Isolation and partial characterization of a parapoxvirus isolated from a skin lesion of a Weddell seal.

A solitary skin lesion was found on the neck of a Weddell seal (Leptonychotes weddellii), chemically immobilized in Queen Maud Land (70 degrees 09'S, 05 degrees 22'E) Antarctica 2001. The lesion was elevated and 3cm in diameter, consisting of partly fresh and partly necrotic tissue, and proliferative papilloma-like structures were seen. Electron microscopy on a biopsy from the lesion revealed typical parapoxvirus particles. Polymerase chain reaction (PCR; B2L gene) generated amplicons of approximately 594 base pairs, comparable to Orf-virus, the prototype parapoxvirus. A comparison of these B2L PCR amplicon DNA sequences with corresponding sequences from other parapoxviruses, showed that the Weddell seal virus resembled isolates from grey seal (Halichoerus grypus) and harbour seal (Phoca vitulina) more than parapoxvirus from red deer (Cervus elaphus), sheep, cattle and Japanese serows (Capricornis crispus). It is thus concluded that the Weddell seal parapoxvirus belong to the tentative seal parapoxvirus species. Since parapox and orthopoxviruses may cause similar clinical diseases, we suggest that the term sealpox should be restricted to the clinical disease, whereas seal parapoxvirus should be used when caused by a parapoxvirus, rather than the general term "sealpox virus". This is the first verified case of parapoxvirus infection in a Weddell seal, and also the first report of any such infections in the Antarctic.

Reference values for serum biochemical parameters in free-ranging harp seals.

BACKGROUND: The harp seal (Phoca groenlandica) is one of the most important predators in the Northeastern Atlantic ecosystem. Establishing biochemical reference intervals is important for evaluating the health status of harp seals kept in captivity and for evaluating the effects of environmental changes on the health of populations in the wild. OBJECTIVE: The purpose of this study was to determine reference values for serum biochemical parameters in wild adult harp seals using readily available current methods. METHODS: Blood samples were obtained from 14 adult female harp seals and 9 suckling pups on the pack ice of the Greenland Sea in early March 1998. Seven seals were humanely killed on the ice by permission of the Norwegian Directory of Fisheries and in conjunction with several other research projects. The seals were sampled within 15 minutes postmortem. Remaining seals were captured alive and sampled via the extradural intravertebral vein. Serum biochemical parameters were measured using a Technicon Axon analyzer and included electrolytes (sodium, chloride, potassium, magnesium, and calcium), substrates (free fatty acids, triglycerides, fructosamine, and glucose), end products (urea and uric acid), and proteins (total protein, globulins, and albumin). Serum protein electrophoresis also was done. Data were tested for normality and reference limits were calculated as mean +/- 1.96 X SD. Results between groups were compared using 2-tailed t-tests. RESULTS: Serum levels of glucose and triglycerides were lower, but serum levels of urea were higher in dead animals than in animals that were captured alive. Serum levels for 7 of 17 parameters were significantly different in pups compared with adults. Separate reference intervals were calculated for adult seals and seal pups. CONCLUSION: Both sampling method and age should be considered when evaluating the results of future analysis of serum parameters in wild and captive harp seals.

Melatonin potentiates the vasoconstrictive effect of noradrenaline in renal artery from newborn hooded seals (Cystophora cristata) and harp seals (Phoca groenlandica).

Isometric tension was recorded by force displacement transducers in ring segments of the inferior branch of the renal artery from newborn hooded seals (Cystophora cristata, n=6), harp seals (Phoca groenlandica, n=3) and domestic pigs (Sus scrofa f. domestica, n=5). Arterial segments were mounted in heated organ baths and exposed to graded concentrations of noradrenaline (NA) and adrenaline (A), alone or together with melatonin. The melatonin concentration in the bath was similar to the plasma concentration normally found in each experimental animal. Melatonin alone did not affect the tension in any of the species, but melatonin potentiated the contraction induced by NA in hooded and harp seal arteries to a maximum of about 25% above the resting, pre-stretch level. The selective melatonin receptor antagonist luzindole reduced this potentiation by 80%. Species-specific concentrations of melatonin did not potentiate the NA-effect in the domestic pig or the A-induced contraction in any of the species. The results indicate that melatonin specifically increases the NA-sensitivity of smooth muscles in renal arteries from newborn seals. It is presumed that similar effects may operate in foetal and maternal seals and may influence their circulation during maternal diving.

