Comparative studies of mitogen- and antigen-induced lymphocyte proliferation in four captive rhinoceros species.

Cellular immune function in four rhinoceros species was evaluated by way of in vitro lymphocyte proliferation responses to mitogenic and antigenic stimuli to establish normative data on white blood cell activity for each species and to identify species-specific differences that might help explain the predisposition of black rhinoceroses (Diceros bicornis) to disease. A cross section of the U.S. rhinoceros population encompassing all four captive species was sampled, including the Sumatran rhinoceros (Dicerorhinus sumatrensis) (n = 3); Indian rhinoceros (Rhinoceros unicornis) (n = 4); African black rhinoceros (n = 16); and African white rhinoceros (Ceratotherium simum) (n = 10). Of the four species evaluated, African black rhinoceroses exhibited the weakest (P < 0.05) lymphocyte proliferative responses to the mitogens: pokeweed (0.1 microg/ml), phytohemagglutinin (0.3 microg/ml), and concanavalin A (5.0 microg/ml). Total cell density at the end of culture was only 70% of that achieved with lymphocytes isolated from African white rhinoceroses, Indian rhinoceroses, and Sumatran rhinoceroses. However, lymphocyte response to bacterial endotoxin lipopolysaccharide was similar (P > 0.05) across species. Antigenic stimulation produced much weaker responses than mitogenic stimulation. No differences (P > 0.05) were observed among rhinoceros species in response to 1 and 10 microg/ml of Leptospira icterohemorrhagiae or Leptospira gryppotyphosa. Lymphocytes from African white rhinoceroses proliferated weakly in the presence of Aspergillus fumigatus filtrate, whereas lymphocytes from the southern black rhinoceros subspecies appeared slightly suppressed in the presence of increasing doses (0.1, 1, and 10 microg/ml) of Aspergillus filtrate. This comparative data set characterizing lymphocyte proliferation in the rhinoceros reveals several differences in immune cell responses among rhinoceros species and provides some evidence that lymphocytes of captive African black rhinoceroses are less vigorous than those of the other rhinoceros species.     

Digestion coefficients achieved by the black rhinoceros (Diceros bicornis), a large browsing hindgut fermenter

In contrast to the grazing white (C. simum) and Indian (R. unicornis) rhinoceros, the black rhinoceros (D. bicornis) is an exclusive browser. Due to the particular fermentation characteristics of browse, one would expect browsers to display both shorter ingesta retention times and lower digestion coefficients on comparable diets than grazers. In order to generate a database to test this hypothesis, we performed digestibility studies in eight black rhinoceroses (D. bicornis) from three zoological institutions, using total faecal collection for the quantification of faecal output. One to three regularly fed zoo rations of roughage, concentrates and varying proportions of browse material were used per animal. Additional data was taken from three hitherto unpublished studies as well as several published sources. When compared with horses on similar rations, black rhinoceroses achieved lower digestion coefficients for organic matter and CF. In general, an increase in dietary CF content led to a steeper decrease in organic matter and GE digestibility in black rhinoceroses than in horses. When comparing available data for rhinoceroses, browsing species showed a steeper decrease in organic matter digestibility than grazing species with increasing dietary cell wall content. Endogenous losses as determined by linear regression analysis were within the range reported for horses and Indian rhinoceroses. The results suggest that the horse is not a useful model animal for evaluating diets for black rhinoceroses energetically. In general, diets fed to captive black rhinoceroses seem to include higher proportions of concentrates than diets for other rhinoceros species, and an increase in browse or roughage would reduce digestion coefficients to levels observed in animals fed natural forage.     

Mineral absorption in the black rhinoceros (Diceros bicornis) as compared with the domestic horse

