not show a normal distribution, we applied a non-parametric Mann-Whitney test (Wilcoxon two sample tests, SAS). We calculated exact p-values because the asymptotic assumptions could not be met with our sample size. Standard asymptotic methods involve the assumption that the test statistic follows a particular distribution when the sample size is sufficiently large. When the sample size is not large, (such is our case) asymptotic results may not be valid, with the asymptotic p-values differing from the exact p-values. Histology: Cross-section of testes (around 10 mm thick) were taken shortly after death, fixed in buffered formaldehyde and later embedded in paraffin. The 10-micron sections were stained with Hematoxylin and Eosin and then observed in the optical microscope. The progress of spermatogenesis, the size of spermatogenic tubules and the proportion of interstitium and Leydig cells to tubules were compared. 244 G. A. Bubenik, J. P. Jacobson, D. Schams, L. Bartoˇs Photographic documentation: Skulls and malformed antlers of intact and cryptorchid bucks were collected since 1994. The shape of antlers, number of points, presence of velvet in mature antlers, shape of the casting surface (the “seal”), coronet malformations, irregular casting lines, presence of periodontal disease and separation of nasal, frontal parietal skull bones were registered. Autopsies: Autopsies were performed on a number of bucks lacking external gonads, search- ing for the ectopic testes in the abdominal cavity or the inguinal canal. 3 Results Periodontal disease, characterized by deep periodontal pockets, which is rare in other cervids of prime age, was present in about 35% of all cryptorchid bucks (Fig. 1). Moderate to severe antler malformations and the impairment of the antler cycle were indicative of hypogo- Fig. 4. Antlers of a bilateral cryptorchid. Note the extremely thick antler base (arrow) and the very sharp antler tips. The antlers were still covered in velvet, when the buck was shot during the rut. The velvet was subsequently lost during the preparation of the skull and antlers Cryptorchism, hypogonadism and antler malformations in black-tailed deer of Kodiak island 245 Fig. 5. Skull and antlers of a bilateral cryptorchid buck. Except for the sharply pointed antler tips (arrow), the antlers appear normal nadism. Numerous signs of altered antlerogenesis were observed in some cryptorchid deer; these included: premature, concave (castration-like) separation of the antler beam from the pedicles (Fig. 2), (BUBENIK, A., 1990), osteofibroma-like proliferation of the suprapedicu- lar tissue, sometime called “cauliflower antlers” (Fig. 3) (BUBENIK, A., 1966), non-polished antlers with extremely sharp points and remarkably thick antler base reminding on “hel- met-peruke” antlers (Fig. 4) (BUBENIK, A., 1966; HARTWIG, 1981). In addition, some antlers of bilaterally cryptorchid deer exhibited very sharp points, but otherwise looked normal (Fig. 5). Finally, several bilateral cryptorchid bucks had completely normal antlers (Fig. 6). Two bucks with unilateral cryptorchism also had normally looking antlers (Fig. 7). Despite a repeated search for intraabdominal or inguinal testes, none were found. The reason for the failure may be their small size and an increased amount of fat found in the abdominal cavity of hypogonadic deer. Several cryptorchid deer exhibited various degrees of impairment of sutures of nasal, frontal and parietal bones. These ranged from a complete separation (Fig. 8) to a visible re- fusion after separation (Fig. 9). 246 G. A. Bubenik, J. P. Jacobson, D. Schams, L. Bartoˇs Fig. 6. A bilateral cryptorchid black-tailed buck. The antlers appear completely normal Fig. 7. Two unilateral cryptorchids. Their antlers also appear normal Cryptorchism, hypogonadism and antler malformations in black-tailed deer of Kodiak island 247 Fig. 8. Frontal view of the skull of Sitka deer with the separated sutures of nasal, frontal and parietal bones. Note the almost complete closure of the wound by newly grown bony tis- sue, especially at the caudal (top) end of the separation gap Because only two unilateral cryptorchids were found in our sample, we decided to compare only nine bilateral cryptorchids with eight control deer. As serum LH (15.7 and 6.6. ng/ml) and T (2.9 and 0.7 ng/ml) concentra- tions in unilateral cryptorchids, as well as antler forms were in the range of control bucks, we con- cluded that the unilateral cryp- torchids were not hypogonadic and therefore they were exclud- ed from both groups. Histological analysis of testes found no difference between the intensity of spermatogenesis or in the size of interstitium or Leydig cells of controls and the unilater- al cryptorchid. Fig. 9. Skull of Sitka deer with an apparent re-fusion of the inter- frontal suture (arrow) 248 G. A. Bubenik, J. P. Jacobson, D. Schams, L. Bartoˇs Intact bucks Bilateral cryptorchids 3.00 25 2.70 2.40 20 2.10 1.80 15 ng/ml 1.50 1.20 10 0.90 0.60 5 0.30 0.00 0 LH testosterone Fig.