Introduction

Although vitamin E occurs in many common foods and feeds, numerous deficiencies have been reported in captive wildlife fed diets based on frozen marine fish". Several of these reports have involved marine mammals(2-6) and birds(7). Vitamin E deficiency has been reproduced experimentally in harp seals (Phoca groenlandica) maintained on a diet of eviscerated herring with no vitamin E supplementation(8).

Vitamin E is a biological antioxidant which is important for cell membrane integrity, enzyme and heme synthesis, and steroidogenesis. Its role in numerous metabolic processes is reflected in the many pathologic manifestations that develop when a deficiency occurs including muscular degeneration, steatitis, liver necrosis, vasculopathy, anemia, and impaired reproductive capacity(9).

Live or fresh fish are in general considered to be an excellent source of vitamin E(1). However, the combination of a relatively high concentration of polyunsaturated fatty acids in marine fish such as herring and mackerel, the frozen storage of the fish for several months before being fed, and the practice of evisceration before feeding contribute to the probability of producing a deficiency. These factors are important because, (a) the antioxidant role of vitamin E increases the requirement as unsaturated fats or dietary fat rancidity increases; (b) vitamin E in fish is oxidized and inactivated even when stored at very cold temperatures (50% loss after 6 months of storage at 20°C) and (c) the viscera of fish are important storage sites of the vitamin.^1,8,10,11

In domestic animals a relationship has been shown between vitamin E and the trace mineral selenium. Both play a role in protecting biological membranes and several of the deficiency diseases can be ameliorated by either selenium or vitamin E supplementation. However, selenium deficiency was not believed to be involved in the disease process of the subject of this report as it is found in high concentrations in fish diets(12).

Case History

A one-year-old male captive born California sea lion (Zalophus californianus) housed in fresh water at the Oklahoma City Zoo in an outdoor exhibit was initially examined because of an acute onset of lethargy, anorexia, and a reluctance to move. The sea lion was unable to support his weight with the foreflippers and experienced difficulty in breathing during attempts at restraint. He died ten hours after the onset of clinical signs. Just prior to death the referring veterinarian reported observing dark colored urine. The animal was sent to the Oklahoma Animal Disease Diagnostic Laboratory, Stillwater, Oklahoma for necropsy.

Postmortem examination showed the sea lion to be in excellent flesh with a considerable amount of fat within internal body deposits. Throughout the body, there were varying degrees of pale muscles surrounded by deep, red muscles. These changes were most prominent on the diaphragm, pectoral muscles and muscles of the thighs. These were characterized by pale yellowish-gray streaks which often were 3 to 8 cm long and appeared to affect whole bundles of muscle. These were bordered by more normal appearing deep red muscles. There were a few deep red 3 to 4 mm diameter foci noted in the myocardium.

Histopathologic changes in all affected skeletal muscle were essentially alike. There were areas in which large groups of muscles were degenerative with a pink coagulum of cytoplasm, some fractured myofibrils along with some small, dark purple foci believed to represent early mineralization of mitochondria. In some places there was complete loss of the sarcoplasma and collapse of the sarcolemma. Within some muscle segments, particularly pectorals and gluteal muscles, there were areas of regenerating myocytes characterized by a more deep blue cytoplasm and proliferation of extremely large oval nuclei with a very prominent nucleolus. In these cells, some cross striations were still present.

In affected sections of myocardium there were patchy areas with nonstaining edema causing separation of muscle bundles along with some hypertrophy of interstitial cells. In a few places a degenerative myocyte was noted running through these areas with a loss of cross striations and having a granular cytoplasm. Within affected sections many of the myocytes had proliferation of nuclei with three or four cells in a row.

Sections of the kidneys were unremarkable with the exception of some tubules within the medulla that contained a brown-to-orange granular pigment believed to be myoglobin.

The histopathologic diagnosis was that of a severe patchy degenerative myopathy and cardiomyopathy, lesions typical of those caused by a vitamin E deficiency.

Discussion

In a captive situation the challenge to be met involves providing a diet which the animal will accept, and one which satisfies the nutrient requirements of that particular species. Although the ideal situation would involve the feeding of fresh fish this is seldom feasible and resorting to frozen sea food is usually the alternative. However, freezing and storage may destroy certain essential nutrients. In general the requirements for the fat-soluble vitamin E have been related to the amount of polyunsaturated fatty acids in the diet(8).

Marine fish are naturally high in unsaturated fatty acids. As an example, a common food for captive marine mammals is herring, Clupea harengus, in which the fatty acid component is about 80% unsaturated. These highly unsaturated lipids are readily susceptible to attack by molecular oxygen, and transform into toxic peroxides and polymerized products Thus, tissues of an animal on an all fish diet are susceptible to in vivo peroxidation if sufficient vitamin E is not available.

Fish tissues are labile and deteriorate rapidly. It has been shown that storage of whole fish may be expected to decrease the amount of vitamin E(14). Freezer temperatures must be kept below -18°C to prevent cell rupture, loss of fluid, rancidity, and inactivation of fat soluble vitamins, however since most fish are frozen before shipping, quality control is difficult(7,9).

The practice of feeding eviscerated fish, in which the gonadal tissues in particular are also removed, further decreases the proportion of vitamin E. Viscera including the gonads may double the concentration of vitamin E per unit(15) weight compared with fish muscle.

