Hypoosmotic Swelling (HOS) as a screening assay for testing in vitro fertility of bovine spermatozoa

Semen from 5 Piedmontese bulls was subjected to the hypoosmotic swelling (HOS) test in order to determine if the results could be correlated to the fertilizing capacity in vitro. Semen was routinely prepared for in vitro fertilization (IVF), with aliquots being sampled after thawing, after separation on a Percoll gradient and after capacitation in a medium containing heparin. The aliquots were added to a fructose-sodium citrate hypoosmotic solution (100 mOsm) and incubated at 37°C for 5 min. At least 200 spermatozoa were observed at x 400 and classified according to the presence or the absence of a swollen tail. After capacitation, spermatozoa were used to fertilize in vitro-matured bovine oocytes (1.5 x106 cells/mL); IVF was performed in Fert-TALP medium supplemented with 6 mg/mL BSA and 10 L/mL heparin in a humidified atmosphere with 5% CO2 and 5% oxygen. Presumptive zygotes were cultured in SOF medium supplemented with 8 mg/mL BSA and amino acids. There were no significant differences in the in vitro fertility of the bulls, but a significant difference was found between bulls in the response to the HOS test. The 3 assays were significantly correlated, while no significant correlation was observed between the percentage of swollen spermatozoa and in vitro fertility. The HOS test does not appear to be sufficiently sensitive to discriminate between semen samples of intermediate fertility like those used in this preliminary research.