Subclinical hypocalcaemia in captive Asian elephants (Elephas maximus)

Papers & Articles Subclinical hypocalcaemia in captive Asian elephants (Elephas maximus) J. H. van der Kolk, J. P. T. M. van Leeuwen, A. J. M. van den Belt, R. H. N. van Schaik, W. Schaftenaar The hypothesis that hypocalcaemia may play a role in dystocia in captive Asian elephants (Elephas maximus) was investigated. The objectives of the study were to measure the total calcium concentration in elephant plasma; assess the changes in parameters of calcium metabolism during a feeding trial; investigate a possible relationship between calcium metabolism and dystocia; and assess bone mineralisation in captive Asian elephants in vivo. The following parameters were measured: total and ionised calcium, inorganic phosphorous and magnesium, the fractional excretions of these minerals, intact parathyroid hormone, 25-OH-D3 and 1,25-OH-D3. Radiographs were taken from tail vertebrae for assessment of bone mineralisation. The mean (sd) heparinised plasma total calcium concentration was 2·7 (0·33) mmol/l (n=43) ranging from 0·84 to 3·08 mmol/l in 11 Asian elephants. There was no significant correlation between plasma total calcium concentration and age. Following feeding of a calcium rich ration to four captive Asian elephant cows, plasma total and ionised calcium peaked at 3·6 (0·24) mmol/l (range 3·4 to 3·9 mmol/l) and 1·25 (0·07) mmol/l (range 1·17 to 1·32 mmol/l), respectively. Plasma ionised calcium concentrations around parturition in four Asian elephant cows ranged from 0·37 to 1·1 mmol/l only. The present study indicates that captive Asian elephants might be hypocalcaemic, and that, in captive Asian elephants, the normal plasma concentration of total calcium should actually be around 3·6 mmol/l and normal plasma concentration of ionised calcium around 1·25 mmol/l. Given the fact that elephants absorb dietary calcium mainly from the intestine, it could be concluded that elephants should be fed calcium-rich diets at all times, and particularly around parturition. In addition, normal values for ionised calcium in captive Asian elephants should be reassessed.

Veterinary Record (2008) 162, 475-479 J. H. van der Kolk, DVM, PhD, DipECEIM, Department of Equine Sciences, Medicine Section, A. J. M. van den Belt, DVM, PhD, Department of Diagnostic Imaging, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands J. P. T. M. van Leeuwen, PhD, Department of Internal Medicine, R. H. N. van Schaik, PhD, Department of Clinical Chemistry, Erasmus University Medical Centre, Rotterdam, The Netherlands W. Schaftenaar, DVM, Rotterdam Zoo, PO Box 532, 3000 AM, Rotterdam, The Netherlands Correspondence to Dr Schaftenaar

DURING the past 20 years the number of Asian elephant (Elephas maximus) calves born annually in European zoos has increased, from 100 in 1987 to 141 in 2007 (R. Belterman, studbook keeper of the European Endangered Species Programme [EEP] for Asian elephants, personal communication), and many anecdotal reports of dystocia have been made to the veterinary adviser of the Taxon Advisory Group (TAG) for elephants. Unfortunately, most of these reports have not been well documented. In the experience of one of the authors (W. S.), two Asian elephants at the Rotterdam Zoo that were suffering from dystocia responded to the intravenous administration of calcium by renewed contractions of the uterus. In addition, several captive elephants within the EEP for Asian elephants appear to have suffered from clinical or subclinical hypocalcaemia. Zoo veterinarians have also observed the unexpected efficacy of even low doses of intravenous calcium ions on the contractility of the uterus during a stagnating parturition (T. Hildebrandt, N. Avni-Magen, personal communications). Furthermore, a retrospective evaluation of the serum progesterone profile in a 32-year-old nulliparous Asian elephant showed that the concentration had decreased to baseline levels at the suspected onset of parturition, suggesting that the fetus had been retained in the uterus for 12 months after parturition began (Thitaram and others 2006), although the elephant’s calcium metabolism was not assessed. Little is known about calcium metabolism in elephants although metabolic bone disease and bone fractures have frequently been reported in juvenile elephants (Emanuelson 2006). The objectives of the present study were first, to measure the total calcium concentration in elephant plasma; secondly, to assess the changes in parameters of calcium metabolism during a feeding trial; thirdly, to investigate a possible relationship between calcium metabolism and dystocia; and fourthly, to assess bone mineralisation in captive Asian elephants in vivo. It was hypothesised that the stagnation of the parturition process observed in captive Asian elephants might have been due to subclinical hypocalcaemia. The Veterinary Record, April 12, 2008

