A comparative study of the sperm nuclear morphometry in cattle, goat, sheep, and pigs using a new computer-assisted method (CASMA-F)

This study was designed to compare the sperm nuclear morphometry of four species of domestic artiodactyls (cattle, sheep, goats, and pigs), using the newly developed automatic computer-assisted sperm morphometry analysis-F. The study was divided into two experiments. In the first experiment, samples from 20 males from each species were collected, diluted, and divided into four sample aliquots. The first was labeled directly with Hoechst 33342, and the others were processed as smears. Between smears, one group was directly labeled with Hoechst after air drying, and the other was fixed either with glutaraldehyde (GLUT), or with methanol, and afterward labeled with Hoechst. Digital images of the fluorescence-labeled sperm were recorded with a digital camera, and at least 200 sperm cells per sample were processed using the Image J analysis open software. Air-drying significantly reduced nuclear sperm dimensions in ruminant species, whereas no effect was observed in pigs. For most of the primary morphometric parameters, the relationship between the four species for the sperm nuclear dimensions can be described as follows: bull > ram ≥ boar > goat. However, ram sperm nuclei had greater width than those of the other species studied. For the secondary morphometric parameters, ram sperm nuclei were clearly less elliptical and elongated and showed greater regularity than in the other studied species. In the second experiment, ejaculates from 10 males per species were used to compare the sperm head morphometric results obtained with the computer-assisted sperm morphometry analysis-F system (using the GLUT treatment as reference) to a more conventional CASMA method (semen smears stained with Harris's hematoxylin and processed with the Integrated Sperm Analysis System [ISAS] commercial software [Proiser R&D SL, Buñol, Spain]). Spermatozoa displayed a bigger size when processed with Harris's hematoxylin than with the GLUT method in all primary sperm head morphometric parameters for the four species studied. A significant correlation was observed between the two methods used in this experiment for all morphometric size parameters in the four species studied. It was concluded that drying and fixation has little effect on sperm nuclear morphometry, with differences between species, and that there are significant variations in size of the sperm nucleus and in the hydrodynamic properties between the four species studied.