Individual variation in field metabolic rate of kittiwakes (Rissa tridactyla) during the chick-rearing period.

Field metabolic rate (FMR), using the doubly labelled water (DLW) method, was measured in free-ranging adult kittiwakes (Rissa tridactyla) early and late in the chick-rearing period at Svalbard, Norway. Individual variation in FMR was analysed by comparing FMR with body mass, sex, nest attendance, chick age, brood size, and basal metabolic rate (BMR). Mean FMR of kittiwakes during the chick-rearing period was 27.0+/-0.9 (SE) W kg(-1), while the individual variation (calculated as coefficient of variation [CV]) in FMR was 24%. Sex, time spent away from the nest, age of the chicks, and brood size contributed significantly to FMR and explained 65% of the variation in FMR. The FMR increased by 32% from early until late in the chick-rearing period. This occurred simultaneously with an increase in the time spent away from the nest. In 15 of 20 pairs, one of the mates had 15% or higher (mean of the 15 pairs, 22%+/-8%) FMR (W kg(-1)) than their partner, even though the mates spent equal amounts of time away from the nest. This indicates an intrapair conflict in FMR. The variation in total FMR of pairs was 40% less than the individual variation, and total FMR of pairs increased with age of the chicks. This indicates that the mates adjust their energy expenditure within a relatively constant FMR determined by the energy needs of the chicks. Individual variation in FMR could not be explained by variation in body mass or BMR. BMR measured late in the chick-rearing period was 26% lower than previous measurements of BMR from the prebreeding and incubation periods. The increase in FMR and simultaneous decrease in BMR caused a 40% increase in metabolic intensity (FMR/BMR) of kittiwakes during the chick-rearing period. It is suggested that the metabolic intensity is not a proper measure of the metabolic load in seabirds.

Physiological effects of seawater intake in adult harp seals during phase I of fasting.

Previous studies have shown that harp seals may drink considerable amounts of seawater. The current study was undertaken to study the physiological responses to bolus administration of seawater. Adult harp seals (Phoca groenlandica) were fasted without access to water for 48 h and then given 1000 or 1500 ml of seawater by a stomach tube. Changes in urine and plasma parameters were thereafter monitored for another 12-20 h. Urine production and urine excretion rate of Na+ and Cl- increased soon after administration and reached a maximum 3-4 h later. Urine osmolality was kept rather stable and high ( approximately 1500 mOsm x kg(-1)) following seawater administration, due to a drop in urine concentration of urea that was proportional to the simultaneous increase in urine concentration of NaCl. Plasma osmolality remained at approximately 340 mOsm x kg(-1), while plasma concentration of urea decreased some 20-25% due to increased excretion of urea when seawater was ingested. Despite bolus administrations of seawater of up to approximately 2% of body mass, homeostasis was maintained and no ill effects observed. It is concluded that the concentrating abilities of the kidneys of harp seals are sufficient to prevent net loss of body water following seawater ingestion. Seawater ingestion may, moreover, increase urinary osmotic space and thus serve as a mechanism to excrete additional urea produced during phase I of fasting.

Experimental evidence of seawater drinking in juvenile hooded (Cystophora cristata) and harp seals (Phoca groenlandica).