MATERIALS AND METHODS Experiment 1 Total calcium concentrations were measured in 43 heparinised plasma samples from Asian elephants that had been stored at –80°C at Rotterdam Zoo. These samples were all obtained from 11 captive Asian elephants, ranging in age from 12 to 36 years (mean [sd] 21·0 [8·6] years), comprising nine cows and two bulls. In addition, total calcium was measured in heparinised plasma samples from three adult captive African elephant (Loxodonta africana) cows, each 18 years old, from another Dutch Zoo. Experiment 2 Calcium metabolism was investigated in a longitudinal study of four Asian elephant cows: one 36-year-old (early) pregnant, lactating female, which had delivered five calves (cow 1), one 16-year-old non-pregnant, lactating female, which had delivered one calf (cow 2), one 20-year-old nonpregnant lactating female, which had delivered two calves (cow 3), and a six-year-old non-pregnant juvenile elephant (cow 4). They were all accustomed to weekly blood sampling. The elephants were fed for periods of three weeks, first with a low calcium ration, then with a high calcium ration, and then again with the low calcium ration. The low calcium ration consisted of grass hay fed ad libitum, and the high calcium ration consisted of alfalfa hay fed ad libitum. Each ration was supplemented with some bread, fruit and lettuces. Blood and urine samples were collected on the first and last days of each period. The grass hay contained 6 per cent ash, 3·6 per cent crude protein, 0·8 per cent crude fat, 35·4 per cent crude fibre, 54·2 per cent other carbohydrates, and 1·8 g/kg (wet weight) calcium, 1·4 g/kg phosphorus, 11·8 g/kg potassium, 0·36 g/kg magnesium and 2·1 g/kg sodium. The alfalfa hay contained 6·7 per cent ash, 11·1 per cent crude protein, 0·9 per cent crude fat, 35·9 per cent crude fibre, 45·4 per cent other carbohydrates, and 7·4 g/kg calcium, 2·3 g/kg phosphorus, 22·2 g/kg potassium, 1·2 g/kg magnesium and 0·73 g/kg sodium. The total calcu-

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Experiment 3 The plasma concentration of ionised calcium was measured in four Asian elephant cows close to parturition. On January 26, 2000, cow 3 delivered a weak calf at term that died 25 minutes after birth without having moved. Stagnation of parturition was suspected on the basis that its plasma progesterone had decreased to below the baseline value (determined from weekly monitoring over previous years) seven days before the calf was born. The same cow delivered a vital calf on May 13, 2003 when it was 17 years of age. A 34-year-old cow (cow 1) delivered a vital calf at term on February 21, 2004. A 17-year-old cow (cow 5) delivered a vital calf on February 13, 1998. A 29-year-old cow (cow 6) delivered a dead calf at term on March 4, 1993 after vaginal vestibulotomy (Schaftenaar 1996). Experiment 4 To evaluate the mineralisation of bone in the elephants from experiment 2, radiographs of tail vertebrae were taken from 3 of them (cows 1, 3 and 4). The animals were trained to present their tails for radiography. In addition, radiographs were taken from the tail of a four-year-old calf (daughter of cow 3), a frozen tail from a 16-year-old bull that had died four years before, and from the frozen tail of a one-day-old male calf. The radiographs were taken with a Gierth HF300 using a mammary film and cassette (exposure data: film focus distance 80 cm, 46 to 50 kV, 60 mA, 0·1 second). Laboratory tests Blood samples were taken from an ear or hindleg vein into heparinised syringes (experiments 1 and 3) or standard 1 ml tubes containing lithium heparin (Greiner Bio-one) (experiment 2). The samples were collected at around 08.00 and placed on ice immediately. The blood was centrifuged for five minutes at 6000 g within 20 minutes after collection and the plasma separated and stored at –80°C until use for the determination of total calcium, phosphorus, magnesium and creatinine on an automated computerised analyser (Menarini). Urine was also collected at around 08.00 and stored at –80°C, until it was thawed for measurements of total calcium, phosphorus, magnesium and creatinine to calculate their respective fractional excretions. The plasma samples were also used for the determination of ionised calcium, intact parathyroid hormone (PTH), 25-OH-D3 and 1,25-OH-D3 (calcitriol). The ionised calcium concentration and corresponding pH of each sample were determined with an ionised calcium analyser (radiometer ABL 725 and ABL 800; Radiometer). Commercial immunoradiometric assay kits designed for the measurement of intact PTH (DSL; Webster), 25-OH-D3 (Diasorin) and 1,25-OH-D3 (Lucron) were used according to the manufacturers’ instructions. Statistical analysis The data were analysed by means of a one-way analysis of variance for repeated measurements. The significance of differences from the basal values was assessed by a paired t test. The strength of the linear association was assessed by calculating the correlation coefficient (r) and testing whether it was significantly different from zero by the two-tailed Pearson product moment correlation test. Significance was

Serum Plasma

1·4

Ionised Ca (mmol/l)

lated composition of the low calcium ration contained 0·2 per cent calcium, 0·15 per cent phosphorus and a calcium/ phosphorus ratio of 1·33. The total calculated composition of the high calcium ration contained 0·85 per cent calcium, 0·30 per cent phosphorus and a calcium/phosphorus ratio of 2·83. Before the feeding trial the elephants were fed a diet with a standard ration, with a total calculated composition containing 0·69 per cent calcium, 0·29 per cent phosphorus and a calcium/phosphorus ratio of 2·38.

1·2 1 0·8 0·6 0·4 7·5

8·0 pH

8·5

defined as P