To test whether mineral recommendations for horses are likely to guarantee adequate mineral provision for black rhinoceroses (Diceros bicornis), we investigated the apparent absorption (aA) of macro- and microminerals in eight black rhinoceroses from three zoological institutions in a total of 32 feeding trials with total faecal collection, with additional data from three unpublished studies (18 feeding trials). Feeds and faeces were analysed for Ca, P, Mg, Na, K, Fe, Mn, Cu, Zn and Co. The resulting aA coefficients, and the linear relationships of apparently absorbable dietary mineral content to total dietary mineral content [per 100 g dry matter (DM)], were compared with data for domestic horses. Rhinoceroses had significantly higher aA coefficients for Ca and Mg (because of a higher calculated 'true' absorption), and lower ones for Na and K (because of calculated higher endogenous faecal losses). High absorption efficiency for divalent cations is hypothesized to be an adaptation to a natural diet of particularly high Ca:P ratio (approximately 14:1); an effective removal of Ca from the ingesta guarantees sufficient P availability at the fermentation site in the hindgut. Higher faecal losses of Na and K are hypothesized to be linked to a higher faecal bulk per DM intake in black rhinoceroses as compared with horses because of a generally lower digestive efficiency. There were no relevant differences in the absorption patterns of microminerals. In particular, there were no discernable differences in Fe absorption within the rhinoceroses for diets with and without tannin supplementation. Several of the zoo diets assessed in this study were deficient in Cu, Mn or Zn, and most contained excessive levels of Fe when compared with horse requirements. The findings of this study indicate that differences in mineral absorption between occur even between species of similar digestive anatomy; that in particular, Ca absorption might vary between hindgut fermenters with Ca:P ratio in their natural diet; that Na might be a particularly limiting factor in the ecology of free-ranging rhinoceroses; that moderate doses of tannins do not seem to markedly influence mineral absorption; and that diets for captive animals should contain adequate, but not excessive mineral levels.     

Clinical implications of rhinoceros reproductive tract anatomy and histology.

Reproductive tracts or tissues from five male black rhinoceroses (Diceros bicornis), two male white rhinoceroses (Ceratotherium simum), two male one-horned Asian rhinoceroses (Rhinoceros unicornis), seven female black rhinoceroses, and six female white rhinoceroses from multiple institutions were examined to characterize their anatomy and histology. Some observations and measurements were obtained from in situ tracts of intact animals before or during necropsy. Formalin-fixed tissues were dissected and examined histologically. Retrospective reproductive data from each rhinoceros was obtained from the institutions of origin. Reproductive histology of these species was similar to that of other mammals. Male accessory gland structure varied among species, and the Asian rhinoceros epididymis was more loosely attached and had larger duct diameters than did the epididymides of the African species. Although histology was typically mammalian, rhinoceros reproductive morphology combined characteristics of several different mammals. Defining this unique morphology of rhinoceroses may help in understanding their reproductive physiology and will effect the development of appropriate reproductive techniques.     

Fecal cortisol metabolite analysis for noninvasive monitoring of adrenocortical function in the cheetah (Acinonyx jubatus).

A radioimmunoassay was validated for quantifying excreted cortisol metabolites in cheetah (Acinonyx jubatus) feces. High-performance liquid chromatography analysis indicated that immunoreactivity was associated with a water-soluble metabolite in fecal extracts from males and females. None of the immunoreactivity corresponded with free cortisol or corticosterone but rather was associated with a more polar, unidentified metabolite. To determine the biologic relevance of excreted immunoreactive cortisol metabolites, cheetahs were exposed to a variety of situations anticipated to increase cortisol secretion. First, to assess acute changes in adrenal activity, adrenocorticotropic hormone (ACTH; 400 IU i.m.) was administered to two adult males and two adult females. Pre-ACTH baseline serum cortisol and fecal cortisol metabolite concentrations varied among individuals. Serum cortisol concentrations were elevated above baseline within 10 min of ACTH injection, followed by corresponding increases in fecal cortisol metabolite concentrations (690-4,194% above baseline) 48 hr later in three of four cheetahs. In the fourth cheetah, a smaller increase (334% above baseline) in fecal cortisol metabolite excretion was observed 96 hr after ACTH injection. Seven cheetah females also were subjected to a variety of potentially stressful manipulations, including immobilization, translocation, and introduction to a male to assess the ability of this technique to detect physiologic changes in adrenal activity. Increased fecal corticoid metabolite excretion was observed 24-72 hr after exposure to these exogenous stressors. Results indicate that adrenocortical activity can be monitored noninvasively in the cheetah through analysis of these metabolites. This technique could be valuable for evaluating, and thus optimizing, environmental and management conditions and for investigating the role of stress in disease pathogenesis and the usually poor reproductive performance of this species in captivity.     

Radiometric assessment of hexose monophosphate shunt capacity in erythrocytes of rhinoceroses.

