Corticosteroid-induced suppression of in vitro lymphocyte proliferation in four captive rhinoceros species.

Captive African black rhinoceroses (Diceros bicornis) are unusually susceptible to several diseases not commonly observed in any of the other three rhinoceros species maintained in captivity. The potential role of corticosteroids (either endogenously produced or exogenously administered) in the development of these sometimes fatal diseases has been questioned. In this study, the suppressive effects of two therapeutic corticosteroids (dexamethasone and hydrocortisone) on in vitro lymphocyte proliferation was examined in four rhinoceros species, including the Sumatran rhinoceros (Dicerorhinus sumatrensis, n = 3), Indian rhinoceros (Rhinoceros unicornis, n = 4), African black rhinoceros (n = 10), and African white rhinoceros (Ceratotherium simum, n = 5). Three blood samples collected from each rhinoceros 1 mo to 1 yr apart provided replicates for the study. Both dexamethasone and hydrocortisone suppressed (P < 0.05) lymphocyte proliferation stimulated by B-cell mitogens (pokeweed and lipopolysaccharide) and T-cell mitogens (phytohemagglutinin and concanavalin A). Suppressive effects of the glucocorticoids differed (P < 0.05) depending on the mitogen used to stimulate the lymphocytes. Overall, dexamethasone was a more potent suppressor of cell proliferation when compared with hydrocortisone (P < 0.05). However, black rhinoceros cell proliferation in response to any of the four mitogens was never completely suppressed, even in cultures containing the highest steroid concentration tested (10(-3) M). The effect of the two corticosteroids differed slightly among the rhinoceros species and subspecies tested, but there was no evidence that eastern or southern black rhinoceros lymphocytes were more sensitive to the suppressive effects of corticosteroids than the other rhinoceros species.     

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.     

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.     

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)     

Gross anatomy and ultrasonographic images of the reproductive system of the Sumatran rhinoceros (Dicerorhinus sumatrensis)

The Sumatran rhinoceros (Dicerorhinus sumatrensis) is the smallest of all the rhino species. It is one of the rarest mammals in the world and is in imminent danger of extinction. A study was carried out on seven wild-caught females, three wild-caught males and one captive born female Sumatran rhinoceros at the Sumatran Rhinoceros Breeding Centre in Sungai Dusun, Selangor, Malaysia, beginning 1990. As a result of the paucity of scientific information on the reproductive biology of the Sumatran rhinoceros, this study was conducted to obtain information, which could assist in the captive breeding of this endangered and near extinct species. The anatomy of the reproductive system was based on two post-mortem specimens and transrectal real-time ultrasonography in six adult females. Genitalia of the Sumatran rhinoceros were similar to those of other species of rhinoceroses. The cervix consisted of several folds, the uterus was bicornuate with a short body and prominent horns and the ovaries were completely covered by the fimbriated end of the fallopian tube. The internal genitalia could be imaged by ultrasonography. The testes were located within a pendulous scrotum. Two lateral projections were located at the base of the penis. A well-defined process glandis was present at the tip of the penis. The accessory sex glands and the testes could be imaged by ultrasonography.     

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.     

Fatty acid status in captive and free-ranging black rhinoceroses (Diceros bicornis)

The fatty acid (FA) patterns of plasma/serum triglycerides (TG), phospholipids (PL) and cholesteryl esters (CE) of captive and free-ranging black rhinoceroses (Diceros bicornis) were investigated. Free-ranging animals (n = 28) stemmed from four different regions. Captive animals sampled included specimens from North American (n = 11) and three different European facilities (n = 6). The European animals were tested on 1-4 different diets, resulting in a total of 15 blood samples. Regardless of differences between the free-ranging animals from different regions, differences between captive and free-ranging animals were relatively uniform: captive animals had higher overall proportions of polyunsaturated fatty acid (PUFA), due to levels of linoleic acid (LA, 18:2n6) that were drastically increased as compared to free-ranging animals. In contrast, levels of alpha-linolenic acid (ALA, 18:3n3) were consistently lower on conventional zoo diets. n6/n3 ratios for TG, PL and CE were 1.6, 10 and 8 in samples from free-ranging animals, respectively, as compared to 4.1-16.3, 16-148 and 40-277 in samples from captive animals. There was a distinct correlation between the proportion of grain-based products (commercial concentrates, plain grains and bread) in the diets of the European animals and the measured levels of n6 PUFA. An animal from a facility with a very low proportion of grain products in the diet nevertheless had high LA readings, most probably due to the use of sunflower oil as 2% (dry matter basis) of its diet. One animal that received a high proportion of grass meal pellets due to an oral disease had increased ALA contents after the diet change. These results allow conclusions on the suitability of diets fed in captivity: the black rhinoceros is prone to several uncommon diseases that have been suspected to be linked to oxidative damage, possibly due to the disposition of this species to excessive iron storage. An unnatural dietary loading with PUFAs would exacerbate this problem. Additionally, n6 FAs are known as precursors of pro-inflammatory mediators, and their overrepresentation could therefore exacerbate any inflammatory processes. Therefore, the current practice of using grain-based feeds as major ingredients in captive rhinoceros diets is discouraged. Diet items containing ALA (a precursor of anti-inflammatory mediators) such as, fresh grass, fresh browse, the respective silages should be included at higher levels in diets for captive black rhinoceroses. Grass meal pellets, although a good source of ALA and linked with high levels of ALA in an animal of this study, must be chosen with care for black rhinoceroses due to their particular proneness for high iron contents.     