Clinical signs associated with a deficiency of vitamin E can be varied and may include:

1.  Lethargy and anorexia

2.  Dyspnea

3.  Muscular weakness

4.  Abnormal molting patterns

5.  Myoglobinuria

6.  Sudden death

A definitive diagnosis of this syndrome is difficult to make owing to the extensive physiologic effects of vitamin E (i.e. tissue antioxidant properties, maintenance of cell membrane integrity, enzyme and heme synthesis, and steroidogenesis)(6). Therefore, several factors (i.e. age, diet, behavior, molting patterns, and clinical and laboratory findings) must be considered when attempting to make a diagnosis of vitamin E deficiency in pinnipeds(6).

In the absence of biochemical or morphologic lesions specific for vitamin E deficiency, a diagnosis in animals showing suspicious clinical signs should be based on cumulative findings including:

1.  A history of a non-supplemented diet of fatty fish, especially(9) mackerel and herring

2.  Hyponatremia(16)

3.  Plasma vitamin E concentrations below 7 to 10 µg/ml

4.  Gross and histological lesions of myopathy and fat necrosis from biopsy specimens of muscle and blubber

5.  Increased serum muscle-enzyme concentrations.

6.  Renal insufficiency due to myoglobinuria(6)

The following have been suggested as a means of preventing vitamin deficiency in captive marine mammals:

1.  Feed only high quality fish that have been quick-frozen and glazed (US Grade A)(6) . To protect the vitamin E content in frozen fish, the fish should be stored at -30°C for maximal periods of 4 months for mackerel 6 to 7 months for herring, and up to 9 months for smelt and capelin(16).

2.  Fish should be stored at the highest humidity possible to prevent dehydration, thawed rapidly to break the ice cake and immediately fed to reduce the loss of nutrients(6).

3.  Feed only whole, rather than eviscerated fish or fillets(15).

4.  Eliminate stressful conditions(6).

5.  Vitamin E supplementation should be established on the basis of the fat content of the fish, the duration of storage and methods of(17) preparation and feeding It has been recommended by several investigators to supplement with 100 Iu of vitamin E per kilogram of fish daily(6,8).

6.  Increase vitamin E supplementation to lactating females to increase vitamin E transfer to young, or parenterally supplement the young pinnipeds with vitamin E at strategic times(6).

The animal of this report showed many of the characteristic signs of vitamin E deficiency: anorexia, lethargy, muscular weakness, dyspnea, and myoglobinuria. The diet had consisted of herring and smelt with no fat-soluble vitamin supplementation. Post-mortem studies including histopathology revealed a severe degenerative myopathy and cardiomyopathy which represented lesions typical of a vitamin E deficiency.

References

1.  Robbins, C.T. Wildlife Feeding and Nutrition. Academic Press, Orlando (1983).

2.  Hubbard, R.C. Husbandry and Laboratory Care of Pinnipeds. In The Behavior and Physiology of Pinnipeds. Appleton-Century-Crofts, New York (1968).

3.  Keyes, M.C. The Nutrition of Pinnipeds. In The Behavior and Physiology of Pinnipeds. Appleton-Century-Crofts, New York (1968).

4.  Wallach, J.D. The Management and Medical Care of Pinnipeds. J. Zoo Ani. Med. 3:45-72 (1972).

5.  Wilson, T.M. Diffuse Muscular Degeneration in Captive Harbor Seals. J. Am. Vet. Med. Assoc. 161: 608-610 (1972).

6.  Citino, S.B., R.J. Montali, M. Bush, et.al. Nutritional Myopathy in Captive California Sea Lion J. Am. Vet. Med. Assoc. 187: 1232-1233 (1985).

7.  Campbell, G. and R.J. Montali . Myodegeneration in Captive Brown Pelicans Attributed to Vitamin E Deficiency. J. Zoo Ani. Med. 11: 35-40 (1980).

8.  Engelhardt, F.R. and J.R. Geraci. Effects of Experimental Vitamin E Deprivation in the Harp Seal. Can. J. Zool. 56: 2186-2193 (1978).

9.  Geraci, J.R. and D.J. St. Aubin. Nutritional Disorders of Captive Fish-Eating Animals: In The Comparative Pathology Animals. Smithsonian Institution Press, Washington (1979).

10. Achman, R.G. and M.G. Cormier. α - Tocopherol in Some Atlantic Fish and Shellfish with Particular Reference to Live-holding Without Food. J. Fish. Res. Bd. Can. 24: 357-373 (1967).

11. Helgebostad, A. and F. Ender. Vitamin E and Its function in the Health and Disease of Fur-bearing Animals. Acta Agric. Scandinavica Suppl . 19: 79-83 (1973).

12. Lunde, G. Analysis of Arsenic and Selenium in Marine Raw Materials. J. Sci. Food Agric. 21: 242-247 (1970).

13. Ackman, R.G. and C.A. Eaton. Some Commercial Atlantic Herring Oils: Fatty Acid Composition. J. Fish Res, Board Can. 23: 991-1006 (1966).

14. Engelhardt, F.R., J.R. Geraci and B.L. Walker. Tocopherol Composition of Frozen Atlantic Herring (Clupea harengus harengus) Tissues and Oils. J. Fish Res. Board Can. 32: 807-809 (1975a).

15. Geraci, J.R. Dietary Disorders in Marine Mammals: Synthesis and New Findings.

16. J. A. Vet. Assoc 179: 1183-1191 (1981).

17. Ackman, R.G. The Influence of Lipids on Fish Quality. J. Food Technol 2: 169-181 (1967).