OBJECTIVES: To measure metabolic rates of the hexose monophosphate shunt (HMPS) in erythrocytes of rhinoceroses, and to test the hypothesis that low concentrations of endogenous ATP in erythrocytes impair HMPS capacity, thereby increasing susceptibility to oxidant-induced hemolysis. ANIMALS: 13 black and 3 white rhinoceroses, free-ranging in several regions of southern Africa, and 1 Sumatran rhinoceros in US captivity. PROCEDURE: HMPS fluxes were measured in rhinoceros erythrocytes with carbon-labeled glucose in the presence and absence of known HMPS activators. RESULTS: Compared with values for human erythrocytes, mean basal state HMPS fluxes were appreciably lower (22 to 46%) in all 3 rhinoceros species studied. Shunt activators increased HMPS rates approximately 5-fold over basal rates in rhinoceros erythrocytes, compared with increases in humans of 10-fold with ascorbate and 15-fold with methylene blue. Stimulated HMPS rates in human erythrocytes were quantitatively 5- to 10-times greater than those observed in rhinoceros erythrocytes. Overall HMPS catabolic rates were completely independent of intracellular ATP concentrations. CONCLUSIONS AND CLINICAL RELEVANCE: HMPS glycolytic and recycling rates and responses to activators are inherently low in erythrocytes from 3 species of rhinoceros, likely contributing to (but not solely responsible for) the high susceptibility of black rhinoceroses to oxidant-induced hemolysis. Slow erythrocyte HMPS capacities were independent of intracellular ATP concentrations, invalidating a current hypothesis regarding the pathogenesis of hemolytic anemia in captive black rhinoceroses. Limitations in HMPS capacities emphasize the importance of protecting rhinoceroses from exposure to drugs, chemicals, toxins, foodstuffs, and other conditions known to increase production of oxidizing metabolites, reactive oxygen species, and free radicals.     

Acute intravascular hemolytic anemia in the black rhinoceros: hematologic and immunohematologic observations

To investigate the syndrome of acute intravascular hemolytic anemia in the black rhinoceros (Diceros bicornis), laboratory techniques used in the differential diagnosis of hemolytic anemias were performed on blood samples from 6 black rhinoceroses: 3 nonrelated healthy rhinoceroses, 1 rhinoceros with iron deficiency anemia, and 2 rhinoceroses with intravascular hemolysis. Osmotic fragility, erythrocyte membrane protein composition, hemoglobin electrophoresis, and hemoglobin stability did not distinguish between healthy and affected (anemia or hemolysis) rhinoceroses. Polyclonal antiglobulin reagents were prepared in rabbits, using whole rhinoceros serum and purified rhinoceros immunoglobulin G. These reagents were nonreactive against erythrocytes of the healthy and iron-deficient rhinoceroses. Reactions with RBC from the rhinoceros with fatal hemolytic anemia indicated increased membrane coating by the third component of complement; this was not evident in a second rhinoceros that survived a hemolytic event.     

Role of excessive maternal iron in the pathogenesis of congenital leukoencephalomalacia in captive black rhinoceroses (Diceros bicornis)

OBJECTIVE: To investigate the possibility that excessive maternal iron (overload) may contribute to development of congenital leukoencephalomalacia in captive black rhinoceroses. SAMPLE POPULATION: Tissue specimens and serum samples from 18 rhinoceroses in 2 kindreds harboring 4 (possibly 5) affected female calves. PROCEDURE: Fresh and archival sera and necropsy tissue specimens were evaluated to determine the nature and extent of iron overload in captive and wild black rhinoceroses as well as other rhinoceros species. RESULTS: Quantitative serum and tissue assays of iron and iron analytes, corroborated by histopathologic findings, indicated that these kindreds carried the greatest body burdens of iron yet found among captive black rhinoceroses. Fourteen of 18 rhinoceroses had the highest serum ferritin concentrations measured among 64 black rhinoceroses in captivity in the United States. Dams of affected calves had serum ferritin concentrations 2 orders of magnitude higher than clinically normal humans, equids, or free-ranging rhinoceroses. A neonatal serum sample from 1 affected female calf had a high ferritin concentration (approx 100-fold increase), but a male sibling of another affected female did not, suggesting a possible sex disparity in fetal response to maternal iron overload. Morphologic hallmarks of hemochromatosis were prominent in dams and grandams of affected calves. CONCLUSIONS AND CLINICAL RELEVANCE: Excessive maternal iron may affect female fetuses more than males, possibly inducing leukoencephalomalacia by catalyzing production of highly toxic hydroxyl free radicals during crucial periods of in utero development. Reduction of maternal iron overload may decrease the probability of developing leukoencephalomalacia and some other disorders commonly affecting rhinoceroses in captivity.     