Urinalysis in three species of captive rhinoceros (Rhinoceros unicornis, Dicerorhinus sumatrensis, and Diceros bicornis)

This study reports urinalysis values for three species of captive rhinoceros (Rhinoceros unicornis, Dicerorhinus sumatrensis, and Diceros bicornis) and evaluates individual and species differences. Repeated urinalysis was conducted on 11 individuals to establish normal reference ranges. Although no individual or species differences existed in urinary values for pH, all species differed in specific gravity. Rhinoceros urine demonstrated many physical and chemical properties similar to that of the horse, but reliability of this comparison was limited. Urinary pH in the rhinoceros was within range of that established for the horse and other large herbivores. However, all rhinoceros species exhibited urinary specific gravities below the lower limit of the normal equine reference range. Comparative urinalysis using an outside laboratory source confirmed the results of this study and illustrated the value of conducting in-house analysis. These results are the first data available on reference ranges for urine parameters in the greater one-horned, Sumatran, and African black rhinoceros and provide a useful diagnostic tool for the veterinary care of individuals in captivity.     

Risk factors associated with a skewed natal sex ratio in captive black rhinoceroses (Diceros bicornis) in the United States.

The skew toward males in the sex ratio of calves is considered to be a problem for the captive population of black rhinoceroses (Diceros bicornis) in North America. This study used a retrospective case-control design to determine whether there was a predisposition toward birth of male calves over female calves and to examine risk factors associated with the occurrence of any detected skewed natal sex ratio in captive black rhinoceroses in the United States. The study population included captive female black rhinoceroses housed in the United States that had given birth to at least one calf of known sex. This study confirmed a skewing of the natal sex ratio toward male calves in the captive black rhinoceros population. The skewed ratio was found in calves born to wild-born dams, for which an increased time in captivity, irrespective of age, was associated with an increased likelihood of a male calf. Dams between 12 and 19 yr of age had a decreased likelihood of a male calf. The data also suggested a possible trend for the southern subspecies of black rhinoceros (Diceros bicornis minor) to be more likely to produce male calves than the eastern subspecies (Diceros bicornis michaeli). No associations were found with the sex of offspring in captive-born dams: however, this lack of association could be a result of low power in the study.     

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.     

Semen collection in rhinoceroses (Rhinoceros unicornis, Diceros bicornis, Ceratotherium simum) by electroejaculation with a uniquely designed probe.

Electroejaculation in rhinoceroses has historically yielded inconsistent results, with the collection of high-quality, sperm-rich samples rare. The goal of this study was to develop a reliable method of electroejaculation in the rhinoceros by designing a rectal probe that appropriately fits the anatomy of this taxon and refining the procedure. A curved probe handle ending in an oblate, ellipsoid head was built using readily available supplies. A combination of rectal massage, penile massage, and electrical stimulation with a specially designed probe was employed in attempts to collect semen on 14 occasions from greater one-horned rhinoceroses (Rhinoceros unicornis; n = 4), black rhinoceroses (Diceros bicornis; n = 2) and a southern white rhinoceros (Ceratotherium simum; n = 1). During 13 of the 14 attempts, ejaculates were collected in multiple fractions. All but one of the ejaculates contained spermatozoa, and seven ejaculates contained good-quality fractions of semen (-60% sperm motility; > or =20 x 106 spermatozoa/ml) suitable for sperm banking and assisted reproduction procedures. Mean (+/-SEM) values for volume, pH, osmolality, and total sperm number for ejaculates containing good-quality fractions (98.2 +/-21.8 ml, 8.5+/-0.1, 290.4+/-6.7 mOsm, and 37.1+/-12.0 x 10(9), respectively) did not differ (P > 0.05) from those containing only poor-quality samples. Urine and/or erythrocyte contamination was not uncommon in fractions of both ejaculate types. Males producing good-quality samples ranged in age from 7 to 34 yr. None of the samples contained > or =75% morphologically normal spermatozoa. Electroejaculation with a uniquely designed probe consistently produced ejaculates in the rhinoceros. However, the production of high-quality samples continued to be challenging, occurring in only 50% of collection attempts. Regardless, the technology has progressed to a stage at which good-quality semen samples can be produced for sperm banking and assisted reproduction, and thereby can be integrated into intensive rhinoceros management strategies for the ultimate survival of this taxon.     

