96 EFFECT OF DIETARY SUPPLEMENTATION WITH TUNA OIL ON THE LIPID COMPOSITION OF BOAR SPERMATOZOA AND CHARACTERISTICS OF SEMEN

In most mammals, docosahexaenoic acid (DHA) is the dominant polyunsaturated fatty acid (PUFA), although, in several species, docosapentaenoic acid is also a major component of the sperm cell membranes. The amount of DHA in spermatozoa is positively correlated with sperm motility. The effect of dietary supplementation with tuna oil (TO) on the lipid and fatty acid composition of boar spermatozoa and the relationship between the changes in composition and boar semen characteristics were studied. Twenty-four boars were distributed in a completely randomized factorial design (2 � 3) with two oil sources (soybean and tuna) and three levels of antioxidant (150, 300, and 450 mg of vitamin E/kg). The diets consisted of a basal diet that was supplemented with 30 g soybean or TO per kg diet. During a period of 10 weeks of feeding the diets, one ejaculate from each boar was collected per week. The sperm was diluted 1:1 with Beltsville thawing solution (BTS) and divided into three portions destined to cooling (5 and 17�C) and freezing. The sperm diluted for cooling at either temperature was stored in plastic bottles for 3 days. After dilution with BTS, the sperm for freezing was centrifuged and rediluted with freezing extender before it was stored in 0.5-mL straws. Thawing was achieved by placing the straws in a water bath (37�C) for 30 s. Motility, vigor, hypoosmotic swelling (host), and morphology were assessed. For determining the fatty acid composition of the spermatozoa and seminal plasma, a sample of 15 mL was taken from each ejaculate shortly after collection and centrifuged for 20 min at 1000g. Sperm motility and vigor were analyzed by placing a sample on a pre-warmed (37�C) microscopic slide, covering with a coverslip, and examining under a light microscope at a magnification of 200�. For host assessment in each case, a volume of 10 µL was mixed with 1 mL hypoosmotic solution (100 mOsm L–1) and incubated for 30 min in a water bath (37�C). After incubation, 50 µL of formol-saline was added to each tube. Sequentially, 20 µL of every sample was smeared on a microscope slide and observed with oil immersion using a phase contrast microscope. A minimum of 200 cells was observed and classified as non-coiled and coiled. Lipid peroxidation was measured using the thiobarbituric acid reaction. Treatment differences for sperm were determined using analysis of variance for means. The proportion of DHA in sperm phospholipid fatty acids increased in semen fatty acid composition after 1 week of feeding TO. The concentrations of the fatty acids were unchanged in the seminal plasma as a result of the diets fed. The proportion of spermatozoa with abnormal morphologies decreased in boars supplemented with TO (P < 0.05). The TO diet showed the lowest level of total antioxidants in the semen (P < 0.05); however, when the diet was supplemented with the higher vitamin E level, an increase in sperm motility and vigor was observed (P < 0.05). Therefore, dietary supplementation with TO alters the lipid composition of the membrane and has a beneficial effect on both cooled and cryopreserved boar spermatozoa by decreasing cold shock during cooling and thus increasing cryosurvival.