High serum concentrations of iron, transferrin saturation and gamma glutamyl transferase in captive black rhinoceroses (Diceros bicornis)

Iron storage disease (haemochromatosis) is thought to be the cause of many disorders unique to captive black rhinoceroses (Diceros bicornis). To establish reliable reference ranges for iron parameters, serum samples from 27 eastern black rhinoceroses (Diceros bicornis michaeli) from a translocation programme in Kenya were analysed and compared with the samples from 17 captive individuals. The transferrin saturation, serum iron concentration and gamma glutamyl transferase were significantly higher in the captive rhinoceroses, but these elevations were not evident when the results were compared with previously published data.     

The influence of dietary tannin supplementation on digestive performance in captive black rhinoceros (Diceros bicornis)

Free-ranging browsers such as the black rhinoceros (Diceros bicornis) consume a diet that contains tannins, whereas the diets offered to them in captivity consist mostly of items known to contain hardly any such secondary plant compounds. Tannins could have potentially beneficial effects, including the chelation of dietary iron (iron storage disease is a common problem in black rhinos). Here, we tested the acceptance, and the consequences on digestion variables, of a low-dose tannin supplementation in captive animals. Eight black rhinoceroses from three zoological institutions were used. Faecal output was quantified by total faecal collection. Diets fed were regular zoo diets supplemented with either tannic acid (T, hydrolysable tannin) or quebracho (Q, condensed tannins); overall tannin source intake increased at 5-15 g/kg dry matter (DM) in relation to regular zoo diets. Adaptation periods to the new diets were >2 months. Additional data were taken from one hitherto unpublished study. Data were compared to measurements in the same animals on their regular zoo diets. All animals accepted the new diets without hesitation. There was no influence of tannin supplementation on digestion coefficients of DM and its constituents, or faecal concentrations of short-chain fatty acids or lactate. Water intake did not increase during tannin supplementation. Should the inclusion of dietary tannin sources be an objective in the development of diets for captive rhinoceroses, moderate doses such as used in this study are unlikely to cause relevant depressions of digestive efficiency and will not interfere with bacterial fermentation in a relevant way.     

Beta-globin chain hemoglobin polymorphism and hemoglobin stability in black rhinoceroses (Diceros bicornis)

To evaluate the syndrome of acute intravascular hemolytic anemia in black rhinoceroses (Diceros bicornis), the hemoglobin of this species was evaluated by use of isopropanol- and heat-stability tests and was further characterized by electrophoretic studies. Samples were obtained from 22 apparently healthy captive North American black rhinoceroses, though 3 of the study animals had survived previous hemolytic events, and 3 others were parents of 3 offspring that had suffered hemolysis. The eastern African (Diceros bicornis michaeli) and the southern African subspecies (D b minor) were represented. Comparative samples were also obtained from 2 white (Ceratotherium simum) and 1 Indian (Rhinoceros unicornis) rhinoceroses. The hemoglobin of all 3 species appeared stable when tested by use of the heat and isopropanol methods. Thus, an unstable hemoglobin does not appear to be involved in the hemolytic crises of captive black rhinoceroses. Black rhinoceros hemoglobin had a striking polymorphism. Thirteen of the samples from black rhinoceroses had a single hemoglobin band, based on results of alkaline electrophoresis. Nine had, in addition to this major band, a slow (more cathodic) minor band that comprised about 10% of the total hemoglobin. Further studies indicated that the major band and the slower minor band may contain globin chains analogous to human beta- and delta-chains respectively; these bands have been tentatively designated B and C. Phenotypes B and BC are common, in a ratio of 4:3. A genetic mechanism is proposed that assumes beta b and beta c gene loci and that beta c-locus-expressed (beta c+) and beta c-locus-inhibited (beta c degrees) are common alleles for the beta c-locus.(ABSTRACT TRUNCATED AT 250 WORDS)     

Reproductive parameters in free-ranging female black rhinoceroses (Diceros bicornis) in Zimbabwe