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.     

Leptospira infection in two black rhinoceroses (Diceros bicornis michaeli).

Two black rhinoceroses (Diceros bicornis michaeli) developed clinical leptospirosis without hemolytic crises. The first rhinoceros presented with peracute depression, anorexia, rear leg trembling, dysuria, glucosuria, gastrointestinal discomfort, and decreased fecal output and died within 12 hr. Necropsy and histopathology revealed lesions within multiple organs. Leptospirosis was diagnosed postmortem based on positive fluorescent antibody staining of liver. The second rhinoceros presented 2 mo later with similar signs. It survived with treatment and was diagnosed with leptospirosis based on serology using microscopic agglutination testing and detection of urinary antigen using a fluorescent antibody technique. Leptospira kirschneri serovar grippotyphosa was postulated as the etiologic agent, with transmission probably occurring through wallow contamination by wild raccoons (Procyon lotor).     

RADIOGRAPHIC PROTOCOL AND NORMAL ANATOMY OF THE HIND FEET IN THE WHITE RHINOCEROS (CERATOTHERIUM SIMUM)

Foot pathology is a common and important health concern in captive rhinoceroses worldwide, but osteopathologies are rarely diagnosed, partly because of a lack of radiographic protocols. Here, we aimed to develop the first radiographic protocol for rhinoceros feet and describe the radiographic anatomy of the white rhinoceros (Ceratotherium simum) hind foot (pes). Computed tomographic images were obtained of nine cadaver pedes from seven different white rhinoceroses and assessed for pathology. A single foot deemed free of pathology was radiographed using a range of different projections and exposures to determine the best protocol. 3D models were produced from the CT images and were displayed with the real radiographs to describe the normal radiographic anatomy of the white rhinoceros pes. An optimal radiographic projection was determined for each bone in the rhinoceros pes focusing on highlighting areas where pathology has been previously described. The projections deemed to be most useful were D60Pr‐PlDiO (digit III), D45Pr45M‐PlDiLO (digit II), and D40Pr35L‐PlDiLO (digit IV). The primary beam was centered 5–7 cm proximal to the cuticle on the digit of interest. Articular surfaces, ridges, grooves, tubercles, processes and fossae were identified. The radiographic protocol we have developed along with the normal radiographic anatomy we have described will allow for more accessible and effective diagnosis of white rhinoceros foot osteopathologies.     

Attempted in vitro maturation and fertilization of postmortem Sumatran rhinoceros (Dicerorhinus sumatrensis) oocytes

A study was conducted opportunistically to evaluate the potential of rescuing immature oocytes from the ovaries of the Sumatran rhinoceros postmortem. Recovered oocytes (n = 30) were placed in maturation culture for 36 hr and inseminated with frozen-thawed homologous spermatozoa. After culture, evaluation of nuclear maturation status revealed that a large number of oocytes were degenerated (n = 21), but nine oocytes were assessed at the germinal vesicle (n = 3), metaphase I (n = 3), and metaphase II (n = 3) stages. Frozen-thawed Sumatran rhinoceros spermatozoa were capable of binding to the zona pellucida of in vitro matured oocytes, but no fertilization or cleavage resulted. In conclusion, relatively large numbers of oocytes can be obtained by ovarian follicular aspiration postmortem in the Sumatran rhinoceros, and some of these oocytes are capable of achieving nuclear maturation in vitro. However, additional studies are required to improve maturation success and achieve fertilization in culture.     

Remote assessment of stress in white rhinoceros (Ceratotherium simum) and black rhinoceros (Diceros bicornis) by measurement of adrenal steroids in feces.