This study was undertaken to measure whether young harp seals (Phoca groenlandica) and hooded seals (Cystophora cristata) drink seawater and, if so, to investigate how the excess salt load is handled. Blood and urine samples were collected from hooded seal pups (n = 3) and harp seal pups (n = 3) after 2 weeks of freshwater exposure, at intervals during 3 weeks of seawater exposure and, finally, after 2 weeks of re-exposure to fresh water. Total water turnover, as measured by injection of tritiated water, was 2200 ml x day(-1) and 3300 ml x day(-1) in hooded seals and harp seals, respectively. The extent of mariposia was taken as the difference between total water turnover and influx of water through food (free and metabolic water) and respiratory water exchange. Seawater drinking amounted to 14% and 27% of total water turnover (rH2O) for the hooded seals and harp seals, respectively. Further evidence of mariposia was obtained from an increase in the excretion rate of the urine osmolytes Na+, Cl- and Mg2+, during the period of seawater exposure. It is concluded that water influx due to seawater drinking can not be excluded as a source of error when estimating food consumption of free-ranging harp seals and hooded seals, by use of labeled water techniques.

The effect of body fat on basal metabolic rate in adult harp seals (Phoca groenlandica).

Three adult harp seals (Phoca groenlandica) were fed different daily amounts of capelin (Mallotus villosus), and their body composition determined by use of the tritiated water method at different levels of fattening. Basal metabolic rate (BMR) was measured after 5 days of fasting by indirect calorimetry, and was on average 1.1 W.kg-1 when 45% of body mass (BM) was fat and 2.3 W.kg-1 when body fat was reduced to 13% of BM. This suggests that body fat contributes little to BMR in these animals. It follows, that predictions of BMR on the basis of BM is questionable in seals, in which body fat may change seasonally between 20 and 60% of BM.

Do minke whales (Balaenoptera acutorostrata) digest wax esters?

Mammals are known to utilize wax esters with an efficiency of less than 50%. The purpose of the present study was to examine whether or not minke whales (Balaenoptera acutorostrata), which at times may eat considerable amounts of wax-ester-rich krill, represent an exception to this general pattern. Samples of fresh undigested forestomach, as well as colon, contents were obtained from minke whales (n 5) that had been feeding on krill (Thysanoessa inermis) for some time. The samples were analysed for dry mass, energy density, lipid content and the major lipid classes, including wax esters. The concentrations of wax esters were compared with previous estimates of dry-matter disappearance of the same type of prey using an in vitro technique, to calculate the dry-matter digestibility of wax esters (DMDwax). Wax esters contributed 21% of the energy and 47% of total lipids in the krill diet. The energy density of gut contents decreased by 50% after their passage from forestomach to the end of the colon. The DMDwax was 94.1 (SD 2.8)% (n 5). This high DMDwax and the occurrence of fatty alcohols, one of the products of wax-ester hydrolysis, in faeces show that minke whales are very efficient digesters of wax esters and absorb most of the energy-rich products of this process.

Determination of levels of polychlorinated dibenzo-p-dioxins, dibenzofurans, biphenyls and pesticides in harp seals from the Greenland Sea.

The levels of polychlorinated dibenzo-p-dioxins (PCDD), dibenzofurans (PCDF), biphenyls (PCB) and compounds of the DDT group were determined in individual seal blubber and brain samples from ten male and one female harp seals caught in the Greenland Sea. No data from this region and from harp seals have been reported before. The age of the animals varied between 1 and 18 years. PCDD/PCDF concentrations in the blubber (4-10 pg/g TEQ wet weight (w.w.), Nordic model) were somewhat lower than in ringed seals from the Arctic. Non-ortho substituted PCB (CB 77,126 and 169) showed TEQ levels in the blubber which were about 4-10-times higher. CB 77 dominated in most samples, but in a few cases CB 169 was the most abundant congener. p,p'-DDE (average 760 ng/g w.w.) and di-ortho substituted PCB concentrations (2560 ng/g w.w. expressed as sigma PCB) in the blubber were comparable to those found in ringed seals from the European and Canadian Artic. Levels of PCDD/PCDF in brain correspond to the detection limit (about. 0.1-0.6 pg/g w.w.) for most congeners. With one exception, PCB amounts in brain (w.w.) were one to two orders of magnitude lower. A highly significant correlation was found between age, p,p-DDE and di-ortho PCB as well as between single PCDD and coplanar PCB congeners. No relationship was observed between levels of PCDD/PCDF, age and di-ortho-PCB.