Samples and data were collected from twenty-eight female black rhinoceroses (Diceros bicornis) during translocation efforts carried out by the Department of National Parks and Wildlife Management in Zimbabwe. Biological data were collected, cytological examination of vaginal smears was performed, and serum concentrations of follicle stimulating hormone, luteinizing hormone, progesterone, oestriol, and 17-beta-oestradiol were determined by radio-immuno-assay. Prolactin levels were determined for 3 pregnant animals, 1 of which was sampled before and after parturition. Vaginal cytology was not found to be helpful for indicating the oestrous cycle stage for the black rhinoceros, but progesterone and 17-beta-oestradiol levels were found to be useful indicators of pregnancy and possibly of oestrous cycle stage as well.     

Influence of dietary endophyte (Neotyphodium coenophialum)-infected tall fescue (Festuca arundinacea) seed on fecal shedding of antibiotic resistance-selected Escherichia coli O157:H7 in ewes

The objectives were to determine the effects of short-term feeding of a toxic endophyte (Neotyphodium coenophialum)-infected tall fescue seed (Festuca arundinacea, cultivar 'Kentucky 31') on fecal shedding and intestinal concentrations of Escherichia coli O157:H7 and the concentrations of prolactin, cortisol, and NEFA in experimentally inoculated ewes. Twelve ewes (mean BW = 46 +/- 2 kg) were fed a diet containing either high endophyte-infected (HI-E) or low endophyte-infected (LO-E) tall fescue seed for 7 d. Each diet consisted of 50% (as-fed basis) tall fescue seed. Ewes were experimentally inoculated with antibiotic resistance-selected E. coli O157:H7 on d 1 of the feeding treatment, and fecal shedding of inoculated pathogens was monitored daily on d 2 to 6. On d 7, ewes were weighed and euthanized, and tissues and contents were sampled from the ileum, cecum, and rectum for quantitative enumeration of E. coli O157:H7. Urine was collected at euthanization to determine total ergot alkaloid concentrations. Ewes fed HI-E had lower (P < 0.001) DMI than did ewes fed LO-E (0.8 and 1.6 +/- 0.1 kg/d of DMI for HI-E and LO-E ewes, respectively); consequently, there was a tendency (P = 0.06) for HI-E ewes to lose 0.3 +/- 0.4 kg of BW/d and LO-E ewes to gain 0.2 +/- 0.4 kg of BW/d during the 7 d. Urinary ergot alkaloids were increased (P < 0.001) in ewes fed HI-E (47.8 +/- 9.4 ng/mg of creatinine) compared with those fed LO-E (6.2 +/- 9.4 ng/mg of creatinine). Prolactin tended (P = 0.06) to be decreased in ewes fed HI-E (7.2 +/- 7.0 ng/mL) compared with those fed LO-E (27.7 +/- 7.0 ng/mL). Fecal shedding of E. coli O157:H7 tended (P = 0.06) to be increased in HI-E ewes [5.4 cfu (log10)/g of feces] compared with LO-E ewes [4.5 cfu (log10)/g of feces]. The population of E. coli O157:H7 in luminal contents from the ileum, cecum, and rectum did not differ (P > 0.36) between treatments. Treatment did not influence (P = 0.30) the occurrence of E. coli O157:H7 in cecal or rectal tissues; however, ileal tissues from HI-E ewes tended (P = 0.12) to have an increased incidence of E. coli O157:H7. Concentrations of NEFA tended (P = 0.12) to be greater in HI-E ewes than in LO-E ewes, whereas cortisol was similar (P = 0.49) for HI-E and LO-E ewes. We conclude that short-term feeding of HI-E tall fescue seed may alter the concentrations of prolactin and NEFA, and may increase fecal shedding of E. coli O157:H7 in experimentally inoculated ewes.     

Oral, nasal, and cutaneous eosinophilic granulomas in the black rhinoceros (Diceros bicornis): a lesion distinct from superficial necrolytic dermatitis.