This study monitored fecal cortisol and corticosterone levels in 14 black rhinoceroses (Diceros bicornis) and in seven white rhinoceroses (Certotherium simum) under various conditions of captivity, including translocation. Free cortisol and free corticosterone were measured in methylene chloride extracts of feces, using high-performance liquid chromatography (HPLC). The extraction-assay method was validated for quantitative measurement of these hormones by mass spectroscopy analysis, chemical derivitization, and radiolabel tracking and recovery. Both cortisol and corticosterone were extractable from feces and routinely detectable by HPLC. In three nonstressed, captivity-adapted white rhinoceroses monitored across 21 days of routine activity, fecal cortisol ranged from 2.0 to 7.3 ng/g dry feces and corticosterone from 4.0 to 10.8 ng/g dry feces, with no observable trend. Matched plasma, urine, and fecal samples in these rhinoceroses yielded corticosterone:cortisol ratios of 2.0:1.0, 2.7:1.0, and 2.2:1.0, respectively. Both black rhinoceroses (n = 5) and white rhinoceroses (n = 4) exhibited higher fecal cortisol (6.9- to 10.0-fold) and corticosterone (3.2- to 4.5-fold) levels in association with restraint-translocation than in limited free-roaming conditions. In five black rhinoceroses monitored across 6 wk after release from translocation, fecal levels of both cortisol and corticosterone decreased significantly between week 1 and weeks 4-6. In general, cortisol and corticosterone paralleled each other, with cortisol exhibiting a greater range of response. Measurement of either hormone in feces appears to be reliable for adrenal axis monitoring in the white and the black rhinoceroses.     

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.     

Reference cardiopulmonary physiologic parameters for standing, unrestrained white rhinoceroses (Ceratotherium simum).

Chemical restraint is an important tool for the management and medical care of both captive and free-ranging rhinoceroses. Current anesthetic protocols for the white rhinoceros (Ceratotherium simum) are reported to cause varying degrees of hypertension, tachycardia, muscular stiffness and fasciculation, acidosis, and, most importantly, respiratory depression with resulting hypoventilation, hypoxia, and hypercapnea. To assist in the assessment and development of new and improved anesthetic techniques for the white rhinoceros, the following cardiopulmonary reference parameters for standing, unrestrained white rhinoceroses were generated (mean +/- standard error [minimum maximum]): heart rate = 39 +/- 0.8 beats/min (32-42), respiratory rate = 19 +/- 0.6 breaths/min (16-23), corrected indirect systolic blood pressure = 160 +/- 2.9 mm Hg (146-183), corrected indirect diastolic blood pressure = 104 +/- 2.3 mm Hg (88-117), corrected indirect mean blood pressure = 124 +/- 2.2 mm Hg (108-135), end tidal CO2 = 45.1 +/- 0.7 mm Hg (41.7-48.0), rectal temperature = 36.8 +/- 0.1 degrees C (36.6-37.2), arterial blood pH = 7.391 +/- 0.007 (7.346-7.431), arterial partial pressure of oxygen = 98.2 +/- 1.4 mm Hg (90.2-108.6), arterial partial pressure of CO2 = 49.0 +/- 0.9 mm Hg (44.4-53.7), base excess = 3.5 +/- 0.4 mmol/L (1.9-5.9), bicarbonate = 29.3 +/- 0.4 mmol/L (27.3-32.2), and arterial hemoglobin oxygen saturation (SaO2) = 97.2 +/- 0.1% (96.6-98.0).     

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.     

Germany/Australia Index of Sperm Sex Sortability in Elephants and Rhinoceros

Flow cytometric sexing of spermatozoa followed by application in artificial insemination or in vitro fertilization provides a unique opportunity to predetermine the sex of offspring and might enhance the conservation management of endangered species in captivity such as the elephant and rhinoceros. To obtain an indication of the sortability of spermatozoa from these species, the relative DNA differences between X and Y chromosome bearing spermatozoa (fresh, frozen thawed, epididymal) from three rhinoceros species [white ( Ceratotherium simum ), black ( Diceros bicornis ), Indian ( Rhinoceros unicornis )] and both elephant species, the Asian and the African elephant ( Elephas maximus, Loxodonta Africana ), were determined through separation of spermatozoa into X and Y chromosome bearing populations, using a modified high speed flow cytometer. The head profile areas of spermatozoa from all five species were measured using light microscopy. By multiplying the relative DNA differences and the head profile areas, the sperm sorting indices were calculated to be 47, 48 and 51 for white, black and Indian rhinoceros respectively. The calculated sorting index for the Asian elephant was 66. In the African elephant, we determined the highest sorting index of 76. These results indicate the practicability of flow cytometric sex sorting of spermatozoa from the tested rhinoceros species and both elephant species. The lower sorting indices in rhinos indicate that sex sorting of spermatozoa from the rhinoceros will be more challenging than in elephants.