Moose (Alces alces) found dead (FD) and hunter-killed (HK) in 1995 on the north slope of Alaska (Colville River drainage) were evaluated for heavy metal and mineral status. Compared to previous reports for moose and domestic cattle, and data presented here from Alaska moose outside the Colville River area, levels of copper (Cu) were determined to be low in hoof, hair, liver, kidney, rumen contents, and muscle for these north slope moose. Iron (Fe) was low in muscle as well. These findings, in conjunction with evidence of poor calf survival and adult mortality prompted investigation of a mineral deficiency in moose (serum, blood, and hair) captured in the spring of 1996 and 1997. Captured males had higher Ca, Zn and Cu levels in hair than captured females. Female moose hair samples were determined to be low (deficient) in Cu, Ca, Fe, and Se with mean levels (ppm) of 2.77, 599.7, 37.4, and 0.30, respectively. Serum Cu level was low, and to a lesser degree Zn was deficient as well. Whole blood (1997 only) was marginally deficient in Se and all animals were deficient in Cu. Based on whole blood, sera and hair, Cu levels were considered low for moose captured in spring 1996 and 1997 in the Colville River area as compared to published data and other populations evaluated in this study. Low levels of ceruloplasmin activity support this Cu deficiency theory. Evidence indicates that these moose are deficient in Cu and other minerals; however, the remote location precluded sufficient examination of animals to associate this apparent deficiency with direct effects or lesions. Renal levels of Cd increased with age at expected levels.
Species: Moose (Alces alces) Tissues/matrices sampled: Liver, kidney, muscle, hair, hoof, and rumen contents from hunter-killed or found-dead moose; serum, whole blood and hair from captured moose. Tissues/rumen contents analyzed for metals and trace minerals, including: Ag, Al, As, B, Ba, Be, Bi, Ca, Cd, Co, Cr, Cu, Fe, K, Li, Mg, Mn, Mo, Na, Ni, P, Pb, S, Sb, Se, Si, Sn, Tl, V and Zn. Other parameters evaluated in captured animals included: serum ceruloplamin, fecal parasites, serology, pregnancy-specific protein B.
Aerial surveys: Aerial surveys were conducted annually in spring (prior to capture operations in 1996 and 1997) and fall (composition count) to determine changes in the total population, calf production (calves per 100 cows), and other population dynamics within a prescribed area (count area) that includes regions of the Colville, Chandler, and Anaktuvuk Rivers on the north slope of Alaska (near Umiat, Alaska) for Game Management Unit 26 (A). This prescribed area is bordered by longitudes 154° W and 151° W, and latitudes 68.5° N and 69.5° N. Sampling of found dead (FD) and hunter killed (HK) moose: Moose carcasses (15 of a reported 30) along the Chandler River and associated drainages were examined and sampled for cause of death on August 1 and 2, 1995. The carcasses were in very poor post mortem condition and of limited diagnostic value. However, fat reserves in viscera and bone marrow indicated depleted energy reserves (Kistner et al., 1980). Predation and other signs of trauma were not obvious. Three intact moose (not scavenged) were sampled (liver, kidney, muscle, hair, lower leg, rumen contents, and feces) and remaining carcasses were visually examined grossly. Bull moose harvested in the Colville and Chandler River regions in the fall of 1995 were sampled by cooperating hunters. Hunter-collected tissues were frozen, or preserved in neutral buffered 10% formalin. Frozen samples include kidney, liver, heart, muscle, rumen contents, feces, skin and hair, and jaw. Tissues placed in formalin include liver, kidney, lung, tongue, testicle, rumen or other stomach sections, spleen, intestine, and muscle. Hunters scored the amount of subcutaneous, perirenal, and pericardial fat to comment on general condition. All hunter-killed animals were reported in good condition with fat present in all areas specified. Liver, kidney, and muscle samples were sent to Mississippi State University, College of Veterinary Medicine, Diagnostic Toxicology Laboratory for trace mineral analyses described by O'Hara et al. (1995). Samples were analyzed by atomic absorption (AA) spectrophotometry (Perkin Elmer 5000; Norwalk, Connecticut USA) with a graphite furnace and continuous background corrections. Results are reported in ppm wet weight (ww). The formalin-fixed specimens were sent to a certified pathologist (Dr. Kathy Burek, Alaska Veterinary Pathology Services, Eagle River, Alaska USA). Sampling of immobilized moose: In April 1996 and 1997, moose from the Colville River area were captured, examined, collared with VHF transmitters, evaluated for pregnancy, serologic evidence of disease, mineral status, fecal indicators of parasites, and standard blood indices. Moose were captured using a helicopter and chemical immobilization as described in O'Hara et al. (1998). Blood samples were taken by syringe from the jugular vein for serum and blood collection. Bundles of hair were removed by pulling full-length strands straight up from the dorsal midline at the shoulder. Elemental analysis of hair, serum and whole blood Samples of hair, serum and whole blood were submitted frozen to the Diagnostic Laboratory, College of Veterinary Medicine, University of Illinois for elemental analyses. Aliquots of 1.0 +/- 0.1 g were digested with 5.0 ml of concentrated nitric acid, evaporated to 1.0 ml and diluted with 