Oral, nasal, and cutaneous lesions resembling eosinophilic granulomas (EGs) were observed in eight captive black rhinoceroses (Diceros bicornis). Oral lesions were observed in all affected animals and occurred most often behind the prehensile lip. The typical clinical history of affected animals included oral bleeding or epistaxis from multilobulated, fungating, proliferative masses with areas of ulceration. Histologically, lesions were characterized by prominent infiltrates of eosinophils and rare foci of collagen degeneration on a background of marked submucosal or dermal neovascularization. Mucosal or epidermal hyperplasia was also present sometimes in association with distinctive epithelial degenerative changes consistent with superficial necrolytic dermatopathy of black rhinoceroses. Lesions of EG and superficial necrolytic dermatopathy were interpreted as being concurrently manifested in these cases. EG lesions spontaneously resolved over periods of 30 days-7 mo and were recurrent in three animals. Lesions were responsive to treatment with corticosteroids or to local cryotherapy with liquid nitrogen. Two animals treated with systemic corticosteroids died of disseminated fungal infections, emphasizing that corticosteroids should be used cautiously in black rhinoceroses.     

Noninvasive monitoring of stress hormone levels in a female steller sea lion (Eumetopias jubatus) pup undergoing rehabilitation.

Steller sea lions (Eumetopias jubatus) rarely strand in areas monitored by humans, and there is little published data on the diseases, parasites, nutritional state, and stress levels of Steller sea lions in the wild. In May 2002, a female Steller sea lion pup (EJS-02-01) was sighted separated from her mother after strong storms in Southeast Alaska. After 5 days of observations, EJS-02-01 was transferred to the Alaska SeaLife Center (ASLC) in Seward, Alaska. During 11 mo of rehabilitation at ASLC, body weight was monitored and opportunistic fecal samples (n = 86) were analyzed for corticosterone concentrations. Fecal corticosterone concentrations ranged from 15 to 3,805 ng/ g for EJS-02-01. Peak corticosterone values reflected responses to acute stressors during rehabilitation. EJS-02-01 was successfully released at Gran Point, Alaska, in April 2003. Fecal corticosterone assay monitoring provided a valuable tool to monitor various stressors and is useful in monitoring long-term situations like rehabilitation.     

Changes in fecal progestagen profile after excretion in miniature pigs

The aim of this study was to evaluate whether the fecal progestagen (progesterone and its metabolites) levels of miniature pigs would change after excretion at room temperature. Our initial investigation focused on the correlations between the fecal progestagen concentrations with and without ether extraction and between the plasma progesterone and fecal progestagen concentrations in order to develop an enzyme-linked immunosorbent assay (ELISA) for fecal progestagen without ether extraction. There were significant correlations between fecal progestagen concentrations with and without ether extraction (r=0.880) and between fecal progestagen concentrations without ether extraction and plasma progesterone (r=0.763). The fecal progestagen concentration obtained by ELISA without ether extraction was almost identical to that obtained with ether extraction. These results validate the ELISA method without ether extraction, which was therefore used for the latter experiment. Fecal samples collected from the pigs were preserved for 0-24 h at room temperature, and then their fecal progestagen concentrations were measured. The fecal samples preserved for 0 to 24 h were analyzed by high performance liquid chromatography (HPLC) and ELISA. The concentrations of all samples significantly increased with time after preservation. The progestagen concentration of fresh feces (0 h) with high progestagen concentration (>1000 ng/g) increased significantly after 3 h. The concentration increased significantly after 12 h for fresh feces containing about 500 ng/g progestagen. HPLC analysis is showed that the fecal progesterone concentration, but not its other metabolites, doubled 24 h after excretion compared with the concentration at 0 h. These results suggest that dynamic changes in the profile of progesterone metabolites occur in feces after excretion.     

Piroplasm parasites of white rhinoceroses (Ceratotherium simum) in the Kruger National Park, and their relation to anaemia

As part of a larger survey to map the geographical distribution of Babesia and Theileria parasites in the southern African rhinoceros population, white rhinoceroses were sampled during routine immobilisations in the Kruger National Park. Polymerase chain reaction (PCR) and reverse line blot (RLB) hybridisation assays were used to screen for the presence of piroplasms and complete blood counts were used to assess associated changes in clinical parameters. Of the 195 rhinoceroses sampled, 71 (36.4 %) tested positive for the presence of Theileria bicornis, with no significant change in the haematological parameters measured, while 18 (9.2 %) tested positive for Theileria equi. None of the rhinoceroses sampled tested positive for Babesia bicornis, a parasite associated with mortalities in black rhinoceroses.     