Digestibility of krill (Euphausia superba and Thysanoessa sp.) in minke whales (Balaenoptera acutorostrata) and crabeater seals (Lobodon carcinophagus).

Apparent digestible efficiency (% DE) was studied by use of dietary Mn as an inert marker, in minke whales (Balaenoptera acutorostrata) and crabeater seals (Lobodon carcinophagus) which had been eating krill. Median % DE in minke whales (n5) eating krill of the genus Thysanoessa sp. (energy density (ED) 23.8 kJ/g) was 93 (range 87-93). Median % DE in crabeater seals (n6) eating krill of the species Euphausia superba (ED 20.8 kJ/g) was 84 (range 79-85), which is significantly lower than the % DE of krill in minke whales (P = 0.008). Since the chemical composition in E. superba and in Thysanoessa sp. is similar, it is suggested that the complex multi-stomached system of minke whales, which contains both chitinase (EC 3.2.1.14)-producing as well as several other types of bacteria, is superior to the single-stomached system of crabeater seals with regard to krill digestion. It is worth noting, however, that the % DE of krill in the crabeater seal is still very high.

In vitro digestibility of different prey species of minke whales (Balaenoptera acutorostrata).

Information on diet composition, daily energy expenditure, energy storage and the utilization of energy in the prey are important factors when evaluating the food consumption of minke whales (Balaenoptera acutorostrata) during their summer stay in northern waters. The purpose of the present study was in this context to obtain information on the digestible energy (DE) of different prey selected by minke whales. An in vitro three-stage digestion technique, simulating the different compartments of the digestive system, has been developed. The initial step simulated the anaerobic microbial fermentation of substrate in the forestomach. The next stage included the addition of pepsin (EC 3.4.23.1)-HCl, simulating ventricle enzymic decomposition, and finally, in the third step, fresh extract from duodenal contents was used to simulate enzymic intestinal degradation of the remaining components of the food. The inoculum was normally obtained from animals which had recently eaten the prey to be tested. In such tests we obtained a dry matter disappearance (DMD) and a DE for herring (Clupea harengus) of 80.4 (SD 5.0)% (n 18) and 92.1 (SD 3.7)% (n 16) respectively, and a DMD of krill (Thysanoessa sp.) of 83.4 (SD 4.9)% (n 6). The DMD of krill was reduced to 73.8 (SD 7.3)% (n 8) while the DE was 70.6 (SD 10.4)% (n 7) when inoculum from whales which had recently eaten cod (Gadus morhua) and haddock (Melanogrammus aeglefinus) was used. These results indicate a high digestibility of the most common species of prey in these animals, and also that the whales have little difficulty in changing from one prey species to another.

Water flux and early signs of entrance into phase III of fasting in grey seal pups.

Body water loss and turnover rate were measured in fasting newly weaned grey seal (Halichoerus grypus) pups without access to water for 52 days, by use of bolus injections of deuterated water. Total body water (N) was 41.8 +/- 2.3 (SD)% of total body weight at day 3 and 44.5 +/- 5.9 (SD)% at day 51 of fasting (P greater than 0.05), while the water content of body core decreased from 73 to 61% over the same period. The average water efflux rate in the same period was 362 +/- 17 (SD) ml d-1, the reduction in N over the same period contributing 35% to this end. The biological half-time of deuterated water was 38.2 +/- 3.3 (SD) d. Plasma osmolality increased from a stable level of 329 +/- 11 (SD) mosmol kg-1 to 445 +/- 11 (SD) mosmol kg-1 after day 38 of fasting. Plasma urea was the major contributor to this increase, in spite of a steadily decreasing urine urea concentration (Nordøy et al. 1990) throughout the entire experimental period. It is concluded that grey seal pups endure 52 d of fasting without intake of water with a minor dehydration of body core, while they become hyperosmotic due to increased catabolism of protein after 38 d when sparing of the insulating blubber layer in preparation for life in cold water seems to occur.