5.0 ml of concentrated HCl for 30 minutes. Hydrogen peroxide (30%) was added in 1 ml increments until effervescence ceased or until a volume of 10.0 ml was reached. The sample was quesced to a volume of 25 ml with deionized water. Tissue samples were processed in a similar fashion to hair except 2.0 +/- 0.1 g was digested, without addition of HCl or peroxide. Serum samples were prepared by adding 0.5 ml to 9.5 ml of 3 M HCl and left at room temperature for 30 minutes and the supernatant analyzed. Digests and supernatants were analyzed with the Thermo Jarrell Ash IRIS AP Inductively Coupled Plasma (ICP) Spectrometer. Quality control samples include SPEX Chemical Reference standards (SPEX Chemical, Metuchen, New Jersey, USA), Spex Chemical Spike Solutions, Spex Chemical QC Solutions, a Dade Monitrol Chemistry Control (VWR Scientific Inc., San Franciso, California USA) and an in-house bovine serum control. Spiked samples were included with each set of digested or diluted samples. The working standard mixtures were analyzed at the beginning and end of each run, and the calibration curve determined from the first set of standards as per manufacturers instructions (Thermo Jarrell Ash). The QC samples following the standards included Spike-1 samples, monitrol samples, in-house serum control, diluted QC-21 samples (10 and 1 ppm), and QC-11 samples. For each ten samples a QC-21 and/or QC 11 sample were run. Values for QC samples should fall within 10% of the known or historic value, if not, the samples are reanalyzed. Tissue levels for moose from Fairbanks, Nome and Nowitna, Alaska (n = 14) were determined and used for comparison to the Colville River population and were analyzed by the Environmental Trace Substances Research Center (ETSR), University of Missouri (Columbia, Missouri USA). In a 100 ml beaker, 15 ml of HNO3 and 2.5 ml of concentrated HCl were added to 0.5 g of sample then gradually heated. Once fumes were no longer evident the sample was cooled and 2.0 ml of HCl was added and heated, then brought to 50 ml. As and Se were measured using a Varian VGA-76 hydride generation accessory on a Perkin-Elmer Model 603 AA using appropriate standards and the manufacturer’s instructions. Other element levels were determined by ICP analyses using a Jarell-Ash Model 1100 Mark III. The instrument is standardized with a series of 7 standards containing 36 elements. Quality control samples were used and standards were measured every 10-15 samples and if measures drifted more than 5% the instrument was recalibrated. Ceruloplasmin (CP) analysis: Ceruloplasmin activity was assayed by oxidation of o-dianisidine after the method of Schosinsky et al. (1974). Blood serum (0.05ml) oxidation rates were determined in duplicate as the difference in absorbance between 15min and 25min. Activity was expressed as the rate of oxidation (umol.min-1.L-1 or IU.L-1). This unit of activity is equivalent to 0.2836mg.dL-1 of ceruloplasmin in human serum (Schosinsky et al. 1974). Pregnancy testing Pregnancy-specific protein B (PSPB) was determined by Biotracking Inc. (Moscow, Idaho USA) for these moose as described in O'Hara et al. (1998) with a competitive radioimmunoassay that measures serum PSPB by determining the amount of 125I-PSPB bound. If less than 93% of the 125I-PSPB binds (displaced by unlabeled PSPB in the serum), then unlabeled PSPB is present and consequently the diagnosis of pregnancy (Sasser et al., 1986). This technique has been used in other moose studies (Haigh et al., 1993; Stephenson et al., 1995). Age determination Age determination was conducted by a contract laboratory (Matson's Laboratory, Milltown, Montana USA) using cementum analysis of the central (primary) incisor. Pairs of matched teeth were submitted for comparison and quality control. When estimates differed between matched teeth, the estimate with the highest score of reliability was used. Statistics and calculations Summary statistics, simple regression analyses, and student t-tests were conducted using Microsoft Excel (7.0) for Windows. Mineral contents of serum and hair in males, non-pregnant and pregnant females were compared by ANOVA (SYSTAT 8.0, SPSS, Chicago, Illinois USA) following transformation to square roots to meet assumptions of normality and homogeneity of variances (Zar, 1974; Wilkinson and Coward 1998). Pairwise comparisons of transformed means were performed with Bonferroni's adjustments. Interactions between untransformed mineral contents and CP activity were tested by Pearson correlations followed by Bonferroni adjustments for pairwise comparisons. Factor analysis of the correlation matrix was used to project two principal components from the rotated matrix (SYSTAT 8.0). Means are calculated in two forms for untransformed data: the arithmetic mean for those matrices with all samples with detectable levels (> MDL) for an element, and half the MDL level is used for samples that are < MDL to calculate the mean. “Normal” values were converted to ppm wet weight (ww) using the following formulas. Whole blood “normal” values for moose are from Bubenik (1997) and were converted: P 4.9 (+/-1.4) mg/dl or mg/100ml or 10 mg/kg (assume fluid density of 1); 49 (+/- 14) ppm; Ca 10.4 (+/- 1.0) mg/dl, 104 (+/-10) ppm; Fe 150.2 (+/- 40.3) mg/dl, 1502 (+/- 403) ppm; Na 137.8 (+/- 6.2) meq/l = mmol/L / 0.435 = 316.8 (+/- 14.3) mg/100ml or 3168 +/- 143 ppm; K 5.3 +/-1.4 meq/l = mmol/L / 0.2258 = 104.0 +/- 6.2 mg/100ml or 1040 +/- 62 ppm.
See methods above.