Assessment of adrenocortical activity by non-invasive measurement of faecal cortisol metabolites in dromedary camels (Camelus dromedarius)

The aim of this study was to determine whether glucocorticoid production could be monitored non-invasively in dromedary camels by measuring faecal cortisol metabolites (FCMs). Five Sudanese dromedaries, two males and three females, were injected with a synthetic adrenocorticotropic hormone (ACTH) analogue. Blood samples were collected pre- and post-ACTH injection. Faeces were sampled after spontaneous defecation for five consecutive days (2 days before and 3 days after ACTH injection). Baseline plasma cortisol values ranged from 0.6 to 10.8 ng/ml in males and from 1.1 to 16.6 ng/ml in females, while peak values after ACTH injection were 10.9–41.9 in males and 10–42.2 ng/ml in females. Peak blood cortisol values were reached between 1.5 and 2.0 h after ACTH injection. The concentration of FCMs increased after ACTH injection in the faeces of both sexes, although steroid levels peaked earlier in males [24 h; (286.7–2,559.7 ng/g faeces)] than in females [36–48 h; (1,182.6–5,169.1 ng/g faeces)], reflecting increases of 3.1–8.3- and 4.3–8-fold above baseline levels. To detect chromatographic patterns of immunoreactive FCMs, faecal samples with high FCM concentrations from both sexes were pooled and subjected to reverse phase high performance liquid chromatography (RP-HPLC). RP-HPLC analysis revealed sex differences in the polarity of FCMs, with females showing more polar FCMs than males. We concluded that stimulation of adrenocortical activity by ACTH injection resulted in a measurable increase in blood cortisol that was reliably paralleled by increases in FCM levels. Thus, measurement of FCMs is a powerful tool for monitoring the adrenocortical responses of dromedaries to stressors in field conditions.     

Effect of venipuncture site and anticoagulant on selected hematologic values in black rhinoceros (Diceros bicornis).

Paired blood samples were collected from the ear and radial vein of four captive healthy adult black rhinoceroses (Diceros bicornis). Samples were collected using heparin or ethylenediaminetetraacetic acid (EDTA) as an anticoagulant. Packed cell volume (PCV) and total protein (TP) values were compared between samples drawn from the two venipuncture sites and treated with the two anticoagulants to determine whether statistically significant variation occurred. No significant difference in the grouped values was observed when venipuncture sites (ear and radial vein) were compared using the same anticoagulant (heparin). However, when comparing different anticoagulants (EDTA and heparin) used to collect blood from the radial vein, the grouped-heparinized samples had higher mean PCV and TP values than did the EDTA-treated samples. These differences may be important when performing serial sampling in a sick rhinoceros and suggest that the choice of anticoagulant should be consistent, although selection of venipuncture site may be less important when monitoring selected hematologic values in black rhinoceroses.     

Validation of a fecal glucocorticoid metabolite assay for collared peccaries (Pecari tajacu)

The possibility of assessing endogenous adrenal activity in the collared peccary (Pecari tajacu) was tested by using an adrenocorticotropic hormone (ACTH) challenge in a fecal glucocorticoid metabolite (FGM) assay. Feces were collected from 12 captive adult male peccaries beginning 48 hr prior to challenge; six of these animals received the challenge as an ACTH injection and the other six were injected with saline solution. Feces collection ended 120 hr after injections. As a control, feces were collected for eight consecutive days from another six adult male peccaries that remained in their original mixed-sex herds in semiconfined paddocks. All feces samples were freeze-dried, extracted by an ethanol vortex method, and assayed for glucocorticoids by means of an enzyme immunoassay. FGM concentrations were compared between the treatments by a repeated measures analysis of variance (ANOVA) followed by a post hoc Tukey test. The assay is reliable but, instead of the usual proportion of 1:50 in ethanol (fecal mass:solvent), 1:10 is recommended for best extraction of FGM. Baseline FGM concentrations were similar among the ACTH, saline, and control treatments (29.7 +/- 11.2 ng/g(-1) dry feces) during the 48 hr before the challenge. The ACTH group reached an FGM excretion peak at 24 hr post-treatment, followed by a decline, while in the control and saline groups FGM levels remained relatively constant. Therefore, the fecal glucocorticoid metabolite assay reflects endogenous adrenal activity in the collared peccary and is a powerful tool for noninvasive stress monitoring in peccaries.