Glucose and ketone body turnover in fasting grey seal pups.

Concentration and metabolic replacement (turnover) rate of glucose and ketone bodies were determined at intervals during a 52 day postweaning fast in five grey seal (Halichoerus grypus) pups, using bolus injections of radiotracers. Blood glucose was maintained at a high level throughout the fast, while beta-hydroxybutyrate increased 26 times from day 3 to day 37, whereafter it by and large was maintained. Glucose replacement rate decreased to 56% of the day 9 value at day 37 and dropped further to only 32% of the day 9 level at day 52 in two seals, while in another 2 seals it increased at this late stage. The average ketone body replacement rate ranged between 8.6 and 13.8 mumols min-1 kg-1, but did not change significantly (P greater than 0.05) during the fasting period. These results suggest a reduced gluconeogenesis from protein and increased production of ketone bodies, which may in part replace glucose as energy source during fasting.

Depressed metabolism and low protein catabolism in fasting grey seal pups.

Grey seal pups (Halichoerus grypus) were collected at the time of weaning (mid-October) and fasted for 52 days at thermoneutrality in separate cages. Body weight decreased exponentially, while metabolic rate dropped 45% from an average of 2.95 +/- 0.15 (SEM) W kg-1 at day 2 of fasting to a stable level of 1.62 +/- 0.06 (SEM) W kg-1 from day 10 to day 47 of fasting. Respiratory quotient was low, indicating extensive catabolism of triglycerides, while plasma cortisol was fairly stable at 110 +/- 8 (SEM) nmol l-1 throughout the fasting period. Daily urinary output decreased from 236 +/- 20 (SEM) ml day-1 at day 2 to a stable value of 87 +/- 6 (SEM) ml day-1 between days 8 and 50 of fasting. The urine was analysed for urea, uric acid, creatinine, ammonia, total nitrogen and osmolality. Urea was always the principal excretory end-product, amounting to between 70 and 80% of the total excreted nitrogen. The urine was moderately concentrated (range 770-1300 mosmol kg-1). Total excreted urinary nitrogen decreased by 68% from 3.7 +/- 0.7 (SEM) g day-1 to 1.2 +/- 0.4 (SEM) g day-1 between days 2 and 50. The urinary nitrogen was used to calculate the daily amount of protein being oxidized and its energy content was compared with the measured basal metabolic rate of individual animals. Approximately 6% of the energy expended by grey seal pups during the post-weaning fast is derived from oxidation of protein. It is concluded that a rapid depression of basal metabolic rate and extensive blubber catabolism enable grey seal pups to endure prolonged periods of fasting without any apparent signs of discomfort or stress.

Energy sources in fasting grey seal pups evaluated with computed tomography.

Grey seal pups (Halichoerus grypus) were collected at the time of weaning (early November) and starved for 31 days at thermoneutrality. During starvation body weight decreased linearly, whereas metabolic rate was stable at 1.58 +/- 0.13 (SD) W X kg-1. Metabolic rate as related to body weight was significantly higher (P less than 0.01) than predicted by Kleiber (The Fire of Life, New York: Kreiger, 1975). Respiratory quotient averaged 0.67 +/- 0.03 during the 1st wk but increased to an average of 0.76 +/- 0.04 during the final week of fasting. Body composition was evaluated three times during starvation by use of computed tomography. The area of blubber and skeletal muscle in three transverse (thoracic, abdominal, pelvic) sections of the animals decreased on average 27.6 +/- 4.8 and 18.6 +/- 8.5%, respectively, from days 3 to 31 of fasting. Caloric content of blubber and skeletal muscle was determined by bomb calorimetry, and the caloric content of catabolized tissue was estimated. Based on this information it was calculated that approximately 94% of the energy expended by grey seal pups during the first 4 wk of the postweaning fast is derived from their subcutaneous deposits of fat.